Design, Synthesis and Characterization Of

Home , Dicarbonate, Diphosphorus tetraiodide, Heptacene, Nile red, Prodan (dye), Radioluminescence

DESIGN, SYNTHESIS AND CHARACTERIZATION OF

FLUORESCENT DYES AND

LIQUID CRYSTAL SEMICONDUCTORS

A dissertation submitted to Kent State University

in partial fulfillment of the requirements for the

degree of Doctor of Philosophy

by Alexander N. Semyonov

August 2006

Dissertation written by

Alexander N. Semyonov

B.S., Saratov State University, 1997

Ph.D., Kent State University, 2006

Approved by

Robert J. Twieg Dissertation Committee Chair

Carmen C. Almasan Dissertation Committee Member

Brett D. Ellman Dissertation Committee Member

Peter Palffy-Muhoray Dissertation Committee Member

John L. West Dissertation Committee Member

Accepted by

Oleg D. Lavrentovich Chair,

Chemical Physics Interdisciplinary Program

John R. D. Stalvey Dean, College of Arts and Sciences

ii TABLE OF CONTENTS

LIST OF ABBREVIATIONS...... vii

LIST OF FIGURES ...... xii

LIST OF SCHEMES...... xv

LIST OF TABLES...... xxii

CHAPTER I. DPP FOR SINGLE-MOLECULE SPECTROSCOPY...... 1

1.1. Single-Molecule Spectroscopy and its Applications ...... 1

1.2. Dye Requirements for Single-Molecule Spectroscopy...... 7

1.3. Review of known DPP chemistry...... 17

1.3.2. Chemical Properties...... 32

1.3.3. Physical Properties...... 44

1.4. Newly prepared DPP dyes ...... 60

1.4.1. Alkylation of DPPs ...... 69

1.4.2. Action of bases on DPP ...... 72

1.4.3. Halogenation of DPPs...... 77

1.4.4. Substitution of halogen by amine in DPPs ...... 80

1.4.5. Extension of the conjugation chain of DPP ...... 83

1.4.6. DPPs with hydrophilic solubilizing groups ...... 87

1.4.7. Hydroxy-functionalized DPPs ...... 88

iii 1.4.8. DPPs with a cysteine-reactive maleimide moiety...... 99

1.4.9. N-Arylated DPPs ...... 102

1.4.10. Physical Properties of Newly Prepared DPPs...... 105

1.4.11. Conclusion ...... 115

CHAPTER II. CYSTEINE-SPECIFIC FLUORESCENT TAGS: NILE RED –

MALEIMIDE AND DCDHF – MALEIMIDE...... 116

2.1. Introduction to molecular probes and tags...... 116

2.2. Design of the Probes ...... 122

2.3. Synthesis ...... 130

2.4. Results and Discussion ...... 140

2.5. Conclusion ...... 149

CHAPTER III. ORGANIC LIQUID CRYSTAL SEMICONDUCTORS...... 150

3.1. Introduction...... 150

3.2. Polyacenes...... 161

3.2.1. Anthracenes...... 162

3.2.1.1. 2,3,6,7-Tetraalkoxyanthracenes...... 163

3.2.1.2. 2,3,6,7-tetraalkoxy-9,10-dialkyltetracenes ...... 169

3.2.1.3. 1,2,3,4,5,6,7,8-octaalkylanthracenes...... 171

3.2.2. Tetracenes ...... 172

3.2.3. Pentacenes...... 175

3.3. Iodoarenes...... 195

3.3.1. Why Iodine?...... 195

iv 3.3.2. Direct iodination ...... 198

3.3.3. Iodo-de-diazoniation...... 200

3.3.4. Halogen exchange...... 202

3.4. Liquid Crystal Semiconductors...... 204

3.4.1. HAT Discotic Liquid Crystals ...... 204

3.4.2. Nitrated HAT5 Discotic Liquid Crystals ...... 209

3.4.3. Conclusions...... 211

CHAPTER IV. EXPERIMENTAL PART...... 212

4.1. General Instrumentation and Techniques ...... 212

Measurement of Fluorescence Quantum Yield...... 212

Measurement of Fluorophore’s Photostability...... 215

Gas Chromatogrphy – Mass Spectroscopy (GC-MS)...... 217

HPLC-MS ...... 219

Nuclear Magnetic Resonance (NMR)...... 220

Thermal Analysis: DSC and TGA ...... 221

M.Braun SPS...... 222

High Pressure Reactors...... 222

UV-Vis...... 223

IR...... 224

4.2. Synthetic Procedures...... 225

CONCLUSIONS ...... 355

REFERENCES...... 357

v APPENDIX A...... 393

APPENDIX B ...... 403

APPENDIX C ...... 417

vi LIST OF ABBREVIATIONS

I A Absorbance (value), lg 0 I

Ab absorbance (subscript)

Ac acetyl, CH3CO–

Ac2O acetic anhydride

AcOH acetic acid

AFM atomic force microscopy

APCI atmospheric pressure chemical ionization

BHT 4-t-butylhydroxytoluene

Am amyl, pentyl

t-AmOH tert-amyl alcohol, 2-methyl-2-butanol

ca. circa, approximately

cf. confer, compare with

Chx cyclohexyl

Δ delta, difference between two values

Δλ Stokes’ shift

Δx refluxing solvent or at solvent’s refluxing temperature

CAS RN Chemical Abstracts Service Registry Number

CDI 1,1′-carbonyldiimidazole vii Cys cysteine

d doublet

DBU 1,8-diazabicyclo[5.4.0]undec-7-ene

DCC 1,3-dicyclohexylcarbodiimide

DEAD diethyl azodicarboxylate

DIB 1,4-diiodobenzene

DMAc dimethylacetamide

DMAE or deanol — N,N-dimethylaminoethanol

DMAP 4-(dimethylamino)pyridine

DMF dimethylformamide

DPP 2H,5H-dihydropyrrolo[3,4-c]pyrrole-1,4-dione

DSC differential scanning calorimetry

Em Emission

ESI electro-spray ionization

ε molar extinction coefficient, l·mol–1·cm–1

f femto, 10–15

FRET fluorescence resonance energy transfer

FCS fluorescence correlation spectroscopy

Fu furyl

HAT-5 hexapentyloxytriphenylene

HMDS 1,1,1,3,3,3-Hexamethyldisilazane

HMPA hexamethylphosphoramide

viii ID internal diameter

i, iso prefix to denote isomeric branching at the terminus of a substituent

kF total rate of fluorescent decay

kisc intersystem crossing rate

knr total rate of non-radiative decay

kT decay rate from T1 to S0

λ wavelength

L, l liter

l, ℓ length

lg decimal logarithm

ln natural logarithm

m meter

MI maleimide residue, 1H-pyrrole-2,5-dione-1-yl

mm Hg pressure unit of millimeters of mercury

mol mole, NA number of species

μ micro, 10–6

23 NA Avogadro’s number, 6.0245·10 of structural units

25 nD refractive index at 25 °C for center of sodium D-line doublet (589 nm)

n nano, 10–9

p pico, 10–12

P pressure, followed by value and units: P = 2 mm Hg

Ph phenyl

ix PPA polyphosphoric acid, 2H3PO4•P2O5 ppb parts per billion ppm parts per million

PPTS pyridinium 4-(para)-tolenesulfonate

PTFE poly(tetrafluoroethylene), Teflon®

RET resonance energy transfer q quartet r.t. room temperature s singlet sec- secondary sec second

σp absorption cross-section

S0 ground singlet state

S1 first excited singlet state

SMS single-molecule spectroscopy

STM scanning tunneling microscopy

Suc succinyl acyl residue, (CH2CO)2 t-, tert- tertiary t triplet

TGA thermal gravimetric analysis

THF tetrahydrofuran

THP tetrahydropyran-2-yl

x TMEDA N,N,N′,N′-Tetramethylethylenediamine

TriMEDA N,N,N′-Trimethylethylenediamine

Ts tosyl, 4-toluylsulfonate (4-methylbenzenesulfonate)

TsCl tosyl chloride, 4-toluylsulfonyl chloride

UV-Vis ultra-violet and visible (spectra, properties or data) y yocto, 10–24

τ1 fluorescence lifetime

T1 first excited triplet state

δ chemical shift

ν wavenumber

ΦF fluorescence quantum yield

φ b photobleaching quantum efficiency

[####-##-#] CAS Registry Number

xi LIST OF FIGURES

Figure 1.1. Jabłoński Diagram...... 3

Figure 1.2. Irreversible photobleaching (at 5.6 sec) of single Cy5 molecule, immobilized

on a glass slide...... 10

Figure 1.3. X-ray molecular structure of DPP and DPP-Me...... 45

Figure 1.4. Crystal structure of (a) DPP – triclinic and (b) DPP-Me – orthorombic...... 46

Figure 1.5. Overlap of the two molecules along the stacking axis: (a) DPP and (b) DPP-

Me...... 46

Figure 1.6. DSC of (a) crude and (b) conditioned DPP...... 49

Figure 1.7. TGA traces of crude DPP (red) and Br-DPP (blue)...... 50

Figure 1.8. Visible spectrum of DPP as NMP solution (absorption) vs. solid (reflection).51

Figure 1.9. Absorption (S), fluorescence in CHCl3 solution (F), and solid state

fluorescence (SF) of DPP and 4-t-Bu-DPP...... 55

Figure 1.10. 1H and 13C NMR spectra of 3,6-diphenyl-2,5-diallylpyrrolo[3.4-c]pyrrole-

1,4-dione...... 58

Figure 1.11. 1H and 13C Assigned chemical shifts for 3,6-diphenyl-2,5-diallylpyrrolo[3.4-

c]pyrrole-1,4-dione...... 58

Figure 1.12. Effect of fast vs. slow work-up on the purity of crude DPP...... 68

xii Figure 1.13. Single molecules of DPP-Me 23 imaged in a PMMA film, excited at

wavelength of 488 nm with an intensity of 0.85 kW·cm–2...... 106

Figure 1.14. Photostability of compounds 32, 33, 64, and 65...... 107

Figure 1.15. Photostability of compounds 32, 91, 92, and 93...... 108

Figure 2.1. Generalized structure of a bioconjugate fluorescent molecular probe...... 117

Figure 2.2. Common dye classes, used in molecular probes...... 122

Figure 2.3. Nile Red absorption (Ab) and emission (Em) spectra in dioxane...... 125

Figure 2.4. DCDHF-6, imaged at single molecule level in PMMA film...... 127

Figure 2.5. Fluorescence intensity of 123 decreases with solvent polarity increase...... 142

Figure 2.6. Ribbon model of GroEL homotetradecamer...... 144

Figure 2.7. Ribbon model of GroEL protein...... 145

Figure 2.8. Fluorescence change after addition of (1) MDH, (2) GroES, (3) nucleotide.144

Figure 2.9. Effect of various addition orders on the fluorescence intensity...... 145

Figure 2.10. Proposed scheme for the formation of symmetric/asymmetric complex of

GroEL/GroES with ADP/AlFx...... 147

Figure 3.1. Conductivity domains of metals, semiconductors, and insulators...... 151

Figure 3.2. Conduction electron concentrations in different materials...... 154

Figure 3.3. Temperature dependence of conductance electron concentration in Ge and Si.154

Figure 3.4. Effect of doping impurity (Sb, donor) concentrations on the resistivity of Ge

as a function of inverse temperature...... 155

Figure 3.5. Band structure of an intrinsic inorganic semiconductor...... 156

Figure 3.6. Mobility of (a) ultrapurified and (b) conventionally purified perylene...... 159

xiii Figure 3.7. TGA Analysis of (a) tetracene from TCI America; (b) crude pentacene 163;

twice...... 176

Figure 3.8. Crystal structure of (a) pentacene and (b) 1,2,3,4-tetrafluoro-6,13-bis(2-

diisopropulsilylethynyl)pentacene...... 181

Figure 3.9. Interatomic distances in a-DIB unit cell...... 196

Figure 3.10. Intermolecular iodines’ p-orbital overlap in crystalline a-DIB...... 197

Figure 3.11. Significance of various HAT-n compounds represented as number of

publications for each member of the homologous series...... 205

Figure 4.1. Photodegradation of compound 93 and its exponential fit...... 215

xiv LIST OF SCHEMES

Scheme 1.1. Farnum’s synthesis of DPP...... 18

Scheme 1.2. Condensation of benzonitrile with alkyl succinate...... 21

Scheme 1.3. Self-condensation of alkyl succinate under basic conditions...... 21

Scheme 1.4. Synthetic approaches to the intermediates 2 and 3...... 24

Scheme 1.5. Stobbe condensation of a Schiff base with alkyl succinate...... 26

Scheme 1.6. Preparation of DPP from benzylaniline...... 26

Scheme 1.7. Mechanism of DPP formation from benzonitrile and succinate...... 29

Scheme 1.8. Condensation of succindiamide with N,N,-dimethylbenzamide...... 30

Scheme 1.10. The sole example of an unsymmetrical N,N′-diaryl DPP...... 31

Scheme 1.11. The sole example of aliphatic amine reaction with furo[3,4-c]furane-1,4-

dione...... 31

Scheme 1.12. Routes to 3,6-diphenyl-furo[3,4-c]furan-1,4-dione...... 32

Scheme 1.13. Reactive sites of DPP...... 33

Scheme 1.14. Various reactions of DPP...... 33

Scheme 1.15. N-Alkylation of DPP...... 34

Scheme 1.16. Acylation of DPP with di-tert-butyl dicarbonate...... 35

Scheme 1.17. Acylation of DPP with di(2-methyl-3-buten-2-yl) dicarbonate and

subsequent decomposition...... 35

xv Scheme 1.18. N-Hydroxymethylation of DPP...... 36

Scheme 1.19. Reactions of N,N′-bis(hydroxymethyl) DPP...... 37

Scheme 1.20. DPP bromination and chlorination products...... 38

Scheme 1.21. Mechanism of DPP bromination...... 38

Scheme 1.22. Sulfonation of DPP...... 39

Scheme 1.23. Replacement of O in –NH–C=O by C...... 39

Scheme 1.24. Replacement of O in –NH–C=O by N...... 40

Scheme 1.25. Replacement of O in –NH–C=O by N–CN (cyanoimination of DPP)...... 40

Scheme 1.26. Nitration and aromatic nucleophilic substitution of chlorine in DPP...... 41

Scheme 1.27. Aromatic nucleophilic substitution of bromine in DPP...... 42

Scheme 1.28. Pd coupling of Br-DPP with CO in presence of calcium formate...... 42

Scheme 1.29. Pd coupling of Br-DPP with CO in presence of MeOH...... 42

Scheme 1.30. Pd coupling of Br-DPP with CO in presence of butylamine...... 42

Scheme 1.31. Stille coupling polymerization of DPP...... 43

Scheme 1.32. Suzuki coupling polymerization of DPP...... 43

Scheme 1.33. Design of red-shifted DPP chromophores...... 63

Scheme 1.34. Preparation of sterically hindered di-alkyl succinates...... 64

Scheme 1.35. Preparation of 4-R-benzonitriles: R= F (13), Br (14), OMe (15)...... 65

Scheme 1.36. Preparation of 4-aminobenzonitriles...... 65

Scheme 1.37. Alkylation of DPP...... 71

Scheme 1.38. Step-wise alkylation process of DPP...... 72

Scheme 1.39. High temperature basic degradation of DPP...... 76

xvi Scheme 1.40. Michael addition of OH– to DPP-Me...... 77

Scheme 1.41. Bromination reactions of DPPs...... 80

Scheme 1.42. Indirect approach to iodo-DPPs...... 81

Scheme 1.43. Conversion of Br-DPP-Pr to CN-DPP-Pr...... 82

Scheme 1.44. Heck coupling between X-DPP-Me with 4-tert-butyl- and 4-acetoxy-

styrenes...... 83

Scheme 1.45. Preparation of styrenes...... 84

Scheme 1.46. Pd-catalyzed coupling between styrene 57 and Br-DPP-Pr 44...... 85

Scheme 1.47. Pd-catylized coupling between styrene 59 and Br-DPP-Pr 44...... 85

Scheme 1.48. Preparation of thien-2-yl zinc reagents 62 and 63. Negishi coupling

between Br-DPP-Pr 44 and thien-2-yl zinc chloride 62...... 86

Scheme 1.49. Negishi coupling between Br-DPP-Pr 44 and 5-(4-(N,N-di-n-

hexylamino)phenyl)-thien-2-yl zinc chloride 63...... 87

Scheme 1.50. Introducing sulfonic acid group into DPP structure...... 88

Scheme 1.51. Proposed direct introduction of alcohol functionality into DPP by

alkylation with ω-halo-α-alcohols...... 89

Scheme 1.52. The sole successful example of DPP alkylation with ω-halo-α-alcohol in

presence of t-BuOK...... 90

Scheme 1.53. Attempted hydroboration route to alcohol-functionalized DPPs...... 91

Scheme 1.54. Cu/CuI-cat. coupling of 2-(ethylamino)ethanol with I-DPP-Pr 46...... 92

Scheme 1.55. Alkylation approach to alcohol-functionalized DPPs with protection-

deprotection of the hydroxy group...... 93

xvii Scheme 1.56. Preparation of alkylating reagents with alcohol functionality, protected

with a THP protecting group...... 93

Scheme 1.57. Reaction of THP- protected alcohols 71–73 with DPP...... 95

Scheme 1.58. Preparation of alkylating reagent 75 with alcohol functionality, protected

with a benzoate ester...... 95

Scheme 1.59. Reaction of benzoate ester protected alcohol 74 with DPP and subsequent

removal of the protecting group...... 96

Scheme 1.60. Preparation of complimentary mono-alcohol functionalized DPPs...... 97

Scheme 1.61. Direct alkylation of DPPs with ω-halo-α-alcohols in presence of Cs2CO3.98

Scheme 1.62. Bis(maleimide) DPP derivative...... 100

Scheme 1.63. An attempt towards maleimide mono-functionalized DPP 90...... 101

Scheme 1.64. Direct copper-mediated N-arylation of isatins...... 102

Scheme 1.65. Direct copper-mediated N-arylation of DPP 1...... 103

Scheme 1.66. Aromatic nucleophilic substitution on DPP...... 104

Scheme 2.1. Fluorophores conceptually similar in structure...... 124

Scheme 2.2. Examples of new DCDHF dyes...... 126

Scheme 2.3. The target fluorescent tags with fluorophores, hook, and spacer of choice.128

Scheme 2.4. Proposed attachment of the maleimide hook via N-(2-hydroxyethyl) group.129

Scheme 2.5. Proposed synthesis of Nile Red derivative 99 and attachment of the

maleimide hook via phenol functionality...... 130

Scheme 2.5. Preparation of Nile Red Phenol...... 131

Scheme 2.6. C6-Spacer attachment to Nile Red Phenol...... 131

xviii Scheme 2.7. Syntheses of maleimides...... 132

Scheme 2.8. Syntheses of maleimides from maleic anhydride...... 133

Scheme 2.9. Attempted Mitsunobu reactions between Nile Reds 105 and 106 and various

hydroxy-functionalized maleimides...... 134

Scheme 2.10. Attempted N-alkylation of maleimide according to a reported protocol. .134

Scheme 2.11. Simple alkylations and acylations of Nile Red Phenol 115...... 135

Scheme 2.12. Diels-Alder adduct of furan and maleic anhydride Fu-MA...... 137

Scheme 2.13. Synthesis of protected Nile Red – Maleimide 122...... 137

Scheme 2.14. Retro Diels-Alder deprotection of 122...... 138

Scheme 2.15. Synthesis of maleimide-tagged DCDHF-2V 124...... 139

Scheme 2.16. Synthesis of maleimide-tagged DCDHF-6 125...... 140

Scheme 3.1. Target polysubstituted n-acenes...... 162

Scheme 3.2. Various approaches to 2,3,6,7-tetraalkoxyanthracene via 2,3,6,7-tetraalkoxy-

9,10-anthraquinone...... 164

Scheme 3.3. Attempted self-acylation of veratroyl chloride...... 165

Scheme 3.4. Ortho-lithiation approach to 138...... 166

Scheme 3.5. Oxidation and cyclization of 138 into anthraquinone 130...... 167

Scheme 3.4. Attempted preparation of 133...... 167

Scheme 3.5. Preparation of 2,3,6,7-tetraalkoxy-9,10-dihydroanthracenes...... 168

Scheme 3.6. Conversion of methoxy groups into hydroxyl functionalities...... 169

Scheme 3.7. Synthesis of 2,3,6,7-Tetraalkoxy-9,10-dialkylanthracenes...... 171

xix Scheme 3.8. Preparation of 1,2,3,4,5,6,7,8-octaalkylanthracenes by Pd-catalyzed ring

extension reaction...... 171

Scheme 3.9. Peralkylation of 9,10-dihydroanthracene with heptyl bromide...... 172

Scheme 3.10. Synthesis of tetracene 157...... 173

Scheme 3.11. Synthesis of 2,3-Bis(decyloxy)tetracene 162...... 174

Scheme 3.12. Synthesis of pentacene...... 177

Scheme 3.13. Preparation of pentacenequinone 164 via 1,4-anthracenequinone 167.....182

Scheme 3.14. Synthesis of the key intermediates for 1-fluoropentacene...... 183

Scheme 3.15. Synthesis of 2-fluoropentacene...... 185

Scheme 3.16. Retrosynthetic analysis of alkoxypentacenes...... 187

Scheme 3.17. Ortho-lithiation approach to 4,5-dimethoxyphthalaldehyde 158...... 188

Scheme 3.18. Three-step synthesis of 158 from veratrole...... 191

Scheme 3.19. Synthesis of 2,3,9,10-tetramethoxypentacene...... 192

Scheme 3.20. Soluble and stable pentacene ethers by Anthony...... 193

Scheme 3.21. Attempted synthesis of 2,3,9,10-tetrahexylpentacene 183...... 193

Scheme 3.22. Reversible Diels-Alder adduct of pentacene...... 194

Scheme 3.23. Preparation of 2,4,6,8-tetraiodoglycoluril...... 198

Scheme 3.24. Iodoarenes prepared by direct iodination...... 200

Scheme 3.25. Iodo-de-diazoniation approach to some iodotoluenes...... 201

Scheme 3.26. Routes to 1,4-diiodonaphthalene...... 203

Scheme 3.27. Preparation of 2,3-diiodonaphthalene ...... 204

Scheme 3.28. Oxidative trimerization of 1,2-dialkoxybenzene to HAT-n...... 207

xx Scheme 3.29. Main by-products of HAT-n synthesis by oxidative trimerization...... 207

Scheme 3.30. Synthesis of MN-HAT-5 and TN-HAT-5...... 210

xxi LIST OF TABLES

Table 1.1. Typical SMS fluorophores...... 17

Table 1.2. DPP compounds prepared by Reformatsky reaction...... 20

Table 1.3. DPP compounds prepared by aromatic nitrile – succinate ester condensations.24

Table 1.4. DPP compounds prepared by step-wise condensations...... 26

Table 1.5. Solubilities of various DPPs in mol·liter–1...... 48

Table 1.6. Influence of 3,6-substituents and crystal structure on absorptive properties of

DPP...... 52

Table 1.7. Fluorescence data for several DPPs in chloroform...... 53

Table 1.8. Fluorescence in solution and solid state of some N-phenyl substituted DPPs. 54

Table 1.9. Fluorescence quantum yields of some N-mono-aryl and N,N′-diaryl substituted

DPPs...... 56

Table 1.10. 1H Chemical shifts of the NH group of DPP in different solvents and at

various temperatures...... 59

Table 1.11. 13C Chemical shifts of DPP, its mono- and di-anion...... 59

Table 1.12. DPP preparation reactions...... 65

Table 1.13. N-Alkylated DPPs...... 72

Table 1.14. Halogenated DPPs...... 81

Table 1.15. Aminated DPPs...... 82

Table 1.16. Photostability of several DPPs...... 109

xxii Table 1.17. Symmetrical 4,4’–Disubstituted DPPs...... 110

Table 1.18. N– and N,N′-substituted DPPs...... 112

Table 2.1. Amine (R2–NH2) reactive groups for molecular probes...... 118

Table 2.2. Thiol (R2–SH) reactive groups for molecular probes...... 120

Table 2.3. Optical properties of maleimide-containing fluorescent tags...... 141

Table 3.1. Carrier mobilities m in some crystalline inorganic semiconductors at room

temperature, in cm2/V·s...... 153

Table 3.2. Key differences between metals, semimetals, intrinsic inorganic

semiconductors, and insulators at room temperature...... 153

Table 3.3. Carrier mobilities (of electrons e– and holes p+) in some organic molecular

crystals at room temperature...... 158

Table 3.4. 2,3,6,7-Tetraalkoxy-9,10-dialkylanthracenes...... 170

xxiii

CHAPTER I.

DPP FOR SINGLE-MOLECULE SPECTROSCOPY

1.1. Single-Molecule Spectroscopy and its Applications

Spectroscopy (Latin spectrum – image, impression, from specĕre – to look; and

Greek σκοπια, – observation, from σκοπειν – examine, look at) is a physical method of study of matter based on observation of radiation intensity as a function of frequency,

wavelength or other parameter (m/z, polarization, energy, etc.). The radiation may be of

electromagnetic (i.e. optical spectroscopy), sonic, or particulate (mass spectroscopy)

origin. The electromagnetic radiation may be emitted, absorbed, reflected, scattered or

transformed into another form. Luminescence (Latin lūmin, lūmen – light, genitive case

lūminis, and -ēscentem, from -ēscĕre – ‘beginning to assume a certain state’) is a physical

phenomenon, where matter, after being excited, emits light in excess of thermal radiation,

and that emission lasts significantly longer than the oscillation period of the light being

emitted. Luminescence differs from scattering, reflection, deceleration emission

(Bremsstrahlung), Vavilov-Cherenkov radiation, or parametric transformation of light. If

the excitation is electromagnetic, the phase or polarization of the emitted light does not

1 2

correlate with the phase of the excitation light, resulting, for example, in the depolarization of the luminescence in isotropic solutions of otherwise randomly oriented fluorophores. There are several types of luminescence, classified by the excitation sources – light (photoluminescence), ionizing radiation (radioluminescence), X-ray, electrical field (electroluminescence), beam of electrones (cathodoluminescence), mechanical force (triboluminescence), crystallization1, or (bio)chemical reaction (chemo-

and bio-luminescence). Photoluminescence splits into phosphorescence and

fluorescence. Fluorescence is short (10–9 to 1 sec) light emission due to transitions

between excited and ground states of the same multiplicity, usually singlets (S1ÆS0). In that it is different from phosphorescence, which is usually long (1 sec to days) light emission due to forbidden (i.e. low probability) transitions between excited and ground states of different multiplicity, usually the excited triplet state and the ground singlet state. Fluorescence quantum yield ΦF is a ratio of number of photons emitted to the

number of photons absorbed by the same amount of the substance.

Conventional spectroscopy observes a large number of molecules at once and the spectral signal is averaged over the statistical ensemble. Single-molecule spectroscopy (SMS) is a relatively new (first SMS experiment was performed in 19892),

yet well-elaborated technique, which detects fluorescence from one single molecule.3

This allows the full distribution of optical values (fluorescence intensity, fluorescence decay, emission spectrum, diffusion coefficient, fluorescence anisotropy, polarization, and lifetime) to be recorded and molecular heterogeneity in optical properties, as well as many molecular time-dependent state changes to be measured on a molecule by molecule

basis. Single-molecule spectroscopy is not the only spectroscopic technique with nanoscale resolution and yoctomole (10–24 mole) detection limits. Single electrons and

ions have been confined in electromagnetic traps4 and their spectra have been recorded.

The spatial resolution of scanning tunneling microscopy (STM) or atomic force

microscopy (AFM) is much higher. However, the trapped species are bound in their

motion by the trap’s potential and to date, no single molecule has been cooled sufficiently

to be bound in an electromagnetic trap.

Figure 1.1. Jabłoński Diagram.5

To specifically detect one single molecule, the molecules should be distanced far enough from each other and the excitation beam should be focused at the probe volume of such dimensions that at a given dilution it contains only one molecule. The first task is achieved by dilution, and the second – by using a laser beam in conjunction with confocal microscopy, near-field scanning microscopy, or other detection methods.6 For a successful SMS experiment, a combination of small (10-100 μm3) probing volume, low fluorophore concentration, and exact laser beam frequency tuning need to be optimized

for any given fluorophore and optical set up. That assured, the signal-to-noise ratio for a given fluorophore, generally depends (variable for different detection methods) on its quantum yield ΦF, absorption cross-section σp, and (implicitly, via detector averaging

time) – on photostability, expressed in terms of photobleaching quantum efficiency, also

called branching ratio φb. The medium where the fluorophore is dispersed also has certain

requirements: it should be transparent in the frequency range of interest, be impurity-free to minimize elastic (Rayleigh) or inelastic (Raman) scattering, and has minimum dark state emission and background fluorescence.

Single-molecule spectroscopy has found numerous applications. First of all,

from a theoretical point of view, it permits the direct comparison of models drawn from considerations of individual molecules and their properties. For example, it was SMS that established the fact that reaction pathways for different molecules (of the same substance) are not predetermined, but still deliver the same product. That is, reactions proceed heterogeneously on multidimensional energy landscapes and individual molecules follow different pathways in the phase space. Since these different pathways can occur on a wide range of time scales, from femtoseconds to seconds, individual reaction rates for single

molecules of the same substance differ significantly.7,8

From a practical point of view, the most promising application for SMS is the

task of single strand DNA sequencing by detecting fluorophore-labelled individual bases as they are being assembled by polymerase (direct method) or sequenced by exonuclease

(reverse method) into or from an immobilized single DNA molecule. The main idea here is that the residence time of a freely diffusing single fluorescently labelled nucleotide

molecule is in the range of few microseconds. After being incorporated into the single

DNA strand, their residence time increases to several milliseconds, becoming sufficiently long for unequivocal fluorescent identification, presuming that each nucleotide base is labelled with its own, easily distinguishable fluorophore. By this method, the DNA sequence is directly retrieved from the detected signal sequence. Although this goal of

DNA sequencing has not been achieved yet as a whole, each step in this scheme has been shown to work on single-molecule level.8 In 2005 NIH gave nine grants totalling

$25,000,000 to implement this process as well as other approaches to reduce the cost of

genome sequencing below $1,000.9 Most biologically relevant SMS experiments have

been conducted in vitro. The ultimate dream, however, is to bring the SMS expertise to a

level which would allow non-invasive in-vivo analysis of living cells in their native

conditions.

An ability to precisely and specifically label several different sites in a

macromolecule allows utilization of such a technique as fluorescence resonance energy

transfer (FRET or RET), a physical phenomenon involving long-range dipole-dipole

interactions, known from 1948.10 FRET relies on nonradiative transfer of electronic

excitation from an excited donor to a ground state acceptor, which depends on the

distance between the two fluorophores (and decays as R–6), the spectral overlap of the

donor emission and the acceptor absorption (overlap integral), the refractive index of the

media, the donor quantum yield, and the relative orientation of the two fluorophores.

Thus, in combination with SMS, FRET is ideally suited for structure elucidation and

monitoring of conformational changes in biomolecules.11 Fluorescence correlation

spectroscopy (FCS) calculates the autocorrelation of the fluorescence intensity fluctuations to follow time-dependent dynamics. Near-field single-molecule optical microscopy, confocal fluorescence microscopy operates with 0.5–1.0 fL probe volume12.

SMS was recently employed in a very elegant method, called by the authors anti-Brownian electrophoretic trap (ABEL), to pin-point, trap, position and manipulate nanoscale objects (down to 20 nm) in solutions at ambient temperatures.13 Two-photon excitation (TPE) 14,15,16 is a second-order, non-linear process with extremely small cross

sections, typically on the order of 10–50 cm4. The molecular excitation rate depends

quadratically on the laser beam intensity. This method was used for two-color

colocalization of different dyes.17 Colocalization is an implementation of single-

molecule imaging to biological objects. Since the fluorophore molecules are much

smaller than the wavelength of the light they emit, they serve as point sources of light.

With the help of modern detection techniques and mathematical processing of the resulting point-spread functions, the emitting center may be localized with high accuracy and has been used to follow the motion of individual motor proteins and the diffusional trajectories of labelled lipids in membranes.8 The colocalization technique gives resolution of a few tens of nanometers and closes up the gap between far-field optical microscopy with resolution of 200 nm and up, and FRET with resolution of ca. 2 to 8 nm.

The single biggest application of SMS nowadays is molecular imaging. The

first optical image of single pentacene molecules diluted in terphenyl was performed by

W. E. Moerner at cryogenic temperatures.2,18 Since then huge progress has been made

and single molecules are now imaged even in cells (living or, more often, immobilized).

Some imaging methods in biology rely on autofluorescent biomolecules,19,20,21,22 the most famous among them being the green fluorescent protein (GFP),23 while majority of

imaging methods require labelling24 of nucleic acids, proteins, DNAs or RNAs with

fluorescent tags. The power of the molecular imaging technique may be exemplified by

the recent real-time monitoring of the infection pathway of single viruses.25 A single,

seven-nanometer (7·10–9 m) long, single-walled carbon nanotube has been imaged at a

resolution, comparable to that of scanning tunneling microscopy.26 The tags, or labels,

used for molecular imaging, have to fulfill a number of requirements.

1.2. Dye Requirements for Single-Molecule Spectroscopy

A fluorescent dye, to be successfully employed in an SMS experiment, should

possess several properties. Specifically, it should possess:

• large absorption cross section σp

• high quantum yield ΦF

• short fluorescence lifetime τ1

• high photostability = small φb

• high Stokes shift

• weak bottlenecks into dark states, i.e. small kISC

If the SMS experiment to be conducted is in biological media – on biomolecules, in living or immobilized cells, in addition to the above requirements, several additional requirements should be added (this does not imply the dye should fulfill all the requirements above and below all the time or simultaneously):

• cell permeability

• high site binding specificity

• non-interference with the biological functions of the substrate being labeled

• emission in red region of spectrum

• high solvatochromism

Absorption cross-section σp is directly proportional to the molar extinction

A()λ ε coefficient ε and may be calculated from it: σ in cm2 per molecule isσ ()λ = = , p ⋅ l N N A where A is the absorbance for an optical path length l (cm), N is the number of molecules

per unit volume, and NA is Avogadro’s number. High σp at the excitation wavelength

means that the photons of the incident (exciting) light beam are efficiently absorbed and

background signals from unabsorbed photons are minimized. Since σp and ε are directly related, everything that affects one is also true in changing the other. Typical SMS fluorophores have ε > 20,000 L·mol–1·cm–1.

Quantum yield ΦF, defined in section 1.1, is a number, characteristic in how

efficient a fluorophore is in emission. An ideal fluorophore emits same number of photons (counting as particles) it absorbed, producing a quantum yield of exactly unity.

This is the case when the radiationless decay to the ground state S0 is much slower than

radiative decay. The energy of the emitted fluorescent photons, however, is always lower

than the energy of absorbed photons because of relaxation processes (Stokes’ losses). A

good fluorophore should possess high (as close to unity as possible) fluorescence

quantum yield ΦF, but at least higher than 0.1. A product of the fluorophore’s quantum

yield and its molar extinction coefficient at the excitation wavelength is called

brightness.27

The fluorescence lifetime τ1, or lifetime of the excited state is the average time

the molecule spends in the excited state prior to decay to the ground electronic state. This time should be fairly short (in the range of 0.5–5 nanoseconds)27 to provide as many

excitation-emission cycles per unit time, as possible. It is during this time span that the fluorophore acquires information about its environment.

Common dye molecules employed in SMS can emit up to a million photons before irreversible photobleaching occurs. A quantitative measure of photobleaching is

photobleaching quantum efficiency, also called branching ratio φ b, defined as

probability per optical absorption event to irreversibly generate a non-fluorescent

product. Photostability is the reciprocal of photobleaching quantum efficiency. Low

photostability of a fluorophore in practical SMS experiments results in low observation

and averaging times available to the experimenter (Fig. 1.2.).8 Photobleaching in a single,

instantaneous step is also an indirect proof that the fluorescence observed from the probe volume is coming from only a single molecule. Photodestruction of fluorophores is one of the most important yet least understood processes that affect the application of

fluorescence in biology.28 Up to date they are best described as “elusive photochemical

reactions, mainly photo-oxidation”.27,29 Depending on the time scale of the detection

method and the processes studied, photostability varies from desirable to essential. For

example, surface-immobilized fluorophore-labelled biomolecules should have long

photosurvival times, while dyes for photo histogram studies are less sensitive to

photobleaching of individual molecules as they come into and pass out of the probe

volume. Photostability may generally be increased by careful exclusion of oxygen from

the solution containing the fluorophore.30

Figure 1.2. Irreversible photobleaching (at 5.6 sec) of single Cy5 molecule, immobilized

on a glass slide.8

Generally, the higher the power of the laser pumping the fluorophore, the higher

the fluorescence response, until the corresponding transition is saturated. Saturation

intensity is at a maximum when there are no strong bottlenecks, like intersystem

crossing (ISC) from the excited singlet state S1 into the excited triplet state T1. During

the lifetime of the triplet state T1, no emission or absorption happens (thus, the name of

dark state), resulting in saturation of the emission rate and decrease of the absorption

cross section. Thus, another requirement for a good SMS fluorophore is small value of

kisc (rate of intersystem crossing) and large value of kT (decay rate from triplet to ground

state). One of the molecular structural classes, satisfying this requirement, are planar aromatic compounds.31

For biologically oriented applications, it is desirable for the fluorophore to

absorb and emit in the red region of the visible spectrum (above 600 nm), since there are

only few compounds of biological origin32 that show intrinsic absorption and emission in

this region. Also, the major source of the background in SMS is scattering of the

excitation light. The intensity of the scattered light decays approximately as fourth power

of the wavelength, thus, red-shifted dyes give less background33. Non-interference with biological functions is essential if the fluorophore’s intended use is in live cells or the molecules, which will be tagged with such fluorophore, will undergo biochemical transformations. For example, if the nucleotides for determining of DNA sequence are labelled with fluorescent tags, those tags should not affect the ability of the nucleotides to

undergo synthesis under polymerase action or sequencing by exonuclease. In fact, most natural DNA polymerases and exonucleases have been found very sensitive to such structural changes in nucleotides as dye labelling and discriminate unlabelled nucleotides against labelled ones. To circumvent this problem, mutant enzymes were employed. Non- interference also means that the dye should be non-toxic.34

High site binding specificity of the fluorescent tag necessitates from the end- purpose of such labelling. If a tag will attach to a wrong part of the molecule to be imaged, the image may provide wrong information about that molecule. Binding here means either covalent bonding to the substrate molecule, i.e. tagging, or mere preference to position (adsorb, adhere) itself within certain parts of the substrate molecule (probing) due to structural, steric, conformational or other peculiarities. For example, it is known that for a dye to specifically remain in the vesicle of a cell, it is often sufficient to have long normal alkyl chains in its structure. The unbound fluorophores create high background fluorescence and must be washed out (usually by gel filtration) from the labelled system as thoroughly as possible. An ideal solution to this problem would be a tag, which becomes fluorescent only after binding to the substrate. Delivery of the dye inside the cells (cell-loading) and then specifically to the sites to be labelled constitute a problem in itself, current solutions of which are direct microinjections or utilizations of liposomes.

The Stokes’ shift35 of the fluorophore is the difference between the emission

maximum and the absorption maximum, Δλ=λem–λabs. The energy decrease has many

causes, the common ones being (a) the rapid (10–12 sec) relaxation of the excited molecule to the lowest vibrational level of S1 and (b) fluorescent decay to the higher

vibrational levels of S0. Therefore, the wavenumber of the Stokes shift is a direct measure

of the vibrational energies of the molecule. Other physical phenomena, like interactions

with solvent, excited-state reactions and energy transfers, may contribute to the Stokes’

shift as well. The high Stokes’ shift makes the fluorophore’s emission bathochromically

shifted, i.e. the larger it is, the more the emission maximum is shifted into the red region of the spectrum, which allows easy discrimination of the fluorescence signal from

Rayleigh scattering and increases signal-to-noise ratio.27 Large Stokes’ shift also results

in smaller overlap integral of absorption and emission spectra, thus minimizing

concentration related self-quenching or RET homotransfer of the fluorophore.

Solvatochromism is a phenomenon involving dependence of the absorption or

emission maximum on the solvent polarity the dye or fluorophore is dissolved in.

Negative solvatochromism corresponds to a hypsochromic shift; positive

solvatochromism corresponds to a bathochromic shift with increasing solvent polarity.

The origin of the phenomenon is the same for both absorption and fluorescence and lies

in the dipole-dipole interactions (distortions) of the ground (for absorption) and excited

(for fluorescence) electronic states of the molecules with the molecules of the solvent.

Thus, solvatochromism may serve and is used indeed to assess the polarity of different

media. There is, however, an important difference between solvatochromism in

absorption and the one in emission. Absorption of light by a molecule is an instantaneous

process: it takes femtoseconds (10–15 sec) to occur. Thus absorption can provide

information only on the average ground state of dye molecules and on the solvent

molecules immediately adjacent to them. Absorption spectra are not sensitive to

molecular dynamics and the absorptional solvatochromism, regardless of number of

molecules, averages the information from the surrounding solvent shell by the very

nature of the absorption process. On the contrary, a fluorophore spends some time (10–9

–8 to10 sec) in its excited state and it is during this time τ1 that it gathers information about

its environment. This time scale allows an excited fluorophore molecule to collide with other species and/or reflect conformational changes in the molecule. Solvent relaxation, a process of reorientation of polar solvent molecules to minimize dipole-dipole interactions with the solute, occurs during a picosecond (10–12 sec) time span. Most of the

fluorophores have larger dipole moment in the excited state than in the ground state and

the solvent molecules have sufficient time to reorient themselves around the excited state

dipole, lowering its energy and resulting in a bathochromic Stokes’ shift. Thus,

solvatochromism of emission spectrum is much more sensitive to the solvent polarity

than solvatochromism of absorption spectrum of the same compound. An important

parameter for a fluorophore to sense its environment polarity is therefore its dipole

moment. Nonpolar fluorophores, like most aromatic hydrocarbons, are non- or weakly

sensitive to the solvent polarity.

When combined with single-molecule spectroscopy, solvatochromic

fluorophores can sense the local polarity of their immediate (i.e. of nanometer size)

environment – be it the closest coordination sphere in a micelle or the closest part of the

biomolecule it is confined or bound to. Since the local polarity of biomolecules often

depends on their conformation states, and the latter often dynamically change and affect

the very biological functions thereof, solvatochromic fluorescent probes and tags are

powerful tools for determination and monitoring of structural changes in biomolecules.

The possibility of successful monitoring of the conformational dynamics of individual

biomolecules with temporal resolutions comparable to those of new molecular dynamics

simulations36 raises the hope that it might be possible to relate conformational dynamics

directly with enzyme activity. Such studies would require site-specific labelling of the active site of enzyme with a fluorophore, which is selectively quenched (perhaps by an amino acid) in close proximity.

For a covalently binding fluorescent tag, an important variable to consider in its design is the length of the spacer or linker between the fluorophore and the hook or reactive group, which after labeling determines the proximity of the fluorophore to the substrate. According to the opinion of Weiss and Kapanidis,27 fluorescent tags with

small, short, and rigid spacers are preferred since they tend to be less perturbative to their

local environment and ensue in fewer fluctuations of the fluorescent properties of the

single fluorophore due to unknown and uncontrolled conformational changes.27 Flexible spacers, on the other hand, provide more sensitivity (more pronounced changes in fluorescence parameters) to the local environment and are also more desirable in FRET experiments. Extreme care must therefore be taken to separate out fluorophore dynamics from the biological dynamics.28

The pool of chemical reactions that provide site-specific covalent binding of the

fluorescent labels (not necessarily detected at single-molecule level) to biosubstrates is

referred to as “bioconjugate” or “bioconjugation” chemistry. There are currently many

fluorescent dyes available with a wide range of physico-chemical parameters to choose

from.37 And yet the need for new SMS fluorophores is growing: “current dye-based

fluorescent technologies do not stand up to the challenge” (of study of individual and rare

biological processes in the living cells). The development of new fluorescent probes with

superior photophysical properties is needed.28 It is also necessary to elaborate new

labelling strategies and “devise chemistries that render the reagent fluorescent only after incorporation to the site of interest, thus minimizing the background of unincorporated reagent that will otherwise overwhelm the SMS signal.”27 Many (i.e. thousands) existing

fluorophores have never been tested for performance and applicability in SMS. There are

also no solid rules for rational design of SMS-compatible fluorophores. The

photophysical parameters of the fluorescence are media dependent as well as fluorophore

dependent. The fluorophores are not easily tunable to the desirable precision, and change

of one parameter (e.g. fluorescence wavelength) often affects the other (e.g. quantum

yield). An emerging solution to fine-tuning fluorescent probes are fluorescent

semiconductor nanocrystals, or quantum dots,38,39 which have broad excitation spectra, narrow and tunable emission spectra, long fluorescence lifetimes, and high photostability.

They have recently been used to monitor individual eight nanometer step motions of two molecular motors, kinesin and dynein, in vivo.40 However, their bioconjugation

chemistries are immature yet.27 Quantum mechanics and computational chemistry allow

calculations of both electronic and vibrational energy levels of a given molecule in the

gas phase with very good reproducibility and often with good accuracy. Yet the mere fact

of a molecular compound being fluorescent or not is still not reliably predicted by

modern theory.

From a molecular structure point of view, synthetic fluorophores are typically

aromatic organic compounds. With the exception of the lanthanides, individual atoms are

generally nonfluorescent in condensed phases.41 The quantum yield and lifetime can be

altered by factors which affect either of the rate constants – kF and knr. A molecule can be

nonfluorescent because of fast internal conversion or because of slow emission rate. It is empirically known that the presence of a nitro-group in aromatic compounds makes knr large and thus quenches (i.e. decreases) the fluorescence. The quenching may be caused by intermolecular processes as well. For example, halogens such as Cl, Br, and I and other heavy atoms quench fluorescence via spin-orbital coupling and intersystem crossing to the triplet state. As a result, molecules containing heavy atoms, specifically bromine and iodine, are often phosphorescent (rather than fluorescent), and the fluorescence quantum yields of fluorophores in chlorinated and other heavy-atom containing solvents are generally lower.

Table 1.1. Typical SMS fluorophores.27

1.3. Review of known DPP chemistry

The first representative of the dihydropyrrolo[3,4-c]pyrrole-1,4-dione (DPP) class of dyes, 3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione (1) was discovered in

1974 by Farnum42 in an attempt of a Reformatsky reaction on benzonitrile, ethyl

bromoacetate, and zinc to obtain 2-azetinone. Instead of the latter, a red pigment of structure 1 was obtained in poor yield (Scheme 1.1).

N O O

OEt a + HN NH NH X Br O O

Scheme 1.1. Farnum’s synthesis of DPP. a) Zn/Cu, toluene, 110°C, several hours.

This new class of dyes turned out to be highly insoluble materials and excellent

pigments43: highly thermally stable, photostable and fluorescent in solid state. Soluble

derivatives of DPP are reported to possess high quantum yields,44,45,46,63 large Stokes

shifts46, and high photostability47. Solubilized DPP derivatives have found applications in

“soluble (latent) pigments”48,49,50, polymers51,52,53, photorefractive,54 photoconductive,55 electrochromic,56 electronic57,58 materials, dendrimers59,60, and liquid crystals61,62. The reported photophysical characteristics63 of the DPP family of dyes would appear to lend

itself to the field of single-molecule spectroscopy. The demands on dye stability for

applications in single-molecule spectroscopy are particularly high64 and the amazing

thermal stability of 1 up to 500°C was encouraging. The sole previous study of single-

molecule fluorescence of a DPP dye involves a diphenyl DPP core covalently imbedded

in a dendrimer in polymer films65. We decided to further examine the diphenyl DPP class

of fluorescent dyes, which might be utilized in single-molecule biological studies. Simple

diaryl DPP’s have solution absorptions in the range of 470-520 nm and emission in the range of 508–540 nm. Since still longer wavelengths of fluorescence are desirable

(biological media are more transparent at longer wavelengths and with less background autofluorescence), we also wanted to prepare DPP dyes with longer fluorescence wavelengths by extending the conjugation of the core and by introducing donor or acceptor groups at the termini of the conjugated system. The only systematic review of

DPP chemistry in the open literature is from Iqbal66. There are also two recent reviews of

DPPs as a class of pigments.67,68 Otherwise, virtually all DPP chemistry has been

described in patents, where over a thousand DPPs are described. Here most important

extract of that chemistry pertinent to our work follows.

The nomenclature of the DPPs is derived from the core bicyclic heterocyle DPP,

named by IUPAC 1957 system, rule B-3 or rule B-4.1(b) – Chemical Abstract system.

Accordingly, the DPP may be termed as

1 H H N 2 R 2 O 5 N 3 2 2 O 3 N 3 B-3 1 CA R1 4 1 1 4 3 R 2 3 + 0 1 3 c 6 b R2 6 1 4 3 R1 d N O 45 N O 4 5 N a H e H R

DPP 1: R1–DPP–R

► 1,4-diketo-2H,5H-dihydropyrrolo[3,4-c]pyrrole (obsolete);

► 2H,5H-dihydropyrrolo[3,4-c]pyrrole-1,4-dione;

► 3,7-diazabicyclo[3.3.0]octa-4,8-dien-2,6-dione.

For derivatives of 3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-diones, we will use the simplified notation of R1–DPP–R, where R1 is the substituent on the phenyl ring

of 1, and R is the substituent on nitrogen. If no position of R1 is specified, it is assumed to

be at the default 4- (para) position.

Since the Reformatsky reaction of benzonitrile gives only variable 5-20% yields

of DPP with various byproducts, Farnum’s approach to DPP preparation is only of

historical interest. Some DPP derivatives prepared by this method69,70 and their

absorption properties are presented in the Table 1.2.

Table 1.2. DPP compounds prepared by Reformatsky reaction.69

H O N R

N O H

R λmax, nm in NMP H 475, 504

Et2N 554, 512 Br 515, 480

CN 535, 500

F 464, 500

OMe 512, 475

Me2N 554, 512 3-COOMe 465, 505

The mechanistic study of the Reformatsky reaction and a retrosynthetic analysis of 1 led Iqbal et al. in 1986 to propose another preparative approach to DPP, the condensation of aromatic nitriles with succinate esters71 (Scheme 1.2). Since alkyl

succinates undergo self-condensation under basic conditions (Scheme 1.3), optimum

conditions for condensation with aryl nitriles must be followed to minimize unwanted

byproducts. First, the alkyl group in the succinate ester should have large steric volume.

Second, the order and rate of reagent addition is of utmost importance.

N O H O N OAlk + a OAlk

N O O H

Scheme 1.2. Condensation of benzonitrile with alkyl succinate.

a) strong base (e.g. t-BuOK), protic solvent (e.g. t-AmOH).

O O O

COOAlk OAlk AlkO Base + OAlk AlkO AlkOOC O O O

Scheme 1.3. Self-condensation of alkyl succinate under basic conditions.

The rate of succinate ester self-condensation decreases in the order of alkyl

substituent bulkiness: Me > Et > i-Pr > t-Bu > t-Am. Thus, dimethyl and diethyl

succinates are least preferred for condensation with aromatic nitriles, and t-amyl succinate is the ester of choice. Self-condensation is a bimolecular process, which rate depends quadratically on the ester concentration. By maintaining a low concentration of

the ester, one decreases both the rates of ester – nitrile and ester – ester condensation, yet the latter is decreased to a greater extent. From the practical point of view, this means very slow addition of the t-amyl succinate by a syringe metering pump to the reaction

mixture of the base and aromatic nitrile. Some exemplary DPP representatives prepared

by this method are given in the Table 1.3. A recent patent claims that this condensation

may be conducted in an essentially “solvent-free” way.72

This new condensation of aromatic nitriles with alkyl succinates resembles the

classical Stobbe condensation of an aldehyde or Schiff base with succinic acid esters

(Scheme 1.5).73 The resulting hexahydropyrrolo[3,4-c]pyrrole-1,4-diones, obtainable also

from N-benzylaniline instead of Schiff bases, may be dehydrogenated with 2,3-dichloro-

5,6-dicyanobenzoquinone (DDQ) to the corresponding DPP compounds (Scheme

1.6).74,75 The Schiff base of aniline and 2-formylpyridine was condensed with

diethylsuccinate to give 3,6-bis(pyridin-2-yl)tetrahydropyrrolo[3,4-c]pyrrole-1,4-dione in

36% yield. N-benzylaniline was condensed with dialkylsuccinate to give 15% of

hexahydropyrrolo[3,4-c]pyrrole. The latter two compounds could be aromatized with

DDQ to yield 30% (4.5% total) of N,N′-diarylated DPP. This approach, however, gives poor yields at both stages and requires separation of the desired compounds from various byproducts.

The reaction in Scheme 1.2 must proceed via several steps. The corresponding intermediates 276 and 377 were separated and the overall mechanism was suggested66,78 as depicted in Scheme 1.6. The enamino compound 2 can be isolated at low temperature (–

78°C) condensation of diethyl succinate dianion with nitriles (both aliphatic and

aromatic) and later converted to lactam 3 with sodium methoxide in methanol. Both 2 and 3 condense with another equivalent of nitrile providing DPP. Asymmetric dialkyl, diaryl, or alkyl-aryl DPPs may be obtained by this step-wise procedure (Table 1.4).

Aliphatic nitriles yield much lower yields of DPPs in the condensation with 3, compared to their aromatic counterparts. Evidently, this is caused by the presence of α-hydrogens in

the structure of aliphatic nitriles, which, upon removal (deprotonation) by the strong base,

decreases the electrophilicity of the nitrile and engages them into side reactions. This fact

is also reflected in the number of known 3,6-diaryl DPPs relative to 3,6-dialkyl DPPs: out

of 1,190 known DPPs, 1,140 (96%) contain 3,6-diphenyl-2H,5H-dihydropyrrolo[3,4-

c]pyrrole-1,4-dione core as a substructure.79 Also noteworthy is that a Beilstein search for

the same substructures yields only 68 DPP compounds (5.7%), 52 of which have

diphenyl DPP core as a substructure.80 This, in turn, reflects the fact that over 95% of

information on DPP compounds is found in patents, most of which belong to the Ciba-

Geigy Specialty Chemicals Company, Inc., headquarters in Basel, Switzerland. The total

number of original, primary literature sources dealing with DPPs is close to a thousand

(STN: 945; Beilstein: 37; references may overlap).

The intermediates 2 (dialkyl 2-[amino(phenyl)methylene]succinate) and 3 (3- alkoxycarbonyl-2-phenyl-pyrrolin-5-one) may be obtained as presented in the Scheme

1.4. The dianion of the succinate ester is much more stable, less prone to self- condensation and thus is preferred to a more accessible mono-anion.

O R O 1 R1 COOR d e – OR H N OR 2 HN _ OR 59% OR 60%

2 O 3 O O

a 58% c

O O O

b COOR OR a – OR R1 OR _ OR COOR O O

Scheme 1.4. Synthetic approaches to the intermediates 2 and 3. a) LDA, THF, –70°C; b) R1COCl, Et3N; c) AcONH4, AcOH, 100°C, 16 hrs; d) R1CN, ZnCl2, THF, –70°C, 2 hrs.

H O Table 1.3. DPP compounds prepared by aromatic nitrile – succinate N

R1 71 ester condensations. R2 N O H

# R T, °Ca Addition time, hrsb Yield, %

1 4-Tol 97-99 3¾ 23.4 2 3-Cl-Ph 89-91 2 56.8 3 4-Cl-Ph 88-91 2 39.5 4 4-MeOOC-Ph 89-91 2¼ 6.6 5 3-CN-Ph 89-91 2¼ 77.5 6 4-CN-Ph 90-91 2½ 80.0 7 1-Np 95-97 2 4.5 8 2-Np 96-97 1¾ 24.2

9 3-CF3-C6H4- 105-110 3 56.8 10 6-AmO-3-Py 105-110 3 65.0

11 4-CF3-C6H4- 105-110 2.5 44.9

12 4-CN-C6H4-C6H4- 105-110 3 36.7

13 4-C6H4-C6H4- 105-110 2.5 10.0

14 4-t-Bu-C6H4- 105-110 2 55.2

15 3,4-Me2-C6H3- 105-110 2 52.4 16 2-Fu 90 1 17.9

17 4-Me-C6H4- 105-110 2 41.8 18 3-thienyl 85 1 42.0

19 3,5-Cl2-C6H3- 85 1 70.4

20 4-Me2N–C6H4– 120 2 3.7

a Reaction temperature; b Total addition time of diethyl succinate for a 0.2 mole reaction of aromatic nitrile.

N O O N OEt Base + OEt N N N O O

Scheme 1.5. Stobbe condensation of a Schiff base with alkyl succinate.

Cl Cl Cl

O N O a b N Cl NH Cl Cl Cl N O N O

Cl Cl

Scheme 1.6. Preparation of DPP from benzylaniline.

a) t-BuOK, DMF, –10°C; b) DDQ, dichlorobenzene.

Table 1.4. DPP compounds prepared by step-wise condensations.76,77,81

H O N

R2 N O H

# R1 R2 Yield, % M.p. °C UV λmax (lg ε)

1 Me Me 14 >250 380 (4.16), 392 (4.2)a

2 Me Pr 24 >250 382 (4.1), 398 (4.1)a

3 Me Ph – – 433 (4.0), 450 (4.0), 550 (2.5)b

b 4 Ph 4-Ph–S–C6H4– 74 – 380 (4.1), 480 (4.5), 518 (4.6)

b 5 Ph 4-CN–C6H4– 80 – 271, 310, 485, 520

b 6 Ph 4-Cl–C6H4– 74 – 471 (4.4), 510 (4.5)

b 7 C11H23 C11H23 – 247–250 385 (4.0), 402 (4.0)

8 Me i-Pr 11 – 383 (4.1), 397 (4.2)a

b 9 Me C11H23 10 – 382 (4.2), 398 (4.2)

c 10 Pr 4-Cl–C6H4– 52 – 440 (4.0), 460 (4.0), 630 (2.8)

b 11 4-Ph–C6H4– 4-Cl–C6H4– 70 336 (4.2), 488 (4.5), 524 (4.6)

b 12 Ph 2-Cl–C6H4– – – 459 (4.2)

b 13 Ph 4-Me–C6H4– 49 – 307, 312, 472, 507

b 14 4-Cl–C6H4– 3-CN–C6H4– 87 – 288, 308, 450, 480, 513

b 15 Ph 3-CN–C6H4– 61 – 289, 305, 445, 478, 512

b 16 4-CN–C6H4– 3-CN–C6H4– 27 – 280, 310, 490, 521

Table 1.4. (Continued).

# R1 R2 Yield, % M.p. °C UV λmax (lg ε)

b 17 4-pyridyl 4-Cl–C6H4– 76 – 268, 308, 483, 517

18 Ph 1-Np 28 – 470 (4.2), 493 (4.2)b

19 Ph 6-MeO-1-Np 2 – 500 (4.4)b

b 20 Ph 2-Me–C6H4– – – 453 (4.3), 481 (4.3)

b 21 Ph 2,5-Me2–C6H4– 6 – 456 (4.3), 482 (4.3)

b 22 Ph PhCH2– 17 – 381 (4.1), 466 (3.1)

b 23 Ph PhCH2CH2– – – 438 (4.2), 459 (4.2)

24 Ph 9-phenanthryl 39 – 472 (4.2), 492 (4.2)b

25 Ph i-Pr 31 306–307 437 (4.1)d

26 Ph Cyclohexyl 11 subl. > 400 439 (4.2), 460 (4.2)d

d 27 Ph Ph2CH 29 295–296 443 (4.25), 466 (4.2)

28 Ph 2-Norbornyl 15 subl. > 400 430 (4.2), 449 (4.2)d

29 Ph cyclohex-3-en-yl 42 358–360 –

a) in methanol; b) in NMP; c) in DMF; d) in DMSO

N O

OAlk Base + COOAlk COOAlk OAlk H N COOAlk 2 HN HN O O O O OAlk OAlk

2 3

O COOAlk Base COOAlk COOAlk HN HN HN + HN NH O O NH O NH2 O

Scheme 1.7. Mechanism of DPP formation from benzonitrile and succinate.

Another synthetic route to DPP, although strategically similar to the one elaborated by Iqbal, is condensation of succindiamide 4 with N,N′-dimethylbenzamide diethylacetal 5 depicted in Scheme 1.8.82 In this reaction conversion to DPP proceeds only partially (generally ~30%) and although the byproduct, succinylbenzamidine 6, may be converted to DPP with t-BuOK (total yield 60%), this reaction offers no apparent overall advantage.

EtO NMe2 H O O O EtO N

NH2 Base N NMe + + 2 NH2 N NMe2 N O H O O

4 5 1 6

Scheme 1.8. Condensation of succindiamide with N,N′-dimethylbenzamide diethylacetal 5.

A different approach, devised by Langlas47, comprises condensation of furo[3,4- c]furane-1,4-diones, e.g. 3,6-diphenyl-1H,4H-furo[3,4-c]furan-1,4-dione 7 with aromatic amines in the presence of DCC in trifluoroacetic acid (Scheme 1.9). This approach gives the only known example of unsymmetrically substituted N,N′-diaryl DPP (Scheme 1.10), though in very low yield (0.3% from 7). Aliphatic amines are much more reluctant to react with 7, and do so only in the presence of 4-(dimethylamino)pyridine to form only traces of the corresponding DPPs (Scheme 1.11).

R %

O O H 46 a O O + H2N N N R R R 4-Me 35

O O 2,3-Me2 56 7

4-t-Bu 52

Scheme 1.9. DCC-driven condensation of furo[3,4-c]furane-1,4-dione with amine.

(a) DCC, CF3COOH, CHCl3, r.t., 3 days.

O O O

CH3 a b O O N O N N CH3 15% 2%

CH3 O O O

Scheme 1.10. The sole example of an unsymmetrical N,N′-diaryl DPP.

a) 1 eq. aniline, DCC, CF3COOH, CHCl3, r.t., 3 days; b) 4-tert-butylaniline, DCC,

CF3COOH, r.t., 3 days.

O O CH3 CH3 NH2 a O O + H3C N N CH3 H3C

H3C H3C O O

Scheme 1.11. The sole example of aliphatic amine reaction with furo[3,4-c]furane-1,4-

dione. (a) DCC, DMAP, CF3COOH (yields ‘trace’ amounts).

The 3,6-diphenyl-1H,4H-furo[3,4-c]furan-1,4-dione83 7 can be prepared

(Scheme 1.12), in turn, (a) from benzoylacetic acid ester via its oxidative dimerization84 to a mixture of meso- and racemic forms of 2,3-dibenzoylsuccinic acid diesters with subsequent thermal cyclization85; and (b) from 1,6-diaryl-1,3,4,6-tetraoxohexane86 via its oxidation with dinitrogen trioxide (N2O3) to 1,6-diaryl-2,5-bis(diazo)-1,3,4,6-

tetraoxohexane87 and subsequent thermal decomposition of the latter neat (25–50% yield) or in toluene (59% yield). The laborious preparation procedures of the lactone 7 make

this seemingly straightforward route to N-aryl disubstituted DPPs somewhat troublesome.

Moreover, the harsh condition of the oxidative coupling in the route (a) and vigorous thermolysis step in the route (b) exclude the presence of sensitive functional groups. In fact, there are merely five known 3,6-diphenylfuro[3,4-c]furan-1,4-diones (and 3,7-

dioxabicyclo[3.3.0]octa-4,8-dien-2,6-diones in general) in the Beilstein database: 4,4′- dimethyl- [76695-70-0]; 4,4′-dichloro- [76695-71-1]; 4,4′-dimethoxy- [76695-69-7]; and the aforementioned 2,2′-dimethyl- 8.

O O O O O O O RO O a RO b O d N2 c O O O 13% O OR OR 56% 59% N2 51% O O O O O O O

Scheme 1.12. Routes to 3,6-diphenyl-furo[3,4-c]furan-1,4-dione.

a) Na/Et2O, I2; b) thermolysis 300°C; c) N2O3, CH2Cl2, –30°C; d) thermolysis 100°C.

1.3.2. Chemical Properties

3,6-Diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-diones undergo several types of reactions. The Scheme 1.13 shows the points of attack by nucleophiles and electrophiles.

Specific examples of each of such attack follow and are also illustrated on Scheme 1.14.

E+

NFG

HN NH

EFG

X – Nu

Scheme 1.13. Reactive sites of DPP. X – halogen; EFG – electrophilic functional group;

NFG – nucleophilic functional group

O OR Alk H N N O N O S

H3C O O O N O S N N Alk S S H P RO Cl P RO O S S

AlkOTs H C 3 O Cl H N H Br H O N N O O Cl Br2 Cl Cl 2

O N O O H Cl N N HO O H H Br S

O POCl3 Oleum CH2OH2SO4 H N CH2OH O N N OPOCl O O 2

N H O O N O N H S CH2OH O OH Scheme 1.14. Various reactions of DPP.

Alkylation of one or both of the amidic nitrogens is usually performed in high- boiling solvents — nitrobenzene (at 200…205°C) or DMF (at 140°C) — with alkyl (Me or Et) 4-toluenesulfonate and potassium carbonate to yield 40–68% of N,N′-dialkyl DPP.

Reaction of n-butyl bromide with preformed DPP sodium salt for 20 hrs at 60°C, then 2 hrs at 100°C gave a mixture of mono- and dibutylated products (m.p. 250–252 and 123–

124°C correspondingly). Benzylation was achieved in 47% yield88,89. It is noteworthy that only N-alkylation takes place, with no O-alkylated products corresponding to the lactim tautomeric structure ever reported.74 The N,N′-dialkylated DPP is strikingly more soluble compared to its dihydro precursor: at 25°C one liter of DMF dissolves a mere 110 mg of DPP vs. 3300 mg of DPP-Me.66 The solubility of mono-alkylated DPPs is in between these two.

OAlk HO O O O Alk N N AlkOTs NH X NH N + N N HN K2CO3 N N Alk Alk OAlk OH O O O

Scheme 1.15. N-Alkylation of DPP.

Acylation of the amidic nitrogens is performed essentially in the same way as alkylation, in this case using acyl chlorides. For example, benzoylation of DPP proceeded in 38% yield. 1,4-Diketo-3,6-diaryl-2,5-bis(alkyloxycarbonyl)pyrrolo[3,4-c]pyrroles may also be obtained in high (20–90%) yields by action of di-alkyl dicarbonates in presence of

DMAP (Scheme 1.16).90,91 Thus acylated DPPs represent a very elegant invention of so-

called “soluble pigments”, for they easily lose their N-substituents (and, thus, solubility)

upon thermolysis and revert back to highly insoluble 2H,5H-dyhydro DPP pigments

(Scheme 1.17).92,93,94

CH3 H3C H C 3 O H O O N O O O N + a t-Bu t-Bu O O O N O H N O

O O

CH3 H C CH 3 3

Scheme 1.16. Acylation of DPP with di-tert-butyl dicarbonate.

a) DMAP, DMF or THF, r.t., 4-12 hrs.

CH2

CH3

CH3 O O H2C O

O H C O O a 3 HN O H C N NH + 3 N CH3 O O O CH3 O O O CH CH3 2

CH3

CH2 b

Scheme 1.17. Acylation of DPP with di(2-methyl-3-buten-2-yl) dicarbonate and

subsequent decomposition.

a) DMAP, DMF or THF, r.t.; b) 126°C, neat.

Arylation of the amidic nitrogens hitherto was possible only indirectly and led

mostly to mono-arylated DPPs – see Scheme 1.10.47

Hydroxymethylation of the amidic nitrogens has been performed with para-

formaldehyde in concentrated (90–96%) aqueous sulfuric acid at 20…30°C to give 2,5-

bis(hydroxymethyl)-3,6-diarylpyrrolo[3,4-c]pyrrole-1,4-diones in unspecified yield. If

the temperature was allowed to rise above 40°C, considerable concurrent sulfonation

ensued. The diol obtained may be reacted in the same pot to condense with two additional equivalents of the same or different DPP, resulting in DPP trimers (Scheme 1.18). The diol also reacts with quinacridone (5,12-dihydroquino[2,3-b]acridine-7,14-dione) and aniline – Scheme 1.19.95,96,97,98

R1 R R R

O O O O NH NH a N O OH b N HO N N HN N N O O O HN O O

R1 R R R R 1

Scheme 1.18. N-Hydroxymethylation of DPP. a) (CH2O)n, H2SO4, 20…30°C; b) R1–

DPP, H2SO4.

O O O N a N N OMe b NH MeO HO OH NH N N N O O O

Scheme 1.19. Reactions of N,N′-bis(hydroxymethyl) DPP. a) MeOH, 4-TsOH, Δx; b)

aniline, DMF, Δx.

The result of the free halogen action on the parent DPP depends on the nature of

the halogen. Chlorine adds to the bicyclicDPP to give tetrachloro adduct (Scheme 1.20).

Bromine in CCl4 gives a mixture of two brominated products and unreacted starting

material. Bromination with gaseous bromine by the method of Buckles and Wheeler99,100 gives much better results, producing 3,6-di-(4-bromophenyl)-pyrrolo-[3,4-c]-pyrrole-1,4- dione 9 in 87% yield (Scheme 1.21), but the final product still contains some monobromo compound as evidenced by elemental analysis (Br found/calc 33.5/35.8)101 and

our own experience on alkylation of such a product. Bromination of DPP with N-

bromosuccinimide in sulfuric acid has been reported. Action of iodine and fluorine has

not been documented to our knowledge.

O Cl Cl OMe O O O Cl NH Br Cl MeOH 2 NH 2 NH NH HN HN HN HN O O O Cl Cl MeO Cl O

9 1

Scheme 1.20 DPP bromination and chlorination products.

Br Br OMe O O O Br O O Br2 MeOH NH NH NH NH NH + + HN HN HN HN HN O O O MeO Br Br Br O Br Br O MeO Br

Scheme 1.21. Mechanism of DPP bromination.

DPP undergoes smooth sulfonation in fuming sulfuric acid (oleum). There is information, scattered over claim sections of several patents, claiming control over degree of DPP sulfonation by varying sulfuric acid concentration, temperature, and reaction time. Thus, degree of sulfonation has been claimed to vary from zero to four

SO3H groups per DPP molecule. The disulfonic acid 10 is soluble in water and alcohols

(first three homologs) and forms salts with alkali earth metals, used as rheology- improving (viscosity reducing) additives to other DPP pigment compositions.

H H O O N N fuming HO3S

H2SO4 SO3H N N O 10 O H H

Scheme 1.22. Sulfonation of DPP.

The nucleophilic attack of the Lawesson’s reagent (or P4S10 in HMPA) at the

amidic carbonyl results in an overall replacement of the carbonyl oxygen with sulphur.102

The resulting 1,4-dithio-DPP is much more susceptible to carbanion attacks and affords overall dicyanomethylation, unattainable directly.

H H O S N N a

N N O S H H b NC SEt N N CN c

N N EtS EtS H

Scheme 1.23. Replacement of O in –NH–C=O by C. a) Lawesson’s reagent;

b) EtI, K2CO3, Me2CO; c) H2C(CN)2, THF, r.t.

Phosphorylation of the amidic carbonyl with POCl3 results in an unusually

stable (its chloride salt form has m.p. 235–237°C with decomposition) adduct (Scheme

1.24), which may subsequently be reacted with such N-nucleophiles as various aromatic

amines to provide 1-mono-arylimino derivatives of DPP or with sodium sulfide to give mono-thioketo derivative.103

O O O a + H b NH N NH - HN HN Cl HN O OH N R

Scheme 1.24. Replacement of O in –NH–C=O by N. a) POCl3;

b) aniline, R= H, 4-Cl, 4-CN, 4-NO2, 2-Br, 2-COOAlk.

One or both oxygens in the DPP heterocycle may be replaced by cyanimino groups under action of bis(trimethylsilyl)carbodiimide in presence of titanium (IV) tetrachloride (Scheme 1.25).104

CH3 H3C CH3 Si O N CN N CN HN N TiCl4 HN HN + + NH C NH NH O N O NC N

Si H3C CH3 CH3

Scheme 1.25. Replacement of O in –NH–C=O by N–CN (cyanoimination of DPP).

Aromatic nucleophilic substitution of chlorine at para position in the phenyl ring of several unsymmetrical N-unsubstituted 2H,5H-dihydro DPPs has been performed with pyrrolidine and dimethylamine.105 In the same patent there is also an example of successful nitration, followed by substitution of chlorine in the other phenyl ring (Scheme

1.26). The resulting 3-(4-nitrophenyl)-6-(4-dimethylaminophenyl)-2,5-dihydropyrrolo-

[3,4-c]pyrrole-1,4-dione is dark blue-violet pigment.

H C Cl 3 Cl N CH3

O O O

a HN NH b HN NH HN NH

O O O

+ + - N - N O O O O

Scheme 1.26. Nitration and aromatic nucleophilic substitution of chlorine in DPP.

a) KNO3, H2SO4, 0–5°C, 1 hr.; b) Me2NH, NMP, 180°C, 10 hrs in autoclave.

Aromatic nucleophilic substitution of bromine at the para position in the phenyl

ring(s) of parent 2H,5H-dihydro DPPs has been conducted with dimethylamine,

pyrrolidine, piperidine, and morpholine in 45–70% yields.106 The compounds obtained

have been claimed to be useful photoconductive substances.

H H N N O O Br N NMP, pyrrolidine

N Br O O N N H H

Scheme 1.27. Aromatic nucleophilic substitution of bromine in DPP.

Palladium-mediated coupling was shown to be an efficient transformation of bromine in the phenyl rings of DPP to carboxy (Scheme 1.28), alkoxycarboxy (Scheme

1.29) or alkylaminocarboxy (Scheme 1.30) functionality in 78–93% yields.107

H H N N O O Br PdCl2, CO, PPh3 HOOC

(HCOO)2Ca, NMP COOH Br O O N N H H

Scheme 1.28. Pd coupling of Br-DPP with CO in presence of calcium formate.

H H N N O O Br PdCl2, CO, PPh3 CH3OOC

MeOH, Et3N, NMP COOCH Br O 3 O N N H H

Scheme 1.29. Pd coupling of Br-DPP with CO in presence of MeOH.

H H N N O O Br PdCl2, CO, PPh3 BuNHOC

n-BuNH2, NMP CONHBu Br O O N N H H

Scheme 1.30. Pd coupling of Br-DPP with CO in presence of butylamine.

Appropriately di-functionalized DPPs can form oligomers,60 polymers,51,52,54 and dendrimers,59,63 as exemplified on Schemes 1.31 and 1.32.

CH2— N O TfO TfO OTf + + Bu Sn SnBu OTf 3 S 3 R O N N

a R =

N O

S S O N x y O SO n

51 Scheme 1.31. Stille coupling polymerization of DPP. (a) Pd(PPh3)4, LiCl, 1,4-dioxane.

C H C H 6 13 6 13 C6H13 O N (HO)2B B(OH)2 + Br + Br Br Br H13C6 H C N O 13 6 H13C6

C6H13 C6H13 C6H13 O N C6H13 x 1-x H13C6 H13C6 N O H13C6 H13C6 n

52 Scheme 1.32. Suzuki coupling polymerization of DPP. (a) Pd(PPh3)4, K2CO3, toluene.

1.3.3. Physical Properties

The DPP and DPP-Me molecules, the crystalline molecular geometry structures of which are shown in Fig. 1.3, both belong to point group Ci, and are not entirely planar.

The phenyl rings are twisted in the same direction, out of the plane of the planar

heterocyclic system by 7°±1° in DPP and by 31°±1° in DPP-Me. The DPP molecules

align in nearly the same molecular plane and parallel to each other due to intermolecular

hydrogen bonding. By contrast, the DPP-Me molecules are arranged in a herringbone

fashion along the stacking c axis. In DPP, the interatomic distances H(1)–H(N) and H(5)–

O(1) are 2.19 and 2.31 Å, respectively. These distances are considerably shorter than the

sum of the van der Waals radii of the corresponding atoms: 2.4 and 2.6 Å, respectively.

The C(2)–C(4) distance consequently becomes shorter than the value expected for the carbon–carbon single bond of 1.54 Å. The observed value of 1.455 Å is much shorter, for example, than the value of 1.496 Å found in biphenyl. The present bond shortening is presumably caused by well-delocalized π-electrons in the pyrrolo[3,4-c]pyrrole-1,4-dione

chromophore which gives some double-bond character to the C(2)–C(4) bond. A similar bond shortening also operates in DPP-Me. The C(2)–C(4) bond is 1.460 Å. This is slightly longer than in DPP but still much shorter than in biphenyl.108

Figure 1.3. X-ray molecular structure of DPP and DPP-Me.108

In DPP there are chains of intermolecular hydrogen bonds along the ·110Ò

direction in the molecular plane between the N–H group of one molecule and the

carbonyl oxygen of the neighboring one. There are van der Waals contacts along the c axis. In DPP-Me, by contrast, molecules face each other alternately, forming a dimeric stacking structure. The overlap of the two molecules along the stacking axis is shown in

Fig. 1.5. There is significant overlap between the heterocyclic ring systems in DPP causing π-π interactions (interplanar spacing 3.36 Å). On the other hand, no such overlap is observed in DPP-Me. Instead, there are π-π contacts between the heterocyclic ring and

the phenyl ring. On the other hand, DPP-Me molecules are mainly associated together by

van der Waals forces. Because of this, the molecular arrangement is very different in DPP compared with DPP-Me.

(b) (a)

Figure 1.4. Crystal structure of (a) DPP – triclinic and (b) DPP-Me – orthorombic.108

(a) (b)

Figure 1.5. Comparison of overlap of the two molecules along the stacking axis:

(a) DPP and (b) DPP-Me.108

The astonishingly low solubility (see Table 1.5) of the parent 2H,5H-dihydro-

DPPs in most organic solvents (except, e.g. sulfuric acid and trifluoromethanesulfonic acid) due to intermolecular hydrogen bonding and π-π interactions, results in impurity trapping during their precipitation immediately upon their formation in the preparation reactions. These trapped impurities are very difficult to remove by recrystallization. Even such high boiling solvents as nitrobenzene and dimethylacetamide at their reflux

temperatures dissolve so little DPP (ca. several hundred milligrams per half a liter) that recrystallization is an impractical technique for preparative scale. A mixture of diphenyl ether and biphenyl has been used109 to change the morphology of DPP polycrystalline powder for dyeing purposes, yet it still isn’t suitable even for gram-scale recrystallizations. The purification method of choice is salt formation/dissolution with strong base (NaOH, KOH) in water — polar organic solvent mixtures (DMSO, DMF), followed by re-acidification/precipitation.110 This strategy has also been reflected in step-

wise (methanol followed by water followed by acetic acid) protolysis of DPP dianion

during work-up after the preparation reaction. In this case the formation of undesirable

by-products and degradation products can be suppressed greatly with a simultaneous

improvement in the coloristic properties.111 In our own experiments we found that, since

N-alkylation of DPP almost always forms a mixture of mono- and di-alkylated products,

the purification method of choice is chromatography, performed after the N-alkylation

step.

The thermal stability of parent 2H,5H-dihydro-DPPs is largely determined by

the lattice crystal structure and thus depends strongly on the crystal forms and

polymorphic modifications. Conditioning of crude DPP in high boiling solvents allows,

apart from partial purification, a change of the morphology of the polycrystalline

particles and an enhancement of the thermal stability. For example, boiling crude DPP in

a mixture of diphenyl ether and biphenyl at 245–260°C for 0.5…1 hr, results in the change of the DSC thermogram as depicted on Fig. 1.6 and particle morphology change

to flake-like or platelet (determined by scanning electron microscopy and powder X-ray, not shown).109

Table 1.5. Solubilities of various DPPs in mol·liter–1.46

H H t-Bu H N O N O N O t-Bu t-Bu t-Bu t-Bu O N O N O N H H H t-Bu [54660-00-3] [84632-59-7] [107680-82-0]

–7 –6 –1 CHCl3: 8.4·10 CHCl3: 6.4·10 CHCl3: 1·10

PhMe: 8·10–8 PhMe: 1.6·10–7 PhMe: 2.8·10–4

MeOH: 2.3·10–6 MeOH: 2.6·10–6 MeOH: 5.8·10–4

DMSO: 3.7·10–3 DMSO: 1.3·10–3 DMSO: 3.9·10–4

t-Bu O O N N N O t-Bu t-Bu

t-Bu t-Bu N O N O N O t-Bu [96159-17-0] [107680-85-3] [107711-05-7]

–1 –1 –2 CHCl3: 1.3·10 CHCl3: 5.0·10 CHCl3: 5.1·10

PhMe: 2.1·10–3 PhMe: 4.4·10–2 PhMe: 4.2·10–3

MeOH: 3.1·10–4 MeOH: 7.8·10–4 MeOH: 3.6·10–5

DMSO: 2.7·10–3 DMSO: 1.3·10–3 DMSO: 2.4·10–5

Figure 1.6. DSC of (a) crude DPP and (b) conditioned DPP.

The thermal stability of 2H,5H-dihydro-DPPs in general may be characterized as

“high”, and that of several specific representatives — as not less than “amazing”. On Fig.

1.7 there are TGA traces of DPP and Br-DPP, showing that these two substances sublime, without melting or decomposition (as determined by DSC, not shown), continuously up to 452° and 500°C (!) correspondingly. The continuous endothermic signal indicates the heat absorption due to sublimation. The observed residue after sublimation is due to contaminants, trapped by DPP during its preparation reaction. Re- sublimed samples of DPP give no residue after sublimation and their TGA traces reach zero.

100 ––––––– As-1-130.000 ––––––– AS-2-11 Br-DPP.sav

65.43%

60 90.23% Weight (%) Weight 40 DPP

452.86°C 33.38%

20 499.97°C 10.41%

Br–DPP

0 100 200 300 400 500 600 700 Temperature (°C) Universal V4.0C TA

Figure 1.7. TGA traces of crude DPP (red) and Br-DPP (blue).

The UV-Vis optical absorption properties of parent 2H,5H-dihydro-DPPs are

noticeably different in the crystalline form and in solution (Fig. 1.8). In the crystalline

state the absorptive (and reflective) properties are determined not only by the molecular

structure, but to a great extent by the crystal structure and intermolecular interactions of

the chromophore molecules. Thus, DPP in very dilute DMF solution is yellow, and in solid state — red. This bathochromic shift of absorption maximum from solution to the

solid state is quite common for other classes of dyes and pigments. What is interesting in

case of DPP is that the introduction of substituents into the phenyl rings causes

bathochromic shifts in their solution spectra, yet in the solid state there is a hypsochromic

shift upon substitution of the meta- position (Table 1.6).67

1 NMP solution Solid DPP

Solid Absorbance Solution

0 400 450 500 550 600 650

Wavelength, nm Figure 1.8. Visible spectrum of DPP as NMP solution (absorption) vs. solid (reflection).67

Table 1.6. Influence of 3,6-substituents and crystal structure on absorptive properties of

DPP.66

H O N

N R O H

a b R Color λmax in NMP λmax solid state Δλmax ε

H yellow-red 504 538 34 33,000

4-Br blue-red 515 555 18 35,000

3-Cl orange 512 528 16 27,000

3-CF3 orange- 509 518 9 21,000

yellow

4-NMe2 violet-blue 554 603 51 81,500

a Determined as shade of plasticized PVC pigmented with 0.2% of corresponding DPP.

b Reflectance.

Fluorescence of parent 2H,5H-dihydro-DPPs is also very much phase- dependent. Bathochromic shifts of fluorescence emission maxima up to 40 nm are observed in solid samples. Vapor-deposited polycrystalline films and vapor-grown single crystals of DPP have emission maximum shifted even further bathochromically, at 634 nm.112 This probably can be explained by more regular network of hydrogen bonds in

single crystal relative to polycrystalline powder, formed by abrupt precipitation. Intensity

of solid-state fluorescence of polycrystalline DPP increases with the size of the grains.

No data on solid-state fluorescence quantum yield have been found in the reviewed literature.

Fig. 1.9. Intensity of solid state fluorescence increases with the size of the DPP grain.112

NMR properties in terms of 1H and 13C chemical shifts of some known DPPs are

presented on Fig. 1.10 and 1.11. Table 1.10 shows solvent and temperature dependence

of amidic proton chemical shift.113 The proton chemical shift increases with polarity of

solvent and appears around 11 pm, which is characteristic of a hydrogen-bonded proton.

As the temperature increases, the thermal motion disrupts hydrogen bonding and δ moves upfield.

Table 1.7. Fluorescence data for several DPPs in chloroform.46

R O N

N R O 1 R

a # R1 R λmax(lg ε) DMSO λmax λEm Δλ ΦF

290sh, 304(4.11), 318sh, 496 509 13 – 1 H H 450sh, 471(4.38), 505(4.50)

Ref.47 496 509 13 –

311br(4.13), 449sh, 3 3,5-(t-Bu)2 H 500 513 13 0.63 473(4.40). 507(4.55)

4 2-Me H 430sh, 480sh, 453(4.29) 448 518 70 0.64

5 2-Me Me 442(4.20) 439 489 50 0.95

291(4.12), 486(4.24) 474 523 49 0.54 6 H Me Ref.45 470 520 50 0.9±0.05

7 4-t-Bu Me 307br(4.21) 485 528 43 0.53

8 3,5-(t-Bu)2 Me 305(4.11), 475sh, 489(4.27) 484 525 41 0.56

9 4-Me Allyl Ref.45 468 521 53 0.97

a) Stokes’ shift in chloroform.

Table 1.8. Fluorescence in solution and solid state of some N-phenyl substituted DPPs.47

R N N R

# R λmax(lg ε) DMSO λEm CHCl3 Δλ λEm Solid

1 H 484(3.941), 464(3.923) 520, 555sh 36 530sh, 580

2 4-Me 488(4.357), 470(4.330) 521, 549sh 51 526sh, 581

3 2,3-Me2 492(5.722), 469(5.698) 524, 555sh 55 563

4 4-t-Bu 489(5.471), 467(5.447) 519, 553sh 52 546, 570sh

Figure 1.9. Absorption (S), fluorescence in CHCl3 solution (F), and solid state

fluorescence (SF) of DPP and 4-t-Bu-DPP.47

Table 1.9. Fluorescence quantum yields of some N-mono-aryl and N,N′-diaryl substituted DPPs.114

R3 R1 N O

O N

R 4

Solvent # R1 R2 R3 R4 λmax(lg ε) λEm ΦF

488.5(5.42), a 47 1 H H 4-t-Bu-C6H4–H 521, 561sh Ref. 466.7(5.39) CHCl3

D 2 H H 4-NO2-C6H4–4-NO2 470(4.33) 516 0.02

3 H H H H 498(4.23) D 520 0.44

4 H H H 4-MeO 472(4.17), 500(4.20) D 521 0.03

D 5 H H H 4-CF3 467(4.17), 493(4.19) 519 0.43

6 Cl Br H 4-MeO 481(4.43), 511(4.46) D 532 0.09

7 H H Me H 468(4.11) DCM 521 0.43

8 H H Me 4-MeO 471(4.19) DCM 522 0.23

DCM 9 H H Me 4-CF3 470(4.11) 520 0.48

10 Cl Br Me 4-MeO 487(4.18) DCM 533 0.15

DCM 11 H H PhCH2– H 468(4.11) 519 0.42

DCM 12 H H PhCH2– 4-MeO 470(4.27) 522 0.27

DCM 13 H H PhCH2– 4-CF3 468(4.12) 518 0.43

Solvent # R1 R2 R3 R4 λmax(lg ε) λEm ΦF

DCM 14 Cl Br PhCH2– 4-MeO 488(4.28) 532 0.05

D 15 H H 4-NO2-C6H4– 4-MeO 470(4.26) 515 0

D 16 Cl Br 4-NO2-C6H4– 4-MeO 483(4.43) 513 0

17 H H Ph H 488(4.27) DCB 524 0.29

18 H H Ph 4-MeO 490(4.24) DCB 520 0.12

DCB 19 H H Ph 4-CF3 486(4.29) 520 0.32

a λEm solid = 506, 543sh

Solvent: D = DMSO; DCM = CH2Cl2; DCB = 1,2-Cl2C6H4.

CH H3C 3

O CH3 O N 1 H NMR (CDC13): 7.75 (d, 4H);

N 7.48-7.50 (m, 6H); 1.40 (s, 18H). O H C 3 O

H C CH 3 3

Figure 1.10. 1H and 13C NMR spectra of 3,6-diphenyl-2,5-diallylpyrrolo[3.4-c]pyrrole-

1,4-dione.45

Figure 1.11. 1H and 13C Assigned chemical shifts for

3,6-diphenyl-2,5-diallylpyrrolo[3.4-c]pyrrole-1,4-dione. 45

Table 1.10. 1H Chemical shifts of the NH group of DPP in different solvents and at

various temperatures.113

Chemical shift, δ, ppm Solvent 25°C 50°C 75°C 100°C

Dioxane-d8 10.88 10.58 10.38 10.20

DMF-d7 11.08 10.81 10.57 10.43

DMSO-d6 11.35 11.14 10.96 10.76

Table 1.11. 13C Chemical shifts of DPP, its mono- and di-anion.

δ, ppm in DMSO-d6 #Ca DPP DPP– DPP2–

1 162.4 172.0 172.5

1a 110.7 117.9 118.7 O 3 NH

3 144.0 155.3 155.4 HN 1 1a O 4 4 127.7 134.4 135.6 5

6 5 128.9 128.0 127.6 7 6 127.7 127.6 127.5

7 131.7 128.5 127.6

a Numeration is shown on the right for carbon atom referencing only and not for

nomenclature purposes.

1.4. Newly prepared DPP dyes

The optical and fluorescent properties of DPP dyes, specifically their high photostability, large Stokes’ shift, small size of the molecule, and high fluorescence quantum yields prompted us to explore them in the SMS, as well as design and preparation of a series of new fluorophores of this class. The small size (both in molecular weight and bulkiness) of the DPP chromophore, we believe, would allow it to permeate more easily through the cell membrane. The targeted new DPP fluorophores should posses the following properties (separately or combined):

• red-shifted (with respect to DPP) absorption and fluorescence

• high (compared to 2,5-dihydro DPPs) solubility

• functionalization: for covalent labeling and for SMS in polymers

The need for the red shift in absorption and fluorescence transferred into the

following structural proposals. From the electronic color theory of organic dyes115,116 and molecular orbital theory117 it follows that absorbance wavelength bathochromically shifts

as the length and degree of the conjugation chain in the dye increase (within certain

limits). Electron-donating and electron-accepting groups, introduced at certain places of

the conjugation chain and capable of participating in the delocalization of electronic

density (i.e. groups with mesomeric effect), also shift the absorbance bathochromically.1

Thus, structure-wise, bathochromically shifted absorption and emission can be achieved

by either extension of the length of a conjugated chain or by increase of the degree of conjugation, or both. Application of these considerations to the DPP structure yields structural modifications, one of which is depicted in the Scheme 1.33.

The property aim of solubility is based on the crystal structure and established trends in solubilities of known DPP compounds, reviewed above. Rupture of the intermolecular hydrogen bonding network by introduction of substituents at one or better both lactam nitrogens, greatly increases the solubility of DPP compounds. Thus, we planned to make N-alkylated DPPs and compare their solubilities. The single-molecule spectroscopy requires relatively low solubility of the fluorophores. Simple dimethylation increases the solubility of DPP in chloroform seven orders of magnitude. Thus, even the solubility of mono-N-substituted DPPs might suffice for our purposes. We have prepared and compared solubilities of N,N′-dimethyl, dipropyl, dihexyl, didecyl, didodecyl, and dibenzyl DPPs and some of their mono- analogs. It turned out that for purification by recrystallization the C3 chain was the most convenient one: methylated DPPs with

solubility in chloroform of order ~0.1 M required much higher solvent volumes for

recrystallization, while the didodecyl derivative (~0.8 M chloroform solubility) was too

soluble. For this reason, most of the compounds prepared here have propyl substituents

on the lactam nitrogen(s).

1 These statements are very simplified generalizations and should not be considered as all-cases rules.

The functionalization was first planned to be achieved via pre-functionalization of nitriles used as starting materials in the condensation with dialkyl succinate. However, this approach proved to be futile, at least in our hands and required exploration of post- factum functionalization (after the formation of the DPP core), which is discussed below.

The parent DPP 1 and Br-DPP 9 are used in commercial pigment mixtures, manufactured by Ciba-Geigy Specialty Chemicals Co., and, according to acknowledge-

ments in several papers,52,60 can be obtained from the European Research Division, in

Switzerland, as individual compounds as a favor. However, our communications with the

U.S. division of Ciba-Geigy (Ann Cardillo, CE) resulted in no fruitful answer. Thus, we

had to learn and prepare the DPP precursors ourselves. Down this road we also learned

about nitrile functionality tolerance in the DPP preparation reaction.

Alk N O N Alk Alk N O N Alk

Alk = C6H13: AS-2-69b

Alk N N O Alk Alk N N

O Decrease Energy *

Alk Shift Bathochromic Alk = Et: AS-3-18 Δ π−π Conjugation Length Increase Length Conjugation

Alk N Alk N O

O N Alk N Alk Alk = Bu: AS-3-22

Scheme 1.33. Design of red-shifted DPP chromophores.

The sterically hindered di-alkyl succinates have been prepared (Scheme 1.34):

di-iso-propyl succinate [924-88-9] 11 — by esterification of succinic acid with isopropyl alcohol in presence of sulfuric acid; and di-tert-amyl succinate [77106-39-9] 12 — by

trans-esterification of diethyl succinate with tert-amyl alcohol (2-methyl-2-butanol) in the

presence of lithium alkoxide.118

O O O O

O a OH OEt b O O OH OEt O 11 O O O O 12

Scheme 1.34. Preparation of sterically hindered di-alkyl succinates.

(a) i-PrOH, H2SO4 (cat.), az. H2O removal. (b) t-AmOH, Li, fractional EtOH removal.

The following aromatic nitriles have been prepared: 4-fluorobenzonitrile 13, 4-

bromobenzonitrile 14, 4-methoxybenzonitrile 15 — from corresponding aldehydes by a corrected procedure of Wang119 (Scheme 1.35); 4-(pyrrolidin-1-yl)benzonitrile 16, 4-(N-

n-hexylamino)benzonitrile 17, and 4-(N,N-di-n-hexylamino)benzonitrile 18 — from 4-

aminobenzonitrile; 4-(N,N-di-n-butylamino)benzonitrile 19 — from 4-fluorobenzonitrile

(Scheme 1.36). The Wang procedure was corrected (and thus improved), taking two equivalents of phthalic anhydride instead of the specified one equivalent, since two equivalents of water are formed overall – one at the oxime formation step, and another – at the dehydration of the latter, resulting in much better conversion and higher yield.

Also, a higher boiling point solvent, NMP, was used instead of acetonitrile, resulting in shorter reaction times.

OH H N CHO CN a b

– H2O – H2O R R R

Scheme 1.35. Preparation of 4-R-benzonitriles: R= F (13), Br (14), OMe (15).

(a) NH2OH, Et3N. (b) phthalic anhydride, NMP.

CN CN CN CN CN a b c d

N N F NH2 HN Alk Alk Alk

16 17 R=C6H13: 18, Bu: 19

Scheme 1.36. Preparation of 4-aminobenzonitriles. (a) 1,4-diiodobutane, i-Pr2NEt, NMP.

(b) C6H13Br, HMPA. (c) C6H13Br, K2CO3, NMP. (d) Bu2N, Py, HMPA.

The results of the reactions between prepared or commercial nitriles with dialkyl

succinates are summarized in the Table 1.12. Firstly, only four out of eleven nitriles gave

any tangible amount of DPP upon condensation. Secondly, for those nitriles, from which

we did obtain corresponding DPPs, we were consistently obtaining much lower yields

than specified in the patents or literature. Because of this, we performed some optimization of the DPP preparation reaction, based on the considerations from the literature, patents, and our own.

Table 1.12. DPP preparation reactions. R # N O H N O H 1 OAlk a R + 4-F 20 OAlk R 4-CN 21 O N R O H 4-Me 22

Alk in Lit. Yield Ref. or R Yield, % (AlkO)2Suc2 [CAS RN] if N/A Et 23 11.5 71, 59 46 H i-Pr 48 65 71 t-Am 56 82 71 Et 12 no prep. F i-Pr 66 [84632-57-5]69,120 Br Et 0 81.7 72, 43 121 MeO Et traces 6.6 46,51 Ph Et traces 10 71 71 Bu2N Me, Et, i-Pr, t-Am traces Me2N: 3.7

C6H13NH Et 0 – 71 (C6H13)2N Et 0 Me2N: 3.7 122 NO2 Et 0 [186967-03-3]

4-C8H17–C6H4– Et 0 –

4-C11H23O–C6H4– Et 0 –

N no prep. Et 0 from nitrile 4-CN t-Am 35 80 71 Me Et, i-Pr, 12 23.4 71

t-Am 38 48.6 71

Since self-condensation of dialkyl succinates slows down with the bulkiness of the alkyl moiety, we switched in our DPP preparations from commercially available diethyl succinate to bulkier di-iso-propyl succinate and even more bulkier di-tert-amyl succinate. In going through this succinate ester series, the yield of the parent DPP 1 increased in our hands from 23 to 48 to 56%. The rate of the succinate ester addition to the reaction mixture also affects the yield of DPP dramatically. For example, in our hands

4-Tol-DPP 22 wouldn’t form in tangible amounts if the 4-toluonitrile had been added to the reaction mixture from the onset at once. However, with a syringe pump metered addition rate of 3.4 ml/hr and use of (t-AmO)2Suc2, we obtained the latter in 38% yield.

Considering the stoichiometry of the condensation reaction, one mole of a succinate ester

reacts with two moles of nitrile. Yet if the succinate ester self-condenses in a parallel

competing reaction, the initial 1:2 ester to nitrile ratio will leave us with some unreacted

nitrile, which was indeed detected by gas chromatography. This consideration led us to

using an excess of succinate ester: the optimal ratio of di-tert-amyl succinate ester to

nitrile was determined to be 0.57 and 0.8:1.16≈0.7 for di-iso-propyl succinate.

Another important improvement, contributing to the purity of the crude DPP is

the rate of hydrolysis. Since the dianion DPP2– is quite soluble in the reaction medium

(especially compared to solubility of DPP), abrupt acidification of the latter causes, to our presumption, occlusion, trapping, and co-precipitation of impurities, which was proved by TGA analysis of the crude DPP (Fig. 1.12): as opposed to clean sample (recrystallized or sublimed), crude DPP does not sublime completely at the end of TGA run, and leaves

a residue. Slow and step-wise increase of the acidity of the reaction mixture allows slower crystallization of DPP, resulting in less sublimation / TGA residue.

––––––– As-1-130.000 ––––––– As-1-130.001 100 ––––––– AS-2-70.000

Weight (%) Weight crude, fast ppt 40 34.52%

crude, slow ppt 20 17.53% 10.07% recr. DMAc 0 0 100 200 300 400 500 600 Temperature (°C) Universal V4.0C TA

Figure 1.12. Effect of fast vs. slow work-up on the purity of crude DPP.

Concurrently with our learning about the above DPP preparation reaction itself,

we also learned that our initial plans to prepare functionalized and derivatized DPPs by

functionalizing or derivatizing the starting nitriles were substantially compromised by our

inability to reproduce many reactions, claimed in the patents, as well as to force other

nitriles to give corresponding DPPs. The “nitrile pre-functionalization” approach became

even less attractive after our several failures to either alkylate certain DPPs or to separate discrete products from such alkylation.

1.4.1. Alkylation of DPPs

The preferred method for alkylation of DPP in the patents is the action of methyl or ethyl tosylates (4-tolylsulfonates) and potassium carbonate on DPP in DMF or nitrobenzene at their boiling point. Alkylation with alkyl halides, especially with their higher homologs (e.g. butyl), gives much less satisfactory yields, as well as considerable amount of mono-alkylated DPP in addition to di-alkylated one. We tried to reproduce the reported alkylation reactions, tried new or modified procedures, and prepared several new mono- and di- alkylated DPPs. The methyl and ethyl tosylates are commercially available, and iso-propyl tosylate 36 was prepared by a modified procedure of

Waldron.123

Methylation and ethylation with corresponding alkyl tosylate proceeds almost

quantitatively and, given enough alkylating reagent and time, the di-alkylated products

are formed in predominance. Separation of mono-methylated DPP, however, was

reported recently by Zambuonis.124 The alkylation reaction (Scheme 1.37) proceeds via

several steps (Scheme 1.38). Firstly, (1) DPP goes from crystalline state to solution,

where (2) it gets deprotonated to DPP– or (3) DPP2–, depending on the strength of the

base used, and either of those anions then (4) attacks alkyl halide (or tosylate), producing

mono-alkylated DPP, followed by (5) subsequent attack onto second equivalent of R–X,

producing di-alkylated DPP. This scheme suggests that the overall rate of the alkylation

may depend on the DPP concentration (low solubility may be a limiting factor) in a given

solvent, solubility of the base in the same, and the stationary concentration of the DPP

anion(s).

If the base used for deprotonation is weak (e.g. K2CO3), and the solvent is nitrobenzene, the stationary concentration of DPP anions is low, and the reaction rate is limited by the solubility of DPP and DPP– in the solvent. In this case high reaction

temperature (200°C) is necessary to complete the reaction. If the base is strong, the

equilibrium may be shifted towards almost complete (t-BuOK) or complete (NaH, KH) deprotonation of the dissolved DPP, and limited now only by the solubility of DPP– and

DPP2– in the solvent, thus permitting lower reaction temperatures to be employed — in

the range of 60 to 120°C. The solubility of the ionized DPP species is highest in mixtures

of polar organic solvents and water (e.g. DMF:water ≈ 5:1), but our alkylation attempts in

such solvent mixture never went to completion (even with addition of phase transfer

reagents, like Aliquat 336) or gave satisfactory yields. Employment of dry DMF, DMAc,

or DMSO, however, allowed complete consumption of the starting material. The

discussion of the possible reasons for such behavior is found below, where basic

degradation of DPP is discussed. The reactivity of the alkylating agent is also an

important factor — alkylating agents with higher reactivity provide not only shorter

reaction times, but also somewhat higher yields, presumably because shorter reaction

times reduce the degradation of DPP, caused by the base. For this reason, addition of

catalytic amounts of potassium iodide to the reaction mixture whenever using alkyl

bromides as alkylating reagents, is advisable. The change in the structure of the

alkylating agent, which hinders SN2 mechanism and favors SN1 mechanism, seems to

prevent N-alkylation of DPP under the conditions studied – compare results for

compounds 26–28 in the Table 1.13 with the other entries. Some additional examples of

N-alkylation reactions, as well as effect of base (Cs2CO3 instead of K2CO3) are discussed below with the functionalized DPPs.

The solubility of the prepared alkyl-DPPs follows the following trend: within the pair, di-alkylated DPP has generally higher solubility than mono- analog. As the length of the alkyl chain increases, however, the solubility of the mono-alkyl DPP in DMF becomes comparable to that of di-alkyl one. The solubility in non-polar solvents (hexane, toluene) or in solvents of low polarity (ether) also increases with the length of the alkyl chain. For example, neither DPP-Me, nor DPP-Et is considerably soluble in ether. The next homolog, however, DPP-Pr, dissolves in THF, and DPP-Hx is soluble in ether, while

DPP-C12 dissolves in neat hexane at room temperature.

O O O DMAc HNHN + RX+ Base RRNN+ R NNH or NMP O O O

Scheme 1.37. Alkylation of DPP. X = OTs for R = Me, Et, i-Pr;

X = I, Br, or Cl for R = i-Pr, n-Pr, C6H13, C10H21, C12H25, PhCH2.

O O O O Base Base – – – HNHN HNHN NNH NN (1) (2) (3) O O O O

crystal solution

RX RX (4) (4)

O O O Base – RX R NNH R NN RRNN (5) O O O

Scheme 1.38. Step-wise alkylation process of DPP.

1.4.2. Action of bases on DPP

The study of base action on DPP was prompted by two reasons: relatively low and sometimes inconsistent yields in alkylation reactions, and contemplation of the possibility to obtain bis-lactone 7 upon hydrolysis of DPP. The bis-lactone 7 has been used in the (limited) preparation of N-arylated DPPs and easy access to it would be quite handy.

Table 1.13. N-Alkylated DPPs.

R1 N O

O N R2

# R1 R2 X Base Yield M.p. °C Comments

[96159-17-0] 1H δ: 7.89 85 230…231 23* Me Me OTs K2CO3 (m, 4H), 7.54 (m, 6H), lit. 68 lit. 236-238 3.34 (s, 6H).

[138369-76-3] 1H δ:

9.05 (s, 1H), 8.31 (m, 24‡ Me H OCOMe Et3N 23 + 46% of 23 2H), 7.92 (m, 2H), 7.54

(m, 6H), 3.41 (s, 3H).

[96159-13-6] Recr. 3.6 / 25* Et Et OTs K2CO3 40 229–230 40 DMF or 100 BuOH

26 i-Pr i-Pr 27 267 [849767-61-9] I, OTs NaH 27 i-Pr H 46 330

only traces of product 28 t-Bu t-Bu I K2CO3 0 – detected

29 n-Bu n-Bu MeONa, 18 123–124 [96159-01-2] Br 30 n-Bu H t-BuOK 32 250–252 [96159-00-1]

31† 2-EtHx 2-EtHx – – – – [132029-46-0]

32 Pr Pr 62 189 Sol. toluene I t-BuOK 33 Pr H 30 276 Sol. CHCl3

34 C6H13 C6H13 8 134 Sol. toluene Br, I t-BuOK 35 C6H13 H 16 252 Sol. toluene

36† C10H21 C10H21 Br t-BuOK 42 117 [132029-47-1]

Sol. Et2O, toluene, 37 C12H25 C12H25 47 114 I t-BuOK hexane

38 C12H25 H Sol. toluene

[96159-02-3] 1H δ: 7.75

(d, 4H), 7.49-7.43 (m,

39* PhCH2 PhCH2 51 290–292 6H), 7.30 (t, 4H), 7.24 Br, Cl t-BuOK (t, 2H), 7.19 (d, 4H),

4.99 (s, 4H).

40 PhCH2 H 32 344

209 [96159-07-8] 41* Allyl Allyl 42 Br t-BuOK lit. 216–217

42 Allyl H 46 309

* known compound † These compounds have been claimed in the patent,125 but no preparation, characterization or properties have been reported. ‡ Preparation and separation is given in the patent.124

A dilute (1 mg/ml) solution of DPP in a 5M solution of sodium hydroxide in mixture of DMAc and water (5:1) has a deep crimson color, characteristic of DPP2– anion. However, left at room temperature for five days, the color fades out completely, indicating degradation of the chromophore. It is entirely reasonable to suggest then that in any reaction of DPP, employing base, there will be a competing reaction of degradation.

More interestingly, even fully alkylated DPP, for example, DPP-Pr 32, subjected to the same conditions, outlives parent DPP not for much longer.

DPP 1 was subjected to a basic aqueous hydrolysis with potassium hydroxide in a closed high-pressure steel reactor at 120°C to give the sole aromatic degradation product — benzoic acid (Scheme 1.39). Lower reaction temperature does not affect any changes on DPP within 24 hrs. The mechanism of such transformation is clear up to the sym-dibenzoylsuccinic acid decarboxylation, whereafter some oxidant needed, probably atmospheric oxygen left in the vessel, or oxidation by means of disproportionation can be proposed, to get the detected product.

The fact that di-alkyl DPPs undergo degradation was also noticed during the study of the optical properties of DPP-Me 23 at Ciba-Geigy by Jin Mizuguchi. He found that upon sodium hydroxide titration of DPP-Me solution in DMSO–H2O, the color of the

solution disappears, creating a new UV peak at 380 nm, though reversibly: titration back

with hydrochloric acid restores the absorption peak at 475 nm. Mizuguchi proposed

(based on 1H NMR evidence, not supplied though) a Michael addition of hydroxide ion to

the 1-position of the DPP ring, which could be also viewed as 4-position of α,β-

unsaturated amide. The Michael addition product is proposed to be stabilized by an

intramolecular hydrogen bond, as depicted in Scheme 1.40. The quantitiy of sodium hydroxide, required to form such adduct, is “slightly more than theoretical amount, and further addition of sodium hydroxide brings a complete discoloration of DPP-Me solution

(decomposition) due to subsequent reactions.”113 It is possible that these unspecified

subsequent reactions are lactam hydrolysis, similar to what we observed in case of basic

degradation of DPP 1. No further study in this direction has been conducted.

H H OOH N O O N a Ph

– O Ph N O N HO H H

H H H N O N O N O Ph Ph Ph Ph Ph Ph – – HOOC OOC HO OOC HN– NH2 O

O O H H – O O COO N N Ph Ph Ph Ph Ph – – 2CO Ph COO 2 Ph Ph – – O OOC OOC HN O H2N

Scheme 1.39. High temperature basic degradation of DPP. (a) KOH, EtOH, H2O, 120°C,

6 hrs.

O – N OH N O O H+ N H – O O N

Scheme 1.40. Michael addition of OH– to DPP-Me.113

1.4.3. Halogenation of DPPs

Since the “nitrile pre-functionalization” approach to the functionalized DPPs has

“kicked the bucket”, at least in our hands, we moved on to the post-functionalization, that

is modification of the DPP after the core bicyclic system has been formed. The

bromination of the parent DPP by gas-phase bromine is known. When repeated as

prescribed,101 however, it gave a material, which upon per-alkylation (with methyl tosylate or propyl iodide, NaH, DMAc) produced a mixture of at least three compounds,

which have very close Rf on TLC. Apparently, they correspond to three precursor DPPs:

unreacted parent DPP 1, as well as mono- and di-brominated ones. Bromination,

conducted with neat liquid bromine or with a solution in CCl4 did not give cleaner

product — in full accordance with the patent’s statement. However, when we applied the

neat liquid bromine treatment after the gas-bromination, the resulting product gave upon

alkylation a good yield of Br-DPP-Pr, though still requiring purification by

chromatography. In our search for a cleaner bromination method, we investigated the bromination of N,N′-dialkylated DPPs and found a peculiar trend (Scheme 1.41): DPP-

Me 23 undergoes bromination both in gas phase and in solution smoothly and cleanly,

while the higher N,N′-dialkylated homologs are destroyed by bromine even under careful exclusion of light. For example, Pr-DPP 32 gives, upon treatment with bromine solution in chloroform even at –30°C, a black tarry product, deficient of the DPP chromophore system and its characteristic absorption and fluorescent properties. This degradation of the DPP chromophore was observed for R = propyl and hexyl, while R = Me gives brominated product (Br-DPP-Me) even in higher overall yield, than if DPP is brominated first and methylated second. Alkylation of brominated DPP was conducted similar to that for the parent DPP — see details in the Experimental section.

Since the intended destiny of the bromine in brominated DPPs was to use them as reactive sites for subsequent Pd-catalyzed couplings, and since it is known that iodo-

arenes generally give better yields in such reactions, we also aimed to introduce iodine

atoms into phenyl rings of the DPP. Iodine monochloride reacts neither with parent DPP

1, nor with alkylated DPPs — neither in gaseous phase, nor in solution, nor neat and thus

the 4,4′-diiododiphenyl DPPs are not available by direct iodination with ICl.1 However,

we found that the bromine atoms at the 4-positions of the benzene rings in both parent

and alkylated DPPs can be easily replaced by iodine using an excess of KI/CuI in

DMAc126 at 180°C, thus producing the desired iodo-DPP indirectly.

The cyano group is often considered as a pseudo-halogen. For example,

dicyanogen (CN)2 has both physical and chemical properties similar to that of a diatomic

halogen. Likewise, reaction of the same aryl bromides 43 and 44 with KCN/CuCN gives the nitrile derivatives 47 (X=CN, Table 1.14) in good yield (Scheme 1.43), which was inaccessible by the attempted alkylation of the 3,6-bis(4-cyanophenyl) DPP 21. In

contrast to successful bromine-to-iodine halogen exchange, treatment of Br-DPP-Pr 44

with cesium fluoride127 did not give any substitution by fluorine. The obtained

halogenated DPPs are summarized in the Table 1.14.

1.4.4. Substitution of halogen by amine in DPPs

Aromatic nucleophilic substitution of chlorine in N-unsubstituted 2H,5H-

dihydro DPPs is known. We extended the scope of this reaction to N-substituted DPPs

with bromine atoms in the aromatic system. Reactions of Br-DPP-Me 43 and Br-DPP-Pr

44 with pyrrolidine, piperidine, dibutyl- and dihexylamines have been performed to give

the corresponding diaminated DPPs (Scheme 1.44), which are deep red in color and give

beautiful crimson fluorescence. The cyclic amines give generally higher yields than

aliphatic di-n-alkylamines. The latter also give a mixture of mono- and di-substituted

products (e.g. 51 and 52, 53 and 54), in the mono-product dominates. And if the reaction

with the cyclic amines can be pushed to a completion (no mono-product on TLC) by

1 Electrochemical iodination and iodination with other «I+»-genic species was not explored.

longer reaction time and higher temperature, the same strategy in the case of di-n-

alkylamines isn’t as fruitful, though addition of HMPA and use of large excess of the

amine increases the yield somewhat.

Pr Pr O a N N O X Br Br N O O N Pr 20 Pr 8

62% bb37% H H N O N O a Br 94% Br N N O O H H 1

46% c 85% c

O N O a N Br 96% Br N O N O 19 7 Scheme 1.41. Bromination reactions of DPPs.

(a) Br2 (gas), (b) PrI, K2CO3, DMAc, (c) MeOTs, K2CO3, DMAc

Table 1.14. Halogenated DPPs – yield indicated is that from parent compound (R=X=H).

R N O X X O N R

# X R Yield % M.p. °C

9 Br H 94 >450

43 Br Me 46 (from 9) 96 (from 23) >350

44 Br Pr 37 247

45 I Me 37 dec. 355

46 I Pr 68 260.5

47 CN Pr 83 227

O O N a N Br I Br I 43 O N 45 O N

Pr Pr O N a N O Br I Br I N 44 O 46 O N Pr Pr Scheme 1.42. Indirect approach to iodo-DPPs. (a) KI, CuI, DMAc, 180°C.

Pr Pr O N a N O Br NC Br CN N 44 O 47 O N Pr Pr

Scheme 1.43. Conversion of Br-DPP-Pr to CN-DPP-Pr. (a) CuCN, KCN, DMAc, 180°C.

Table 1.15. Aminated DPPs.

R R O O N R1 N T °C N Br R + Amine R 1 Br Solvent 1 N N O N O R1 R R

# R Amine T °C Solvent R1 Yield %

48 Me NH 140 DMAc, HMPA (CH ) 74 2 4

49 Me NH 140 DMAc, HMPA (CH2)5 67

50 Pr NH 200† DMAc (CH ) 32 2 4

51 n-Bu 8

Pr n-Bu2N 180 DMAc mono- 52 11 aminated

53 n-C6H13 11

Pr n-C6H13N 180 DMAc, HMPA mono- 54 21 aminated

† in a sealed vessel

1.4.5. Extension of the conjugated chain of DPP

To make longer-wavelength absorbing and emitting DPPs, we extended the conjugation length of the fluorophore. For this purpose we explored and applied some

Pd-catalyzed coupling reactions. Firstly, we used the Jeffery modification128 of Heck

coupling of DPP aryl halides (bromides and iodides) with various styrenes. Trial

reactions with commercially available 4-tert-butylstyrene and 4-acetoxystyrene went well

with both methylated iodo- 43 and bromo- 45 DPPs (Scheme 1.44).

X R N O N a O + O N O N X R = tert-Bu: 55

X = Br or I AcO: 56 R

Scheme 1.44. Heck coupling between X-DPP-Me with 4-tert-butyl- and 4-acetoxy-

styrenes. (a) Pd(OAc)2, TDA-1, K2CO3, DMAc.

Encouraged by this fact, we prepared several other styrenes: 4′-

(diethylamino)styrene 57, 1,4-divinylbenzene 58, and 4'-(2-[(4-

dibutylamino)phenyl]vinyl)styrene 59 from the corresponding aldehydes by Wittig

reaction (Scheme 1.45).

CHO CHO Bu aab ; N Bu N N CHO Et Et Et Et 59 57 58 d

c HOOC OHC CHO COOH

+ – Scheme 1.45. Preparation of styrenes. (a) Ph3P Me I , t-BuOK, DMSO. (b) Pd(OAc)2, (o-

Tol)3Ph, Et3N, DMAc. (c) CH2(COOH)2, Py. (d) quinoline, Ph2O, Cu, hydroquinone, Δx.

The prepared styrenes 57 and 59 were coupled with Br-DPP-Pr 44 (Scheme 1.46 and 1.47 correspondingly) to give DPPs 60 and 61 with long conjugation chains and amine donors, as well as propyl solubilizing groups. The DPP 61 was rather difficult to purify from numerous colored by-products and it was obtained eventually in a state of purity, which gave a good NMR and mass spectra, but eluted at least two additional minor components on HPLC. The physical and optical properties of these compounds are discussed below.

The Negishi coupling between Br-DPP-Pr 44 and either thien-2-yl zinc chloride129 62 or 5-(4-(N,N-di-n-hexylamino)phenyl)-thien-2-yl zinc chloride 63 gave

another pair of DPPs with an extended conjugation chain — 64 and 65 (Schemes 1.49

and 1.50).

O N Et Br + 2 N Br Et O N 44a 57

N N O

O N N

Scheme 1.46. Pd-catalyzed coupling between styrene 57 and Br-DPP-Pr 44.

N O Bu Br 44 N 59 + Bu Br O N

Bu N Bu N O

O N Bu N Bu 61

Scheme 1.47. Pd-catylized coupling between styrene 59 and Br-DPP-Pr 44.

C6H13 Br N S H S C6H13 a b

C6H13 Mg S N Li S Br C6H13 c c

C6H13 Zn N S Zn S Cl C6H13 62 Cl 63

N O Br 44 62 + Cl–Zn Br S O N

N O

S S O N

Scheme 1.48. Preparation of thien-2-yl zinc reagents 62 and 63. Negishi coupling

between Br-DPP-Pr 44 and thien-2-yl zinc chloride 62.

(a) Mg, THF. (b) n-BuLi, THF. (c) ZnCl2. (d) Pd(PPh3)4, THF.

N O C6H13 Br 44 N 63 + Cl–Zn S Br C6H13 O N

H13C6 N O N S C6H13 H13C6 N N S O C6H13

Scheme 1.49. Negishi coupling between Br-DPP-Pr 44 and 5-(4-(N,N-di-n-

hexylamino)phenyl)-thien-2-yl zinc chloride 63. (a) Pd(PPh3)4, THF.

1.4.6. DPPs with hydrophilic solubilizing groups

Apart from the known 4,4′-bis(hydroxysulfonyl) DPP derivative 10, there are no

water-soluble DPPs, to our knowledge. Even for compound 10 there was no preparation

procedure or characterization given in the original paper.68 Since biological applications require dye solubility in polar solvents and aqueous buffers, we reproduced the sulfonation of DPP (Scheme 1.22), worked out the separation of the product from the

reaction mixture, characterized compound 10, and prepared several other derivatives,

containing the hydroxysulfonyl group. For this purpose we alkylated the DPP’s lactam’s

nitrogen(s) with either 1,3-propanesultone or 1,4-butanesultone (Scheme 1.50). The DPP

with two sulfonic acid groups 66 was not isolated in crystalline form due to difficulties in its crystallization — it is not soluble in non-polar solvents and does not crystallize from polar ones (ethanol, water, acetone), once dissolved in them. DPPs with only one hydroxysulfonyl group 67 and 68 did not tend to crystallize on cooling either — their purification was achieved either by chromatography (68) or by acid re-precipitation (67).

O O S O O O O O O + HN N S S N N S O b O O OH O HO OH O O HN NH 66 67

O Pr O Pr a N S O N O OO b O N O N H HO O 33 S 68 O

Scheme 1.50. Introducing sulfonic acid group into DPP structure.

(a) t-BuOK, PrI, DMAc. (b) sultone, t-BuOK, DMAc.

1.4.7. Hydroxy-functionalized DPPs

For the purpose of incorporation of DPP fluorophore into a polymer chain

(performed by another researcher — Wonhee Jeong), we needed bis- and mono-alcohol

functionalized complimentary DPP pairs. The complementation here means similarity in the position and reactivity of the hydroxy groups in the bis- and mono- functionalized compounds. Based on our alkylation experience of various DPPs, we contemplated introduction of the alcohol functionality via alkylation of DPP with ω-halo-α-alcohols

(Scheme 1.51). However, this approach, when implemented with alkali metal alkoxides or hydrides as bases, either failed completely or worked on a sole example of already mono-alkylated DPP only. As a possible explanation for these failures, we might speculate that strong base t-BuOK with its conjugate acid t-BuOH correspondingly

weaker than any n-CnH2n+1OH primary aliphatic alcohol, abstracts the hydroxyl’s proton

in competition with deprotonation of lactam’s nitrogen. The formed alkoxide, although

sufficiently strong to reversibly deprotonate DPP in turn, might undergo intra- or inter-

molecular side-reactions, as well as to add to the DPP bicyclic system by Michael type

(similar to hydroxy anion, cf. Scheme 1.40), which would be impossible for the tert-

butoxide ion due to its bulkiness.

O O O t-BuOK, DMAc HN NH HO (CH2)N N (CH2)OH+ HN N (CH2)OH X OH n n n O (CH2) O O n X = Cl, Br, I

Scheme 1.51. Proposed direct introduction of alcohol functionality into DPP by

alkylation with ω-halo-α-alcohols.

O O a N NH N N (CH2)OH8

O O 40 69

Scheme 1.52. The sole successful example of DPP alkylation with ω-halo-α-alcohol in

presence of t-BuOK. (a) t-BuOK, DMAc, 8-bromo-1-octanol.

Since the di-allylation of 3,6-bis(4-methylphenyl)pyrrolo[3,4-c]pyrrole-1,4- dione (Tol-DPP 22) has been reported to proceed in 75% yield45, we pursued a

hydroboration approach (Scheme 1.53) to alcohol functionality via mono- and diallyl

DPPs (41 and 42), supported by two literature examples130 of terminal alkene

hydroborations in the presence of a cyclic amide. The attempted hydroboration reactions with both BH3•THF and 9-BBN did not yield any alcohols on the DPP-containing

substrate. Moreover, those reagents destroyed the fluorophore’s core when the reaction

was forced by heating. This destructive action of boranes is definitely characteristic of the

DPP heterocycle, rather than the allyl termini, for the two above boranes similarly

destroy on warming a THF or CH2Cl2 solution of Pr-DPP 32. No analysis of the

degradation products has been conducted and the criteria for the DPP core destruction

were the loss of the DPP’s UV-Vis absorption and fluorescence.

O O O a HNHN NN + HNN

O O O 1 41 42 bb

HO O O

NN HNN

O OH O OH

Scheme 1.53. Attempted hydroboration route to alcohol-functionalized DPPs.

(a) t-BuOK, allyl bromide. (b) BH3•THF or 9-BBN.

The first success in alcohol functionalization of DPP has been achieved with a method, discovered and reported by Lu and Twieg:131 copper-catalyzed coupling of aryl

iodides with sec-amines. We coupled I-DPP-Pr 46 with 2-(ethylamino)ethanol in presence of copper and copper (I) iodide in DMAc (Scheme 1.54). No DMAE, essential as a solvent for the reaction success in other cases, can be used when this specific amine, similar to DMAE, is used as a reactant. Although the mono-coupling product has been detected on TLC while monitoring the progress of the reaction, the major product was 70.

Perhaps, one could use 1:1 ratio of I-DPP-Pr and 2-(ethylamino)ethanol to get predominantly mono-coupling product, but the optical absorbtive properties of them

would be different due to electron-donor properties of the amino group being introduced.

Therefore, we moved back to the utilization of lactam nitrogen(s) alkylation reaction, where the optical properties of the mono- and di- products do not differ significantly and where they could sometimes be obtained in equal amounts, relatively easily separated and the mono-product then can be re-alkylated with a different reagent.

I OH HO N O O a N + HN N N O N I O N OH 46 70

Scheme 1.54. Cu/CuI-cat. coupling of 2-(ethylamino)ethanol with I-DPP-Pr 46.

(a) Cu, CuI, DMAc, 80°C.

Revisiting the alkylation methodology, we resorted to a protection-deprotection approach to introduce the terminal alcohol functionalized groups (Scheme 1.55). Thus, we have prepared the THP- protected C6 alcohols 71–73 (Scheme 1.56) and benzoate

ester-protected C6 alcohol 74 (Scheme 1.57) with a halide (or tosylate) on the other terminus of the alkyl chain.

O O O X OPG HN NH + (CH ) HN N (CH ) OPG + PGO (CH )N N (CH ) OPG 2 n 2 n 2 n 2 n O X = Cl, Br, I O O

Deprotection

O O O

HO (CH ) NN Alk HN N (CH )OH HO (CH )N N (CH )OH 2 n 2 n 2 n 2 n O O O

Scheme 1.55. Alkylation approach to alcohol-functionalized DPPs with protection-

deprotection of the hydroxy group. PG = Protecting Group.

OH a O O b O O

OH OH OTs 71 e c

OH O O O O d I X X X = Br 72 73 = I 73

Scheme 1.56. Preparation of alkylating reagents with alcohol functionality, protected with a THP protecting group. (a) dihydropyran (DHP), THF, Dowex 50WX8-100. (b) 4-

toluenesulfonyl chloride (TsCl), Py, Et2O. (c) HBr 48% aq. or HI 55% aq. or P2I4, CS2.

(d) DHP, THF, Dowex 50WX2-100. (e) P2I4, CS2.

The tosylate 71 reacted with DPP 1 in presence of t-BuOK in DMAc very slowly up to 180°C: even after four days most of the starting material was not consumed.

When the reaction temperature was raised to 200°C, the starting material was consumed in three hours, however, these harsh conditions seemed to destroy the THP protection and side-reactions ensued, for very little target material was obtained. The same reaction performed with bromide 72 went under more mild conditions and that with iodide 73 — was complete in twelve hours at 40°C (Scheme 1.57). The THP-protected DPP diol 74

turned out to require rather strong conditions for the removal of the THP groups.

Reaction with pyridinium 4-tolenesulfonate (PPTS) in ethanol resulted in no deprotection

at all. Acetic acid removed the THP group only partially. Trifluoroacetic acid, used in

amount of 10% with respect to the substrate, did give the target compound, though as

mixture with at least three other ones with very similar retention factors. The deprotection was confirmed by 13C NMR spectrum of the mixture, but our separation attempts did not

yield pure individual compounds. Thus, we resorted to the alkylation with a benzoate

ester-protected iodo-alcohol 75.

The iodide 75 alkylated DPP 1 at 60°C in four hours to give 76, with a little of

mono-alkylated product 77. Same reaction, repeated with mono-propyl and mono-

dodecyl DPPs 33 and 38 yielded corresponding precursors of DPP mono-alcohols 78 and

79. The removal of the benzoate ester moiety by classical methods132 also encountered

with some difficulties. Potassium hydroxide 1% solution in ethanol did perform

deprotection, but very slowly and the deprotection was never complete at room temperature.

O O O O O a O HN NH + N N X O O 71– 73 O O 1 74

HO (CH2)6 N N (CH2)6 OH

Scheme 1.57. Reaction of THP- protected alcohols 71–73 with DPP.

O O OH a O b O

OH OH 75 I

Scheme 1.58. Preparation of alkylating reagent 75 with alcohol functionality, protected

with a benzoate ester. (a) PhCOCl, Et3N, THF. (b) KI, H3PO4, PPA.

When pushed by heating, however, the dye underwent degradation (presumably of the same nature as described in Chapter 1.4.2) in parallel with deprotection. A much milder deprotecting reagent, potassium cyanide in methanol,133 successfully tested firstly on the simplest ester — ethyl benzoate, gave a clean deprotection of 77 to 81 on a ten milligram scale, but scaling up this reaction met with the same problem of the dye degradation. Eventually we found that excess (over recommended catalytic amount) of

titanium (IV) tetra-iso-propoxide in methanol at temperatures quite a bit over those

usually employed for such a reaction (180°C vs. usual r.t. to 60°C) and for much longer

reaction time (days vs. usual hours) gave almost quantitative deprotection of 76 to 80

without any side reactions.

O O H a N N O HN NH +

O N O O N 1 76 77

+ O O

O O O

O b c

I 75 O O

HO (CH ) N N (CH ) OH 2 6 2 6 HN N (CH2)6 OH

O O 80 81

Scheme 1.59. Reaction of benzoate ester protected alcohol 74 with DPP and subsequent

removal of the protecting group. (a) t-BuOK, DMAc. (b) MeOH, Ti(i-PrO)4, 180°C. (c)

KCN, MeOH.

H O R N O N O O a +

O N I O N R R = C3H7 33 74 78 = C H 79 O 12 25 38 R b N O O

O N R = C3H7 82 OH = C12H25 83

Scheme 1.60. Preparation of complimentary mono-alcohol functionalized DPPs. (a) t-

BuOK, DMAc. (b) MeOH, Ti(i-PrO)4, 180°C.

Having prepared a set of haloalcohols — with free and protected hydroxy groups

— we continued to experiment and search for reaction conditions, which would allow direct alkylations of DPPs with ω-halo-α-alcohols. The obvious first parameter we varied

was the base — one, which wouldn’t degrade the DPP core and be sufficiently strong to

at least partially deprotonate lactam’s nitrogen(s). Eventually we found that cesium

carbonate does the magic we searched for. On Scheme 1.61 we depict the di- and mono-

alkylation reactions of DPP 1 and mono-Pr-DPP 33 with 10-iodo(bromo)decan-1-ol,

prepared similar to the corresponding C6 analogs on Scheme 1.56. The reaction runs best

in strictly anhydrous polar aprotic solvents (NMP, DMAc) and can be complete even at

room temperature, though with DPP 1 itself it will take many days, and only six hours at

160°C. The alkylation of mono-Pr-DPP 33 with 10-iodo-1-decanol and cesium carbonate,

due to its higher solubility, will be complete in 6 hours at much lower temperature of

60°C.

The preparation of ω-halo-α-alcohols was deemed necessary, despite the fact that ω-bromo-α-alkanols are commercially available, for the GC-MS and NMR analysis

of the commercial samples on our shelves proved their inferiority. For example, GC-MS

analysis of 8-bromo-1-octanol from Aldrich’s batch #09915TG revealed at least three

major components therein, with 8-bromo-1-octanol accounting to only 56% of total

integration area. 13C NMR of the same sample, in turn, gave twenty-four discrete signals, also suggesting a mixture of three individual compounds (24/8).

O O HO (CH2)10 X HNN Pr HO (CH2)10 NNPr Cs2CO3 O O

33 84

O O HO (CH2)10 X HNHN HO (CH2)10 NN (CH2)10 OH Cs2CO3 O O 1 85

Scheme 1.61. Direct alkylation of DPPs with ω-halo-α-alcohols in presence of Cs2CO3.

Mono-alkylation of DPP 1 is possible with 1.2 eq. of I-C10-OH. Cs2CO3 alone in

DMAc does degrade Pr-DPP on prolonged heating too.

1.4.8. DPPs with a cysteine-reactive maleimide moiety

Maleimide group has been widely utilized in the bioconjugate chemistry as a

specific binding site for the cysteine residue of proteins. In part it is due to low

abundance of the Cys unit, in part – due to high binding specificity of maleimide to the

latter. Following the successful application of maleimide-containing Nile Red dye (see

Chapter 2) for imaging of GroEL chaperonine, we were about to exploit the DPP

chemistry learned to prepare maleimide-functionalized DPPs. The furan-protected

maleimide alkylating reagent 86 (see its preparation in Chapter 2) has been used to di-

alkylate DPP 1, followed by thermal removal of furan (retro Diels-Alder reaction) to give

bis-functionalized DPP derivative 88 (Scheme 1.62).

An attempt was made to prepare mono-functionalized maleimide derivative 90,

possessing a sulfonic acid group for improved solubility in polar medium. A one-pot,

mixed, stepwise alkylation of DPP 1 was performed — firstly with 1,3-propanesultone,

followed by 2-(6-iodohexyl)-3a,4,7,7a-tetrahydro-4,7-epoxy-1H-isoindole-1,3(2H)-dione

86 to give the product 89 (Scheme 1.63). However, the subsequent retro Diels-Alder

reaction, performed on pure isolated substrate 89 in neat refluxing xylene, gave an

intractable brown tar, missing both the characteristics of DPP dye (by absorption

13 spectroscopy) and C signal of maleimide C3-carbon altogether. Apparently, the

100

maleimide subunit does not tolerate the simultaneous presence of strongly acidic group in the same molecule, at least at elevated temperature involved, and, we speculate, had polymerized right upon deprotection.

O O I HNHN + O N

O O 1 86 a O

O N N O O

O O N N O

b 87 O O

N N O O

O O N N

88 O

Scheme 1.62. Bis(maleimide) DPP derivative. (a) t-BuOK, DMAc, 100°C. (b) xylene, Δx.

101

1 2 O O O O I HNHN + S + O N O O O 1 86 a

O N b N O O tar

N 89 O SO H b 3 O

N 2 3 1 N O O

90 O N

SO3H

Scheme 1.63. An attempt towards maleimide mono-functionalized DPP 90. (a) t-BuOK,

DMAc, 100°C. (b) xylene, Δx.

102

1.4.9. N-Arylated DPPs

Langhals and Potrawa47 have suggested that absence of ‘benzylic’ hydrogens on

the lactam’s nitrogens of DPP should considerably improve the photostability of DPP dyes. They have prepared a couple of N-arylated DPPs and characterized their

spectroscopic (NMR, UV-Vis, and fluorescence) properties, yet the “high photostability”,

claimed in the paper title, has not been discussed, measured or shown otherwise. This

argument, however, seemed reasonable to us, and since there was no general approach to

N-arylated DPPs, with only few examples, prepared indirectly, have been known, we

explored ‘direct’ approaches to N-arylated DPPs. The first approach was inspired by a

work of Copola134 on direct copper-mediated N-arylation of isatins (Scheme 1.64).

H Br O R Yield, % N CuO O + O H 92 N DMF 4-F 97 O R 4-MeO 71 R 3,5-Me2 81 Scheme 1.64. Direct copper-mediated N-arylation of isatins.134

The reactions of bromo-arenes with DPP in the presence of copper (I) oxide in

DMAc were performed at 120–180°C (Scheme 1.65). In the reactions with 1-bromo-4-

(perfluorobutylsulfonyl)benzene (4′-bromophenyl perfluorobutyl sulfone) and with 1- bromo-4-fluorobenzene in addition to the bis-adducts 91 and 94 we also separated mono- adducts 92 and 95. When monitoring the reaction of 1-bromo-3,5-

103

bis(trifluoromethyl)benzene with DPP under the same conditions, the mono-adduct’s spot on TLC is also detected, is major at some point of time and most surely the reaction can be stopped at that time and the mono-adduct separated as well, but we drove the reaction to completion and separated only the target compound 93.

H O Br N O N O CuO + HNHN + DMAc O N O O R N 1

R R

Di-arylated Yield Mono-arylated Yield

4-C4F9–SO2– 91 32% 4-C4F9–SO2– 92 18% R = 3,5-(CF3)2 93 37% –

4-F 94 12% 4-F 95 37%

Scheme 1.65. Direct copper-mediated N-arylation of DPP 1.

The second approach to direct N-arylation was based upon our hypothesis of

utilizing DPP anion(s) in nucleophilic aromatic substitution of halogen.135 The preformed

DPP2– dianion was reacted with 1-fluoro-4-nitrobenzene, and Pr-DPP– anion — with 1- fluoro-2,4-dinitrobenzene to give the corresponding N-arylated compounds 96 and 97

(Scheme 1.66).

104

NO2

O F N O a HNHN + O O N NO2

NO2 Pr O F N O NO2 a HNN Pr + O O N NO2 O2N

97 NO2

Scheme 1.66. Aromatic nucleophilic substitution on DPP. (a) t-BuOK, DMAc.

These two direct approaches to N-arylated DPPs are, to our knowledge, new for this class of dyes and have never been reported before. They allow easy, one-step access to compounds, which could potentially be prepared before only via laborious multi-step synthesis. The photostability of the prepared N,N-diarylated DPPs turned out to be superior, compared to their N,N-dialkyl analogs and is discussed below.

105

1.4.10. Physical Properties of Newly Prepared DPPs

The physical properties of the newly prepared DPPs are summarized in the

Tables 1.17 and 1.18. There are several generalizations that can be drawn from these

data. The melting points of N,N′-di-substituted DPPs are always lower than those of their

N-mono-substituted counterparts, which is in agreement with all previous observations.

Melting points of N,N′-dialkylated DPPs decreases as the length of the alkyl chain grows.

Optical absorption spectra of N,N′-di-substituted and N-mono-substituted DPPs are quite similar and are almost not affected by the nature of the substituent — be it aromatic or aliphatic. The substitution of the DPP’s phenyl rings, however, brings about considerable changes into the electronic spectra of these compounds. Extension of the conjugation chain causes a bathochromic shift in both emission and absorption spectra, so does the introduction of electron-donating groups at the termini of the phenyl rings. The quantum yields vary, but generally are high to excellent and always higher than 0.5. The Stokes’ shifts span from 28 nm to a good 84 nm. Photostability of some of the prepared compounds has been measured semi-quantitatively as the time required for a sample

solution to completely fade out under UV radiation of λ=360 nm. For most of the DPPs

this time interval falls within several days, while for DPP 1, DPP disulfonic acid and

N,N′-diarylated DPPs no complete fading even after two weeks of irradiation has been

observed. Several DPPs have been utilized (by the group of Prof. W. E. Moerner) in

single-molecule imaging and have been shown to be suitable for such applications (Fig.

1.13), though their photostability remains to be still further improved. We are looking

106

forward now to test the more photostable DPPs under SM conditions and see the improvement over the total observation time.

Figure 1.13. Single molecules of DPP-Me 23 imaged in a PMMA film, excited by 488

nm with an intensity of 0.85 kW cm-2. A 6.2 x 6.2 μm region of the sample is shown.

Photostability of seven prepared DPPs has been measured quantitatively (see

experimental part for details) as the slope tangence of fluorescence (or absorbance) decay

vs. time under constant UV radiation flux of 1.2 mW/cm2 at λ=360 nm. The graphs

below (Fig. 1.14 and 1.15) show that under same exposure conditions, mono-alkylated

DPP 33 is ca. 5.8 times (calculated as ratio of two slope tangences: –0.176/–0.0306) more prone to photodecomposition than its di-alkylated analog 32. Likewise, compound 65 is ca. 2.1 times (–0.0574/–0.0272) less photostable than its counterpart 64.

107

1.0

0.8

0.6

C6H13 H C 13 6 N 65 33 64 32 N

Intensity, a.u. Intensity, O 0.4 O S N S

N N O O O N O N 0.2 H N S S O O N N C6H13 H13C6 0.0 0102030 Exposure time, hrs

Figure 1.14. Photostability of compounds 32, 33, 64, and 65.

The photostability of N,N′-diarylated DPPs compared to that of their N,N′- dialkylated analogs seems to depend on the nature of the N-aryl substituent (Fig. 1.15): while compound 91 has substantially higher photostability than 32, both 92 and 93 do not differ significantly from 32 in the fluorescence decay rate. Thus, the assumption of

Langhals and Potrawa47 on higer photostability of N,N′-diarylated DPPs might be not

quite valid. This matter requires, however, additional investigation, for the above

photostability experiments were conducted in chloroform as solvent, which undergoes

108

photochemical decomposition on its own, and may thus level, rather than differentiate photostability of the fluorophores.

1.0

0.8

0.6 O

F(F2C)4O2S N N SO2(CF2)4F

93 92 32 91 O Intensity, a.u. 0.4

CF3

F3C

O N N 0.2 O O

N N O

CF3

F3C 0.0 0 10203040506070

Exposure time, hrs

Figure 1.15. Photostability of compounds 32, 91, 92, and 93.

The quantitative data on photostability of compounds 32, 33, 64, 65, 91, 92, and

93 are summarized in the Table 1.16.

109

Table 1.16. Photostability of several DPPs.*

# Structure tg α |α|, °

Pr 32 N O –0.03059 1.95 O N Pr

H 33 N O –0.17591 11.09 O N Pr

64 Pr N O S –0.0272 1.73 S O N Pr

65 Pr H13C6 N O N S C6H13 H13C6 N –0.0574 3.65 N S O C6H13 Pr

91 O

F(F2C)4O2S N N SO2(CF2)4F –0.01682 1.07 O

92 O

F(F2C)4O2S N NH –0.05223 3.3 O

F3C O CF3 N N –0.08765 5.56

F3C O CF3

* tg α is the slope and |α| is the angle of the linear fit of the photodecomposition curves.

R1 N O R Table 1.17. Symmetrical 4,4’–Disubstituted DPPs. R O N R1

# R R1 yield, % m.p., °C λmax, absorption, nm λmax, emission, nm ΦF

142 H H 56 420 50466 508, 545b –

21 CN H 35 443 dec. 501, 538 556, 603a 0.63b

22 Me H 38 354 dec. 274, 472, 508 514, 557 0.66b

66 a c 10 HO3S– H 77 – 268, 474, 505 525, 566 0.63

Me Pr 60 186 306, 474 530, 575a 0.64

47 CN Pr 83 227 285, 496 568 0.65

43 Br Me 46 >350 275, 305, 491 541, 587a 0.83

44 Br Pr 37 247 275, 304, 478 541, 587a 0.86

45 I Me 37 dec. 355 276, 318, 495 544, 590a 0.89

46 I Pr 68 260.5 276, 317, 479 550, 592a 0.86 110

56 4-AcO-C6H4-CH=CH– Me 6 357.7 279, 335, 510 582 0.73

55 4-t-Bu-C6H4-CH=CH– Me 50 295 280, 337, 515 587 0.75

48 4-Pyrr– Me 74 324 dec. 282, 385, 548 575 0.95

50 4-Pyrr– Pr 32 267 273, 384, 539 577 0.94

a 53 (C6H13)2N– Me 11 132 282, 385, 548 575, 620 0.92

60 4-Et2N-C6H4-CH2=CH– Pr 49 247 334, 538 620 0.72

64 S Pr 84 281 334, 362, 532 616, 690a 0.96

S 65 a (H13C6)2N Pr 32 278 241, 359, 541 615, 693 0.82

a) Shoulder of the main peak, determined as minimum of the second derivative. b) In dimethylformamide; c) In ethanol; d) Yield from parent DPP or Br-DPP. 111

R N O Table 1.18. N– and N,N′-substituted DPPs.

O N R1

# R R1 yield, % m.p.°C λmax, absorption, nm λmax, emission, nm ΦF

23 Me Me 85 231 265, 294, 476 522 0.9044, 0.54

32 Pr Pr 62 189 289, 466, 488 528, 568a 0.76

33 Pr H 30 276 265, 465, 488 523, 563a 0.69

a 68 C4H9SO3H Pr 9 oil 264, 465, 486 524, 565 0.73

a b 67 C3H7SO3H H 61 270 465, 486 525, 566 0.77

a 34 C6H13 C6H13 8 134 472 530, 572 0.74

a 35 C6H13 H 16 252 262, 464, 481 522, 565 0.71

a 36 C10H21 C10H21 42 117 268, 467 529, 573 0.65

a 37 C12H25 C12H25 47 114 284, 474 529, 573 0.66

a 76 C6H12OCOPh C6H12OCOPh 23 111 475 527, 563 0.97 112

a 77 C6H12OCOPh H 8 194 467, 493 522, 563 0.96

a 80 C6H12OH C6H12OH 15 – 474 525, 567 0.94

81 C6H12OH H 63 – 467, 486 – –

a 83 C6H12OH C12H25 28 127 468, 486 522, 563 0.89

69 C8H17OH PhCH2– 60 107 264, 486, 490 514, 557 0.97

a 85 C10H21OH C10H21OH 6 143 475 525, 566 0.87

a 84 C10H20OH Pr 16 105 467, 490 530, 561 0.76

a C10H21OH H 10 166 468, 493 523, 563 0.86

45 a 41 CH2=CH–CH2– CH2=CH–CH2– 42 209 476 525, 566 0.92

a 42 CH2=CH–CH2– H 46 309 467, 495 518, 560 0.97

a 40 PhCH2 H 32 344 263, 295, 468 525, 566 0.95

† a 88 MI –C6H12– MI–C6H12– 7 dec. 264, 464 525, 565 0.67

‡ a 89 Fu-MI –C6H12– C3H7SO3H 15 165 470, 489 524, 565 –

Et Et 20 – 281, 369, 534 – – 70 HO N HO N

113

a 91 4-C4F9-SO2-C6H4 4-C4F9-SO2-C6H4 32 262 271, 475 515, 555 0.91

a 92 4-C4F9-SO2-C6H4 H 18 320 461, 485, 490 513, 554 0.93

a 93 3,5-(CF3)C6H3– 3,5-(CF3)C6H3– 37 298 251, 315, 475 520, 552 0.95

96 4-NO2–C6H4– 4-NO2–C6H4– 12 200 268, 470 – –

a 94 4-F–C6H4– 4-F–C6H4– 12 344 271, 475 520, 566 –

95 4-F–C6H4– H 37 404 271, 475 – –

97 2,4-(NO2)2-C6H4 Pr 25 dec. 263, 298, 466 non-fluor. –

†MI = maleimide residue, 1H-pyrrole-2,5-dione-1-yl.

‡ Fu-MI = 3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2-yl.

a) Shoulder of the main peak, determined as minimum of the second derivative

b) In ethanol 114

115

1.4.11. Conclusion

A series of new fluorophores for single-molecule spectroscopy applications have

been designed and prepared based on derivatives of dihydropyrrolo[3,4-c]pyrrole-1,4-

dione (DPP) with substantially higher solubility, longer absorption and fluorescence

emission wavelengths, and a wide range of functionalization. A range of reliable N, N′- alkylation, aryl halogenation and organometallic mediated C-C bond formation reactions has been developed and conducted. A cysteine-reactive fluorescent tag and a variety of hydroxy-functionalized and water-soluble DPP derivatives have been synthesized. Two new methods for preparation of N,N′-diaryl-substituted DPPs have been identified and explored. The fluorescence properties of the new DPPs were studied in detail and some of the fluorophores have been successfully imaged at the single-molecule level. The quantum yields were measured and vary from moderate to high and Stokes shifts as large as 84 nm were observed. Photostability of seven new DPPs was quantified and compared with the structures of the fluorophores.

CHAPTER II.

CYSTEINE-SPECIFIC FLUORESCENT TAGS:

NILE RED – MALEIMIDE AND

DCDHF – MALEIMIDE.

2.1. Introduction to molecular probes and tags

The fluorescent tags, or bioconjugate molecular probes are a subclass of

fluorescent labels, discussed briefly in the introduction to the Chapter 1. Their

characteristic feature is a specific covalent bonding of the probe’s reactive site to the certain region(s) of a biomolecule or of a living cell due to a chemical reaction, resulting

in overall binding of the probe to the substrate. The specificity of the overall binding is determined by that of the chemical reaction (called here conjugation, meaning binding), which creates the new covalent bond. The structure of all bioconjugate probes may generalized as it is depicted on Fig. 2.1: a fluorescent dye, connected to a reactive group with a spacer of variable length.

Some of the commonly used hooks, or the reactive groups with their

corresponding bioconjugation chemistry, are summarized in Table 2.1 and Table 2.2.

Some of the dyes commonly used as fluorescent molecular probes (but not necessarily

116 117

Et2N N O Dye

Spacer

N O O Hook

Figure 2.1. Generalized structure of a bioconjugate fluorescent molecular probe.

generally for SMS) and their absorbtive and fluorescence ranges are summarized as a chart in Fig. 2.2.139

As can be seen from the examples in the Tables 1.1 and 1.2 and Fig. 2.2, the

pool of the commercially available fluorescent dyes is very diverse, yet it exploits a small

number of fluorophore’s structural templates (of xanthenes — fluorescein, rhodamines;

of chromans — coumarins; of BODIPY — difluoroboradiazaindacene, of polynuclear

aromatic hydrocarbons — pyrene, perylene, coronene, prodan, of conjugated or fused

diazoles — NBD, 7-nitrobenza-2-oxa-1,3-diazoles, POPOP, 1,4-bis(5-phenyloxazol-

2yl)benzene, PPD, 2,5-diphenyl-1,3,4-oxadiazole, 2-pyridyl-2-phenyl-1,3-oxazoles;

cyanines … to name some of them). Few of them fulfill some of the requirements for the

SMS dyes, listed in Chapter 1. Specifically, the demand for dyes with high quantum yield

and photostability remains unfulfilled with many commercial dyes.

Table 2.1. Amine (R2–NH2) reactive groups for molecular probes.

Reactive group Structure Product Example benzophenone-4-isothiocyanate, and NCS

R1 NH C NH R2 Isothiocyanate R1–N=C=S S HOOC

+ Me2NO NMe2 [243670-15-7], and O R C O NH R O Succinimidyl Ester R C O N 1 2 1 O O O N O O O O O O O SO3H SO3H O O O N Sulfosuccinimidyl R1 C O NH R2 R C O N HO3S 1 O Ester (STP) O N O O O O O O O

118

Reactive group Structure Product Example Alexa Fluor 488 [247144-99-6] derivative: O F FF O F Tetrafluorophenyl R C O NH R 1 2 HOOC R1 C O F Ester (TFP) O O F FF + H2N O NH2 SO3H SO3H O O N3 R C O N R C O NH R Carbonyl Azide 1 3 1 2 O O O

AcO O OAc Dansyl chloride,136 and

SO2Cl

Sulfonyl Chloride R1–SO2Cl R1–SO2–NH–R2 SO3H

+ N ON

119

Table 2.2. Thiol (R2–SH) reactive groups for molecular probes.

Reactive group Structure Product Example BODIPY137 [400885-92-9] derivative O O Iodoacetamide + I I NH2 R2 S NH2 N - N B F F NH O

O N O O O H Maleimide R N R N 1 1 HOOC S R2 O O

+ Me2NO NMe2 BODIPY 138 [615574-25-9] and Br

Alkyl Halide R1–CH2–X R1–CH2–S–R2 + N - N B F F

120 121

Figure 2.2. Common dye classes, used in molecular probes.139

To create a bioconjugate fluorescent molecular probe (fluorescent tag), suitable for SMS detection, one needs to connect a suitable, reliable hook to an SMS-effective dye with an appropriate spacer. Sometimes the spacer is missing (or of zero length) and the

reactive group connects directly to the fluorophore moiety. If the subsequent chemical

reaction creates a longer conjugation chain, the fluorophore’s absorption and emission

wavelengths may change. For example, dansyl chloride (5-(dimethylamino)naphthalene-

1-sulfonyl chloride, λmax Ab ≈ 350 nm) does not absorb or fluoresce in the visible region

until it reacts with an amine (λmax Em ≈ 520 and is solvent-dependent). A short spacer may

122

facilitate resonance energy transfer, while a very long one may cause steric problems and report local polarity falsely, separating the fluorophore and the site of reactive group attachment. The optimal length of a spacer is contigent upon specific application.

2.2. Design of the Probes

We designed and prepared several fluorescent tags with a thiol- (mercapto, or sulfhydryl, –SH) reactive maleimide group. The synthesis of two DPP-Maleimide fluorescent tags has been attempted, and one of them succeeded – see the details in the

Chapter 1, compounds 88 and 89, 90.

Choice of the dye moiety. Structurally new or modified dyes rarely give improvements in several parameters, all crucial for fluorescence detection, at once. For example, the new BODIPY dyes have excellent quantum yields, high extinction coefficients (~80,000 M–1·cm–1), and thus — good brightness, but possess small Stokes’

shift and are not very sensitive to the local polarity (that is, their fluorescence is not

solvatochromic). The cyanine dyes give useful far-red and near-infrared absorption and

emission maxima, but also suffer from small Stokes’ shift (~30 nm). The small Stokes’

shift results in (a) large background signal due to the scattering of the excitation light

(especially important factor in the SMS experiments), and in (b) large overlap integral,

which in turn ensues significant self-quenching of the fluorophore (fluorescein is a

classically notorious example).

Such is the case with many new SMS dyes as well. This is another reason for

further research and diversification of the available pool of the dyes. In Scheme 2.1 a

123

series of fluorophores are depicted, which resemble each other structurally. Lakowicz41 comments on them (except fluorescein, acridine, and coumarin): “At present, these dyes are less used in biochemistry owing to their lack of water solubility, their tendency to aggregate, and the lack of conjugatable forms”.[41, p 75]. One of the depicted fluorophores, Nile Blue140,141 [2381-85-3], has a close relative from the same oxazine

class, called Nile Red [7385-67-3]. Nile Red has an extinction coefficient142,143 of 38,000

–1 –1 M ·cm at λmax Ab = 519.4 nm, its fluorescence maximum (λmax Em = 580 nm in dioxane,

Fig. 2.3) is highly solvent-dependent, thus exhibiting solvatochromism144. The Stokes’

shift of Nile Red in dioxane is thus 61 nm, and its quantum yield is 0.7.145 The red emission, large solvatochromism,146 and fair quantum yield — all these properties make

this fluorophore attractive to incorporate it into a bioconjugate form, sensitive to and

aimed to measure the local polarity in the biomolecules. We picked Nile Red Phenol147

[188712-75-6], a known phenolic derivative of Nile Red with λmax Ab = 551 nm, λmax Em =

632 nm (in MeOH), to use the existing hydroxyl group as a site for attachment of the thiol-reactive group via an appropriate spacer.

124

R – COO

+ N ON – O OOEt2NOO Rhodamines Fluorescein Coumarine 1

N N

+ + Me2N N NMe2 Et2N O NEt2 Me2N S NH2

Acridine Orange Oxazine 1 Azure A OH

N N N

+ Et2NNHO 2 Et2NOO Et2NOO Nile Blue Nile Red Nile Red Phenol

Scheme 2.1. Fluorophores conceptually similar in structure.

Another fluorophore we utilized belongs to the newly invented148 DCDHF class

of dyes.149,150 DCDHF1 stands for dicyanodihydrofuran and is an informal abbreviation

of the heterocyclic system it is based upon. Its proper name is 3-cyano-2-

dicyanomethylene-4-R-5,5-dimethyl-2,5-dihydrofuran, and proper CAS name 3-cyano-4-

R-5,5-dimethyl-2(5H)-furanylidene propanedinitrile accordingly. Several representatives

1 «DCDHF» not to be confused with another common individual fluorophore, 2-(2,7- dichloro-3,6-diacetyloxy-9H-xanthen-9-yl)-benzoic acid with CAS RN [4091-99-0], also known as 2,7-dichlorodihydrofluorescein diacetate.

125

of this class of dyes are depicted on Scheme 2.2. DCDHF-6, [402490-54-4] individual

molecules, incorporated into a polymethyl methacrylate (PMMA) film, have been

successfully imaged by W. E. Moerner and his group (Fig. 2.4).150

1,0

0,8 Ab Nile Red Em 0,6

0,4 Intensity, a.u. Intensity,

0,2

0,0

200 300 400 500 600 700 800

Wavelength, nm

Figure 2.3. Nile Red absorption (Ab) and emission (Em) spectra in dioxane.

In PMMA polymer film, DCDHF-6 has a fluorescence quantum yield of ΦF =

0.92 and, on average, emits more than 1.2·106 total photons before irreversible

photobleaching occurs. In addition, this dye is stable against “blinking” or flickering in

126

emission, with approximately 85% of the molecules showing no blinking behavior on the

100-ms time scale of the measurement.150

In toluene solutions, however, the quantum yields of the DCDHF dyes were

found to be almost an order of magnitude lower. This result suggests that there is an

environmentally sensitive path through which the molecule can return nonradiatively to

the ground state. Quantum mechanical calculations of the electronic structure of this

system suggest the presence of two excited-state minima — one radiative and the other

nonradiative — accessed through different intramolecular twists in the excited state. The

twist leading to nonradiative decay has an environmentally dependent energy barrier,

resulting in a DCDHF fluorescence quantum yield that varies with local environment.151

Therefore, these dyes are promising candidates as fluorophores in single-molecule probes for local environment viscosity, rigidity, or polymer-free volume. 150

NC NC NC NC CN CN (H13C6)2N O O (H13C6)2N NC NC CN DCDHF-6 DCDHF-2v R O NC NC NC NC CN CN DCDHF (H13C6)2N O S O (H13C6)2N S

DCDHF-Th-6 DCDHF-Th-6v

Scheme 2.2. Examples of new DCDHF dyes.

127

Figure 2.4. DCDHF-6, imaged at single molecule level in PMMA film.152

The color saturation towards red indicates increase in fluorescence intensity.

The peaks correspond to locations of single molecules.

Choice of the reactive group. The amine-reactive groups (mostly acylating reagents) are quite indiscriminative, and most of them also react with hydroxy and mercapto functionalities in the biomolecules. Thiol-reactive groups are much more selective and for this reason they are used to prepare fluorescent peptides, proteins, and oligonucleotides for probing biological structure, function, and interactions. Maleimides are excellent reagents for the thiol groups, for they do not react with methionine, histidine, or tyrosine, as iodoacetamides sometimes do.153 Reaction of maleimides with

amines usually requires a higher pH than reaction of maleimides with thiols. However,

hydrolysis of the maleimide to an unreactive maleamic acid can compete significantly

with thiol modification, particularly above pH 8. Furthermore, once formed, maleimide

128

adducts can hydrolyze to an isomeric mixture of maleamic acid adducts, which may cause a significant change in the fluorescence properties of the conjugate,154 or they can

ring-open by nucleophilic reaction with an adjacent amine to yield crosslinked

products.155 Because of their overall best selectivity, the maleimide reactive group is the

hook of choice for thiols.

Choice of the spacer. Since there are many views on the spacer length role and

how it affects the fluorescence properties, it is hard to predict and assess, which spacer

length is “right”. Since lengthier and flexible spacers tend to amplify the probe’s

sensitivity to the local environment effects, we have chosen a flexible alkyl chain as our

spacer. In our opinion, C6 length of such a spacer is somewhat the optimal or at least a

good compromise to start with.

The resulting target fluorescent probes are depicted in Scheme 2.3. For the Nile

Red fluorophore, we contemplated other sites of the hook attachments, as depicted on the

Schemes 2.4 and 2.5, yet these approaches have not been implemented, and the Nile Red

Phenol with the hook attached via hydroxy functionality at the 2-position is described below.

OH O NC NC CN O NH R N ; O NH Spacer O Spacer NOO O Nile Red Phenol Maleimide (MI) DCDHF Maleimide (MI)

Scheme 2.3. The target fluorescent tags with fluorophores, hook, and spacer of choice.

129

a b NO c N

NH OH NOHEt NOH 2 NOO [621-31-8] g OH f 98 OH

d,e N N or or N

NOO NOO NOO O O O O O O N O N N O O O O O O O HO I CH2 N N N HO 100 O 101 102 O O O

Scheme 2.4. Proposed attachment of the maleimide hook via N-(2-hydroxyethyl) group.

156 (a) oxirane, MeOH. (b) NaNO2, H2SO4, 0–5°C. (c) 1-naphthol, EtOH. (d) Bu4NMnO4.

(e) MI–CH2–OH (100), Ph3P, DEAD. (f) MI–(CH2)5–COOH (101), CDI. (g) MI–(CH2)6–

I (102), K2CO3.

130

a NO b N

NOH NOH N OO 99 [41175-50-2] O c

N O O O HO N N 103 O N OO

Scheme 2.5. Proposed synthesis of Nile Red derivative 99 and attachment of the

157 maleimide hook via phenol functionality. (a) NaNO2, HCl. (b) 1,6-

dihydroxynaphthalene, DMF. (c) MI–(CH2)6–OH (103), Ph3P, DEAD.

2.3. Synthesis

Nile Red Phenol (9-diethylamino-2-hydroxy-5H-benzo[a]phenoxazin-5-one)

105 was prepared according to the literature procedure, starting from 3-N,N-diethyl- aminophenol (Scheme 2.5). Nitrosation was performed with sodium nitrite in concentrated aqueous hydrochloric acid to give 5-(diethylamino)-2-nitrosophenol 104 as a hydrochloride salt.158 The latter was condensed with 1,6-dihydroxynaphthalene in boiling DMF to give a mixture of different dyes (crude yield 70%), which was purified by several consecutive chromatographies to give Nile Red Phenol 105 in 35% yield.

131

OH OH 2 1 3 a NO b 11 N 4 10 + 12 9 5 Et2NOHEt NOH OH 8 6 2 Et2NOO 7 104 105

Scheme 2.5. Preparation of Nile Red Phenol. (a) NaNO2, HCl, 0–5°C. (b) DMF, Δx.

A hexamethylene spacer was then attached to Nile Red Phenol by either alkylation of the phenol group with 6-bromo-1-hexanol (path a on Scheme 2.6), or by

Mitsunobu reaction with 1,6-hexanediol (path b on Scheme 2.6).

HO HO O OH O

a b N N N 77% 26% Et2NOO Et2NOO Et2NOO 106a 105 106b

Scheme 2.6. C6-Spacer attachment to Nile Red Phenol.

(a) 6-bromo-1-hexanol, K2CO3, KI (cat), DMF. (b) Ph3P, DEAD, THF, 1,6-hexanediol.

A series of maleimides have been prepared (Scheme 2.7 and 2.8) for subsequent attachment to Nile Red and DCDHF dyes: N-(hydroxymethyl)maleimide 107 — from maleimide and formaldehyde,159 N-(iodomethyl)maleimide 108 — from 107 and

160 161 diphosphorus tetraiodide P2I4, prepared according to a published procedure; maleimid-2-yl-acetic acid (6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)acetic acid) 109, 6-

132

(maleimid-2-yl)caproic acid (6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoic acid)

110 — from maleic anhydride and corresponding aminoacids, via dehydration of the

intermediate maleamic acids with acetic anhydride;162 6-(maleimid-2-yl)caproyl chloride

(6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl chloride) 111 — from 110 and thionyl chloride; N-(4-hydroxyphenyl)maleimide163 (1-(4-hydroxyphenyl)-1H-pyrrole-

2,5-dione) 112,164,165 N-(6-iodohexyl)maleimide (1-(6-iodohexyl)-1H-pyrrole-2,5-dione)

113 — from the silver salt of maleimide166 MI–Ag and 1,6-diiodohexane; N-(6-

hydroxyhexyl)maleimide 114 — from MI–Ag and 6-bromo-1-hexanol. Maleimide itself,

although commercially available, is rather expensive ($10 per gram) and was required in plentiful amounts. Thus, it was prepared from maleic anhydride according to the literature procedure in 60% overall yield.167

O O O HO abI NH N N

O MI O 107 108 O

c O d O N I O N Ag 113 O e O N HO MI–Ag 114 O

Scheme 2.7. Syntheses of maleimides. (a) CH2O, NaOH 5% aq. pH = 9. (b) P2I4, CS2,

CHCl3, K2CO3. (c) AgNO3, EtOH, NaOH. (d) 1,6-diiodohexane. (e) 6-bromo-1-hexanol.

133

O O O O NH b, d O + 2 N HO (CH ) HO 2 5 O O c, d 110

a, d e OON O O O OH O N N OH Cl O O 109 112 111

Scheme 2.8. Syntheses of maleimides from maleic anhydride. (a) glycine, AcOH. (b) 6-

aminocaproic acid, AcOH. (c) 4-aminophenol, AcOH. (d) Ac2O, AcONa. (e) SOCl2.

Following a reported protocol for the preparation of N-alkylated maleimides by

the Mitsunobu reaction,168 a series of such reactions between Nile Red Phenol with (106)

a 6-hydroxyhexyl spacer, or without (105) such a spacer as one reagent, and maleimide per se, N-(hydroxymethyl)maleimide 107, (4-hydroxyphenyl)maleimide 112, or N-(6- hydroxyhexyl)maleimide 114 as other reagent has been performed (Scheme 2.9), but no tangible amounts of the expected products have been obtained from any of the attempted reactions.

Benzyl alcohol has been reported169 to react with maleimide under Mitsunobu

reaction conditions. We tried to reproduce this reaction, as well as with 1-hexanol, 1,6-

hexanediol, and 1,4-benzenedimethanol (1,4-bis(hydroxymethyl)benzene) but without

avail: although some of the expected product was separated in each case (Scheme 2.10),

most of the starting materials were recovered and the reaction was very messy overall.

134

HO O OH

N 106 OR 105 N

Et NOO Et2NOO 2 + MI O O 107 107 N XOH OR NH 112 112 114 O O EtOOC + Ph3P + NN COOEt O O N X O N O O O N N

Et NOO Et2NOO 2

Scheme 2.9. Attempted Mitsunobu reactions between Nile Reds 105 and 106

and various hydroxy-functionalized maleimides.

R % EtOOC Ph3PNN+ + R OH C5H11– 7 COOEt O Ph– 2 O R N HO + OH + HN O 17 O

HO–(CH2)5– 0

Scheme 2.10. Attempted N-alkylation of maleimide according to a reported protocol.169

135

Likewise, no reaction has been detected upon treatment of Nile Red Phenol 105 with either 109 or 110 neither under Mitsunobu conditions, nor under action of 1,1-

carbonyldiimidazole and DMAP in THF or DMAc. Overall with 105, we observed only two successful Mitsunobu reactions: with 1,6-hexanediol in 26% yield (Scheme 2.6, b) and acylation with simple caproic acid in 17% yield. The product obtained in the latter reaction, was identical to the one (118) obtained by acylation with caproyl chloride.

To check the actions of simple alkylating and acylating reagents on 105, the latter was methylated to give 115 in 99% yield, alkylated with ethylene carbonate to 116, with 1,6-diiodobenzene — to 117, and acylated with caproyl chloride to 118 (Scheme

2.11).

O OH O

a b N N N 99% 63% Et2NOO Et2NOO Et2NOO 115 105 118 c d 42% 79% HO I O O

N N

Et2NOO Et2NOO 116 117

Scheme 2.11. Simple alkylations and acylations of Nile Red Phenol 115.

(a) MeI, K2CO3, DMF. (b) AmCOCl, DBU, DMF. (c) ethylene carbonate, K2CO3, DMF.

(d) I–(CH2)6–I, K2CO3, DMF.

136

Apparently, the maleimide functionality interferes with and deteriorates the alkylations and acylations.170 Thus, we considered a strategy of protecting the maleimide

moiety before its introduction and deprotecting it later. Protecting groups for maleimide’s

can be a diene, entering into a [2+4] cycloaddition reaction, and which can be easily

removed later by a retro Diels-Alder reaction.171 At first, we attempted to employ the

adduct of maleimide with cyclopentadiene as a protective group.172 However, it turned

out to be difficult to deprotect these adducts later — temperatures above 200°C

sometimes are required.173 Then we spotted an approach of Clevenger and Turnbull,

utilizing an adduct of furan and maleimide, which could be deprotected in 1–2 hours in

boiling anisole in most cases.174 This approach can possibly be complicated somewhat by two diastereomeric adducts formed — endo (m.p. 126–128°C) and exo (m.p. 162°C). If necessary, the endo-isomer (or mixture thereof, m.p. 130–132°C) can be converted into thermodynamically more stable exo-isomer by recrystallization from a high-boiling solvent.175

Following this strategy, the adduct of furan and maleic anhydride Fu-MA 119

(exo-cis-7-oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic anhydride) was prepared.176 It was then converted into Fu-MI 120 (exo-cis-7-oxabicyclo[2.2.1]hept-5-ene-2,3- dicarboximide), best following the modified procedure of Zawadowski177 to obtain the

product in crystalline form: the internal reaction temperature must be held strictly at 125

± 5°C for 45 ± 5 min. Then, after evaporation of ca. 40% of the water on a rotovap,

boiling ethanol was added in such amount as to completely dissolve the material. This

137

modification yielded 94% yield of crystalline material, compared to the original 40% reported.

O H O H O a b O + O O O NH O H O O H O 119 120

Scheme 2.12. Diels-Alder adduct of furan and maleic anhydride Fu-MA.

(a) Et2O, 90°C or r.t., then recr. (b) NH3·H2O, 125°C, 45 min.

H O H O a O NH O N I H O H O 120 121

OH H O

N + O N (CH2)6 I b Et NOO H O 2 H O 105 121 O N O I O H O H O N b N + O NH Et2NOO Et2NOO H O 122 117 120

Scheme 2.13. Synthesis of protected Nile Red – Maleimide 122.

(a) I–(CH2)6–I, K2CO3, Me2CO. (b) K2CO3, DMF.

138

The Fu–MI adduct 120 was then alkylated (in DMF or, better, acetone178) with either 1,6-diiodohexane to give 121, which was in turn used for subsequent alkylation of Nile Red Phenol 105, or with 117. In either case, the product was the furan-protected maleimide derivative of Nile Red Phenol 122. The furan-protected maleimide derivative of Nile Red Phenol 122 was then deprotected to the target Nile Red Maleimide 123 by refluxing in a mixture of xylene and dichloromethane (added for solubility reasons) with a short Vigreaux column and no reflux condensor — to condense and return xylene, but allowing furan to escape. Physical and optical properties of the Nile Red Maleimide 123 and its application in protein structure and function elucidation are described below.

H O O

O N N O O H O O a N N

Et2NOO Et2NOO 122 123

Scheme 2.14. Retro Diels-Alder deprotection of 122. (a) xylene, CH2Cl2, Δx.

After the advantage of double bond protection of the maleimide was realized and proved efficient on the Nile Red dye, the DCDHF Maleimide tags 124 (Scheme 2.15) and 125 (Scheme 2.16) syntheses were rather straightforward. These two tags are derived from DCDHF-2V and DCDHF-6 correspondingly (Scheme 2.2) and also differ in the place of the spacer and maleimide attachment — via the 2′-hydroxy group in 124 and via the 4′-amino group in 125. The synthesis of 124 by alkylation of the precursor with a DCDHF subunit already in place (upper path on Scheme 2.15) was not successful, and thus the introduction of the DCDHF heterocycle was performed after the protected maleimide

139

subunit had been introduced (lower path), followed by deprotection of the maleimide hook.

NC NC CHO H O OH CN a O + O N Et2N I H O NEt2 OH 121 CN

+121 b CN b O CN NC NC NC NC CHO CN CN O a c O 4' 1' O Et2N Et2N 3' 2' O O NEt2 O 124 H N OON OON O O HH H O

Scheme 2.15. Synthesis of maleimide-tagged DCDHF-2V 124.

(a) 3-cyano-2-dicyanomethylene-4,5,5-trimethyl-2,5-dihydrofuran, Py, AcOH, 60°C. (b)

Fu-MI-C6-I 121, K2CO3, Et2CO, Δx. (c) xylene, Δx.

140

HO HO H O H O a b OH + O NH O N

H O H O

120 57% c OH OTs

NH2 H O H O d OTs NH O N O N 38% H O H O + NC NC H7C3 NC NC CN N CN e O F O O H 48% N O

O H NC NC f O CN N 4' 1' N 3' 2' O 72% ZL-1-48W 125 O

Scheme 2.16. Synthesis of maleimide-tagged DCDHF-6 125.179

(a) TsCl, Py, THF. (b) Fu-MI 120, K2CO3, DMF, 45°C. (c) TsCl, Py.

(d) 1-PrNH2, Py, THF, Δx. (e) 4′-F-DCDHF, Py. (f) xylene, Δx.

2.4. Results and Discussion

The optical properties of the prepared tags 123, 124, and 125 are summarized in

Table 2.3. The emission spectra, quantum yield measurements, and all biological studies

were performed in the group of Prof. W. E. Moerner, Prof. Arthur L. Horwich, and Prof.

Judith Frydman at Stanford University.

141

Table 2.3. Optical properties of maleimide-containing fluorescent tags.

Solvent # Structure λmax Ab, nm (ε) λmax Em ΦF O

N DCM PhMe O 536 564 0.83 123 O 547 MeOH 635 MeOH – N (3.62·105) 656 water <0.1%

Et2NOO

NC NC CN O 589 DCM O DCM 124 Et2N 5 720 0.73 N (8.35·10 ) O O

NC NC H C 7 3 CN N O 496 DCM DCM 125 O 516 0.12 (9.27·105) N

The longer conjugated path in the chromophore subunit of 124 gives

correspondingly red-shifted absorption and emission maxima, compared to that of 125.

The difference in the quantum yield between the two compounds is 5.8 times, with 124

being noticeably more efficient emitter. Maleimide-tagged Nile Red 123 showed very

strong solvatochromism, especially in its fluorescence properties: besides the

142

bathochromic shift in the emission maximum, its quantum yield drops 300 times when measurements are done in water (as hydrochloride salt), compared to toluene (Fig. 2.5).

Decreasing x=1 (Pure Methanol) 1.5e+6 x=0.85 Hydrophobicity x=0.65 x=0.5 1.3e+6 x=0.35 x=0.25 x=0.2 1.0e+6 x=0.18 x=0.15 x=0.13 7.5e+5 x=0.1 x=0.06 x=0.03 5.0e+5 x=0 (Pure Water)

2.5e+5 Fluorescence Intensity (a.u.) Intensity Fluorescence

0.0 550 575 600 625 650 675 700 725 750 Wavelength (nm)

Figure 2.5. Fluorescence intensity of 123 decreases with solvent polarity increase.419

The Nile Red Maleimide tag 123 has been employed in covalent bonding and labelling of a single-cysteine mutant of GroEL (аCys261, Fig. 2.6 and 2.7), whose cysteine subunit is located inside the folding cavity at the apical region of the protein.

Bulk fluorescence spectra of Cys261-NR were measured to examine the effects of binding of the stringent substrate, malate dehydrogenase (MDH), GroES (cofactor of

GroEL), and nucleotide on the local environment of the probe. The sequence of changes in local hydrophobicity of GroEL due to binding of GroES, substrate, and different nucleotides was investigated using fluorescence spectroscopy. Changes in local polarity

143

were monitored by fluorescence intensity as a function of time. In order to observe the sequence dependence, various reagents such as GroES, substrate, and nucleotide or nucleotide mimics were added in different orders.

A typical set of kinetic scans showing the relative peak intensity as a function of time is shown in Fig. 2.8 for one specific sequence of adding reagents, MDH-ES-nt.

Since the preliminary study of fluorescence intensity showed linear dependence of intensity on hydrophobicity the intensity changes on Fig. 2.8 may be quantified and related to specific binding/conformational changes.

For example, when GroES-cofactor is added before ADP/AlFx, GroEL would

begin in the TT state, since nucleotides have not been added and GroES does not strongly

bind to GroEL without them. After adding ADP/AlFx, however, GroES can bind to

GroEL, and form either a symmetric complex or an asymmetric complex (known as the

“bullet complex”). The large fluorescence increase induced by MDH addition suggests

that the asymmetric bullet complex seems to prevail, leaving open a binding site for unfolded MDH in the trans ring. This might be explained by a reduced binding affinity of

ADP/AlFx to the trans ring when GroES is already bound to the cis ring (ADP/AlFx bullet

complex).

We have observed a variety of local polarity changes upon the addition of

substrate MDH, nucleotide (nt), and GroES. For the most cases, a large shift to less

polarity is observed when unfolded MDH is added, while addition of nucleotide and

GroES produce shifts toward increasing polarity. For sequences in which GroES is added last, we observe competition between GroES and substrate for the binding sites of the

144

apical domain (Fig. 2.9). For the special case in which the substrate was added last and the nucleotide was the transition state mimic ADP/AlFx, the fluorescence changes

depended upon the order of addition of the first two components, the nucleotide and

GroES. This behavior is consistent with the formation of a doubly-capped symmetric

complex when ADP/AlFx is added first.

Intriguing results were obtained for the two sequences in which MDH was the

last reagent added and the nucleotide used was ADP/AlFx (Fig. 2.9). Specifically, if

ADP/AlFx was added after GroES (ES-ADP/AlFx-MDH), the fluorescence increase

induced by MDH (“the MDH effect”) was large (20 ± 1%, Fig. 2.9 a), while if ADP/AlFx was added before GroES (ADP/AlFx-ES-MDH), the MDH effect was much smaller than

in all other cases (7 ± 2%, Fig 2.9 b). On the other hand, when the nucleotide added is

either ATP or ADP, the MDH effect was independent of the order of adding nt and ES

(i.e., for the sequences nt-ES-MDH and ES-nt-MDH). In these two situations, the

fluorescence increase induced by MDH was ~31% and ~28% for ATP and ADP,

respectively.

145

Figure 2.6. Ribbon model of GroEL homotetradecamer.

The monomer unit is shown in color.

These observations may be interpreted as a competition between GroES and the substrate for the binding sites of the apical domain, and the degree of this competition, which ultimately induces the substrate release into the cavity, is determined by the nucleotides. For the special case in which the substrate was added last and the nucleotide was the transition-state mimic ADP/AlFx, the fluorescence changes depended upon the

order of addition of the first two components, the nucleotide and GroES.

146

Cys

Figure 2.7. Ribbon model of GroEL protein.

The position of the Cys subunit is shown with an arrow.

1.4 ATP Adding ADP/AlFx 1.3 ADP GroES Adding Nucleotide

1.2

Decreasing 1.1 Hydro- phobicity

Relative Peak Intensity Peak Relative 1.0 Adding MDH

0.9 0 1000 2000 3000 4000 Time (sec)

Figure 2.8. Fluorescence change after addition of (1) MDH, (2) GroES, (3) nucleotide.

147

(a) (b) 1.10 1.10

GroES-ADP/AlFx-MDH ADP/AlFx-GroES-MDH 1.05 1.05 Adding Adding GroES 1.00 1.00 ADP/AlFx MDH effect Adding (Decreased) 0.95 ADP/AlFx 0.95 Adding GroES Adding 0.90 0.90 MDH

0.85 0.85 Relative Peak Intensity MDH Effect Relative Peak Intensity Peak Relative Adding MDH 0.80 0.80 0 1000 2000 3000 4000 5000 0 1000 2000 3000 4000 5000 Time (sec) Time (sec) (c) (d)

ADP/AlFx - GroES (1:1 molar ratio) - MDH 1.04 GroES-ADP/AlFx(45 min)-MDH 1.04 Adding Adding GroES 1.00 1.00 ADP/AlFx Adding Adding 0.96 0.96 GroES ADP/AlFx 0.92 0.92 Adding Adding MDH MDH 0.88 0.88 Relative Peak IntensityRelative Peak Relative Peak Intensity MDH Effect MDH Effect

0.84 0.84 0 1000 2000 3000 4000 5000 6000 0 1000 2000 3000 4000 5000 6000 Time (sec) Time (sec)

Figure 2.9. Effect of various addition orders on the fluorescence intensity.

On the basis of these results, we can understand the behavior as follows. First, a substrate like MDH can bind to the GroEL apical domain in GroEL, causing increased hydrophobicity. Although GroES exists in the solution, it cannot bind tightly to the

GroEL apical domain due to the absence of nucleotides. After addition of nucleotides,

GroES can finally bind to GroEL, and the nucleotide type determines the extent of the conformational changes of the apical domain by GroEL-GroES complex, inducing MDH release into the cavity. Furthermore, as noted alone, the fluorescence decay after the final addition in Fig. 2.9 a shows a fast change followed by a slow drop. The common feature

148

of all these sequences is that the compact GroEL-GroES complex formation can only start at the end of the sequence and, therefore, the displacement of MDH by GroES should always occur at the end. The schematic explanation of these sequences is depicted on Fig. 2.10.

(a)

GroES ADP/AlFx MDH ADP/AlFx ADP/AlFx

TT state No GroES binding RT state MDH binding TT state (Bullet Structure) to the trans-ring (b)

ADP/AlFx GroES MDH ADP/AlFx ADP/AlFx ADP/AlFx

ADP/AlFx ADP/AlFx ADP/AlFx

TT stateRR state Symmetric complex No MDH binding (American Football)

Figure 2.10. Proposed scheme for the formation of symmetric/asymmetric complex of

GroEL/GroES with ADP/AlFx. (a) When GroES is added first, GroES can only bind after adding ADP/AlFx, and the binding affinity of ADP/AlFx to the trans ring may be reduced because of GroES binding to the cis ring, thus preventing formation of the symmetric complex. (b). When ADP/AlFx is added first (before GroES), hindered MDH binding to the GroEL can be explained by the formation of the symmetric football complex.

149

2.5. Conclusion

We have designed, prepared and characterized fluorescent probes with two different fluorophores — Nile Red and DCDHF, with a hexamethylene spacer and a maleimide reactive group. One of the probes has been utilized in local polarity probing and conformational changes elucidation experiments after covalently binding to a single- cysteine mutant of GroEL chaperonin of E. coli. Large solvatochromism and polarity dependence of the prepared probe, combined with high site binding specificity, allowed to study conformational changes of GroEL chaperonin.

CHAPTER III.

ORGANIC LIQUID CRYSTAL SEMICONDUCTORS.

3.1. Introduction

The electrical conductivity is a physical manifestation of charge transport phenomenon in materials. The physical value of electrical conductivity σ is the quantitative measure of this phenomenon — total charge, transported across a unit cross- section area per second per unit electric field applied. Depending on their electrical conductivity values, all materials historically and rather arbitrarily can roughly be divided into the classes of conductors (metals), semimetals, semiconductors, and insulators.

Conductors are materials with σ between 106 and 104 Ohm–1·cm–1. Semiconductors have σ values between 104 and 10–10 Ohm–1·cm–1. Insulators (dielectric materials) have conductance below 10–10 Ohm–1·cm–1. Schematically these classes may be represented as depicted on Fig. 3.1.180

Semimetals are materials, whose electrical properties lie in between of metals and semiconductors. Semimetals have considerably lower electrical conductivity due to lower carrier density, compared to metals and much weaker temperature dependence of electrical conductivity on temperature compared to semiconductors. Semimetals also possess electrical conductivity at the absolute zero of temperature (which is

150 151

characteristic of metals), while semiconductors are insulators under the same conditions.

Examples of semimetals are graphite, α-As, α-Sb, Bi, Po, At. Structurally, most semimetals are anisotropic heterodesmic crystals.181 That is, the chemical bonds differ by their energy (and sometimes by type) in different directions of the crystal.

Figure 3.1. Conductivity domains of metals, semiconductors, and insulators.180

The classification into the four named classes of the materials by the values of electrical conductivity does not reflect, however, the fundamental differences between them, which are much more complex than simply the conductance values. The sign of temperature dependence of the conductivity and whether or not the material conducts at absolute zero are the ultimate criteria to delineate metals from insulators. While there is no sharp, well-defined border between semiconductors and insulators (for

151 152

semiconductors are insulators), the physical properties of typical representatives of each class are markedly different.

The general phenomenological equation for conductivity is182:

σ = μ + μ Ne e Pe h , (3.1) where N and P are the concentrations of negative (electrons) and positive (holes) charge carriers of charge e. The mobility μ is the magnitude of the drift velocity v (that is, average velocity, attained due to electric field) of a charge carrier per unit electric field E:

v μ = , (3.2) E The mobility is defined to be positive for both electrons and holes, though their drift velocities are opposite in a given field. The mobility in SI is expressed in m2/V·s and in practical units, cm2/V·s.

Table 3.1 gives experimental values of the mobility at room temperature for some inorganic semiconductors. The highest mobility observed in a bulk semiconductor is 5×106 cm2/V·s in PbTe at 4 K.182

Table 3.2 summarizes the key differences between metals, semimetals. semiconductors, and insulators. The conductivity of metals decreases with temperature, while conductivity of semi-conductors — increases, and is strongly dependent on temperature. For example, Fig. 3.2 shows the concentration of electrons in different materials, and Fig 3.3 — the dependence of electron concentration (and thus, conductivity) on temperature in Ge and Si. 182

153

Table 3.1. Carrier mobilities μ in some crystalline inorganic semiconductors at room temperature, in cm2/V·s. 182

Crystal Electrons Holes Crystal Electrons Holes

Diamond 1800 1200 GaAs 8000 300

Si 1350 480 GaSb 5000 1000

Ge 3600 1800 PbS 550 600

InSb 800 450 PbSe 1020 930

InAs 30000 450 PbTe 2500 1000

InP 4500 100 AgCl 50 –

AlAs 280 – KBr (100 K) 100 –

AlSb 900 400 SiC 100 10–20

Table 3.2. Key differences between metals, semimetals, intrinsic inorganic semiconductors, and insulators at room temperature.

Parameter Metal Semimetal Semiconductor Insulator

Concentration of charge carriers, N, per structural unit ~1 10–2…10–5 10–6…10–10 <10–10 N, per cm3 1023 1021…107 1013…1017 <1013 Energy gap, eV 0 ~0 >0…6 >6

Mobility, μ, cm2/V·s ~10† 105…107 >0…105 ~0

Temperature dependence, dσ/dT – – + +

σ, Ohm–1·cm–1 106…104 104…102 104…10–10 ~10–10

† The notion of mobility has little use in metals. Calculated from Al σ by (3.1).

154

Figure 3.2. Conduction electron concentrations in different materials.182

Figure 3.3. Temperature dependence of conductance electron concentration in Ge and Si.182

155

Figure 3.4. Effect of doping impurity (Sb, donor) concentrations on the resistivity of Ge

as a function of inverse temperature.183

The band theory of inorganic semiconductors is very well elaborated.184 It explains the properties of semiconductors by an energy gap (forbidden zone) between the populated (filled) valence band and the unpopulated (vacant at 0 K) conduction band

156

(Fig. 3.5). At temperatures above 0 K a fraction of the electrons may be thermally excited from the valence band into the conductance band, creating a conducting electron and a hole, capable of hole-type conduction. The theory gives the following expression for the product of concentration of electrons and holes in an intrinsic semiconductor:

⎛ − E ⎞ = 3 g NP CT exp⎜ ⎟ , (3.3) ⎝ kBT ⎠ where C is some constant, which depends on material T is temperature, Eg is energy gap

(also depends on temperature), and kB is the Boltzmann constant. From this formula it follows that the concentration of charge carriers rapidly increases with temperature (Fig.

3.3), in striking contrast to metals, where, though the charge carriers’ density stays about the same as the temperature rises, the conductivity slowly decreases due to increasing electron-phonon interactions (scattering on the vibrations of the crystal structure units).

Conductance Band Conductance Band a b

e–

Forbidden Band E Forbidden Band Eg

g Energy Energy

e– p+ e–

Valence Band Valence Band k T = 0 Kk T > 0 K

Figure 3.5. Band structure of an intrinsic inorganic semiconductor.

157

According to the band gap theory, a semiconductor is a material which is an insulator at 0 K (has zero charge carriers at absolute zero of temperature), but whose energy gap is small enough (0…6 eV, typically 0…2 eV) for thermal excitations of electrons to the conductance zone, resulting in observable electrical conductivity at T > 0

K.184

Organic semiconductors185 are fundamentally different from inorganic ones.

All solid inorganic semiconductors are coordination compounds, while organic semiconductors are molecular crystals or glasses and have a different mechanism of conductivity. The origin of the conductivity in organic semiconductors lies in the multicentered, delocalized, conjugated π-bonds of organic molecules. The conjugation gives delocalization energy gain, resulting in a decrease of HOMO-LUMO gap for a given molecule. In conjugated polymers this energy gap may be comparable to thermal energy kT. The conductivity in organic semiconductors is primarily due to electron transitions between discrete levels (not merged into bands or zones) of molecular orbitals with different energies. The band theory, developed for crystalline inorganic semiconductors, is often applied to the organic ones and frequently gives correct predictions,186 but may not be quite adequate187 or not even valid for molecular structures, where electronic levels cannot be considered as collectivized between molecules — at least at room temperature.188

The mobility of charge carriers in many of the highest mobility organic semiconductors at room temperature is of order of 1 cm2/V·s and strongly depends on: purity (assay of the material and absence of impurities/dopants), state (crystal or

158

amorphous — in the latter case the mobility in them decreases to 10–3…10–5 cm2/V·s), lattice defects (if the material is crystalline), crystallographic axis direction (if crystalline and structurally anisotropic), or director (if liquid crystal). Table 3.3 gives several representative values of electron and hole mobilities in organic molecular crystals.189

Table 3.3. Carrier mobilities (of electrons e– and holes p+) in some organic molecular crystals at room temperature: μ, cm2/V·s.189

Crystal e– p+ Crystal e– p+

Benzene 1.5 – Perylene 2.0 –

Naphthalene 0.63 1.5 1,4-Diiodobenzene – 12.0

Anthracene 1.73 1.13 Terphenyl 0.34 0.8

Durene 8.0 5.0 Phenazine 0.29 –

Biphenyl – 0.42 Azulene 0.2 0.19

Pyrene 0.7 0.7 Phenothiazine 5.0 –

Tetracene – 0.85 Anthraquinone 0.02 –

1,4-Dibromonapthalene 0.017 0.66 Benzophenone 0.16 –

The purity of organic semiconductors is of no less importance than in the case of inorganic ones. For example, on Fig. 3.6 mobility data are shown for two monocrystalline samples of perylene: (a) ultrapurified (with zone refining of metallic potassium treated perylene), and (b) conventionally purified.190 The two samples

159

obviously have very different temperature dependences. Only the ultrapure perylene shows an increase in mobility almost up to 100 cm2/V·s.191

Figure 3.6. Mobility of (a) ultrapurified and (b) conventionally purified perylene.191

The liquid crystalline state of the matter is a mesomorphic192 (μεζο — intermediate, between, and μορφοσ — form, beauty, outward appearance) phase with a long-range orientational order and either partial positional order or complete positional disorder.193 The molecular crystalline structure of liquid crystals (LC) is intermediate between that of three-dimensional crystals and one of liquids. According to their nature, they may well be termed as anisotropic liquids.194,195 Consequently, liquid crystals possess the properties of both crystals and liquids.196,197 Liquid crystals have two main subclasses: thermotropic and lyotropic. The thermotropic LCs are compounds that form a

160

mesophase by heating a crystal or cooling an isotropic liquid. The lyotropic LCs form the mesophase by dissolving an amphiphilic mesogen in suitable solvent, under appropriate conditions of concentration and temperature. There are also amphotropic mesogens, which can exhibit both thermotropic and lyotropic properties. We will be dealing here only with thermotropic LCs.

Columnar liquid crystals are compounds (mesogens) that under suitable conditions of temperature, pressure and concentration can form a mesophase, wherein molecules are stacked in columns. Discotic liquid crystals are mesogens with relatively flat, disc- or sheet-shaped molecules. All discotic mesogens can form columnar mesophase(s).

Liquid crystal semiconductors (that is, organic molecular crystalline semiconductors in a liquid crystal state) benefit from the (a) order and anisotropy and (b) easy processability of the LC phase. The long-range orientational and (in some cases) short-range positional order of the organic molecules in LC phase cause the columnar

LCs to generally show higher mobilities than the same compound in liquid phase.198

More ordered (compared to nematic) smectic phases of calamitic (rod-like) liquid crystals and the columnar phases of discotic liquid crystals show electronic and hole transport199 up to 0.1 cm2/V·s 200 by either a hopping or (in the most ordered systems) a band transport mechanism.201,202 The conductivity obtained is usually highly anisotropic — 1000:1 parallel to perpendicular with respect to the director.203

The organic semiconductors represent an attractive area of research from theoretical and — especially — from a practical viewpoint. The mobilities measured in

161

new organic and liquid crystalline semiconductors pose a challenge to the classical band theory and thus stimulate new models, insights, and theories on the mechanism of charge transport in molecular crystals. Application-wise, organic semiconductors are candidates to compete with and possibly replace amorphous silicon in industry. The obvious advantages are the ease of processability, absence of grain boundaries, spontaneous self- alignment between interfaces, and an ability to self-repair due to their inherent dynamic nature.199

We studied the synthesis, purification, trace analysis, thermal properties, phase diagrams, and mobilities of several low molecular weight semiconductors and a few liquid crystalline semiconductors. The syntheses of tail-functionalized polyacenes are provided below, followed by iodoarenes and triphenylene derivatives.

3.2. Polyacenes

Our targets were new n-acenes (n=3 anthracene, n=4 tetracene, and n=5 pentacene) with two, four, six, or eight substituents (Scheme 3.1). We expected that some of them might exhibit liquid crystalline and semiconducting properties in analogy to the behavior of other multiple tail decorated polynuclear hydrocarbons such as triphenylene and hexa-substituted benzene esters.

162

R R R R R R R

R R R R R R R R

Scheme 3.1. Target polysubstituted n-acenes. R = alkyl or alkoxy.

3.2.1. Anthracenes

For many years anthracene was one of the best known organic semiconductors.204 It was obtained in pure enough form to observe a cyclotron resonance at 2 K.205 We aimed to modify the anthracene core to obtain materials, that could posses a columnar mesophase. Nematic liquid crystals containing 2-mono-substituted and 2,6- disubstituted anthracene moieties in the core have been long known.206 Recently, their charge transport properties in the nematic phase have been reported to be about 2·10–3 cm2/V·s, which is quite high for a disordered nematic phase and may be explained by some π-stackings, induced at the short range by the mesophase structure.207 (In the nematic phase there is only long-range orientational order, and no positional order.)

Nematic 9,10-bis(phenylethynyl)anthracenes were shown to posses luminescent208 and fluorescent209 properties. Some 2,6,9,10-tetrasubstituted anthracenes for liquid crystals have been patented.210 1,4,5,8-Tetrasubstituted anthraquinones and anthracenes showed smectic mesophases.211,212 1,2,3,5,6,7-Hexasubstituted anthraquinones are thermotropic

163

liquid crystals, which form columnar mesophases.213,214,215 1,2,3,4,5,6,7,8-

Octa(alkanoyloxy)-substituted ester derivatives of anthraquinone also show columnar mesophase.216 2,3,6,7-Tetra-n-alkylanthracenes are known, non-mesogenic compounds with m.p. 88 (n-C3), 97 (n-C5), and 84 °C (n-C7). 2,3,6,7,9,10-Hexa-n-heptylanthracene has m.p. 74 °C and does not exhibit any mesophases ether.217 2,3-Dialkoxyanthracenes have been prepared, though with difficulties and on a small scale, as efficient organogelators.218 We aimed to synthesize some of the 2,3,6,7-tetraalkoxyanthracenes,

2,3,6,7-tetraalkoxy-9,10-dialkylanthracenes, and 1,2,3,4,5,6,7,8-octaalkylanthracenes.

3.2.1.1. 2,3,6,7-Tetraalkoxyanthracenes

The symmetrical structure of 2,3,6,7-tetraalkoxyanthracene and the increased reactivity of anthracene core’s meso (9- and 10-) positions naturally prompt to trace it in the retrosynthetic analysis to the corresponding symmetrical 2,3,6,7-tetraalkoxy-9,10- anthraquinone. 2,3,6,7-Tetramethoxy-9,10-anthraquinone (130, R=Me) is a known compound [5629-55-0], which may be approached, in turn, via several routes (Scheme

3.2):

(1) from meta-hemipinic (4,5-dimethoxyphthalic) acid anhydride [4821-94-7]

126 by Friedel-Crafts acylation (a) of veratrole with subsequent internal dehydration (b) of the veratroyl-veratric acid 127 in polyphosphoric or sulfuric acid; 219

(2) from veratrole and formaldehyde condensation (c) product 129,220 after the oxidation (g) of the latter with Cr(VI) reagents; and

(3) from the bis(trimethylsilyl) ether of 2,3-butanedione dienol 132 221 by its double [4+2] addition (d) to benzoquinone, hydrolysis/oxidation218 (e) of the adduct 133,

164

and methylation (or alkylation, in general) of the 2,3,6,7-tetrahydroxy-9,10- anthraquinone 134.

O O MeO OMe a MeO OMe MeO O + + CH2O MeO OMe MeO COOH OMe MeO O 126 127 128 b, i, f c O RO OR h RO OR g, i, f MeO OMe

RO OR RO OR MeO OMe R = Me 131 R = Me 130 O 129 f O O O OTMS d TMSO OTMS e HO OH + OTMS TMSO OTMS HO OH 132O O 133 134 O

Scheme 3.2. Various approaches to 2,3,6,7-tetraalkoxyanthracene via 2,3,6,7-tetraalkoxy-

9,10-anthraquinone. (a) AlCl3, PhCl. (b) PPA. (c) H2SO4. (d) EtOH, Δx. (e) NaOH, EtOH,

O2. (f) R–Hal, K2CO3, Me2CO. (g) CrO3, H2SO4. (h) (ChxO)3Al, ChxOH, Δx.(i) BBr3,

CH2Cl2, or Py·HCl melt.

meta-Hemipinic acid [577-68-4], despite its symmetrical and simple structure, turned out to be a not easily accessible compound and required a multi-step preparation.222 We tried to prepare 130 directly by double Friedel-Crafts self-acylation of

3,4-dimethoxybenzoyl chloride 135 (scheme 3.3), but only traces of the target compound were separated out and most of the starting material was recovered. Obviously, the

165

reactivity of acyl-containing benzene ring towards electrophilic substitution is not sufficient, even despite of the two methoxy groups, present in the same ring.

O O MeO Cl OMe a MeO OMe + Cl MeO OMe traces MeO OMe 135 O 130 O

Scheme 3.3. Attempted self-acylation of veratroyl chloride. (a) AlCl3, PhNO2.

Given the commercial availability of the veratric aldehyde (3,4- dimethoxybenzaldehyde) 136 and our experiments in ortho-lithiation reactions en route to

4,5-dimethoxyphthalic dialdehyde (vide infra), we devised a route to 130 starting from

136 (scheme 3.4 and 3.5). First, we (a) brominated veratric aldehyde to 137 with bromine in chloroform. Interestingly, this reaction never went to completion if exactly one equivalent of bromine was used and required ca. 1.2 eq. of bromine, though the excess of the reagent did not produce any dibromo or other byproducts. The aldehyde was (b) protected as either a 1,3-dioxolane or a dimethyl ether (the latter is depicted in the scheme), which was subjected to (c) halogen-lithium exchange with n-butyl lithium and the organolithium compound was condensed with another equivalent of veratric aldehyde to give, after acidic work-up, aldol 139. To avoid the protection of the aldehyde as an extra step in this synthesis, we resorted to (e) an in situ protection by a lithium diethylamide, preformed in the same flask, with almost no decrease in the overall yield.

Then we realized that the bromination step could also be dropped if the amine we used

166

for in situ protection of carbonyl, will be modified to act as ortho-director in the direct lithiation step. Thus, (d) N,N,N′-trimethylethylenediamine (TriMEDA) was employed.

O O O MeO MeO MeO H a H b O

MeO MeO Br MeO Br 136 137 138

d e – + – + O Li O Li MeO MeO N N N MeO MeO c

c c

– + – + O Li O Li O MeO MeO MeO N N O N MeO Li MeO Li MeO Li

42% 33% + 135 34%

OH MeO OMe

MeO CHO OMe 139

Scheme 3.4. Ortho-lithiation approach to 138. (a) Br2, CHCl3. (b) HC(OMe)3, Dowex

50W-X8-100, MeOH. (c) BuLi, TMEDA. (d) BuLi, TriMEDA. (e) BuLi, Et2NH.

The aldol 139 was oxidized by potassium dichromate in acetic acid into veratroylveratric acid, which was then cyclized (without purification) into 130 by heating in sulfuric acid. The overall yield of 130 from 136 was mere 5%.

167

OH O O MeO MeO OMe OMe a MeO OMe b

13% MeO CHO OMe MeO COOH OMe MeO OMe O 139 130

Scheme 3.5. Oxidation and cyclization of 138 into anthraquinone 130.

(a) K2Cr2O7, AcOH. (b) H2SO4 (conc.)

The Diels-Alder condensation of 2,3-bis(trimethylsilyloxy)-1,3-butadiene 132 with benzoquinone didn’t look attractive and promising either, for the employment of the same reagent in the similar [4+2] addition to 1,4-napthoquinone produced only 33% of the desired product accompanied by many byproducts, and didn’t scale up well.218

Nevertheless, we prepared disilyl ether 132 223 and introduced it into reaction224 with benzoquinone (Scheme 3.4). As determined by GC, the known mono-adduct224 was formed predominantly, with only small amount of 133 (identified by its mass spectrum).

Close monitoring of the reaction by GC-MS revealed that the first cycloaddition proceeds fast, and benzoquinone (the limiting reagent) disappears after 8 hrs of reflux in benzene.

Further heating results in formation of dark brown-black tar, insoluble in chloroform, which presumably is a polymerization product of 132. This route has not been pursued any further.

O O O OTMS a TMSO TMSO OTMS + + OTMS TMSO TMSO OTMS 45% O 6% 132 O O 133

Scheme 3.4. Attempted preparation of 133. (a) PhH or EtOH, BHT (0.5 mass %), Δx.

168

Condensations of formaldehyde with veratrole, 1,2-bis(hexyloxy)benzene 140 and 1,2-bis(dodecyloxy)benzene 141 gave the corresponding 2,3,6,7-tetraalkoxy-9,10- dihydroanthracenes 142 [26952-97-6], 143, and 144 (Scheme 3.5). The dihydroanthracene 142 turned out to be quite resistant to oxidation and either did not undergo any oxidation (Ag2O, KMnO4 in H2SO4) or (with CrO3 in AcOH, K2Cr2O7 in

H2SO4) gave an inseparable mixture of compounds, wherein no target anthraquinone has been detected. After many trials, the only successful oxidant that gave any tangible, yet not preparatively valuable, amount of 130 was KMnO4 in aq. NaOH. Later we found that this strange oxidation resistance has been already described in the literature.225 If formaldehyde is substituted for higher aliphatic aldehyde homologs, the condensation with veratrole in presence of nitriles226 produces anthracenes, rather than dihydroanthracenes, and the former can be oxidized to 130 by K2Cr2O7 in AcOH in 40% yield.227

RO a RO OR + CH2O RO RO OR R = Me veratrole 142

C5H11 140 143

C12H25 141 144

Scheme 3.5. Preparation of 2,3,6,7-tetraalkoxy-9,10-dihydroanthracenes. (a) CH3SO3H.

Both 2,3,6,7-tetramethoxy-9,10-anthraquinone 130 and its 2,3,6,7-tetramethoxy-

9,10-dihydroanthracene 142 were successfully deprotected into tetrahydroanthracenes

169

145 and 146 by Py·HCl melt and BBr3 in CH2Cl2 correspondingly (Scheme 3.6). From these four compounds only 142 (perhaps due to its non-planar structure) has sufficient solubility in common organic solvents to be characterized by NMR. Compound 130 was

1 characterized by its H NMR spectrum in hot DMSO-d6 and comparing its melting point

(348 °C) to literature values (340…346 °C).219

O O MeO OMe a HO OH

MeO OMe HO OH O O 130 145 MeO OMe b HO OH

MeO OMe HO OH 142 146

Scheme 3.6. Conversion of methoxy groups into hydroxyl functionalities.

(a) Py·HCl melt. (b) BBr3 CH2Cl2, 0 °C.

228 Aromatization of compounds 142–143 with Bu4NIO4, Pb(OAc)4, and SeO2 did not succeed. Aromatization used for highly alkyl-substituted dihydroanthracenes — with n-BuLi, TMEDA, and MeI 229— was contemplated, but was not performed.

3.2.1.2. 2,3,6,7-tetraalkoxy-9,10-dialkylanthracenes

Several 2,3,6,7-tetraalkoxy-9,10-dialkylanthracenes have been prepared by condensing230 the corresponding 1,2-bis(alkoxy)benzenes (veratrole, 140, 141) with aliphatic aldehydes (acetaldehyde CH3CHO, caproic C5H11CHO, and capric C9H19CHO aldehyde), as depicted in Scheme 3.7 and summarized in Table 3.4.

170

Table 3.4. 2,3,6,7-Tetraalkoxy-9,10-dialkylanthracenes.

# Structure Yield, % M.p., °C

O O 7.4 a 328 147 OOlit. 22231 lit. 323.5,232 316233

C5H11 O O 148 17 a 182 OO C5H11

C5H11 a H11C5O OC5H11 15 149 43 b H11C5OOC5H11 95 C5H11

C5H11 H25C12O OC12H25 150 90 b 36.5 H25C12OOC12H25 C5H11

C9H19 H25C12O OC12H25 151 67 b 32 H25C12OOC12H25 C9H19 a with H2SO4 b with CH3SO3H

171

R1 R1 RO R O OR OH RO a + + RO H O RO R O O R R1 R1

Scheme 3.7. Synthesis of 2,3,6,7-Tetraalkoxy-9,10-dialkylanthracenes.

(a) H2SO4 (aq. 70%) or CH3SO3H, Et2O.

3.2.1.3. 1,2,3,4,5,6,7,8-octaalkylanthracenes.

1,2,3,4,5,6,7,8-Octaalkylanthracenes have been prepared by two methods: (1) aromatic ring extension via Pd-catalyzed reaction234 of 1,2,4,5-tetraiodobenzene with alkynes

(Scheme 3.8, R=C5H11: 6-dodecyne, R=C6H13: 7-tetradecyne) and (2) electrophilic peralkylation of commercially available 9,10-dihydroanthracene with C7 alkyl halide and aluminum chloride in the presence of atmospheric oxygen235 (Scheme 3.9).

R R R R R R I I a R R ++ R R R IIR R R

R = C5H11 152 R R CH 153 6 13

Scheme 3.8. Preparation of 1,2,3,4,5,6,7,8-octaalkylanthracenes by Pd-catalyzed ring

extension reaction. (a) AgOAc, Pd(OAc)2 (cat.), toluene, Δx.

172

C7H15 C7H15 H C C H a 15 7 7 15

H15C7 C7H15 C7H15 C7H15 154

Scheme 3.9. Peralkylation of 9,10-dihydroanthracene with heptyl bromide.

(a) C7H15Br, AlCl3, neat.

None of the anthracene compounds described here exhibited any liquid crystalline properties.

3.2.2. Tetracenes

Although tetracene itself has not shown remarkable semiconducting properties, it is a predecessor of pentacene (vide infra) and a catacondensed1 parent of rubrene.

Rubrene (5,6,11,12-tetraphenyltetracene) shows extraordinary mobility values of 13 cm2/V·s at room temperature 236 and 30 cm2/V·s at 200 K.237 The parent tetracene, although commercially available as benz[b]anthracene, is expensive ($150 per gram) and was prepared (Scheme 3.10) and thoroughly purified before single crystal growth (vide infra). Naphthalene-1,4-quinone was reduced with either zinc dust in acetic acid or

1 Catacondensed (Greek cata: down, under, against; entirely, completely, back) aromatic hydrocarbons have all carbons on the periphery of the ring system (no interior carbons).

173

238 Na2S2O4 in ether/water to 1,4-dihydroxynaphthalene [571-60-8] 155, which was condensed with phthalic dicarboxaldehyde in an ethanolic solution of KOH to tetracene-

5,12-dione 156. The latter quinone was reduced by Meerwein-Ponndorf-Verley method239 with aluminum cyclohexanolate240,241 in anhydrous cyclohexanol in presence

242 of CCl4.

O O OH a OHC + OHC O O 155 OH b O

157 156 O

Scheme 3.10. Synthesis of tetracene 157. (a) Zn, AcOH, Δx. (b) KOH, EtOH, Δx.

2,3-Bis(decyloxy)tetracene was prepared likewise (Scheme 3.11): 4,5- dimethoxyphthalic dicarboxaldehyde [43073-12-7] 158 (preparation see below) was condensed with 1,4-dihydroxynaphthalene 155 in presence of NaOH in ethanol-THF mixture to give 2,3-dimethyltetracene-5,12-dione 159. The latter was heated with molten pyridinium hydrochloride at ~200 °C under an argon atmosphere to give 2,3- dihydroxytetracene-5,12-dione 160, which was alkylated with 1-bromodecane, potassium carbonate, and catalytic amount of potassium iodide in NMP to 2,3-bis(decyloxy) tetracene-5,12-dione 161. This quinone was reduced with cyclohexanol and aluminum

174

cyclohexanolate into 2,3-bis(decyloxy)tetracene 162. In contrast to 2,3- dialkoxyanthracenes, 162 is not an organogelator, is soluble in THF, CH2Cl2, CHCl3, and can be recrystallized from ethanol. Only one crystal to isotropic phase transition and no mesophases have been detected for 162 both on DSC trace and on the hot stage under polarizing microscope at 130–132 °C.

OH O O CHO O a + O CHO O 158 OH 155 159 O b H C 21 10 O O O c HO

O HO O 160 O H21C10 161 d

O 162

Scheme 3.11. Synthesis of 2,3-Bis(decyloxy)tetracene 162.

(a) NaOH, EtOH, THF. (b) Py·HCl melt, 200 °C. (c) C10H21Br, K2CO3, KI (cat.), NMP.

(d) (ChxO)3Al, ChxOH, HgCl2, CCl4, Δx.

175

3.2.3. Pentacenes

Pentacene 163 has attracted much attention as a semiconductor (both in single crystalline, polycrystalline, and amorphous forms) in recent years. In fact, it has established itself as a de-facto standard in the field of organic semiconductors, for it consistently shows very high mobility values243, in structured films — over 1 cm2/V·s.244

163

The conductivity of linear polyacenes was predicted to grow with the number of annealed rings.245,246 And, although the mobility of the best pentacene specimens is already comparable with that of amorphous silicon, there are still at least two problems to be solved: stability and solubility.

All linear polyacenes are aromatic compounds. However, their thermodynamic and kinetic247 stability decreases, while their reactivity increases248,249,250,251 with the number of rings.252 Naphthalene is more reactive than benzene. Anthracene is easily oxidizable at the meso- (9,10-) position to anthraquinone, wherein two separate aromatic rings are more thermodynamically stable than three delocalized ones. Tetracene oxidizes above its melting point. Pentacene solutions are destroyed immediately upon contact with atmospheric oxygen and in its absence neat pentacene can photodimerize.253 Hexacene

[258-31-1] shows similar reactivity and must be handled under an inert atmosphere.

Heptacene [258-38-8] has been long disputed by many authors to exist at all, and the identity of the prepared samples was argued, too. Some believe it is not possible to obtain

176

this compound in pure state.254 Only derivatives of octacene [258-33-3] and nonacene

[258-36-6] are known.255

Commercial pentacene from TCI America ($325 per gram) was found by TGA analysis (Fig. 3.7) to contain up to 34% of impurities and after two years of storage on a shelf in a closed vial was found decomposed. Preparation of pentacene was performed according to Scheme 3.12.

100 ––––––– Tetracene TCI ––––––– AS-2-14 crude pentacene ––––––– Pentacene TCI ––––––– AS-2-14 sublimed twice ––––––– AS-2-14 sublimed once

60 Weight (%) 40 b

20 d

a e

0 20 120 220 320 420 520 Temperature (°C) Universal V4.0C TA Instruments

Figure 3.7. TGA Analysis of (a) tetracene from TCI America; (b) crude pentacene 163;

The main intermediates of the pentacene synthesis, pentacene-6,13-dione [3029-

32-1] (pentacenequinone) 164 and pentacene-5,7,12,14-tetraone [23912-79-0]

(pentacenediquinone) 166 were prepared by (a) base-catalyzed condensation of o-

177

phthalic dicarboxaldehyde with 1,4-cyclohexandione256 and (c) Friedel-Crafts acylation of benzene with pyromellitic anhydride in an autoclave with (d) successive dehydrative cyclization of dibenzoylterephthalic acid in boiling sulfuric acid.257 Both quinones were then reduced with aluminum cyclohexanolate in anhydrous cyclohexanol in presence of

258 CCl4 and HgCl2.

O O CHO OHC a ++ CHO OHC O O 164

b O O

++O O

O O 163

c b

O O O COOH d

HOOC O O 166 O 165

Scheme 3.12. Synthesis of pentacene.

(a) NaOH, EtOH, Δx. (b) (ChxO)3Al, ChxOH, HgCl2, CCl4, Δx.

The quinone 164 was also prepared according to Scheme 3.13: quinizarin (1,4- dihydroxyanthraquinone259) was reduced with sodium borohydride in methanol to 1,4-

178

anthracenequinone 167, required for other reactions (vide infra) and the latter was

260 261 reduced either under sonication with zinc dust in trifluoroacetic acid or with TiCl3 into 2,3-dihydroanthracene-1,4-dione 168. Condensation262 thereof with phthalaldehyde in methanol and pyridine afforded pentacenequinone 164 in 41% yield.

O OH O O a b

O OH O O [81-64-1] 167 168

O O OHC c + OHC O O 168 164

Scheme 3.13. Preparation of pentacenequinone 164 via 1,4-anthracenequinone 167.

(a) NaBH4, MeOH. (b) Zn, CF3COOH, sonication. (c) NaOH, EtOH, Δx.

Purification of the prepared pentacene was performed via vacuum sublimation and single crystal vapor growth in a tube furnace with controlled temperature zones.

Sublimation of the crude product posed significant difficulties, for the crude pentacene contains up to 40% of impurities (see Fig. 3.7, b), which in the process of sublimation form an extremely fluffy residue, and while the sublimation per se is a very effective method of purification (two sublimations remove ~88% of impurities), the inadvertent transport of the fluffy residue on the bottom of a sublimator to the cooled finger may

179

devastate much of the purification effort. To increase the effectiveness of the sublimation, we introduced the following changes into the apparatus setup and procedure:

• the crude material was loaded into the sublimer premixed with either

degreased reduced iron filings (to prevent floating of the fluffy stuff) or coarse

quartz sand (1:1 v/v);

• the layer of the above mixture was covered with a layer of degreased reduced

iron filings;

• the evacuation of the sealed sublimer was performed very slowly, with a

needle valve between the vacuum manifold and the sublimer’s outlet tube;

• the temperature was raised very slowly by means of either an oil bath or a

heating mantle;

• the outer walls of the sublimer were wrapped with a heating glass fiber ribbon

to avoid condensation of the material on parts other than the cooling finger.

The purification was followed by a TGA analysis and the sublimation was repeated in stages until the TGA residue ceased to decrease after another purification stage. The material thus obtained consistently gave ~1…3% residue after a TGA run, though there was no detectable residue after sublimation (performed on neat material, without admixtures). This observation suggested that upon prolonged heating pentacene gradually decomposes even under inert atmosphere. Indeed, TGA analysis of the material from the source zone of the furnace crystal grower, wherein the purified pentacene had spent about three days, showed an increase of the non-volatile residue from 1…3% to ca.

27…30%. In addition, DSC analysis of purified pentacene in a sealed hermetic pan

180

shows an exotherm at 410 °C, immediately following by an exotherm at 411 °C, which may be interpreted as melting with decomposition. Recently a very thorough investigation of pentacene thermal behavior appeared, which states that neat pentacene disproportionates at elevated temperatures.263

Pentacene crystals arrange in the crystal lattice in the order, which is often referred to as “herringbone” packing (Fig. 3.14).264,265 This arrangement diminishes effective face-to-face π-π interactions between the aromatic π-molecular orbitals of pentacene molecules, thus reducing the overlap integral and decreasing the mobility of charge carriers compared to alternative possible packing, wherein pentacene molecules would stack upon each other more or less in columnar arrangement. To facilitate such columnar-like arrangement in the pentacene crystal lattice, we took two approaches. First, we prepared two pentacenes, wherein hydrogen has been replaced with fluorine: 1- fluoropentacene 169 and 2-fluoropentacene 170. A fluorine atom is very similar to a hydrogen atom in many respects: first of all, the size and the polarity of C–F bond. This similarity reflects, for example, in the fact that element “hydrogen” is dot-printed in the main subgroup of the seventh group in the Mendeleev Periodic Table of the Elements, and there were times, when it was considered as one of the halogens only. Our intention was to break the symmetry of the pentacene molecule by replacing one hydrogen atom by one fluorine atom, but not to change significantly the geometrical, electronic, and other properties of pentacene. Fluorine seemed to suit this purpose the best. In fact, after we had completed the synthesis of the two isomeric fluoropentacenes, a paper was published by an Anthony group, implementing similar structural modification approach: they

181

prepared halogenated pentacenes (including 1,2,3,4-tetrafluoro- and 1,2,3,4,8,9,10,11- octafluoro-6,13-bis(2-diisopropulsilylethynyl)pentacenes) and showed that some of those compounds switch in crystal structure from herringbone packing to face-to face stacking.266

H H H H H H H H H H H H H H

H H F H F H H H H H H H H H 170 169

The approach to 1-fluoropentacene 169 included intermediate, similar to the one in the parent pentacene synthesis: 1-fluoropentacene-6,13-dione 173. One approach (a) to

173 was based on the strategy, depicted on Scheme 3.13, namely, we contemplated to condense 2,3-dihydro-1,4-anthracenequinone 168 with 3-fluorophthalic aldehyde 172 and reduce the resulting 1-fluoropentacenequinone 173. Another approach (c) to 173 was based upon Cava’s methodology267 and reactivity of o-quinodimethanes.268,269

a b

Figure 3.8. Crystal structure of (a) pentacene270 and

(b) 1,2,3,4-tetrafluoro-6,13-bis(2-diisopropulsilylethynyl)pentacene.266

182

O O OHC a + OHC O F F O 168 172 173 b c O

+ F O F 167 174 169

Scheme 3.13. Synthesis of 1-fluoropentacene 169. (a) EtOH, Py, KOH. (b) (ChxO)3Al,

ChxOH, HgCl2, CCl4, Δx. (c) DMF, Δ.

Since both 3-fluorophthalic dicarboxaldehyde 172 and 3-fluoro-o- quinodimethane 174 retrosynthetically trace to 2,3-dimethylaniline (2,3-xylidine) 175, we explored both approaches (Scheme 3.14). 2,3-Dimethylaniline was (a) diazotized,271 the diazonium salt was precipitated with fluoroboric acid,272 and thermally decomposed to 1- fluoro-2,3-dimethylbenzene 176. The latter was (b) radically dibrominated273 into the side

274 chains with N-bromosuccinimide (NBS) and benzoyl peroxide (Bz2O2) in CCl4 under

UV irradiation. The presence of two initiators (Bz2O2 and UV light) is due to the very long induction period, which was observed (even in boiling CCl4) when only one initiator was employed. Both 1-fluoro-2,3-dimethylbenzene 176 and 1,2-bis(bromomethyl)-3- fluorobenzene 177 were subjected to radical tetrabromination under conditions similar to the above, with either NBS or bromine, but these reactions has never been observed to run to completion: an inseparable mixture of dibromo-, tribromo-, and tetrabromo- 178

183

isomers was always detected by NMR and GC. We found that this is a general outcome for this type of reaction: “The direct bromination of two aromatic methyl groups is a very unsatisfactory procedure. The yields are low, and in most cases a mixture of isomers of similar properties is formed, from which the isolation of definite compounds is difficult.”275

a b Br c Br Br Br

NH2 F F F Br 175 176 177 178 g e d Br (Br) CHO Br g OH f Br OH CHO F Br F (Br) F F 174 179 172

Scheme 3.14. Synthesis of the key intermediates for 1-fluoropentacene.

(a) NaNO2, HCl, 0–5 °C ; HBF4, Δ. (b) NBS, Bz2O2, hν, CCl4. (c) Br2, AIBN, hν. (d, e)

K2CO3, H2O, Aliquat 336. (f) (COCl)2, DMF, –20 °C, CH2Cl2. (g) NaI, DMF, 70 °C.

Therefore, instead of obtaining 3-fluorophthalic dicarboxaldehyde 172 by (d) hydrolysis of 178, we resorted to (e) hydrolysis of 177 with aqueous potassium carbonate and a phase-transfer reagent into fluorodiol 179 and oxidation thereof with a Swern reagent276,277 into 172. Fortunately, quinodimethanes may be formed from both dibromo- and tetrabromo-o-xylenes. The former may also be reacted with Rongalit® (sodium

184

hydroxymethanesulfinate) to form sultines, which upon heating to 80 °C are cleanly converted into o-quinodimethanes.278,279 When 1,2-bis(bromomethyl)-3-fluorobenzene

177 was heated with 1,4-anthracenequinone 167 and sodium iodide in DMF, a dark brown tar was formed, characteristic for the reactions involving o-quinodimethanes, from which by a series of sublimations, 1-fluoropentacenequinone 173 was separated out in

35% yield. The same product was obtained in a near quantitative yield by a condensation of 3-fluorophthalic dicarboxaldehyde 172 with 2,3-dihydro-1,4-anthracenequinone 168.

Reduction with cyclohexanol and aluminum cyclohexanolate afforded 1-fluoropentacene

169 in 12% yield, which after purification by sublimation dropped to 5%.

Our approach to the 2-isomer of fluoropentacene280 170 employed the commercially available 4-fluorophthalic anhydride and did not required C–F bond formation. The anhydride was condensed with hydroquinone in concentrated sulfuric acid, wherein boric acid had been added to prevent side sulfonation.259 The resulting fluoro-quinizarin 180 was reduced (cf. Scheme 3.13) with sodium borohydride in methanol to 6-fluoroanthracene-1,4-dione 181. The latter was condensed281 with 1,3- dihydro-2-benzofuran-1-ol282 — dihydroisobenzofuran-1-ol [81305-98-8], a tautomer of

2-hydroxymethylbenzaldeyde [55479-94-2] — in glacial acetic acid into 2- fluoropentacenequinone 182, which was then reduced to 2-fluoropentacene 170.

185

O OH O OH O a b O + F F F O OH 180 O OH 181 O

F O + d c O F HO 170 182 O

280 Scheme 3.15. Synthesis of 2-fluoropentacene. (a) H2SO4, B(OH)3.

(b) NaBH4, MeOH. (c) AcOH. (d) (ChxO)3Al, ChxOH, HgCl2, CCl4, Δx.

The crystal structure of and carrier mobility in the prepared fluoropentacenes have yet to be determined. The compounds 169 and 170 were characterized by UV-Vis and IR (HATR sampling) spectroscopies; TGA and DSC thermal analyses, and solid- state, magic-angle NMR, for their solubilities in common organic solvents are very low

(enough for recording UV spectrum) and, more important, these solutions react instantly with atmospheric oxygen, irreversibly forming peroxides. For example, a saturated solution of 1-fluoropentacene in degassed benzene has a spectacular deep blue color, which disappears in four minutes, when this solution, unstirred, is exposed to air.

Next, we attempted a synthesis of functionalized pentacenes with long alkoxy chains, contemplating that such compounds could exhibit liquid crystalline properties.

Liquid crystals, containing tetracene or pentacene unit as a core, are unknown to date, to the best of our knowledge.283 The retrosynthetic analysis of the approaches to 2,3,9,10- tetraalkoxy- 183 and 2,3,9,10-tetraalkoxy-6,13-dialkylpentacenes 184 is shown on

186

Scheme 3.16. The grounds for the step (a) are the known aluminum cyclohexanolate reduction of parent pentacenequinone into pentacene.258 The grounds for the step (c) are the reported addition of phenylmagnesium bromide to pentacenequinone, which could be reduced (b) with potassium iodide in acetic acid to 6,13-diphenylpentacene.284 Alkylation

(d) of 2,3,9,10-tetrahydroxypentacenes 184 was anticipated to go smoothly, since alkylation of 2,3-dihydroxyanthracenequinone was reported.218 Deprotection of the

2,3,9,10-tetramethoxy-6,13-pentacenedione 188 was planned to be performed with either

BBr3 (solubility problems anticipated here, though) or pyridinium chloride in its melt.

2,3,9,10-Tetramethoxy-6,13-pentacenedione 188, the key intermediate in this approach, was to be prepared by proven condensation262 of o-dialdehyde 158 with 1,4- cyclohexanedione. In order to accomplish these transformations first we needed a reliable preparation method for 4,5-dimethoxyphthalaldehyde 158.

The known approaches for 158 included: oxidation285 of 4,5-dimethoxyphthalic alcohol,286 obtained either by reduction287 of 4,5-dimethoxyphthalide [531-88-4],

288,289,222b by oxidative cleavage of the veratrole and formaldehyde condensation trimer290; oxidation of 4,5-dimethoxyxylylene chloride [1134-52-7]291; or from reduction/oxidation of the esters of m-hemipinic acid.

187

R1 O O O O R R RR R R R R O O OO 183R1 184 ab

O HO R1 O O O O RRc RR R R R R OOOO 186 O 185 R1 OH d O O HO OH e O O

HOHO OO 188 O 187 O f O O CHO + O CHO 158 O

Scheme 3.16. Retrosynthetic analysis of alkoxypentacenes.

(a) (ChxO)3Al, ChxOH, HgCl2, CCl4, Δx. (b) SnCl2, HCl. (c) R1MgCl.

(d) RX, K2CO3, NMP. (e) Py·HCl, melt. (f) EtOH, NaOH.

188

O O O MeO MeO MeO H a H b O

MeO MeO Br MeO Br 136 137 138

d e – + – + O Li O Li MeO MeO N N N MeO MeO c

c c

– + – + O Li O Li O MeO MeO MeO N N O N MeO Li MeO Li MeO Li

45% 32% + DMF 34%

MeO CHO

MeO CHO 158

Scheme 3.17. Ortho-lithiation approach to 4,5-dimethoxyphthalaldehyde 158.

(a) Br2, CHCl3. (b) HC(OMe)3, Dowex 50W-X8-100, MeOH. (c) BuLi, TMEDA.

(d) BuLi, TriMEDA 189. (e) BuLi, Et2NH.

None of these approaches seemed satisfactory in terms of overall yields, number of steps, or availability of the starting materials. The most attractive approach from practical point of view, starting from veratrole and formaldehyde condensation, gave ambiguous results and varying products in different literature sources. We draw our

189

attention to the facts that (1) the carboxaldehyde groups in the target 158 are in ortho- positions to each other and (2) there has been significant progress in the area of introduction substituents into a phenyl ring specifically into an ortho-position relative to an existing substituent. This methodology has become known as directed ortho- metallation.292,293,294 Based on this new methodology and its application to similar systems,295,296 we elaborated an ortho-lithiation approach to 158 (Scheme 3.17, cf.

Scheme 3.4).

Commercially available veratric aldehyde (3,4-dimethoxybenzaldehyde) 136 was (a) brominated with bromine in chloroform to 137 and then (b) protected as a dimethyl acetal. Dioxolane protection of the aldehyde functionality with ethylene glycol has been tried as well, but it gave much poorer results on subsequent lithiation- formylation, since it tends to undergo fragmentation reaction (resulting in carboxylate and ethylene) under the metallation conditions.297 The protection serves a two-fold purpose: (1) protect the already present aldehyde functionality from the nucleophilic attack of the organolithium reagent used for ortho-lithiation, and (2) form an ortho- directing group. The pool of the carbonyl-derived ortho-directing groups is represented,

298 299 for example, by CONR2, CONHR, cyclohexylimines, α-amino alkoxides,

300 297,301 CH2OAlk, and CH(OR)2.

The protected bromoaldehyde 138 was subjected to (c) halogen-lithium exchange with n-butyl lithium in THF and the organolithium compound was quenched with DMF to give, after acidic work-up, 4,5-dimethoxyphthalic aldehyde 158. To avoid the protection of the aldehyde as an extra step in this synthesis, we resorted to (e) an in

190

situ protection by a lithium diethylamide, preformed from Et2NH and n-BuLi, with almost no decrease in the overall yield. Then it was realized that the bromination step could also be omitted if the amine we use for in situ protection of carbonyl, will be modified to act as an ortho-director and butyl lithium activator in the direct lithiation step. It is known that N,N,N′,N′-tetramethylethylenediamine (TMEDA) greatly accelerates and facilitates lithiation reactions due to intermolecular complexation with butyl lithium via coordination of lithium with the two nitrogens, polarization of C–Li bond, and forming a monomeric butyl lithium species.302

Thus, (d) N,N,N′-trimethylethylenediamine (TriMEDA) 189 was prepared by alkylation303 of aqueous methylamine solution with N-(2-chloroethyl)dimethylamine hydrochloride, prepared in turn from 2-(dimethylamino)ethanol and thionyl chloride.304

The adduct of its lithium salt to carboxaldehyde has been shown to be both ortho-director and lithiation activator.305 By employing tetrahydropyran (THP) instead of THF as a solvent more stable to n-BuLi action,305,306 we attained 45% yield of 158 in a one-pot reaction from commercially available starting material.

We also tried several other approaches to aromatic o-dialdehydes, tested in the preparation of the parent phthalaldehyde: oxidation of side chains with either CrO3 in presence of acetic aldehyde307,308 or chromyl chloride309 (A. L. Étard reaction), and recently reported oxidative decarboxylation of 1,2-phenylenediacetic acid with solid

310 KMnO4. All these approaches gave less than 16% yields. Later another, more convenient route was elaborated,311 based on Blanc bis(chloromethylation) of veratrole.312,313 Hydrolysis of 1,2-bis(chloromethyl)-4,5-dimethoxybenzene with

191

potassium carbonate and subsequent oxidation287 of the resulting diol with freshly prepared MnO2 provided a better access to 158.

MeO MeO CH Cl MeO CH OH MeO CHO a 2 bc2

MeO MeO CH2Cl MeO CH2OH MeO CHO 158 Scheme 3.18. Three-step synthesis of 4,5-dimethoxyphthalic aldehyde 158 from

veratrole. (a) CH2O, HCl (conc. aq.), HCl (gas), dioxane. (b) K2CO3, H2O. (c) MnO2.

After a reliable approach to 158 was established and preparative quantities (ca.

10 g) thereof were prepared, the synthesis of 2,3,9,10-tetramethoxypentacene-6,13- dione314 188 and 2,3,9,10-tetramethoxypentacene 189 has been performed as depicted in

Scheme 3.19. The solubility of 4,5-dimethoxyphthalic aldehyde 158 in ethanol, even at the boiling point, was much lower compared to the parent phthalaldehyde, and use of

THF as co-solvent was necessary in the condensation step. Reduction of the thus obtained tetramethoxypentacenequinone 188 with aluminum cyclohexanolate gave hitherto unknown 2,3,9,10-tetramethoxypentacene 189 — crystalline compound of dark ruby-red color, only very sparingly soluble in benzene. Its solutions are destroyed upon contact with atmospheric oxygen within several minutes. It was purified for this reason by sublimation in high vacuum and characterized by DSC in a sealed hermetic pan (m.p. 413

°C dec.) and by solid-state magic angle NMR.

192

O O O CHO a O O + O CHO OO O O 188 b O O

O O 189

Scheme 3.19. Synthesis of 2,3,9,10-tetramethoxypentacene.

(a) NaOH, EtOH, THF. (b) (ChxO)3Al, ChxOH, HgCl2, CCl4, Δx.

Quite recently Anthony315 prepared a series of pentacenes, structurally similar to

2,3,9,10-tetramethoxypentacene (Scheme 3.20). Their main distinctive feature is substitution in the meso 6,13-position, which makes them more stable in solution.

Removal of the methyl groups from 2,3,9,10-tetramethoxypentacene-6,13-dione

188 by reaction with molten pyridinium hydrochloride gave 2,3,9,10- tetrahydroxypentacene-6,13-dione 187, which was successfully alkylated with 1- iodohexane and potassium carbonate in NMP to provide 2,3,9,10-tetrahexylpentacene-

6,13-dione 185 (Scheme 3.21). The latter compound did not exhibit mesogenic properties and had m.p. 190–194 °C (microscope, hot stage).

193

O O CHO O a O O Z + Z Z O CHO O O O O b (i-Pr)3Si Li Si(i-Pr)3 Si(i-Pr)3 HO O O c O O Z Z Z Z O O O O OH

Z = CH2, (i-Pr)3Si Si(i-Pr)3

Scheme 3.20. Soluble and stable pentacene ethers by Anthony.315

(a) KOH, EtOH. (b) (i-Pr)3Si–C≡C–Li, THF. (c) SnCl2, THF, MeCN.

O O O O a HO OH

OOHOHO O 188 O 187 b O O O c O O RRRR R R R R OOOO 183 O 185 O2 R = C6H13 R = C6H13

O O RRO R O R OO 183a

Scheme 3.21. Attempted synthesis of 2,3,9,10-tetrahexyloxypentacene 183.

(a) Py·HCl, melt. (b) n-C6H13I, K2CO3, NMP, 180 °C. (c) (ChxO)3Al.

194

The attempted reduction of the 2,3,9,10-tetrahexyloxypentacene-6,13-dione 185 did not succeed, however. We believe that the reduction reaction per se was successful, for (1) we observed a distinct color change of the reaction mixture from yellow, characteristic of pentacenequinone 185 to deep cherry-red, characteristic of tetraalkoxypentacene 183, and (2) complete consumption of the starting material by TLC.

Upon working up the reaction mixture with water, a brick-red precipitate was obtained.

Every attempt of purification thereof by chromatography was unsuccessful, however, for even traces of oxygen caused an almost immediate change of color from cherry red to gray-brown, presumably because of formation of peroxo-compound 183a, similar to the case of pentacene.316,317,318 Small-scale sublimation did not yield any material on the cooling finger either. The possible solutions for this problem would be (1) handling and purification in a glove box, under complete exclusion of oxygen, (2) same-pot preparation of Diels-Alder adduct, which shall be purified in solution and exposed to deprotection under an inert atmosphere later. An example of the second approach has been published by Afzali.319,320

N O S N SO a + O 120–200 °C

Scheme 3.22. Reversible Diels-Alder adduct of pentacene.

319 (a) MeReO3, CHCl3, Δx.

195

3.3. Iodoarenes

3.3.1. Why Iodine?

In general, charge mobility due to a hopping mechanism321,322 depends, amongst other factors, upon the overlap integral of the electronic wavefunctions. For example, for a small polaron hopping model the mobility expression goes as:323

2 2 − ea J Ea μ = e kT , 4E kT kT a h π where e is the carrier’s charge, a is the hopping distance, h is reduced Plank’s constant, k is Boltzmann constant, T is temperature, Ea is activation energy, and J is an overlap integral: = ψ *ψ = ψ ψ . Polynuclear acenes have high mobilities due to J ∫ a b dV a b overlap of their π-orbitals, which significantly increases in case of π-stacking.243,324 Other types of orbitals may contribute to the overlap integral, too. In order for this contribution to be significant, the orbitals should be bulky (diffuse in space), just like the π-orbitals.

The p-orbitals of iodine seem to suite this purpose well: iodine’s van-der-Waals radius is

215 pm (picometers).325,326 For example, the charge mobility in zone-refined 1,4- diiodobenzene (DIB) single crystals has been reported as high as 12, 4, and 1.7 cm2/V·sec in the a, b, and c directions of the orthorombic unit cell,327 which is an order of magnitude higher than the highest mobility values for pentacene reported so far.243,244

196

The computer-aided density functional theory calculations for a series of iodine- containing low molecular weight aromatic organic compounds, made by Prof. Ellman,328 revealed, for the case of 1,4-diiodobenzene, that (1) the shortest iodine-iodine and carbon-iodine distances (1.47 and 1.9 Å) between two nearest DIB molecules is less than the sum of two iodines’ (2×1.4 Å) and iodine–carbon (1.4 + 0.7 Å) Slater radii. This suggests the possible principle role bulky electronic orbitals of iodine may play in the electronic properties of the DIB crystal. Fig. 3.10 shows the intermolecular overlap of the iodine’s p-orbitals in red color. (2) Iodine’s contribution to the HOMO molecular orbital of DIB is the largest.

Figure 3.9. Interatomic distances in α-DIB unit cell.

197

Figure 3.10. Intermolecular iodines’ p-orbital overlap in crystalline α-DIB.329

Therefore we purified some of commercially available iodoaromatic compounds, prepared several of them, and elaborated reliable purification protocols for their use as organic single-crystal semiconductors.

The pool of preparative methods for introduction of iodine atoms into aromatic molecules includes274 iodo-de-diazoniation1 (Sandmeyer reaction of aryldiazonium salt with iodide anion), halogen exchange,330,331,332 iodo-de-metallation333 (of electrophilic metalloorganic species: e.g. reaction of Grignard, aryl lithium, aryl thallium,334 aryl nickel,335,336 or aryl mercury,337 reagents with free iodine), and various direct iodination

1 The organic reaction naming convention, elaborated by Jerry March takes the group being introduced (iodine), and separates it by reaction type (overall formal replacement, thus ‘-de-’) from the group being replaced (diazo group in this case).

198

methods338 (electrophilic attack of «I+» species339 onto the aromatic system), including electrochemical iodination.340,341

3.3.2. Direct iodination

Direct iodination of aromatic compounds is electrophilic aromatic substitution of

+ – – – – hydrogen by «I » species, usually prepared in situ from I /I O,3 I/I O,4 I2/HNO3/H2SO4, or other iodine-containing reagents. We used the above combinations of inorganic iodine salts, as well as tetraiodoglycoluril, benzyltrimethylammonium dichloroiodate, and bis(pyridine)iodonium (I) tetrafluoroborate, prepared according to the literature procedures as described below.

Tetrahydroimidazo[4,5-d]imidazole-2,5-dione 190 was prepared from glyoxal and urea,342,343 brominated to 191,344 and the resulting tetrabromide 191 was converted345

346 347 into 2,4,6,8-tetraiodoglycoluril (I4Glu) 192 and used for iodination in 90% sulfuric or trifluoroacetic346 acid media.

O O O Br Br I I O CHO a HNHN b NN c NN + H N NH 2 2 CHO HNHN BrNN Br IINN O 190 O 191 O 192

Scheme 3.23. Preparation of 2,4,6,8-tetraiodoglycoluril.345

(a) H2O, HCl, pH=1.5…2.0. (b) H2O, Br2, pH=9…10. (c) I2, Ac2O.

199

Benzyltrimethylammonium dichloroiodate (BnMe3NICl2) was prepared from benzyltrimethylammonium chloride and iodine monochloride.348 Bis(pyridine)iodonium

(I) tetrafluoroborate synthesis started from preparation of mercury (II) oxide –

349 tetrafluoroboric acid impregnation on silica gel. This impregnation HgO–HBF4/SiO2

+ – 350 was treated with iodine in dichloromethane to yield Py2I B F,4 which was used for iodination with triflic acid.351 Both reagents are yellow crystalline solids and may be stored, after recrystallization, in pure form on the shelf for at least several months without decomposition.

The following iodoarenes (Scheme 3.24) have been prepared by the direct iodination methods (the reagents used are specified in parentheses): diiododurene352,353

+ – 354 193 (I4Glu/H2SO4/dioxane; Py2I B F,4 KIO4/I2/H2SO4; HIO4·2H2O/I2/H2SO4 ),

355,356 iodopentamethylbenzene [64349-91-3] 194 (KIO4/I2/H2SO4; HIO4·2H2O/I2/H2SO4),

357 1,2,4,5-tetraiodo-3,6-dimethylbenzene [27059-93-4] 195 (HIO4·2H2O/I2/H2SO4),

358,359 358,360 1,2,4,5-tetraiodobenzene 196 (HIO4·2H2O/I2/H2SO4), hexaiodobenzene 197,

2,6-diiodo-4-methylaniline 198 (I2/NaHCO3; ICl; BnMe3NICl2), 2-iodo-4- methylacetanilide 199 (ICl; HIO4·2H2O/I2/AcOH/H2O; BnMe3NICl2).

200

I I I I I I I

I I I I I I I 193I 194 195 196

I I I

I I I I I NH NH I 2 197 198 199 O

Scheme 3.24. Iodoarenes prepared by direct iodination.

Noteworthy, hexaiodobenzene has been extensively studied in the last few years and has been shown to change its crystal361 and molecular362 structure under high pressure, shows insulator-to metal transition and becomes a superconductor at 35GPa and

2 K. Some authors363 present evidence that the increase in conductivity is mainly due to enhanced charge transfer interaction generated by the intermolecular overlap of a 5pz orbital of I and a p-orbital of C, while others364 argue the significance of the overlap integral increase in superconductivity of C6I6, though support this mechanism for iodanil

(that is, iodanil's conductivity does improve from the overlap integral increase, while cause of C6I6 metallization is different).

3.3.3. Iodo-de-diazoniation

The Sandmeyer reaction has been the most widely used method for preparation of iodoarenes, for, in fact, it was the only main general method known to organic

201

chemists up until the 1950s.338 According to this method, still widely employed, an aromatic amine is diazotized into a diazonium salt, which is then subjected to solution of iodide anion, resulting in nitrogen evolution and overall displacement of diazo-group with an iodine atom.274 Modern modifications of the Sandmeyer reaction have been elaborated as well.365 We employed the classical version of Sandmeyer reaction for the synthesis of 3,5-diiodotoluene366,367 [49617-79-0] 200, 3,4,5-triiodotoluene366,367,368

[89677-87-2] 201, and 3,4-diiodotoluene366,369 202 (Scheme 3.25).

BnMe3NICl2 a b

I I I I I I + NH2 NH2 N2 I 198 201 d, e c

a, b

I I I I HN I

O 199 202 200

Scheme 3.25. Iodo-de-diazoniation approach to some iodotoluenes.

(a) NaNO2, H2SO4, 0…5 °C. (b) KI, H2O. (c) H2O, r.t. (d) Ac2O, AcOH, Δx.

(e) HIO4·2H2O/I2/AcOH/H2O.

202

3.3.4. Halogen exchange

Direct iodination of many substrates is either difficult to accomplish, or it gives mixtures or undesired isomer(s). For example, we needed an easy access to highly pure

1,4-diiodonaphthalene370 [36316-83-3] 203 and tried to prepare it by direct electrochemical iodination of naphthalene,340,341 but only incomplete reaction and a mixture of various isomers of different degree of iodination has been obtained. The only reported attempt on direct iodination of 1,5-dialkoxynaphthalene was also negative.371 On the contrary, highly selective bromination of naphthalene has been recently reported.372

Therefore, 1,4-diiodonaphthalene has been prepared by halogen-exchange reactions.

There are several variants of such reaction, involving formation of various organometallic intermediate species of Li, Cu, Ni, Tl and Hg. We explored the lithium, copper, and nickel-mediated reactions (Scheme 3.26). Nickel-mediated halogen exchange of 1,4- dibromonapthalene gave unsatisfactory results to make 1,4-diiodonaphthalene, both in yield and purity. Copper-mediated variant worked well in terms of yield both with and without HMPA, originally reported as solvent. However, the conversion of 93% (after one run) left us with a mixture of dibromo and diiodonaphthalenes, which we failed to separate efficiently. Use of excess of KI and repeated re-subjection of the separated products to the same reaction conditions improved the purity of the product via increase of the conversion rate, but 1-bromo-4-iodonaphthalene was always still detectable by

GC-MS as a major (3% after first run) impurity even after two subsequent runs. Thus, we resorted to lithium-bromine exchange reaction with subsequent quenching of the dilithio- derivative with iodine.371 The use of tert-butyl lithium373 was advantageous over n-BuLi,

203

for the metal-halogen exchange proceeds irreversibly (with iso-butylene escaping as gas) and no similar by-products (like 1-bromo-4-iodonaphthalene) are formed. The main byproduct detected in tiny amount by GC-MS was 1-iodonapthalene, which was easily separable by repeated recrystallization.

Br I Br a b + or c Br I I d or e

Li I I f +

Li 202 I

Scheme 3.26. Routes to 1,4-diiodonaphthalene.

(a) Br2, CH2Cl2, –30 °C. (b) KI, Ni powder, DMF, Δx. (c) KI, CuI, DMAc (HMPA),

160…180 °C. (d) n-BuLi, Et2O, –20 °C. (e) tert-BuLi, Et2O, –20 °C. (f) I2.

A positional isomer of 202, 2,3-diiodonaphthalene374,375 [27715-43-1] 203 has been prepared from 2,3-dihydroxynaphthalene (Scheme 3.27) via a Bucherer reaction,376 followed by diazotation of 2,3-diaminonaphthalene and Sandmeyer reaction. The low yield (18%) of the Sandmeyer reaction may be excused perhaps due to the ortho- relationship of the substituents, which make the intermediate bis(diazo)cation presumably less stable and prone to decomposition.

204

OH NH + a 2 b N2 c I

OH + NH2 N2 I 203

Scheme 3.27. Preparation of 2,3-diiodonaphthalene 203.

(a) NH3·H2O aq. 28%, NaHSO3, 140…160 °C, pressure vessel.

(b) NaNO2, H2SO4, 0…5 °C. (c) KI, H2O.

The prepared iodoarenes have been thoroughly purified by repeated preparative column chromatography, series of careful recrystallizations and zone refinings, and single crystals of some of them were grown either by vapor furnace or Bridgeman methods. The single crystals obtained are currently being studied for charge mobility in the laboratory of Professor Brett Ellman.

3.4. Liquid Crystal Semiconductors

3.4.1. HAT Discotic Liquid Crystals

2,3,6,7,10,11-Hexaalkyloxytriphenylenes (HAT-n) constitute the single most important subclass in the realm of discotic liquid crystals, largely because of their applications as quasi-one-dimensional conductors377 and photoconductors.378 In fact, one compound from this family, 2,3,6,7,10,11-hexapentyloxytriphenylene (HAT-5) [69079-

52-3] 205 has established itself as a de-facto media standard for study of charge mobility in columnar liquid crystals. The chart on Fig. 3.18 shows total number of publications

205

(registered in CAPLUS database as per March of 2006) vs. alkyl chain length of HAT-n compounds. Note that the first three members of this homolog series are not liquid crystals, and the first member has been known for years (from 1965379) and deserved its attention as the series’ progenitor.

140

120 CnH2n+1 O CnH2n+1 100 O H C 2n+1 n O

80 O H2n+1Cn O C H 60 O n 2n+1

CnH2n+1

40 Number of Publications

0 2468101214161820

Alkyl chain length

Figure 3.11. Significance of various HAT-n compounds represented as number of

publications for each member of the homologous series.

Despite being the de-facto research standard, HAT5 (and HAT6) compounds are still not commercially available, one of the possible reasons being the very high degree of purity required for them to be useful, at least as semiconductor applications are

206

concerned. Generally, symmetrical HAT-n compounds are prepared (Scheme 3.28) by

380 381 oxidative trimerization of 1,2-dialkoxybenzene with chloranil in sulfuric acid, FeCl3,

382 383 384,385 386 FeCl3/Al2O3, MoCl5, VOCl3, and electrochemically.

O O O O

O a O +6e– + 6H+ 0 O E O

O O 204 O 205 O

Scheme 3.28. Oxidative trimerization of 1,2-dialkoxybenzene to HAT-n.

(a) VOCl3, CH2Cl2.

Unfortunately, use of any reagents listed above and any conditions in this trimerization reaction leads to formation of at least two by-products, in amounts minimum 3%: mono-hydroxy-pentaalkoxytriphenylene 206 and α-chloro-hexaalkoxytri- phenylene 207 (Scheme 3.29).385 The amounts of these undesired by-products might be kept at minimum if strictly anhydrous conditions are employed, none or only catalytic

381 (0.3%) amount of sulfuric acid used (necessary only when FeCl3 is used as oxidant), dichloromethane used as solvent, for no reaction happens in THF, MeCN or AcOH,385

207

and the reaction is quenched with anhydrous methanol before addition of water to perform the work-up.

OR OR OH OR

RO RO Cl

RO RO OR 206 OR 207 OR OR

Scheme 3.29. Main by-products of HAT-n synthesis by oxidative trimerization.385

After reviewing the available literature on the mechanism of this trimerization, we came to a conclusion that formation of these by-products cannot be avoided by the virtue of the reaction and the nature of the compounds formed. The standard redox potential E0 for 1,2-dipentyloxybenzene (M) oxidation to triphenylene (T) is 1.05 V,387 while E0 of the next step, oxidation of triphenylene (T) to radical-cation (T+•) is 1.0 V.388

The increase in the anodic potential has been shown to give higher charged species,

++•• 0 +++• 0 dications-diradicals T (ΔE 1-2 = 300 mV) and trication-radicals T (ΔE 2-3 = 450

389 0 mV) and even tetracations (E 1-4 = 2.27 V), if electrolytic reaction is conducted in

388 CH2Cl2/CF3COOH, which stabilizes these species. All these cations are highly susceptible to attack by nucleophiles and at temperatures above –70 °C polymerize very quickly. The formation of dication-diradical T++•• is irreversible above –40 °C.388 A couple of other side reactions have been observed as well.390 The electrochemical potentials of all three oxidants traditionally utilized in HAT-n preparation (FeCl3, MoCl5,

208

VOCl3) are high enough to cause the trimerization of dialkoxybenzene, therefore they will inevitably cause side reactions and form by-products. This is also why all attempts to conduct the preparative trimerization of dialkoxybenzene electrochemically have been inferior to chemical oxidants-employing reactions.391 Therefore, we aimed to elaborate a reliable protocol for effective purification of HAT5 prepared with any of the known methods.

We tried all three reported oxidants and found that FeCl3 gives crude product of the poorest quality, while MoCl5, if employed at ambient temperature, gives crude product of the best quality. The high cost of anhydrous molybdenum (V) chloride, however, made vanadyl chloride our reagent of choice.

After having tried many standard purification techniques and combinations thereof, including those developed by other researchers,392 we arrived at the following purification protocol. Firstly, neutral alumina (as opposed to usually employed silica gel) is activated at 450–500 °C under argon atmosphere for 12 hrs, cooled under argon and transferred into a tightly closed container. Second, commercial activated charcoal is subjected to a two-stage activation procedure: (a) under ambient atmosphere at 550 °C for 4 hrs, followed by (b) twelve hours at 700 °C under argon atmosphere, then cooled under argon stream to room temperature and sieved. The fraction 16–35 mesh is collected. Thirdly, the crude HAT-5 is impregnated onto (five times its weight) activated neutral alumina and left in a thinly distributed layer open to air, but in the dark392 for 12 hrs. This process can be shortened by air suction through the bed of HAT-5 impregnated alumina. The impregnation changes color from white to yellow to brown.

209

A flash suction column with a sintered coarse glass at the bottom is then packed in four layers (from the bottom): (1) a 3…4 mm layer of Celite® 545 (to retain fine charcoal particles and avoid quick clogging); (2) 150…170 mm of activated alumina; (3)

30…35 mm of activated carbon; (4) aged HAT-5 alumina impregnation. The solvents

(hexanes and dichloromethane) must be distilled or of residue-free grade. Gradient elution from neat hexane to 35% dichloromethane in hexane yields, after evaporation of solvents, snow-white crystalline HAT-5, which remains snow-white and does not change color (to pink or purple) upon shelf storage.

Samples of HAT-5 solutions in dichloromethane, chloroform, 1-propanol, iso- octane and mixtures thereof intentionally left exposed to open air and especially to direct sunlight turned yellowish in several weeks. Therefore, all recrystallizations of HAT-5 after the chromatographic purifications (a series of three chromatographies usually suffices if conducted as described above) in (distilled and micron-filtered) 1-propanol and chloroform mixtures were conducted under inert atmosphere and in the dark.

3.4.2. Nitrated HAT-5 Discotic Liquid Crystals

The chemical structure of HAT-n liquid crystals has received quite a few modification attempts in the search for wider mesophase, increase of dipole moment, change of molecular symmetry, etc. Apart from unsymmetrical HAT-n,382 several post- derivatizations of trimerized HAT-5 have been reported: nitration to mono-nitro-HAT-n

210

(MN-HAT-n),393 selective mono-dealkoxylation to mono-hydroxy-pentaalkoxytriphenylene, and bromination.394 We attempted synthesis of di- and tri-nitro substituted HAT-5. Only 1,5,9-trinitro-2,3,6,7,10,11-hexapentyloxytriphenylene (TN-

HAT-5) was isolated in pure form and characterized, while two isomeric dinitro-HAT-5 compounds have been detected by HPLC-MS only and have not been individually isolated due to difficulties in their separation and purification.

OR OR OR OR OR OR NO2 NO2 RO a RO b RO NO2

RO RO RO

OR OR O2N OR OR OR OR

Scheme 3.30. Synthesis of MN-HAT-5 and TN-HAT-5. (a) 1 eq. HNO3, CH2Cl2,

CH3NO2, room temperature. (b) 3.2 eq. HNO3/Al2O3, CH3NO2, –20 to room temperature.

Both mono- and trinitro-HAT-5 isomers exhibit columnar liquid crystalline properties: MN-HAT-5: Cr–Colh –37°C, Colh–I 139°C; TN-HAT-5: Cr–Colh 34°C, Colh–

I 142°C. Both compounds are being measured for charge transport properties in their liquid crystal phases in the laboratory of Prof. Brett Ellman.

211

3.4.3. Conclusions

We have attempted preparation of polyacene compounds, aimed to exhibit liquid crystalline properties. None of the synthesized tail-equipped anthracenes, anthraquinones, and pentacenequinones showed expected mesogenic behavior.

Iodine-containing arenes we prepared and purified are to be employed in further study, pertaining to measurements of their mobilities.

A reliable purification protocol for HAT-5 discotic semiconductor has been elaborated.

212

Chapter IV.

Experimental Part

4.1. General Instrumentation and Techniques

Measurement of Fluorescence Quantum Yield

There are several approaches to measure fluorescence quantum yield.41,395 The most simple and widely employed method is a secondary (or relative) method, elaborated originally by Parker and Rees.396 This method is based on comparing the quantum yield of a sample to that of a standard at the same conditions.397 Ideally, the standard and sample should have absorption and emission spectra matched (overlapped) as close as possible and quantum yield of standard should be of approximately same value as that of sample.398

The standard’s emission spectrum and quantum yield ideally should be independent of excitation wavelength, temperature, and concentration. The standard should be stable in solution, be easily purified or commercially available in pure form, and ideally be quenched with oxygen as little as possible. There are very few compounds that fulfill all these requirements. For example, fluorescein’s fluorescence is easily quenched by oxygen and photo bleached, and its emission spectrum depends on pH of the

213

solution. Rhodamine 6G’s quantum yield is temperature dependent.399,400 Thus, the choice of standard is the most crucial step in the whole technique of relative quantum yield measurement. Some good standards are Rhodamine 101,401 perylene, perylene tetracarboxylates – esters and salts,402 pyrene. Good sources of information on common fluorophore’s properties are http://fluorophores.org and Photochem CAD Database. After a standard has been chosen, the following is the sequence:

• The solvents should be of spectrophotometric grade with no inhibitors and as low cut-off as possible. Chlorinated, nitrated and other heavy-elements containing solvents will generally quench fluorescence.

• Deoxygenate the solvent by passing a stream of argon or helium into it and/or sonicate it (in a special degas mode of the sonicator) at the same time.

• Weigh out precisely 0.1–0.5 mg of sample (i.e. both sample and standard) on microbalances (weighing is optional and can be used for simultaneous extinction coefficient determination). Dissolve the sample in 10.00 ml of deoxygenated solvent of choice and perform a series of dilutions to bring the solution’s absorbance below 0.05.

Generally (with compounds having ε of order 105 l·mol–1·cm–1), it means to dilute to some micromolar concentration range ~10–5 M = 1-10 μM.

• Take the UV-Vis spectrum and note the optical density A at the wavelength, which will be used for excitation (λEx). Note: the samples at micromolar concentrations might have very low absorbances at wavelengths other than λmax. When taking absorbances of dilute samples at such wavelengths, big errors are possible. Thus, it is better to measure, for example, 10× times more concentrated solution (A of penultimate solution, i.e. of that

214

in the dilution series, from which the solution for fluorescence measurement is prepared), taking the optical density (which is taken most accurately within 0.1–0.45 range) at the necessary wavelength and calculating the optical density of the final solution by dividing the obtained number by the dilution factor.1

• Ideally, the dilutions should give you such solutions that the absorbances of the standard and the sample solutions should be exactly the same at the excitation wavelength. At any rate, they should be very close to each other and less that 0.05.

• Record fluorescence spectra of the standard and of the sample solutions as soon as possible after each other and after the absorbance measurements and at EXACTLY the same conditions. Note the conditions – temperature, λEx, PMT voltage/gain, excitation monochromator and detector slits. Recording fluorescence of blank solvent(s) to ensure clear fluorescence background is recommended.

• Chose an excitation wavelength ~20 nm hypsochromic (blue-shifted) off the λmax to get most of the emission spectrum. Start collection of emission at least 10 nm (depends on detector slit) bathochromic (red-shifted) from λEx.

• Keep in mind scattering signal at 2λEx, which should be excluded from integration or be the same for standard and sample (that is, not absorbed by both). Keep fluorescence

1 “…and there is difficulty in determining the low absorbances. This latter problem is generally overcome by accurately diluting more concentrated solutions of measured absorbances and/or using long-path-length cells.”397

215

intensity within specification range of the fluorimeter by adjusting concentration and/or

PMT voltage/gain. Integrate both sample and standard fluorescence spectra.

• Plug the values obtained into the equation (4.1):

2 F()λ dλ A ()λ Φ = Φ n × ∫ F F × ref Ex F F ref , (4.1) n2 F ()λ dλ A()λ ref ∫ ref F F Ex which is an approximation of more precise equations:

− ()λ 2 F()λ dλ − Aref Ex Φ = Φ n × ∫ F F × 1 10 F F ref − ()λ or n 2 F ()λ dλ 1−10 A Ex ref ∫ ref F F (4.2) 2 F()λ dλ 1− exp[]− A ()λ ln10 Φ = Φ n × ∫ F F × ref Ex F F ref , n 2 F ()λ dλ 1− exp[]− A()λ ln10 ref ∫ ref F F Ex where ΦF is quantum yield, n – refractive index of solvent, A(λEx) – absorbance at the excitation wavelength, and F(λF) – fluorescence spectrum trace to be integrated. Note that optical density of the standard goes into nominator, while all other values of the standard

— into denominator.

Measurement of Fluorophore’s Photostability

The photobleaching quantum yield φ b is the slope of the least square fit line relating the photobleaching rate of a sample to the excitation rate of the laser.403 The bleaching curves should be recorded at several excitation intensities (that is, five or more fluxes of the excitation light) and fit to an exponential decay functions (see Fig. 4.1).

Often, two exponents and an offset are required to obtain a good fit for all samples and

216

intensities. Using the exponential fit, the bleaching rate is calculated from the initial (t=0) slope, i.e. from the fastest exponential.404

1.6 RT-3-188

1.4

1.2

1.0

0.8

Absorbance 0.6

0.4

y = A + B*x y = a*exp(b*x) 0.2 R^2 = 0.99409 R^2 = 0.98043 A = 1.33525 ± 0.0089 a = 1.40277 ± 0.02219 B = -0.08765 ± 0.00138 b = -0.1116 ± 0.00303 0.0 -20 2 4 6 8 1012141618202224

Exposure, hrs

Figure 4.1. Photodegradation of compound 93 and its exponential fit.

Although it is possible to control the light intensity by varying the distance from a conventional light source (I ~ d–3), it is a suitable mean for only decreasing the flux.

Moreover, in order to see an exponential photobleaching rate, a source of very strong light intensity is required, i.e. a laser.404 Since we did not have a laser source with a tunable intensity, we were able to perform measurements of photobleaching rates for our compounds at a single and low excitation intensity from a conventional UV lamp.

217

The measurements have been performed on chloroform solutions in a standard

1 cm quartz cuvette, illuminated at λ = 360 nm by a UVP 8W UV lamp model 3UV-38, distanced from the cuvette by 4 cm. The intensity of UV radiation was measured with an

International Light photometer model IL1400 to be 1.2 mW/cm2. For most of the samples we observed a linear decay in fluorescence (or absorbance, since the photobleached dyes are colorless — see Fig. 1.15, 1.16). The tangence of the slope of the least square fit gave the following photobleaching rates at the specified flux: 32 (–0.03059), 33 (–0.17591), 64

(–0.0272), 65 (–0.0574), 91 (–0.01682), 92 (–0.05223), 93 (–0.08765).

Gas Chromatography – Mass Spectroscopy (GC-MS)

All GC-MS runs have been performed on a Thermo Electron Trace GC 2000, coupled to a Polaris Q ion trap MS. The injection of a series of samples has been performed with autosampler AI 3000. The compound being injected into Trace GC must be proven (by usual GC or TGA) to be volatile and contain no non-volatile or thermally decomposable matters. That is, no reaction mixtures or crude products with non-volatile impurities may be injected. The specification sensitivity of Polaris Q MS is 10 picogram of decafluorobenzophenone. Both volume and amount/concentration of sample being injected matter, for the injector liner cannot accept more vapor volume than equivalent to

3 μl of liquid. The injector was usually used in split mode (with split liner installed) with split ratios 10…500. Inject no more than 1–100 ng (nanogram) = 10–9 g = 10–6 mg of

218

sample, dissolved in no more than 1 μl (microliter) = 10–3 ml of solvent. To prepare such solutions, use one of the three methods below. Useful to notice is that 1 ng/μl = 1 μg/ml =

1 mg/liter, and 100 ng/μl = 0.1 mg/ml = 1 mg/10 ml.

Method 1. Prepare Solution#1 by dissolving 1 mg in 10 ml. Take an aliquot of 1 ml and dilute to 10 ml of Solution#2. Take an aliquot of 1 ml of Solution#2 and dilute to

10 ml of Solution#3. Inject 1 μl of Solution#3. If no signal appears on GC-gram, inject 1

μl of Solution#2.

Method 2. Prepare Solution A by dissolving 1 mg in 10 ml. Take an aliquot of 1

μl and dilute to 1 ml of Solution B. Inject 1 μl of Solution B. If no signal appears on GC- gram, take an aliquot of 10 μl of Solution A, dilute to 1 ml of Solution C and inject 1 μl thereof.

Method 3. Prepare Solution Z by dissolving 1 μg (0.001 mg) in 1 ml. Inject 1 μl of Solution Z.

The GC column used was Restek #12623 RTX-5MS 30 m × 0.25 mm × 0.25 μm film, crosslinked 5% diphenyl – 95% dimethylpolysiloxane. Column maximum temperature is 350 °C. The carrier gas used was ultra-high purity helium (grade 5.0) at 1 ml/min with vacuum compensation. Injector nominal temperature was 225 °C. Transfer line nominal temperature was always 300 °C. MS ion source nominal temperature was

200 °C. The complete, illustrated, step-by-step instructions for running a GC-MS experiment can be found in Appendix A.

219

HPLC-MS

High-performance liquid chromatography (HPLC) was performed with an

Agilent 1100 HPLC, equipped with an Agilent «Zorbax NH2» column, which has ID 4.6 mm; ℓ= 250 mm; average particle size 5 μm; inverse phase is medium-polar 3-aminopropyldiethoxysilane. Column volume was calculated to be 4.15 ml; void volume was calculated from hexane void time as ~3ml. Typical flow rates were 1…1.5 ml/min. The mobile phase was a combination of usually two or three solvents from the following: iso- octane (2,2,4-trimethylpentane), chloroform, THF, acetonitrile, methanol. All solvents were either of HPLC grade or were distilled twice on a dedicated rotovap and filtered through 0.3 μm PTFE micron filter. When used as standalone instrument, a diode array detector (DAD) was used to register and identify components in the effluent. Normally,

UV-Vis spectrum of the effluent in the range 190–900 nm (in 2 nm resolution) was recorded at the sampling rate of 2 sec. The HPLC-gram’s ordinate axis was set in correspondence with 254 (350), 360 (500), or 400 (500) nm absorption of the effluent

(reference wavelength in parentheses).

Bruker Daltonics Esquire 3000+ ion trap mass spectrometer was used both standalone for identification of single-component analytes by direct infusion and coupled with the Agilent 1100 HPLC for analysis of multi-component mixtures. Atmospheric pressure chemical ionization (APCI) ion source was generally used. In some cases of low-volatile analytes electro-spray ionization (ESI) ion source was employed. Drying gas temperature was usually set to 300 °C, APCI heater’s temperature varied in the range of

300…450 °C. When used as HPLC detector, the ion trap parameter ICC was set to

220

20,000, the signal was averaged over 7 measurements and “rolling average” option was checked on.

Nuclear Magnetic Resonance (NMR)

NMR spectra were recorded for 1H, 13C, and 19F nuclei. Instruments used were:

Bruker AMX 300 and Bruker Biospin Avance 400 spectrometers with 1H base frequencies 300 (7.05 T) and 400 MHz (9.4 T superconducting magnet) correspondingly.

Both instruments were equipped with 5 mm probes. The 5-mm NMR glass tube should be filled with a deuterated solvent solution to a minimum depth of 5.0…5.5 cm (about

0.60…0.75 ml). The amount (concentration) of sample required for a proton spectrum ranges from less than 1 mg/ml to about 20mg/ml (for a compound with Mw ~ 400). Too much sample can result in a loss of resolution or a distorted 1H spectrum. This includes not just the sample of interest, but any proton source such as protonated buffers, residual protonated solvents, and water. About 5mg/ml is sufficient maximum for 1H. For 13C spectra the higher the concentration used the better. The solution should be free from any solid, such as undissolved solute, or dust. Cloudy solutions were routinely filtered through either a Pasture pipette with a tiny cotton swab, or a syringe filter. The following deuterated solvents were used: chloroform-d (CDCl3, used by default, if another solvent not specified explicitly), acetone-d6, dimethylsulfoxide-d6, and trifluoroacetic acid-d

(CF3COOD), prepared from trifluoroacetic anhydride and D2O. Step-by-step instructions to run a 1D 1H and 13C experiments on Bruker AMX 300 can be found in Appendix B.

221

Thermal Analysis: DSC and TGA

Thermal Analysis (TA) Instruments differential scanning calorimeter (DSC) models 2920 and Q10 were used to routinely measure melting points of the samples.

Both instruments were calibrated in temperature and heat capacity against a certified indium standard. Non-hermetic 5 μl aluminum pan was charged with 1…10 mg of sample, topped with corresponding lid and crimped in a press. A blank pan charged with no sample, also crimped with a lid, served as a reference. Liquid samples were loaded into hermetic pans and crimped with corresponding lid. The sample and reference pans were placed onto their corresponding thermocouples inside DSC cell, and the cell was set to constant purge with nitrogen gas at 5 ml/min. Standard heating rate (ramp) was 10

°C/min, if not specified otherwise. On DSC traces endothermal peaks show up, exothermal peaks — down. Melting point, followed by decomposition is reported as melting point onset1 temperature, followed by “dec.”, while decomposition without concurrent melting is reported as “dec.” followed by onset decomposition temperature.

Thermal stability, decomposition temperature and sublimation range were measured on thermal gravimetrical analyzer (TGA) model 2950 by the same manufacturer (TA Instruments). The instrument was calibrated in temperature with nickel standard by determining Curie point transition, and with calcium oxalate standard in

1 Onset m.p. is independent of the sample amount charged into the pan, while peak maximum shifts to bigger temperature values proportionally to the sample mass.

222

mass. Platinum 100 μl sample pan was tared to zero and charged with no more than 1…5 mg of sample. Standard heating rate (ramp) was 10 °C/min, if not specified otherwise.

M.Braun SPS

M.Braun solvent purification system (SPS) is a safer alternative to solvent stills.405 The solvent is purified therein by passage through a bed of activated alumina, contained in a hermetic steel reservoir. Hydrocarbons can also be deoxygenated by a pass through another column with activated copper. Inert gas is used to push the solvent through the columns. Complete, step-by-step illustrated instructions on how to operate

SPS can be found in Appendix C.

High Pressure Reactors

Reactions, required high pressure (e.g. amination with pyrrolidine or Bucherer reaction) were performed either in glass pressure vessel CG-1880 by ChemGlass, or in

Parr Instrument High Pressure Reactor, model 4563, depending on the load and maximum expected pressure.

Glass pressure vessel CG-1880 can be used for reactions (see, for example, 83), where maximum possible pressure does not exceed ~10…15 atm. It should never be filled more than 1/3 of its volume and always be used behind a safety shield. When aggressive media (like concentrated nitric acid) are employed, the Viton® O-ring needs to replaced with a lead (Pb) gasket, custom-cut from a sheet. This glass pressure vessel has

223

also been employed in combination with Parr Instrument High Pressure Reactor 4563, to run reactions in mineral acids, which would severely attack the stainless steel body of

4563 reactor. Such reaction mixtures were placed inside the glass pressure vessel (sealed with its original Teflon® stopper and Viton® O-ring or with a custom-made Pb gasket), which was placed inside the 4563 reactor, filled with sufficient amount of appropriate solvent. The solvent was chosen such that at the reaction temperature the pressure from the solvent onto the glass reactor would approximately match the pressure from the reaction mixture (e.g. water and benzene mixture for reactions in nitric acid).

Parr Instrument High Pressure Reactor 4563 was used with either a Teflon®or glass insert (liner). The Teflon® liner can be used up to 250°C. The glass liner has the same maximum operating temperature as the reactor itself — 350°C, which limit is imposed by PTFE self-sealing gasket of the reactor. Whenever the reactor is heated above

100 °C, cooling water must be set running through the cooling jacket around the stirrer coupling. The thumbscrew on the drop band (encircling the split ring closure) should fit into the slot in the stand’s bracket to prevent slipping of the whole reactor when the stirrer is turned on. Never operate the heating mantle without reactor lowered into it.

UV-Vis

Ultra-violet and visible (UV-Vis) spectra were recorded on Agilent / HP 8453 diode array spectrophotometer. Samples were dissolved in a suitable solvent of photometric grade and diluted down so that their maximum absorbance would fall within

224

0.1…0.45 range (units of optical density). Standard quartz cuvettes of 1.0 cm path-length were generally employed. Blank solvent was measured before the sample spectrum was recorded. According to Bouguer-Lambert-Beer law, A = ε C l , thus for 1 cm path we get:

A A •V[l] • M[g / mol] A •V[ml]• M[g / mol] ε = = = Cl m[g]• l[cm] m[mg]

For a solution of m mg per 10 ml of solution and 1 cm path:

A• M[g / mol] ε[l • mol –1 • cm –1 ] = ×10 . m[mg]

This formula has been used to calculate the molar extinction coefficients from the UV-Vis spectra of solutions with a known concentration of solute.

Bruker Optics Vector 33 Fourier-transform infra-red spectrometer (FT-IR) was used to record infra-red (IR) spectra. Sampling was performed in either of three ways: (1) neat on the MIRacle™ Horizontal Attenuated Total Reflectance (HATR) stage by Pike

Technologies; (2) solution on the MIRacle™ HATR; (3) between KBr plates. If not specified otherwise, IR spectra are reported as neat (both solids and liquids) on MIRacle™

HATR.

225

4.2. Synthetic Procedures

3,6-Diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione, DPP 1.

N O H N O O t-BuOK + O t-AmOH O 12 O N H 1

DPP was prepared by modification of the published procedure.71 A five-liter reactor was topped with a three-neck lid. The central neck was fitted with a mechanical stirrer, and a thermometer/nitrogen inlet adaptor into another neck. The reactor was flushed with nitrogen and charged with tert-amyl alcohol (3 liters). The third neck was fitted with a reflux condenser and the reactor was heated with stirring on a heating mantle until reflux of the solvent (102°C) began. At that temperature potassium tert-butoxide

(560 g, 5.0 mol) was added in portions and heating and stirring continued until all of the t-BuOK dissolved (the temperature rose to 120°C). After the temperature was lowered to

110°C, benzonitrile (206.24 g, 2.0 mol) was added at once. Di-tert-amyl succinate

(258.35 g, 1.0 mol) was placed into a syringe and added by means of a syringe pump during 4 hrs. In the middle of the addition and near the end of the addition, additional portions of tert-amyl alcohol (500 ml) were added to facilitate stirring. After the addition of succinate was complete, the reaction mixture was stirred at 110°C for an additional four hours, and then cooled to 60°C and methanol (1 liter) was added in 100 ml portions, followed by acetic acid (350 ml). The resulting slurry was filtered on two 2-liter Buchner funnels, and then gradually suspended (with help of sonication) in methanol and filtered

226

from methanol (2×2 liters), hot water (2×1 liter), and boiling ethanol (4 liters). The filter cake was dried overnight to give 161.5 g (56%) of 1 as fine red powder. This material was characterized as 2,5-dimethyl-3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione.

M.p. 231°C (lit.74 m.p. 233-234°C); 1H δ: 7.87 (d, 2H, J=8.7 Hz), 7.61 (d, 2H, J=8.7 Hz),

13 3.31 (s, 3H). C δ: 138.35, 130.50, 29.60. ΦF=0.79.

3,6-bis(4-bromophenyl)-2,5-dimethyl-pyrrolo[3,4-c]pyrrole-1,4-dione, Br-DPP 9.

H H N O N O Br2 gas Br Br N N O O H H

A Petri dish (14 cm internal diameter) was filled with finely ground 3,6- diphenyl-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (22.3 g, 0.077 mol). Bromine (124 g, 0.775 mol) was placed into a 50 ml evaporation dish at the bottom of a desiccator (15 cm internal diameter) and topped with a porcelain insert, over which the Petri dish was placed. The desiccator was left with closed lid and slightly open vent in the dark for ten days. After the reaction was complete, the Petri dish was removed, left open to the air for

2 hours, then placed into a vacuum oven and dried at 75°C: for 2 hours at 40 mm Hg, and for 4 hours at 1 mm Hg to remove absorbed bromine and hydrogen bromide and then was weighed out to check the mass gain (19.8 g, 162% of theory). The crude product was suspended in water (300 ml) and ethanol (200 ml). To that vigirously stirred suspension, sodium hydrocarbonate (sat. aq., 150 ml) was added until CO2 evolution ceased. The

227

neutralized suspension was filtered, and gradually washed with water (200 ml), methanol

(200 ml), ether (100 ml) and vacuum-dried to give 32.5 (94%) of 9 as fine red powder.

3,6-bis(4-hydroxysulfonylphenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione 10.

H O H N O N O H2SO4·SO3 HO S O O S OH N O O N O H H

A 200 ml round bottom flask with a magnetic stir bar was charged with DPP

(6.0 g, 20.8 mmol), and oleum (45 g). The reaction mixture was heated in an oil bath at

40°C for 4 hours, cooled in an acetone – dry ice bath, and a mixture of ice (200 g) and dry ice (40 g) was added slowly. The dark red precipitate formed was filtered through fritted glass. The thick paste on the filter was re-suspended (3×) in acetone (50 ml) aided by sonication, and filtered again to allow, after vacuum drying, 7.2 g (77%) of dark-red

1 powder. M.p. > 400°. H (NaOD, DMSO-d6/D2O) δ: 8.2 (br., 2H), 7.75 (br., 2H), 4.55

13 (br.). C (NaOD, DMSO-d6/D2O) δ: 162.8, 147.8, 144.3, 128.7, 128.2, 126.5, 111.5.

UV-Vis (H2O) λmax: 268, 477, 505. Fluorescence (EtOH) λmax: 525, 566sh. ΦF=0.63.

Di-iso-propyl succinate 11.

O O H SO OH 2 4 O + 2 H O OH + 2 2 OH O O O

228

A two-liter, two-neck round bottom flask with a stir-bar was charged with succinic acid (236.1 g, 2 mol), iso-propyl alcohol (480.8 g, 8 mol), benzene (500 g, Note

1), and sulfuric acid (20 ml). One neck was fitted with a nitrogen inlet adapter, and another – with a short Vigreaux column, topped with a Dean-Stark trap and a condensor.

The reaction mixture was heated with a heating mantle (465 W, 50% of 120 V, 195° mantle temperature) at reflux with a slow stream of nitrogen, introduced at the nitrogen adapter inlet, until no more water was separated in the Dean-Stark trap. The trap should be emptied periodically from separated water, total amount of which was ca. 36 ml, 2 mol

(50% of this amount separates during first four hours, the remaining amount – during additional 20 hrs). After azeotropic removal of water ceased, the solvents were removed on a rotovap at reduced pressure in two stages (first at P=20 mm Hg with water aspirator, then at P=1 mm Hg with oil pump). The residue was vacuum-distilled on a rotovap at

25 406 25 P=0.1 mm Hg, yielding 336 g (83%) of colorless liquid, nD = 1.4151 (lit. nD =

1.4177). The residue after vacuum distillation was extracted with hexane (200 ml, Note

2) in a separatory funnel, hexane was removed from the extract on a rotovap and the residue of the extract was vacuum-distilled as above to give additional 90 g of colorless

25 1 δ liquid, nD = 1.4165. The total yield was 381 g (94%). H NMR : 5.02 (quintet, 2H, J =

6.3 Hz), 2.56 (s, 4H), 1.24 (d, 12H, J = 6.3 Hz) — in accordance with lit. data.407

Note 1. The amount of benzene was calculated, based on the azeotropic data

408 from : PhH:H2O = 91:9 (mass), az. b.p. 69°C. To distill out 2 moles of water, 365 g of benzene are required. However, iso-propanol forms a ternary azeotrope with benzene and water, requiring more benzene. Note 2. At the end of the first vacuum distillation,

229

considerable gas evolution and foaming occurs, indicating possible decarboxylation.

Extraction with hexane helps to remove the component susceptible to decarboxylation, accumulated at the end of the distillation.

tert-Amyl succinate 12.

O O

OEt Li O + HO + EtOH OEt O

O O

tert-Amyl succinate was prepared by a modified procedure of a U.S. Patent409. A two-liter, two-neck round bottom flask with a stir-bar was charged with diethyl succinate

(174.2 g, 1 mol), tert-amyl alcohol (970 g, 11 mol), and lithium (wire, 0.7 g, 0.1 mol).

One neck of the flask was fitted with a nitrogen inlet adapter, and another – with a long

(80 cm), efficient, double-section fractionating column, wrapped with asbestos tape for thermal insulation. The column was topped with a Würtz adapter, thermometer, and a condenser. The reaction mixture was heated with a heating mantle to reflux with a slow stream of nitrogen, introduced at the nitrogen adapter inlet, continuously distilling out ethanol with admixture of tert-amyl alcohol. The amount of alcohol distilled off the flask was supplemented by equal amount of tert-amyl alcohol (350 ml of t-AmOH added in

25 total). The progress of the reaction was monitored by nD of the distillate ( nD of EtOH is

25 25 1.3595, nD of t-AmOH is 1.41021, and nD of (EtO)2Suc – 1.4178). After 38 hrs the reaction mixture was cooled down to ~60°C and the alcohols were distilled off on a rotovap (Note 1), leaving 340 ml of brown-green residue. To that residue, hexane (80 ml)

230

was added and the mixture was consecutively washed with water (200 ml), HCl (3M, 80 ml in 200 ml of water), water (250 ml × 5 times), and brine (50 ml), leaving a solution of light yellow color and strong blue fluorescence under λ=365 nm UV lamp, but noticeable under normal light too. The solution was dried with MgSO4, hexane was removed on a rotovap, and the residue was vacuum-distilled twice on a rotovap with oil pump, followed by traditional vacuum distillation, to give 238 g (92%, cf. to 77% in the patent) of

25 1 δ colorless liquid, b.p. 89–100°C at 0.2 mm Hg. nD = 1.4273. H NMR (CDCl3) : 2.5 (s,

2H), 1.8 (quartet, 2H), 1.4 (s, 6H), 0.9 (t, 3H).

Note 1. The distilled alcohol mixture (~670 ml) was washed three times with water (200 ml), then water-brine (1:1, 200 ml), and brine (200 ml). The water-immiscible layer after all washings was dried over CaCl2, then over Na, and vacuum-distilled twice on a rotovap to give 300 ml of recovered t-AmOH.

4-Fluorobenzonitrile 13.

4-Fluorobenzonitrile was prepared similar to 15, starting from 4-fluorobenzaldehyde.

M.p. 35–37 °C (lit.410 m.p. 32–34 °C). B.p. 70°C at 10 mm Hg. Mixture with a commercial sample did not give depression in m.p. 13C NMR δ: 162.9, 131.3, 113.7,

112.7, 104.5.

4-Bromobenzonitrile 14.

4-Bromobenzonitrile was prepared similar to 15, starting from 4-bromobenzaldehyde.

M.p. 112–115 °C (lit.411 m.p. 110–115 °C). Mixture with a commercial sample did not

231

give depression in m.p. IR ν, cm–1: 3100 – 2850, 2240, 1590, 1450, 1260, 1040, 830, 670.

EI-MS: m/z (%): 182 (M+). 13C NMR δ: 162.3, 132.5, 127.8, 117.8, 111.2.

4-Methoxybenzonitrile 15.

OH N CHO CN

+ NH2OH — H2O OMe OMe OMe

4-Methoxybenzonitrile was prepared by a modified procedure of Wang.119 A two-liter round bottom flask with a heavy stir-bar was charged with anisaldehyde (4- methoxybenzaldehyde, 272 g, 2.0 mol), hydroxylamine hydrochloride (166 g, 2.4 mol), triethylamine1 (137 g, 190 ml, 1.35 mol), and NMP (anhydrous, 500 ml). The mixture was stirred for 30 min, and phthalic anhydride (352 g, 2.3 mol) was added at once. The flask was topped with reflux condensor and nitrogen bubbler, and heated to reflux on a heating mantle (mantle temperature 150–170°C) for five hours. The reaction mixture was cooled down, poured into cold water (4 liters), and chilled in a fridge, resulting in crystallization of gray crystals. After 12 hrs in the fridge, the solids were filtered off on a

Büchner funnel, suspended in sodium hydrocarbonate solution (sat. aq.), stirred at 40°C for 30 min to remove phthalic acid, and then recrystallized once from ethanol with activated charcoal (2% wt/v) and four times from ethanol–water mixtures, increasing the

232

water content each time from 10 to 40% in 10% increments, yielding 91.7 g (69%) of 15 as off-white needles. M.p. 57°C, lit.412 m.p. 55–60°C. 13C NMR δ: 162.9, 104.0, 134.0,

114.7, 55.5, 119.2. The mother liquors from all recrystallizations may be diluted with water, chilled in a fridge, and the precipitated oil, after recrystallization from ethanol with activated charcoal, would give additional amount of 15.

4-(Pyrrolidin-1-yl)benzonitrile 16.

CN CN i-Pr NEt I 2 + I N NH2

A 250 ml round bottom flask with a stirbar was charged with 4- aminobenzonitrile (10.0 g, 0.085 mol), 1,4-diiodobutane (26.25 g, 0.085 mol), Hunig’s base (N,N-diisopropylethylamine, 21.88 g, 0.17 mol), HMPA (10 g), and NMP (20 g).

The reaction mixture was stirred at 100°C for two days, cooled down, poured into water

(200 ml), and filtered on Büchner funnel. The residue on filter was recrystallized twice from ethanol (30 and 50 ml correspondingly) to give 8.1 g (55%) of off-white crystals.

413 414 1 M.p. 89°C, lit. m.p. 81 and 89°C. H NMR (CDCl3) δ: 7.4 (d, 2H), 6.5 (d, 2H), 3.3

(t, 4H), 2.0 (m, 4H).

1 The amount of Et3N has been reduced due to the basicity of NMP used as solvent.

233

4-(N-n-hexylamino)benzonitrile 17.

CN CN

+ I

HN NH2 C6H13

4-(N-n-hexylamino)benzonitrile was prepared according to published procedure for a similar (hexadecylamino-) compound.415 A 250 ml round bottom flask with a stirbar was charged with 4-aminobenzonitrile (23.6 g, 0.2 mol), 1-bromohexane (16.5 g, 0.1 mol), and HMPA (200 ml). The reaction mixture was stirred at 120°C under argon for 22 hrs, cooled down, poured into water (300 ml), and extracted with ether (2×100 ml).1 The extract was evaporated from the solvent and the residue was vacuum-distilled to give

18.9 g (93.6%) of colorless liquid, b.p. 167–170°C at 2 mm Hg, crystallizing on standing.

Since TLC of this product (eluent hexane:EtOAc = 2:1) showed presence of some starting material, the distilled material was chromatographed with the same eluent to give

17.7 g (88%) of 17 as white crystals. M.p. 48–49°C. Lit.416 m.p. 38–39°C. 1H NMR

(CDCl3) δ: 7.4 (d, 2H), 6.5 (d, 2H), 4.3 (s, br, 1H), 3.1 (m, 2H), 1.6 (m, 2H), 1.4 (m, 6H),

13 0.9 (t, 3H). C NMR (CDCl3) δ: 151.7, 133.9, 120.9, 112.2, 98.3, 43.4, 31.8, 29.3, 26.9,

22.8, 14.3.

234

4-(N,N-di-n-hexylamino)benzonitrile 18.

4-(N,N-di-n-hexylamino)benzonitrile was prepared similar to 19, starting from 4-

1 fluorobenzonitrile and di-n-hexylamine. Yield 23%. H NMR (CDCl3) δ: 7.4 (d, 2H), 6.6

13 (d, 2H), 3.3 (t, 4H), 1.6 (pentet, 4H), 1.4 (m, 8H), 0.9 (t, 6H). C NMR (CDCl3) δ: 150.8,

133.7, 121.2, 111.3, 96.3, 50.9, 31.8, 29.3, 26.9, 22.8, 14.3..

4-(N,N-di-n-butylamino)benzonitrile 19.

CN CN Py + HN HMPA F N Bu Bu

A 250 ml round bottom flask with a stir-bar was charged with 4- fluorobenzonitrile (12.1 g, 0.1 mol), di-n-butylamine (30 g, 0.23 mol), and pyridine (10 ml). The reaction mixture was heated at 60°C for 24 hrs and monitored by TLC (neat hexane or hexane:ether = 8:2) – no new spots detected. HMPA (15 g) was added and temperature was raised to 100°C. After 48 hrs the reaction mixture was cooled down, poured into water (200 ml), diluted with HCl (3M, aq., 40 ml), and extracted with ether

(2×100 ml). Ether was evaporated on a rotovap and the residue was vacuum-distilled.

The fraction, boiling 150–160°C at 0.4 mm Hg was collected and amounted to 12.0 g

1 To check that diethyl ether does not extract considerable amount of HMPA from water solution, 1.00 g of HMPA was dissolved in 40.0 g of water and extracted with 15.0 g of ether. The ethereal layer was separated, and evaporated to give 11.5 mg of residue.

235

(52%). Lit.417 b.p. 176–179 at 2 mm Hg, which corresponds to 150°C at 0.4 mm Hg. 1H

NMR (CDCl3) δ: 7.4 (d, 2H), 6.6 (d, 2H), 3.3 (t, 4H), 1.6 (pentet, 4H), 1.4 (pentet, 4H),

13 0.9 (t, 6H). C NMR (CDCl3) δ: 150.8, 133.7, 121.2, 111.3, 96.3, 50.9, 29.3, 20.5, 14.2.

3,6-Bis(4-fluorophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione, F-DPP 20.

N O H N O O t-BuOK F + O t-AmOH F N F 13 O 11 20 O H

A five liter reaction vessel was charged with tert-amyl alcohol (2000 ml), potassium tert-butoxide (561 g, 5 mol), topped with a three-neck lid, and flushed with nitrogen. The central neck was fitted with a mechanical stirrer, and a thermometer/nitrogen inlet adaptor into another neck. The third neck was fitted with a reflux condenser and the reactor was heated with stirring on a heating mantle (590 W,

60% of 120 V) to reflux (102°C) until all t-BuOK dissolved. After that a solution of 4- fluorobenzonitrile (140 g, 1.156 mol) in tert-amyl alcohol (warm, 200 ml) was added slowly from an addition funnel, placed on top of the reflux condensor – the temperature of the reaction mixture rose to 113°C. Then a solution of di-iso-propyl succinate (161.8 g,

0.8 mol) in tert-amyl alcohol (150 ml) was added from a syringe, using a syringe pump, at a rate of 40 ml/hr. After all di-iso-propyl succinate had been added, the reflux condensor was replaced with a 15-cm Vigreaux column and a Liebig condensor and the alcohols were distilled out (ca. 800–850 ml of distillate) until nD of the distillate reached

236

20 that of tert-amyl alcohol ( nD = 1.4050). When the distillation had been finished, the heating was continued for 6 additional hours, and then the reaction mixture was cooled down and transferred to a 4000 ml Erlenmeyer flask, equipped with a mechanical stirrer.

To that mixture, methanol (1000 ml) was added in portions of 100 ml so that the temperature did not rise above 60°C. After the methanol addition was complete, acetic acid (350 ml) was added from a syringe pump at a rate of 40 ml/hr, with vigorous stirring

(the viscosity of the mixture increases considerably), followed by water (200 ml), and added in one portion. The suspension obtained was filtered on two 2-liter Buchner funnels, and the thick red mud on the filter was then gradually suspended (with help of sonication) in and filtered from: methanol (2×2 liters), hot water (2×1 liter), and boiling ethanol (4 liters). The filter cake was dried overnight to give 123.85 g (66%) of 20 as fine red powder. Direct characterization was not performed due to low solubility. Alkylation gave an inseparable mixture of products.

3,6-Bis(4-cyanophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione 21.

CN O H N O O NC + O CN N CN O 12 O H 21

A one liter three-neck round bottom flask was fitted with a mechanical stirrer, a thermometer/septum inlet adaptor, and a reflux condenser. The flask was charged with tert-amyl alcohol (250 ml) and heated with stirring on a heating mantle until reflux of the

237

solvent (102°C) began. At that temperature sodium tert-pentoxide (33 g, 0.3 mol) was added in portions and heating and stirring continued until all of the t-AmONa dissolved

(the temperature rose to 110°C). After the temperature was lowered to 110°C, 1,4- dicyanobenzene (25.6 g, 0.2 mol) was added at once. Di-tert-amyl succinate (33.6 g, 0.13 mol) was added fro a syringe pump during 2 hrs period. After the addition was complete, the reaction mixture was stirred at 110°C for four additional hours, cooled to 60°C and methanol (60 ml) was added, followed by acetic acid (30 ml). The resulting slurry was filtered, and then re-suspended in and filtered from ethanol (2×200 ml), hot water (2×100 ml), and boiling ethanol (200 ml). The filter cake was air-dried to allow 23.75 g (35%) of

1 dark red crystals. H NMR (NaOD, DMSO-d9/D2O) δ: 8.64 (m, 2H), 7.75 (m, 2H).

3,6-Bis(4-methylphenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione 22.

H O N O CN O + O O N O H

A one liter three neck round bottom flask was flushed with argon and charged with tertiary amyl alcohol (2-methyl-2-butanol, 320 g). The central neck of the flask was fitted with a mechanical stirrer, the side neck – with a thermometer/nitrogen inlet adaptor, and the third neck – with a reflux condenser. The reaction mixture was heated with stirring on a heating mantle until reflux of the solvent (102°C) began. At that temperature sodium tert-amylate (110 g, 1 mol) was added in portions and heating and stirring continued until all of the t-AmONa dissolved. 4-Toluonitrile (47 g, 0.4 mol) was added at

238

once. While maintainig reaction temperature 105-110°C, di-iso-propyl succinate (AS-2-

09, 54 g, 0.267 mol) was added at vigirous stirring from a syringe via syringe pump at a rate of 20 ml/hr. After the addition was complete, the reaction mixture was stirred at

110°C for four additional hours, and then cooled to 60°C and methanol (50 ml) was added dropwise, followed by acetic acid (50 ml). The resulting slurry was filtered on a

Buchner funnel, and then gradually suspended (with help of sonication) in and filtered from: hot water (400 ml), hot ethanol (2×400 ml) and hot acetone (2×400 ml). The filter cake was dried overnight: 24 g (37.8%) – compare to 56% for parent Ph-DPP-H. UV-Vis

1 (DMAc) λmax: 269, 444, 473, 508; (sodium salt, DMAc): 277, 400, 417, 531, 565. H

NMR (DMSO-d6) δ: 8.4 (d, 2H, J=8.5 Hz), 7.4 (d, 2H, J=8.5 Hz), 2.4 (s, 3H).

2,5-dimethyl-3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione74 23.

H N O N O

O N O N H

A 500 ml round bottom flask with a stir bar was charged with DPP 1 (14.42 g,

0.05 mol) and DMAc (300 ml). To the stirred reaction mixture sodium hydride (2.5 g,

0.105 mol) was added in portions. When the addition was complete, the reaction mixture was heated at 80°C for 30 min and then methyl p-toluenesulfonate (30.0 g, 0.161 mol) and additional dimethylacetamide (70 ml) were added at once. The reaction mixture was stirred at 150°C for 24 hrs, cooled down, poured into water (200 ml), and filtered. The residue on the filter was re-dissolved in boiling chloroform (200 ml), filtered from

239

insolubles and after evaporation of solvent and recrystallization from DMAc gave 13.45 g (85%) of orange-red crystals. M.p. 231°C (lit.46 m.p. 233-234°C); 1H δ: 7.87 (d, 2H,

13 J=8.7 Hz), 7.61 (d, 2H, J=8.7 Hz), 3.31 (s, 3H). C δ: 138.35, 130.50, 29.60. ΦF=0.79.

2,5-diethyl-3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione 25 was prepared similar to 23, starting from DPP 1 and ethyl p-toluenesulfonate. Yield 78%. M.p. 229 °C.

2,5-diisopropyl-3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione 26 was prepared similar to 23, starting from DPP 1 and iso-propyl tosylate. Yield 66%. M.p. 267 °C

(DSC, 20 °C/min).

3,6-Diphenyl-2,5-dipropylpyrrolo[3,4-c]pyrrole-1,4-dione 32.

O O O

HNHN NN + NNH

O O O 1 32 33

A one liter round bottom flask with a stir bar was charged with 3,6-Diphenyl-

2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (20.0 g, 0.07 mol), potassium tert-butoxide

(24.0 g, 0.24 mol), dimethylacetamide (200 ml), fitted with an air condenser and heated at 140°C for 4 hours, distilling out tert-butanol. After distillation ceased, the reaction mixture was cooled down to room temperature and propyl iodide (47.0 g, 0.27 mol) was

240

added dropwise with stirring. When the addition was completed, the reaction mixture was heated at 140°C for 12 hours and monitored by TLC. After 12 hours reaction mixture was cooled down and poured into water (500 ml). The precipitate was collected under suction filtration, air-dried, dissolved in chloroform (200 ml), applied onto silica gel, and chromatographed with dichloromethane.

32. M.p. 189°C (DMAc). 1H δ: 7.82-7.85 (m, 2H), 7.56-7.50 (m, 3H), 3.72 (t,

2H), 1.59 (sextet, 2H), 0.83 (t, 3H). 13C δ: 162.2, 148.7, 131.3, 129.3, 129.0, 128.8,

128.4, 43.6, 23.0, 11.3. UV-Vis λmax, nm: 289, 466, 488. Fluorescence λmax: 528, 568sh.

ΦF=0.76.

33. (3,6-diphenyl-2-propyl-5H-pyrrolo[3,4-c]pyrrole-1,4-dione) M.p. 276°C

1 (DMAc). H (CDCl3/DMSO-d9) δ: 11.00 (s, 1H), 8.53-8.48 (m, 2H), 7.80-7.77 (m, 2H),

7.59-7.50 (m, 6H), 3.75 (t, 2H, J=7.8 Hz), 1.61 (m, 2H), 0.83 (t, 3H, J=7.8Hz). 13C

(CDCl3/DMSO-d9) δ: 163.1, 162.9, 146.7, 131.7, 130.6, 128.8, 128.7, 128.4, 128.2,

109.4, 42.8, 23.1, 11.9. UV-Vis λmax, nm: 265, 465, 488. Fluorescence λmax: 523, 563sh.

ΦF=0.69.

3,6-Diphenyl-2,5-dihexylpyrrolo[3,4-c]pyrrole-1,4-dione 34.

O O O

HNHN H13C6 NNC6H13 + H13C6 NNH

O O O 1 34 35

241

3,6-Diphenyl-2,5-dihexylpyrrolo[3,4-c]pyrrole-1,4-dione 34 was prepared similar to 32 starting from DPP 1 (2.88 g, 10 mmol) and 1-iodohexane (2.12 g, 10 mmol).

Chromatography yielded 0.4 g (8%) of 34 and 0.6 g (16%) of 35.

34. M.p. 134°C (DMAc). 1H δ: 7.82-7.78 (m, 1H), 7.56-7.49 (m, 4H), 3.74 (t,

2H, J=7.8 Hz), 1.58 (pentet, 2H, J=7.8 Hz), 1.18 (m, 6H), 0.82 (t, 3H, J=7.8Hz). 13C δ:

162.9, 148.7, 131.2, 129.0, 128.8, 128.4, 109.9, 42.0, 31.4, 29.5, 26.5, 22.6, 14.1. UV-Vis

λmax, nm: 472. Fluorescence λmax: 530, 572sh. ΦF=0.74.

1 35. M.p. 252°C (DMAc). H (CDCl3/DMSO-d9) δ: 11.00 (s, 1H), 8.53-8.48 (m,

2H), 7.80-7.77 (m, 2H), 7.59-7.50 (m, 6H), 3.80 (t, 2H, J=7.8 Hz), 1.58 (pentet, 2H,

13 J=7.8 Hz), 1.21 (m, 6H), 0.83 (t, 3H, J=7.8Hz). C (CDCl3/DMSO-d9) δ: 163.1, 162.9,

146.7, 131.7, 130.6, 128.8, 128.7, 128.4, 128.2, 109.4, 41.6, 30.9, 29.1, 26.1, 22.1, 13.9.

UV-Vis λmax, nm: 262, 464, 481. Fluorescence λmax: 522, 565sh. ΦF=0.71.

3,6-diphenyl-2,5-didecyl-dihydropyrrolo[3,4-c]pyrrole-1,4-dione 36 was prepared similar to 32 starting from DPP 1 (1.44 g, 5 mmol) and iododecane (2.68 g, 10 mmol). Chromatography with dichloromethane and recrystallization from 1-propanol gave 1.2 g (42%) of orange crystals. M.p. 117°C. 1H (300 MHz, CDCl3) δ: 7.78-7.82 (m,

2H), 7.51-7.55 (m, 3H), 3.74 (t, 2H), 1.19-1.25 (m, 16H), 0.86 (t, 3H). 13C δ: 162.9,

148.7, 131.3, 129.1, 128.9, 128.5, 109.9, 42.1, 33.0, 32.0, 29.6, 29.5, 29.2, 26.9, 25.9,

22.8, 14.3. UV-Vis (CH2Cl2, λmax, nm): 268, 467. Fluorescence λmax: 529, 573sh.

ΦF=0.65.

242

3,6-Diphenyl-2,5-didodecylpyrrolo[3,4-c]pyrrole-1,4-dione 37 was prepared similar to 36, substituting 1-iododecane for 1-bromododecane. Yield 44%. M.p. 114°C.

1H δ: 7.8 (m, 1H), 7.6-7.5 (m, 4H), 3.7 (t, 2H, J=7.8 Hz), 1.6 (pentet, 2H, J=7.8 Hz), 1.3–

1.15 (m, 18H), 0.82 (t, 3H, J=7.8Hz). 13C δ: 162.9, 148.7, 131.3, 129.1, 128.9, 128.5,

110.0, 42.1, 32.0, 29.6 (double intensity), 29.5, 29.2, 26.9, 22.9, 14.3. UV-Vis λmax: 284,

474. Fluorescence λmax: 527, 563sh. ΦF=0.97.

iso-Propyl tosylate.

Na O O ; + S Cl S O OH ONa ONa – NaCl O O

A one-liter round bottom flask with a large stir-bar was charged with iso-propyl alcohol (300 g, 5 mol), THF (200 ml), and sodium (48 g, 2 mol), topped with a reflux condensor, and heated at 80°C for two hours. After all sodium had been dissolved, the heating mantle was replaced with an ice bath and the reaction mixture was cooled to below 5°C. At that temperature tosyl chloride (360 g, 1.89 mol) was added in portions — smaller at the beginning of the addition, larger towards the end, at such a rate that the temperature did not rise above 15-20°C. After the TsCl addition was complete, the reaction mixture was heated at 70°C for an hour, cooled down, and poured into cold water (1000 ml). The layers were separated and the organic layer was gradually washed with water (2×300 ml), HCl (1M, 200 ml), ammonium chloride (sat. aq., 200 ml), dried with MgSO4, and in the vacuum oven. The crude product thus obtained (308 g, 76%) is a dark brown oil and may be used for the alkylation reactions without further purifications.

243

If colorless material is desired, the crude product may be purified either by careful vacuum distillation – on large scale (decomposition occurs if overheated!), or by column chromatography – on small scale.

3,6-Bis(4-methylphenyl)-2,5-dipropylpyrrolo[3,4-c]pyrrole-1,4-dione.

H N O N O

O N O N H A 500 ml round bottom flask with a stir bar was charged with 3,6-bis(4-tolyl)-

2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione 22 (20.7 g, 65.4 mmol), potassium tert- butoxide (18.4 g, 164 mmol), dimethylacetamide (150 ml), fitted with air condenser and heated at 60°C for 4 hours, distilling out tert-butanol. After distillation ceased, the reaction mixture was cooled down to ~65°C and propyl iodide (25 g, 147 mmol) was added dropwise with stirring. When the addition was completed, the reaction mixture was heated at 80°C for 12 hours. The reaction mixture was cooled down and poured into water (500 ml). The precipitate was collected by suction filtration, air-dried (49.7 g), dissolved in chloroform (250 ml), filtered from insoluble matter, applied (37 g) onto silica gel, and chromatographed with hexane:dichloromethane from 8:2 to 1:1 ratio yielding two fractions.

Fraction #1 is Tol-DPP-Pr (3,6-bis(4-methylphenyl)-2,5-dipropylpyrrolo[3,4- c]pyrrole-1,4-dione), recrystallized from DMAc (12 ml) to give 15.9 g (60%) of brown- orange crystals. M.p. 186°C. 1H δ: 7.75 (d, 2H), 7.34 (d, 2H), 3.76 (t, 2H), 2.46 (s, 3H),

244

1.65 (sextet, 2H), 0.87 (t, 3H). 13C δ: 162.7, 148.2, 141.4, 129.5, 128.7, 125.7, 109.6,

43.5, 22.8, 21.5, 11.0. UV-Vis λmax, nm (lg ε): 270 (4.35), 306 (4.16), 474 (4.27).

Fluorescence λmax: 530, 575sh. ΦF=0.64.

Fraction #2 is 3,6-bis(4-methylphenyl)-2-propyl-5-hydropyrrolo[3,4-c]pyrrole-

1,4-dione, recrystallized from DMAc (20 ml) to give 0.68 g (3%) of red-orange crystals.

M.p. 294°C. 1H δ: 8.16 (d, 2H), 7.72 (d, 2H), 7.33 (AB, 4H), 3.80 (t, 2H), 2.43 (s, 3H),

2.44 (s, 3H), 1.65 (sextet, 2H), 0.87 (t, 3H). 13C δ: 162.9, 142.7, 141.5, 138.9, 129.9(CH),

129.6(CH), 128.7(CH), 127.7(CH), 125.6, 125.0, 43.7, 22.7, 21.7, 21.5, 11.1 (other peaks do not show up due to low solubility). UV-Vis λmax, nm (lg ε): 268 (4.47), 304 (4.17),

469 (4.36), 495 (4.36). Fluorescence λmax, nm: 524, 565; ΦF=0.90.

2-Benzyl-3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione 40. H H O N O N

N O N O H

2-Benzyl-3,6-diphenyldihydropyrrolo[3,4-c]pyrrole-1,4-dione was prepared similar to 32 starting from DPP 1 (722mg, 2.5 mmol) and benzyl bromide (428mg, 2.5 mmol). Chromatography with dichloromethane and recrystallization from 1-propanol gave 303 mg (32 %) of 40 as orange crystals. M.p. 344°C. 1H (300 MHz, DMSO) δ:

11.36 (s, 1H), 8.5 (m, 2H), 7.7 (m, 2H), 7.6 (m, 4H), 7.5 (m, 2H), 7.3 (m, 4H), 7.1 (m,

245

1H), 5.0 (s, 2H). UV-Vis (CH2Cl2, λmax, nm): 263, 295, 468. Fluorescence λmax: 525,

566sh. ΦF=0.95.

3,6-diphenyl-2,5-diallylpyrrolo[3,4-c]pyrrole-1,4-dione 41.

O O O

HNHN NN + NNH

O O O 1 41 42

3,6-diphenyl-2,5-diallylpyrrolo[3,4-c]pyrrole-1,4-dione 41 was prepared similar to 32 starting from DPP 1 (5.76 g, 20 mmol), sodium tert-amylate (6.6 g, 60 mmol), NMP

(150 ml), and allyl bromide (8.0 g, 66 mol). Chromatography with neat dichloromethane, followed by dichloromethane : ethyl acetate = 1:1 gave 3.13 g (42%) of 41 and 3.0 g

(46%) of 42.

41. M.p. 209°C. 1H δ: 7.95-7.90 (m, 2H), 7.53-7.51 (m, 3H), 6.0-5.9 (m, 1H),

5.24-5.18 (m, 2H), 4.4 (d, 2H). 13C δ: 162.3, 148.6, 133.4, 131.2, 129.0, 128.7, 128.1,

116.9, 109.9, 44.5. UV-Vis λmax: 476. Fluorescence λmax: 525, 566sh. ΦF=0.92.

42. (3,6-diphenyl-2-allyl-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione) M.p. 289°

(DMAc). 1H (DMSO) δ: 8.5 (dd, 2H), 7.8 (dd, 2H), 7.6 (m, 6H), 5.9 (m, 1H), 5.1 (dd,

1H), 5.0 (dd, 1H), 4.4 (m, 2H). 13C (DMSO) δ: 163.0, 161.8, 146.7, 146.5, 134.3, 132.7,

131.5, 129.5, 129.2, 129.0, 128.4, 128.2, 128.0, 116.7, 111.8, 108.9, 44.1. UV-Vis λmax:

467, 495. Fluorescence λmax: 518, 560sh. ΦF=0.97.

246

3,6-bis(4-bromophenyl)-2,5-dimethyl-pyrrolo[3,4-c]pyrrole-1,4-dione 43.

O N N O Br Br N O O N 23 43

(a) A Petri dish (14 cm internal diameter) was filled with finely ground 3,6- diphenyl-2,5-dimethylpyrrolo[3,4-c]pyrrole-1,4-dione 23 (8.0 g, 0.025 mol). Bromine

(40.0 g, 0.25 mol) was placed into a 50 ml evaporation dish at the bottom of a desiccator

(15 cm internal diameter) and topped with a porcelain insert, over which the Petri dish was placed. The desiccator was left with lid closed and a slightly open vent in the dark for ten days. After the reaction was complete, the Petri dish was removed, left open to the air for 2 hours, then placed into a vacuum oven and dried at 75°C: for 2 hours at 40 mm

Hg, and for 4 hours at 1 mm Hg to remove absorbed bromine and hydrogen bromide and then was weighed out to check the mass gain (5.5 g, 137% of theory). The crude product was recrystallized from DMAc (45 ml) to yield 11.7 g (96%) of ruby red crystals. M.p.

1 13 343°C. H (CDCl3) δ: 7.75 (AB, 2H), 7.65 (AB, 2H), 3.31 (s, 3H). C(CDCl3) δ: 205.3,

132.4, 130.6, 126.8, 126.2, 29.6. UV-Vis λmax, nm: 275, 305, 491. Fluorescence λmax:

541, 587sh. ΦF=0.83.

H O N N O Br Br Br Br O N N H O 9 43

247

(b) 3,6-bis(4-bromophenyl)-2,5-dimethyl-pyrrolo[3,4-c]pyrrole-1,4-dione 43 was also prepared from 9: 3,6-bis(4-bromophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4- dione 9 (9.7 g, 0.0217 mmol) was placed into a 250 ml round bottom flask together with potassium carbonate (30.0 g, 0.217 mol), methyl p-toluenesulfonate (16.0 g, 0.086 mol), and dimethylacetamide (70 ml). The reaction mixture was stirred at 150°C for 24 hrs, cooled down, poured into water (200 ml), and filtered. The residue on the filter was redissolved in boiling chloroform (200 ml), filtered from insolubles and after evaporation of solvent gave 4.30 g (43%) of orange-red crystals with the same characteristics as above.

3,6-bis(4-bromophenyl)-2,5-dipropylpyrrolo[3,4-c]pyrrole-1,4-dione 44 was prepared similar to 43 starting from Br-DPP 9. Yield 37% of Br-DPP-Pr 44 as yellow crystals. M.p. 247°C (DMAc). 1H δ: 7.68 (AA′, 4H), 3.72 (t, 2H), 1.60 (m, 2H), 0.86 (t,

3H). 13C δ: 162.5, 147.4, 132.3, 130.0, 127.0, 125.8, 109.9, 43.4, 22.8, 11.2. ES-MS:

530.8. UV-Vis λmax, nm: 275, 304, 478. Fluorescence λmax: 541, 587sh. ΦF=0.86.

3,6-Bis(4-iodophenyl)-2,5-dimethylpyrrolo[3,4-c]pyrrole-1,4-dione 45.

O N N O Br I Br I N O O N 43 45

248

A 200 ml pear-shaped flask with a magnetic stirbar was charged with 3,6-bis(4- bromophenyl)-2,5-dimethylpyrrolo[3,4-c]pyrrole-1,4-dione (2.37 g, 5 mmol), potassium iodide (12.50 g, 75 mmol), copper (I) iodide (4.75 g, 25 mmol), and dimethylacetamide

(50 ml). The reaction mixture was stirred under argon at 165°C for 115 hours, cooled to

~80°C, and mixed with water (200 ml). The resulting slurry was filtered, washed with water (2×100 ml) and ethanol (30 ml), and air-dried, resulting in 7.7 g of pink powder, which was boiled with chloroform (3×300 ml), and filtered through a #50 filter. The filtrate, after evaporation, gave 1.1 g (37%) of red powder, m.p. 355°C dec. 1H δ: 7.87 (d,

2H, J=8.7 Hz), 7.61 (d, 2H, J=8.7 Hz), 3.31 (s, 3H). 13C δ: 138.35, 130.50, 29.60. UV-Vis

λmax, nm: 276, 318, 495. Fluorescence λmax: 544, 590sh. ΦF=0.89.

3,6-Bis(4-iodophenyl)-2,5-dipropylpyrrolo[3,4-c]pyrrole-1,4-dione 46 was prepared similar to 45 starting from Br-DPP-Pr 44 to give 5.74 g (92%) of red powder.

Recrystallization from dimethylacetamide (5 ml/g) gave 4.25 g (68%) of ruby red crystals. M.p. 260.5°C dec. 1H δ: 7.86 (d, 2H, J=8.7 Hz), 7.53 (d, 2H, J=8.7 Hz), 3.70 (s,

2H), 1.58 (m, 2H), 0.83 (t, 3H). 13C δ: 162.5, 147.6, 138.4, 130.1, 127.9, 110.34, 97.9,

+ 43.8, 23.2, 11.4. ES-MS: 624.7 (M ). UV-Vis λmax, nm (ε): 277 (21520), 317 (18923),

479 (21350). Fluorescence λmax: 550, 592sh. ΦF=0.86.

3,6-Bis(4-cyanophenyl)dihydropyrrolo[3,4-c]pyrrole-1,4-dione 47 was prepared similar to 1, using 1,4-dicyanobenzene instead of benzonitrile. Yield 35% of

249

1 dark red crystals. H (NaOD, DMSO-d9/D2O) δ: 8.64 (m, 2H), 7.75 (m, 2H). UV-Vis

(DMAc) λmax: 556, 603sh. ΦF=0.63.

3,6-bis(4-[pyrrolydin-1-yl]phenyl)-2,5-dimethylpyrrolo[3,4-c]pyrrole-1,4-dione 48.

O N N O Br N Br N N O O N 43 48

A 50 ml pear shaped flask with a magnetic stirbar was charged with Br-DPP-Me

43 (300 mg, 0.63 mmol), dimethylacetamide (3 g), and pyrrolidine (3.0 g, 42 mmol). The reaction mixture was stirred at 140°C for 12 hours (the starting material spot is gone on

TLC after 2 hrs, the di-substituted, lower Rf spot becomes major on TLC after 6 hrs), cooled to room temperature and water (30 ml) was added dropwise. The precipitate was filtered, and air-dried, leaving 213 mg (74%) of dark red crystals. M.p. 324°C dec. 1H δ:

7.94 (d, 2H, J=9 Hz), 6.64 (d, 2H, J=9 Hz), 3.40 (m, 7H), 2.04 (m, 4H). 13C δ: 163.2,

149.4, 147.4, 131.1, 115.3, 111.5, 107.0, 47.4, 30.1, 25.6. UV-Vis (λmax, CHCl3): 240,

282, 385, 548. Fluorescence λmax: 575. ΦF=0.95.

3,6-bis(4-[piperidin-1-yl]phenyl)-2,5-dimethylpyrrolo[3,4-c]pyrrole-1,4- dione 49 was prepared similar to 48, starting from Br-DPP-Me 43 and piperidine. Yield

45%.

250

3,6-bis(4-[pyrrolydin-1-yl]phenyl)-2,5-dipropylpyrrolo[3,4-c]pyrrole-1,4- dione 50 was prepared similar to 48 in a 120 ml Ace Glass pressure vessel starting from

Br-DPP-Pr 44 (625 mg, 1.23 mmol) and pyrrolidine (1.88 g, 26 mmol) at 200°C during

12 hours. Chromatography with hexane:dichloromethane from 2:1 to 1:1 gave 201 mg

(32%) of dark-red crystals. 1H δ: 7.87 (d, 2H, J=9 Hz), 6.65 (d, 2H, J=9 Hz), 3.8 (m, 2H),

3.4 (m, 4H), 2.05 (m, 4H), 0.92 (t, 3H, J=8 Hz). 13C δ: 163.3, 149.5, 147.4, 131.6, 130.7,

129.0, 115.3, 111.5, 49.7, 47.5, 25.5, 23.0, 11.3. UV-Vis (λmax, CHCl3): 273, 384, 539.

Fluorescence λmax: 577. ΦF=0.94.

3,6-bis(4-[N,N-dibutylamino]phenyl)-2,5-dipropylpyrrolo[3,4-c]pyrrole-1,4-dione 51 was prepared similar to 48 starting from Br-DPP-Pr 44 and di-n-butylamine. Yield of 51

8%, 52 – 11%.

3,6-bis(4-[N,N-di-n-hexylamino]phenyl)-1,4-dimethyl-2,5-dihydropyrrolo-

[3,4-c]pyrrole-1,4-dione 53.

H13C6 H13C6 C6H13 C6H13 Br N N

O O O

NN NN+ NN

O O O

43 53 54 Br N Br H13C6 C6H13

251

A 50 ml pear shaped flask with a magnetic stir bar was charged with Br-DPP- 43

(1.0 g, 2.11 mmol), dimethylacetamide (5.0 g), hexamethylphosphoramide (7.8 g) and dihexylamine (1.5 g, 8.1 mmol). The reaction mixture was stirred at 180°C for 96 hours

(the starting material spot was still present on TLC after 36 hrs, the di-substituted, lower

Rf spot becomes major on TLC after 50 hrs, six spots on TLC total), cooled to room temperature and water (60 ml) was added dropwise. The organic layer was extracted with chloroform (150 ml), applied on silica (15 g) and flash-chromatographed: 430 mg of crude material (six spots on TLC). A second chromatography yielded 158 mg (11%) of diaminated product 53 and 257 mg (21%) of monoaminated product 54.

1 53. H (CDCl3) δ: 7.93 (d, 2H), 6.70 (d, 2H), 3.42 (s, 3H), 3.33 (t, 4H), 1.62 (m,

4H), 1.33 (m, 12H), 0.907 (t, 6H). 13C δ: 163.2, 149.9, 148.1, 132.1, 131.8, 131.2, 111.1,

51.2, 31.8, 30.127.4, 26.9, 22.8, 14.2. UV-Vis (λmax): 282, 385, 548. Fluorescence λmax:

575. ΦF=0.92.

1 54. H (CDCl3) δ: 7.93 (d, 2H), 7.66 (d, 2H), 7.53 (d, 2H), 6.64 (d, 2H), 3.36 (s,

3H), 3.33 (t, 4H), 3.30 (s, 3H), 1.62 (m, 4H), 1.33 (m, 12H), 0.91 (t, 6H). 13C δ: 163.3,

162.2, 151.0, 150.6, 131.9, 131.8, 130.6, 127.6, 124.6, 114.2, 111.0, 106.2, 51.3, 31.8,

30.1, 29.4, 27.4, 26.9, 22.8, 14.2.

252

2,5-dimethyl-3,6-bis(4-[2-{4-t-butylphenyl}vinyl]phenyl)-2,5-dihydropyrrolo[3,4- c]pyrrole-1,4-dione (55).

O N O NN+

O O N

I a) In a 50 ml pear-shaped flask with a magnetic stirring bar was placed 2,5-dimethyl-3,6- bis(4-iodophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (4, 109.4 mg, 0.193 mmol), 4-tert-butylstyrene (67.2 mg, 0.42 mmol), potassium carbonate (82.8 mg, 0.6 mmol), TDA-1 (tris(3,6-dioxaheptyl)amine418, 6 mg, 0.02 mmol), and dimethylacetamide

(5 g). The flask was fitted with an oil bubbler and warmed to 100°C in a heating mantle while degassing with argon. After 10 min palladium (II) acetate (1 mg, 0.004 mmol) was added. The reaction mixture was stirred at 100°C for 10 hr, cooled to room temperature, diluted with water (40 ml) and extracted with chloroform (4×60 ml). Montmorillonite clay and magnesium sulfate were added to the combined extracts, filtered, solvent evaporated to small volume and the residue was impregnated onto silica (13 g). Flash- chromatography on 50 g of silica gel with hexane to hexane-dichloromethane (1:1) yielded, after removal of solvent, 54 mg of dark solid, which was washed with pentane to give 51 mg (43%) of dark crystals, m.p.295°C. 1H NMR δ: 7.93 (d, J=9 Hz), 7.65 (d, J=9

Hz), 7.50 (d, J=9 Hz), 7.41 (d, J=9 Hz), 7.24 (d, J=16.2 Hz), 7.12 (d, J=16.2 Hz). ES-MS

253

13 (APCI, CHCl3) m/z: 633.3 (M+1). C δ: 162.7, 151.6, 148.1, 140.5, 134.2, 131.0, 129.7,

127.1, 126.8, 126.7, 125.9, 109.4, 34.9, 31.4, 29.7. b) 2,5-Dimethyl-3,6-bis(4-bromophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (8) reacted as above for 10 hrs at 150°C gave 50% yield of the same compound.

2,5-dimethyl-3,6-bis(4-[2-{4-acetoxyphenyl}vinyl]phenyl)-2,5-dihydropyrrolo[3,4- c]pyrrole-1,4-dione (56).

O O O N O NN+

O N O O O O

A 200 ml pear-shaped flask with a magnetic stir bar was charged with 2,5-dimethyl-3,6- bis(4-iodophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (4, 607 mg, 1.07 mmol), 4- acetoxystyrene (357 mg, 2.2 mmol), TDA-1 (tris(dioxa-3,6-heptyl)amine, 78 mg, 0.24 mmol), potassium carbonate (504 mg, 3.6 mmol), and dimethylacetamide (30 g). The flask was fitted with an oil bubbler and warmed to 80°C in an oil bath while degassing with nitrogen. After 30 min palladium (II) acetate (5 mg, 0.02 mmol) was added. The reaction was stirred at 100°C for 4 hr, cooled down to room temperature, diluted with water (150 ml), filtered, washed with water, and air-dried. The crude product, dark red crystalline material, 535 mg (83%), was dissolved in chloroform (350 ml), impregnated on silica gel (48 g) and flash-chromatographed with dichloromethane yielding, after

254

solvent removal, 40.5 mg (6%) of dark red crystals. M.p. 357.7°C (DSC, 10°C/min). 1H

(CDCl3, DMSO-d9) δ: 7.96 (d, 2H, J=8.7 Hz), 7.74 (d, 2H, J=8.7 Hz), 7.64 (d, 2H, J=8.7

Hz), 7.31 (AB, 2H, J=14.9), 7.1 (d, 2H, J=8.7 Hz), 3.4 (s, 3H), 2.26 (s, 3H). UV-Vis

(λmax): 279, 335, 510.

4-Diethylaminostyrene 57.

O N N

A 500 ml round bottom flask with a stir bar was charged with methyltriphenylphosphonium bromide (11.0 g, 30.8 mmol) and DMSO (70 ml). To that stirred mixture, potassium tert-butoxide (3.5 g, 31.2 mmol) was added at once and stirring continued for 30 min while yellow ylide color was developing. To the stirred reaction mixture, 4-diethylaminobenzaldehyde (5.4 g, 30.5 mmol) in DMSO (30 ml) was added dropwise during 40 min period and stirring continued for 4 additional hours. After the reaction was complete (TLC, hexane:ethyl acetate = 9:1), the reaction mixture was diluted with water (300 ml), extracted with ether (2×100 ml), and chromatographed on silica gel with hexane:ethyl acetate = 9:1 to allow, after evaporation of solvents, 1.82 g

24 1 δ (34%) of pale yellow oil. nD =1.540 (lit. 1.5904). H NMR (CDCl3) : 7.57 (d, 2H, J=9

Hz), 6.95 (m, 1H), 6.90 (d, 2H, J=9 Hz), 5.83 (dd, 1H, J1=17 Hz, J2=1.2 Hz), 5.30 (dd,

13 1H, J1=17 Hz, J2=1.2 Hz), 3.59 (q, 4H, J=7 Hz), 1.42 (t, 6H, J=7 Hz). C NMR (CDCl3)

δ: 147.7, 137.1, 127.8, 125.5, 111.8, 108.8, 44.6, 12.9. ES-MS: 176.2 (M+H).

255

1,4-divinylbenzene 58.

An oven-dried 500 ml three-neck round bottom flask with a stir bar was fitted with a gas inlet, cooled under argon flow and charged with methyltriphenylphosphonium bromide (17.9 g, 0.05 mol) and anhydrous ether (150 ml). The flask was topped with a reflux condenser and a septum. To the above stirred mixture, n-butyl lithium solution (2.5

M in hexanes, 20 ml, 0.05 mol) was added from a syringe during 30 min period. The reaction mixture turned yellow and was stirred at room temperature for 4 hrs. After that time, terephthalic aldehyde (3.35 g, 0.05 mol) in ether (100 ml) was added dropwise from an additional funnel and reaction continued for 24 additional hours. The white precipitate of Ph3PO was filtered off, re-suspended in ether (300 ml) and filtered again. The combined ether filtrates were evaporated on rotovap, applied onto silica gel and chromatographed with neat distilled hexane to yield, after evaporation of hexane, 469 mg

(14%) of clear oil, which crystallizes upon standing to clear plates. M.p. 30°C (lit. m.p.

1 31°C from AcOH-H2O). H NMR (CDCl3) δ: 7.54 (m, 2H, J=1.5 Hz), 6.88 (ddt, 1H,

13 J1=17 Hz, J2=11 Hz, J3=1.5 Hz), 5.93 (d, 1H, J=17 Hz), 5.42 (d, 1H, J=11 Hz). C NMR

–1 (CDCl3) δ: 137.3, 136.7, 126.6, 113.9. IR (HATR, neat solid, ν, cm ): 3088, 3005, 1626,

1508, 1398, 985, 902, 840.

256

N,N-di-n-butyl-4-[2-(4-vinylphenyl)vinyl]aniline 59.

Bu Bu N I N + Bu Bu

A 250 ml round bottom flask with a stir bar was charged with N,N-dibutyl-4- iodoaniline (12.4 g, 37.4 mmol), 1,4-divinylbenzene (#3-8c, 4.9 g, 37.6 mmol), diisopropylethylamine (6 g, 46.5 mmol), tri-o-tolylphosphine (35 mg, 0.11 mmol), and dimethylacetamide (50 ml). This mixture was stirred under argon for 30 min and then palladium acetate (8 mg, 0.03 mmol) was added. The reaction mixture was stirred at

120°C for 5 hrs, cooled down, poured into water, and extracted with chloroform (3 × 100 ml). The combined extracts were washed with water (3 × 100 ml), saturated aq. ammonium chloride (2 × 100 ml), dried with MgSO4, and evaporated on rotavap to give

12.0 g (96%) of pale yellow oil, which crystallized on standing to yellow crystals. M.p.

1 65°C. H NMR (CDCl3) δ: 7.47-7.39 (m, 6H), 7.06 (dd, 1H, J1=16 Hz, J2=3 Hz), 6.90

(dd, 1H, J1=16 Hz, J2=3 Hz), 6.66 (d, 2H, J=8 Hz), 5.77 (d, 1H, J=16 Hz), 5.24 (d, 1H,

J=16 Hz), 3.32 (t, 4H, J=7 Hz), 1.62 (pentet, 4H, J=7 Hz), 1.45-1.35 (m, 4H), 1.00 (t, 6H,

13 J=7 Hz). C NMR (CDCl3) δ: 147.7, 136.7, 129.2, 128.7, 128.2, 127.7, 126.2, 125.6,

124.7, 123.6, 111.7, 111.0, 50.9, 29.6, 20.5, 14.1.

257

2,5-Dipropyl-3,6-bis(4-{2-[4-diethylaminophenyl]vinyl}phenyl)-2,5-dihydropyrrolo-

[3,4-c]pyrrole-1,4-dione 60.

O O N N

N N O N N O 44 60 Br

A 200 ml round bottom flask with a stir bar was charged with 2,5-dipropyl-3,6-bis(4- bromophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (1.05 g, 1.98 mmol), 4- diethylaminostyrene (1.05 g, 6 mmol), triethylamine (2.0 g), and dimethylacetamide (40 ml). The flask was topped with a reflux condenser, argon bubbler and degassed with stirring for 30 min under argon flow. Palladium acetate (2 mg) was added to the degassed reaction mixture. The reaction mixture was then heated at 120°C for 36 hrs. After the reaction was complete (TLC, hexane : ethyl acetate = 7:3), the reaction mixture was cooled down, poured into water (100 ml), extracted with chloroform (3×100 ml), and chromatographed onto silica gel (hexane : ethyl acetate = 7:3, then neat dichloromethane) to allow, after evaporation of solvents, 700 mg (49%) of dark red solid. The crude product was recrystallized from 1-propanol : chloroform (70 : 6 ml) to give 38 mg of dark red crystals. M.p. 247°C. 1H δ: 7.87 (d, 2H, J=8 Hz), 7.62 (d, 2H, J=8 Hz), 7.45 (d, 2H,

J=9 Hz), 7.2 (d, 1H, J=16 Hz), 6.9 (d, 1H, J=16 Hz), 6.70 (d, 2H, J=8 Hz), 3.81 (t, 2H,

J=7 Hz), 3.43 (q, 4H, J=7 Hz), 1.70 (sextet, 2H, J=7 Hz), 1.22 (t, 6H, J=7 Hz), 0.91 (t,

3H, J=7 Hz). 13C δ: 163.0, 147.8, 141.3, 131.2, 129.1, 128.3, 126.1, 126.0, 124.1, 122.6,

258

44.4, 43.7, 22.9, 12.7, 11.2. ES-MS: 719.2 (M+H). IR (HATR, neat solid, ν, cm–1): 2951,

1671, 1586, 1528, 1401, 1359, 1185, 1153, 1084, 1010, 960, 825. UV-Vis λmax, nm (lgε):

333 (4.99), 538 (5.08). Fluorescence λmax: 620. ΦF=0.72.

2,5-Dipropyl-3,6-bis(4-thien-2-yl-phenyl)-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4- dione 64.

O N N O Br S Br S N O O N 44 64

A 200 ml round bottom flask with a stir bar was charged with magnesium turnings (0.20 g, 8.3 mmol), 2-bromothiophene (1.0 g, 6.1 mmol), anhydrous THF (20 ml), and refluxed for 4 hrs. After the Grignard reagent had been formed, its THF solution was decanted from excess of magnesium, and to that solution anhydrous zinc chloride

(2.0 g, 14 mmol) in anhydrous THF (30 ml) was added at once and the reaction mixture was stirred for 1 hr at room temperature while precipitating magnesium chloride. Then a solution of Br-DPP-Pr 44 (0.68 g, 1.28 mmol) in warm THF (120 ml), followed by tetrakis(phenylphospine)palladium (0) (37 mg, 0.03 mmol) were added and the resulting mixture was refluxed for 12 hrs. Chromatographic separation on silica gel with neat dichloromethane gave 1.25 g of dark red solid, which after recrystallization from chloroform-ethanol yielded 576 mg (84%) of ruby red crystals. M.p. 281°C. 1H δ: 7.90

259

(d, 2H, J=8Hz), 7.80 (d, 2H, J=8Hz), 7.45 (dd, 1H, J1=3.5 Hz, J2=1.3 Hz), 7.4 (dd, 1H,

J1=5 Hz, J2=1.3 Hz), 7.14 (m, 1H), 3.81 (t, 2H, J=7.5Hz), 1.70 (sextet, 2H, J=7.5 Hz),

13 0.91 (t, 3H, J=7.5 Hz). C (CDCl3, 50°C) δ: 162.9, 147.8, 143.5, 137.1, 129.5, 128.4,

127.3, 126.2, 124.4, 110.2, 43.8, 23.0, 11.3. UV-Vis λmax, nm: 315, 497. Fluorescence

λmax: 616, 690sh. ΦF=0.96.

3,6-Bis{4-[5-(4-dihexylaminophenyl)thien-2-yl]phenyl}-2,5-dipropylpyrrolo[3,4-c]- pyrrole-1,4-dione 65.

N(C6H13)2 S O N O Br N Br N O N O 44 S 65

(H13C6)2N A 500 ml round bottom flask with a stir bar was charged with 2-(4- dihexylaminophenyl)thiophene (5.5 g, 16 mmol), anhydrous ether (100 ml), fitted with an argon bubbler and cooled to –20°C. To that stirred mixture, butyl lithium (1.6 M in hexanes, 10 ml, 16 mmol) was added dropwise via syringe, the cooling bath was removed, the mixture was stirred for 30 min and cooled to 0°C. A solution of anhydrous zinc chloride (3.5 g, 25 mmol) in anhydrous THF (20 ml) was added at once, the reaction mixture was allowed to warm up to room temperature and stirred for 1 hr. A suspension of Br-DPP-Pr 44 (3.9 g, 7.3 mmol) in anhydrous THF (200 ml) was added at once folowed by tetrakis(triphenylphosphine)palladium (35 mg) and the resulting mixture was

260

refluxed for 24 hrs. After the reaction was complete, the reaction mixture was cooled

down, solvents evaporated on a rotovap, the residue was dissolved in chloroform (200

ml) and washed with HCl (2M aq., 2×100 ml), water (200 ml) and applied onto silica gel.

Chromatography with gradient elution from dichloromethane to chloroform-methanol

allowed, after solvent evaporation, 2.5 g (32%) of dark purple crystals. M.p. 278°C. 1H δ:

7.91 (d, 2H), 7.76 (d, 2H), 7.51 (d, 2H), 7.37 (d, 1H), 7.15 (d, 1H), 6.69 (d, 2H), 3.84 (t,

2H), 3.33 (t, 4H), 1.65 (m, 6H), 1.38 (m, 12H), 0.97-0.87 (m, 9H). 13C δ: 162.9, 148.4,

147.8, 146.9, 140.0, 137.6, 129.5, 127.1, 126.7, 125.5, 125.4, 121.8, 121.6, 112.3, 110.2,

51.3, 43.9, 31.8, 27.5, 27.0, 23.0, 22.7, 14.0, 11.2. UV-Vis λmax, nm (ε): 354 (37652), 539

(47585). Fluorescence λmax: 615, 693 sh. ΦF=0.82.

3,6-Diphenyl-2-(3-hydroxysulfonylpropyl)-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione 67. H H N O N O

O N O N H 1 67 O S HO O A 250 ml round bottom flask with a stirbar was charged with DPP 1 (1.44 g, 5

mmol), potassium tert-butoxide (1.2 g, 10.5 mmol), dimethylacetamide (50 ml), and

heated at 80°C for 30 min while the tert-butanol formed was removed under reduced

pressure. To the cooled reaction mixture, 1,3-propane sultone (1.2 g, 10.0 mmol) was

added dropwise resulting in an exothermic reaction. After the addition was complete (10

min), the reaction mixture was heated at 80°C for an additional hour and cooled down.

261

Water (50 ml) was added and stirred for 2 hrs, the resulting solution was filtered, cooled

in a refrigerator and acidified with cold hydrochloric acid (12M, 30 ml). The acidified

solution upon refrigeration deposited a precipitate, which was filtered off and redissolved

in a mixture of 1-propanol (30 ml), water (2 ml), and hydrochloric acid (12M, 1 ml).

Evaporation of the solvents allowed 1.3 g (61%) of dark brown crystals with blue luster.

1 M.p. 270°C. H (300 MHz, DMSO-d6) δ: 11.0 (s, 1H), 8.5 (m, 2H), 7.82 (m, 2H), 7.57-

13 7.50 (m, 3H), 3.96 (t, 2H), 2.85 (t, 2H), 2.11 (m, 2H). C (75 MHz, DMSO-d6) δ: 162.4,

161.6, 146.0, 145.8, 132.2, 131.0, 129.1, 128.9, 128.7, 127.9, 127.7, 127.5, 110.8, 108.4,

49.0, 40.6, 24.9. UV-Vis (EtOH) λmax, nm: 465, 486. Fluorescence (EtOH) λmax: 525,

566sh. ΦF=0.77.

3,6-Diphenyl-2-(4-hydroxysulfonylbutyl)-5-propylpyrrolo[3,4-c]pyrrole-1,4-dione 68.

Pr Pr N N O O + O S O O N OO O N H HO S 68 33 O A 200 ml round bottom flask with a stir bar was charged with 3,6-diphenyl-2-

propyl-5H-pyrrolo[3,4-c]pyrrole-1,4-dione 21 (1.0 g, 3.3 mmol), dimethylacetamide (20

ml), and sodium hydride (0.42 g, 17.5 mmol). The reaction mixture was stirred at 40°C

for an hour, 1,4-butanesultone (5.0 g, 36.7 mmol) was added dropwise and stirred at

room temperature for 12 hrs. After the reaction was completed, the solvent was rotary

evaporated, the residue dissolved in chloroform (150 ml), applied onto silica gel and

262

1 chromatographed with CH2Cl2:MeOH = 9:1 to give 145 mg of dark brown oil. H δ: 8.05

(d, 2H), 7.33-7.52 (m, 8H), 4.52 (t, 4H), 3.13 (m, 2H), 2.23 (m, 2H), 1.82 (m, 2H), 0.83

(t, 3H). UV-Vis λmax, nm: 275, 465, 486. Fluorescence (EtOH) λmax: 524, 565sh.

ΦF=0.73.

3,6-bis(4-[ethyl(2-hydroxyethyl)amino]phenyl)-2,5-dipropylpyrrolo[3,4-c]pyrrole-

1,4-dione 70.

O N N O OH I N I N N HO O O N 46 70

A 100 ml round bottom flask with a stirbar was charged with I-DPP-Pr 46 (286 mg, 0.458 mmol), 2-(ethylamino)ethanol (10 g), copper powder (5 mg, 0.08 mmol), copper (I) iodide (10 mg, 0.05 mmol), dimethylacetamide (5 ml), fitted with air condenser and heated under argon at 80°C. After 12 hours the reaction mixture was cooled down to room temperature, diluted with chloroform (70 ml) and filtered. The filtrate was washed with water (3×150 ml) in a separatory funnel, dried with MgSO4 and applied onto silica gel. Chromatography with 20 to 80% of ethyl acetate in dichloromethane gave 50 mg (20%) of the product as a dark red oil. 1H δ: 7.8 (d, 4H), 6.8

(d, 4H), 3.8-3.3 (m, 12H), 1.74 (m, 4H), 1.12 (t, 6H), 0.9 (t, 6H). 13C δ: 163.4, 150.0,

263

145.0, 130.8, 129.2, 115.8, 111.6, 107.7, 60.1, 52.4, 45.6, 44.0, 23.1, 12.0, 11.4. UV-Vis

λmax: 281, 369, 534.

6-(tetrahydro-2H-pyran-2-yloxy)hexan-1-ol.

OH OO

OH OH

A 100 ml round bottom flask with a stir bar was charged with 1,6-hexanediol

(8.2 g, 0.07 mol), Dowex 50WX2-100 resin (7 g), and tetrahydrofuran (50 ml). To that mixture, dihydropyran (10.0 g, 0.118 mol) was added dropwise (during 30 min) at room temperature with vigouros stirring. After the addition was complete, stiring was continued for three additional hours. The resin was filtered off and the solvent was evaporated. The colorless oil was impregnated onto silica gel and chromatographed with hexane : ethyl acetate = 4:1 to elute 1,6-bis(tetrahydropyran-2-yloxy)hexane (3.65 g,

18%), followed by hexane : ethyl acetate = 1:1 to elute 6-(tetrahydropyran-2-

25 21 yl)oxyhexan-1-ol (6.32 g, 45%) as colorless oil. nD =1.4777 (lit. nD = 1.457). EI-MS: 203

(M++1). IR (neat) ν, cm–1: 3406, 2938, 2863, 1454, 1441, 1353, 1201, 1138, 1121, 1077,

1026, 982, 868, 813. 1H NMR δ: 4.52 (m, 1H), 3.81 (m, 1H), 3.7-3.6 (m, 2H), 3.57-3.54

(m, 1H), 3.47-3.43 (m, 1H), 3.38-3.32 (m, 1H), 1.83-1.75 (m, 1H), 1.68-1.62 (m, 1H),

13 1.58-1.44 (m, 8H), 1.35 (m, 4H). C NMR (CDCl3) δ: 98.8, 67.4, 62.5, 62.3, 32.6, 30.8,

29.7, 26.1, 25.5, 25.4, 19.5.

6-(tetrahydro-2H-pyran-2-yloxy)hexyl 4-methylbenzenesulfonate 71.

264

O O O O 71 OH OTs

A 200 ml round bottom flask with a stir bar was charged with 6-

(tetrahydropyran-2-yl)oxyhexan-1-ol (10.0 g, 0.05 mol), pyridine (8 g), and ether (100 ml). The reaction mixture was cooled in an ice-acetone bath to 0°C and powdered 4- toluenesulfonyl chloride (12.0 g, 0.063 mol) was added at once. The reaction was allowed to warm up in the bath during 12 hrs and then was filtered off. The precipitate on filter was washed with ether (2×100 ml). The combined ethereal filtrates were chromatographed with hexane : ethyl acetate = 4:1 to elute elute 6-(tetrahydropyran-2- yl)oxyhex-1-yl 4-methylbenzenesulfonate (6.4 g, 36%) as colorless oil. 1H NMR δ: 7.76

(m, 2H), 7.33 (m, 2H), 4.52 (m, 1H), 4.0 (m, 2H), 3.82 (m, 1H), 3.67 (m, 1H), 3.46 (m,

1H), 3.32 (m, 1H), 2.43 (s, 3H), 1.78 (m, 1H), 1.63 (m, 1H), 1.50 (m, 8H), 1.30 (m, 4H).

13 C NMR (CDCl3) δ: 144.7, 133.1, 129.8, 127.8, 98.9, 70.6, 67.3, 62.4, 30.7, 29.5, 28.7,

25.6, 25.4, 25.2, 21.6, 19.7 (16 C). IR: the band 3406 cm–1 is gone.

2-(6-iodohexyloxy)tetrahydro-2H-pyran 73.

O O O O 73 OH I

A 250 ml round bottom flask with a stir bar was charged with diphosphorus tetraiodide (2.4 g, 4.2 mmol) and carbon disulfide (100 ml). The mixture was sonicated until P2I4 dissolved. To that stirred mixture, 6-(tetrahydro-2H-pyran-2-yloxy)hexan-1-ol

(3.3 g, 16.3 mmol) was added at once to form brown precipitate. After one hour, solid

265

potassium carbonate (1.5 g, 0.01 mol) was added and stirring continued for additional 12 hrs. Saturated aqueous potassium carbonate (15 ml) was added, the layers separated. The

CS2 layer was dried with MgSO4 and applied onto silica gel. Chromatography with 10 to

20% of dichloromethane in hexane gave, after removal of solvents, 3.4 g (66%) of

22 transluscent oil. nD =1.5265. (Compare with the refractive index of the s.m. = 1.4589

1 and that of 1,6-diiodohexane = 1.5834). H NMR (CDCl3) δ: 4.51 (m, 1H), 3.81 (m, 2H),

3.68 (m, 2H), 3.48 (m, 2H), 3.34 (m, 2H), 3.16 (t, 2H), 1.8 (m, 2H), 1.5 (m, 4H), 1.35 (m,

13 4H). C (CDCl3) δ: 98.9, 67.5, 62.5, 33.3, 30.5, 29.7, 29.5, 25.7, 25.4, 19.8, 7.05.

3,6-diphenyl-2,5-bis(6-(tetrahydropyran-2-yl)oxyhex-1-yl)pyrrolo[3,4-c]pyrrole-1,4- dione 74.

O O O O O O HN NH + N N I O O 73 O O 1 74

A 100 ml round bottom flask with a stir bar was charged with 3,6-diphenyl-2,5- dihydropyrrolo[3,4-c]pyrrole-1,4-dione (1.44 g, 5 mmol), potassium tert-butoxide (2.0 g,

17.8 mmol), and dimethylacetamide (35 ml). The reaction mixture was sonicated for five minutes, placed onto rotovap and tert-butanol was evaporated under 1 mm Hg vacuum.

To that mixture, 1-(6-iodohexyloxy)tetrahydropyran (3.12 g, 10 mmol) was added dropwise with vigorous stirring at room temperature (~20 min). After the addition was

266

complete, the flask was fitted with air condenser and heated under argon at 40°C. After

12 hours the reaction mixture was cooled down to room temperature, diluted with chlorofrom (150 ml) and filtered. The filtrate was washed with water (3×150 ml) in a separatory funnel, dried with MgSO4 and appled onto silica gel. Chromatography with 40 to 80% of ethyl acetate in dichloromethane gave, after removal of solvents, 1.1 g (34%)

13 of dark red oil. C (CDCl3) δ: 162.5, 131.0, 128.8, 128.6, 128.2, 109.6, 98.7, 67.5, 62.2,

41.7, 30.7, 29.7, 29.5, 26.1, 25.6, 25.4.

6-(benzoyloxy)-1-hexanol.

O OH O

OH OH

A 250 ml round bottom flask with a stir bar was charged with 1,6-hexanediol

(5.0 g, 42.3 mmol), triethylamine (4.3 g, 42.2 mmol), and tetrahydrofuran (30 ml). The mixture was cooled in an ice bath to ~5°C and cold benzoyl chloride (5.95 g, 42.3 mmol) was added dropwise during a 30 min period, with vigorous stirring so that the temperature did not rise above 10°C. After the addition was complete, the reaction mixture was stirred for six additional hours. The precipitate of triethylammonium chloride was filtered off and the filtrate evaporated on a rotovap. The residue was chromatographed first with 20% ether in hexane to elute diester (3.1 g, 22%) and then

1 with neat ether to elute 6-(benzoyloxy)hexan-1-ol (5.6 g, 60%). H (CDCl3) δ: 8.0 (dd,

2H), 7.5 (t, 1H), 7.41 (t, 2H), 4.3 (t, 2H), 3.6 (t, 2H), 2.6 (s, br, 1H), 1.8 (pentet, 2H), 1.6

267

13 (pentet, 2H), 1.4 (m, 4H). C (CDCl3) δ: 167.0, 132.9, 130.4, 129.5, 128.3, 65.0, 62.5,

32.5, 28.7, 25.9, 25.5.

6-(benzoyloxy)-1-iodohexane 75.

O O

OH 75 I

A one liter two neck round bottom flask with a stir bar was charged with phosphoric acid (85% aq., 32g, 0.277 mol), polyphosphoric acid (84%, 2H3PO4×P2O5, 32 g, 0.094 mol), the mixture was swirled until a homogeneous solution was formed, and was allowed to cool to room temperature. To that mixture, potassium iodide (36.5 g, 0.22 mol) and 6-(benzoyloxy)-1-hexanol (48 g, 0.216 mol) were added at once, the flask was fitted with a nitrogen/thermometer adapter, and a nitrogen bubbler. The reaction mixture was heated to 110±10°C on a heating mantle for four hours (monitored by TLC with neat hexane or 10% ether in hexane), cooled down and ether (300 ml) added. The layers were separated and the organic layer was washed with sodium sulfite (10% aq., 2×100 ml), water (2×200 ml), dried with MgSO4, and the solvent was removed on a rotovap. The residue was chromatographed with gradual elution from neat hexane to ether.

Fraction 1 was eluted with neat hexane: 15.4 g, identified as a mixture of 1,6- diiodohexane (traces) and 6-(benzoyloxy)-1-iodohexane. Fraction 2 was eluted with 6% ether in hexane: 43 g (60%) of (benzoyloxy)-1-iodohexane as a colorless liquid.

268

23 nD =1.5494. Fraction 3 was eluted with 20% ether in hexane: 8.4 g. Recrystallized from ethanol (30 ml) to give 6.5 g of white crystals. Fraction 4 was eluted with ether: 4.4 g.

1 Fraction 2: 6-(benzoyloxy)-1-iodohexane. H (CDCl3) δ: 8.0 (dd, 2H), 7.6 (t,

1H), 7.5 (t, 2H), 4.3 (t, 2H), 3.2 (t, 2H), 1.9 (pentet, 2H), 1.8 (pentet, 2H), 1.5 (m, 4H).

13 C (CDCl3) δ: 166.7, 132.9, 130.4, 129.5, 128.4, 65.2, 33.4, 30.2, 28.6, 25.1, 6.9.

1 Fraction 3: 1,6-dibenzoyloxyhexane. H (CDCl3) δ: 8.0 (dd, 2H), 7.6 (td, 1H),

13 7.5 (t, 2H), 4.4 (t, 2H), 1.8 (pentet, 2H), 1.6 (pentet, 2H). C (CDCl3) δ: 166.7, 132.9,

130.4, 129.5, 128.4, 64.9, 28.7, 25.8. M.p. 53°C.

13 Fraction 4: was not identified as a discrete compound. C (CDCl3) δ: 169.7,

166.8, 133.2, 132.9, 130.0, 129.5, 128.4, 65.0, 62.4, 62.3, 33.4, 32.2, 32.2, 30.2, 28.6,

25.8, 25.4, 24.7, 7.1.

3,6-diphenyl-2,5-bis(6-(benzoyloxy)hex-1-yl)-pyrrolo[3,4-c]pyrrole-1,4-dione 76.

O O H N N O HN NH +

O N O O N 1 76 77

O O

A 200 ml round bottom flask with a stir bar was charged with 3,6-diphenyl-2,5- dihydropyrrolo[3,4-c]pyrrole-1,4-dione (1.15 g, 4 mmol) and dimethylacetamide (70 ml).

269

To this reaction mixture, a solution of lithium diisopropylamide (prepared from n-butyl lithium 1.7M, 2.3 ml, and diisopropylamine, 4 ml) in ether (30 ml) was added at once.

The reaction mixture was allowed to stir at room temperature for one hour and 6-

(benzoyloxy)-1-iodohexane (3.12 g, 9.4 mmol) was added dropwise with vigorous stirring at room temperature. After the addition was complete, the flask was fitted with air condenser and heated under argon, gradually increasing the temperature from 80 to

160°C. After 26 hours the reaction mixture was cooled down to –10°C, and ice-cold hydrochloric acid (12M, 15 ml) was added dropwise. The precipitate formed was filtered off, washed with water (3×100 ml), dissolved in chloroform (50 ml), dried with MgSO4 and appled onto silica gel. Chromatography with 10% ethyl acetate in dichloromethane to neat ethyl acetate gave, after removal of solvents, two fractions. Fraction 1 was 76: 0.64 g (23%) of brown-orange crystals. M.p. 111°C (1-propanol). 1H δ: 8.1–8.0 (m, 2H), 7.8

(dd, 2H), 7.6-7.5 (m, 3H), 7.47–7.43 (m, 3H), 4.3 (t, 2H), 3.7 (t, 2H), 1.7 (m, 4H), 1.4 (m,

4H). 13C δ: 166.6, 162.7, 148.5, 132.8, 131.2, 130.4, 129.5, 128.9, 128.7, 128.3, 128.2,

109.7, 64.8, 41.7, 29.3, 28.6, 26.4, 25.5. UV-Vis λmax: 475. Fluorescence λmax: 527,

563sh. ΦF=0.97. Fraction 2 was 77: 0.155 g (8%) of bright orange crystals, m.p. 194°C.

270

3,6-diphenyl-2-(6-(benzoyloxy)hex-1-yl)-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione 77. H N O

HN NH O N

O O 1 77 O A 200 ml round bottom flask with a stir bar was charged with 3,6-diphenyl-2,5- dihydropyrrolo[3,4-c]pyrrole-1,4-dione (0.288 g, 1 mmol), potassium tert-butoxide (0.33 g, 2.9 mmol), and dimethylacetamide (70 ml). The reaction mixture was sonicated for five minutes, placed onto rotovap and tert-butanol was evaporated under 1 mm Hg vacuum. To that mixture, 6-(benzoyloxy)-1-iodohexane (0.71 g, 2.1 mmol) was added dropwise with vigorous stirring at room temperature. After the addition was complete, the flask was fitted with air condenser and heated under argon at 60°C. After 6 hours the reaction mixture was cooled down to room temperature, diluted with chlorofrom (100 ml) and filtered from unreacted starting material. The filtrate was washed with water (3×100 ml) in a separatory funnel, dried with MgSO4 and appled onto silica gel. Chromatography with 20% ether in hexane to neat ether gave, after removal of solvents, 80 mg (11%) of red crystals. M.p. 194°C (1-propanol). 1H δ: 10.1 (s, br., 1H), 8.4 (dd, 2H), 8.0 (dd, 2H),

7.8 (m, 1H), 7.6 (m, 4H), 7.5 (m, 1H), 4.3 (t, 2H), 3.9 (t, 2H), 1.7 (m, 4H), 1.4 (m, 4H).

13 C (CDCl3 or DMSO-d6) δ: 166.3, 162.8, 162.1, 146.7, 133.4, 132.5, 131.2, 129.4,

129.3, 129.2, 129.0, 128.9, 128.4, 111.5, 109.1, 64.9, 41.5, 28.9, 28.4, 26.0, 25.3. UV-Vis

λmax: 467, 493. Fluorescence λmax: 522, 563sh. ΦF=0.96.

271

3,6-diphenyl-2-(6-(benzoyloxy)hex-1-yl)-5-propylpyrrolo[3,4-c]pyrrole-1,4-dione 78 was prepared similar to 79, starting from 3,6-diphenyl-2-propyl-5-hydropyrrolo[3,4- c]pyrrole-1,4-dione 33 and using LDA (prepared in situ from n-BuLi and i-Pr2NH) as a base.

3,6-diphenyl-2-(6-(benzoyloxy)hex-1-yl)-5-dodecylpyrrolo[3,4-c]pyrrole-1,4-dione 79.

H O H25C12 N O N O O +

O N I O N 38 C12H25 74 79 O

A 200 ml round bottom flask with a stir bar was charged with 3,6-diphenyl-2- dodecyl-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione (0.205 g, 0.449 mmol) and dimethylformamide (70 ml, to insure there is no precipitation on cooling). The reaction mixture was cooled in an ice bath and tert-butyl lithium (1.7 M solution in hexanes, 1.0 ml) was added via syringe under argon. (t-BuLi probably reacts with DMAc, but the enolate formed should be a good base as well.) The ice bath was removed and the mixture was stirred at room temperature for 1 hr. 6-(Benzoyloxy)-1-iodohexane (0.250 g,

0.75 mmol) was added at once and the reaction mixture was heated at 80°C for 52 hrs.

After the reaction was complete by TLC (starting material is gone, two new fluorescent spots), it was cooled down to room temperature, diluted with chlorofrom (100 ml), washed with water (3×100 ml) in a separatory funnel, dried with MgSO4 and appled onto

272

silica gel. Chromatography with 10 to 35% ether in hexane to neat ether gave, after

13 removal of solvents, 0.1 g (34%) of orange crystals, m.p. 74°C. C (CDCl3) δ: 166.6,

162.7, 148.7, 148.4, 132.9, 131.1, 130.4, 129.6, 129.0, 128.9, 128.7, 128.6, 128.4, 128.2,

109.8, 109.6, 64.8, 42.1, 32.6, 31.8, 29.5, 29.3, 29.2, 29.0, 28.7, 28.6, 26.7, 26.4, 25.9,

25.6, 22.7, 14.1.

3,6-diphenyl-2,5-bis(6-hydroxyhex-1-yl)-pyrrolo[3,4-c]pyrrole-1,4-dione 80.

N O O

HO (CH2)6 N N (CH2)6 OH N O O 76 80 O

O A 350 ml glass pressure vessel with a stir-bar was charged with 28 (38 mg, 0.05 mmol), anhydrous methanol (50 ml), and titanium triisopropoxide (0.1 ml, 0.34 mmol).

The vessel was closed with a Teflon plug and heated, with stirring at 180°C (mantle temperature) for 72 hrs. Each 24 hr the vessel was cooled and an aliquot was taken to monitor reaction progress by TLC (neat EtOAc as eluent). After the reaction was complete, the reaction mixture was cooled down to room temperature and methanol was evaporated under reduced pressure. The residue was impregnated onto silica gel.

Chromatography with neat ethyl acetate gave 17 mg (64%) of orange crystals. 1H δ: 7.8

(dd, 2H), 7.6-7.5 (m, 3H), 3.8 (t, 2H), 3.6 (t, 2H), 1.65-1.57 (pentet, 2H), 1.50 (pentet,

273

2H), 1.2 (m, 4H). 13C δ: 162.7, 148.7, 131.1, 128.9, 128.6, 128.2, 110.0, 62.7, 41.8, 32.5,

29.4, 26.4, 25.1. UV-Vis λmax: 474. Fluorescence λmax: 525, 567sh. ΦF=0.94.

3,6-diphenyl-2-(6-(hydroxy)hex-1-yl)-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione 81.

H O H N N O

N O O N

77 O 81 OH O

A 50 ml round bottom flask with a stir bar was charged with 3,6-diphenyl-2-(6-

(benzoyloxy)hex-1-yl)-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione (70 mg, 0.1 mmol), potassium cyanide (5 mg, 0.077 mmol), and anhydrous methanol (20 ml). The resulting suspension was heated, with stirring, under argon at 50°C. After two hours, the reaction mixture was cooled down, methanol evaporated, and the residue was chromatographed with 20% ether in hexane to neat ether to neat ether to allow 50 mg (63%) of yellow oil.

1H δ: 8.9 (s, br., 1H), 8.3 (m, 2H), 7.80 (m, 2H), 7.56 (m, 6H), 3.80 (t, 2H), 3.6 (t, 2H),

1.66 (m, 2H), 1.53 (m, 2H). 162.6, 148.4, 132.0, 131.0, 129.1, 128.8, 128.7, 127.7, 109.9,

62.8, 42.1, 32.4, 29.3, 26.4, 25.1. UV-Vis λmax: 467, 486.

3,6-diphenyl-2-(6-(hydroxy)hex-1-yl)-5-propylpyrrolo[3,4-c]pyrrole-1,4-dione 82 was prepared similar to 83, starting from 78. Yield 89%.

274

3,6-diphenyl-2-(6-(hydroxy)hex-1-yl)-5-dodecylpyrrolo[3,4-c]pyrrole-1,4-dione 83.

H25C12 H25C12 N O N O

O N O N 79 83

O OH

A 350 ml glass pressure vessel with a stir-bar was charged with 3,6-diphenyl-2-

(6-(benzoyloxy)hex-1-yl)-5-dodecylpyrrolo[3,4-c]pyrrole-1,4-dione (100 mg, 0.15 mmol), anhydrous methanol (70 ml), and titanium triisopropoxide (0.5 ml, 1.7 mmol).

The vessel was closed with a Teflon screw-in plug and heated, with stirring at 180°C

(mantle temperature) for 72 hrs. Each 24 hr the vessel was cooled and an aliquote taken to monitor reaction progress by TLC (neat EtOAc as eluent). After the reaction was complete, the reaction mixtu.re was cooled down to room temperature and methanol evaporated under reduced pressure. The residue was impregnated onto silica gel.

Chromatography with neat ethyl acetate gave, after removal of solvents, 68 mg (81%) of

83 as orange crystals. 1H δ: 7.8 (dd, 4H), 7.5 (m, 6H), 3.8 (m, 4H), 3.6 (t, 2H), 1.6 (m,

2H), 1.5 (m, 2H), 1.3–1.2 (m, 26H), 0.9 (t, 3H). 13C δ: 162.73, 162.70, 148.7, 148.3,

131.1, 128.9, 128.7, 128.2, 109.8, 109.6, 62.6, 41.9, 41.6, 32.4, 31.8, 29.7, 29.5, 29.4,

29.3, 29.2, 29.0, 26.7, 26.4, 25.0, 22.7, 14.1. UV-Vis λmax: 468, 486. Fluorescence λmax:

522, 563sh. ΦF=0.89.

275

3,6-diphenyl-2-(10-hydroxy-1-decyl)-5-propylpyrrolo[3,4-c]pyrrole-1,4-dione 84.

O O

HNN Pr HO (CH2)10 NNPr

O O 84 33

A 100 ml round bottom flask with a stir-bar was charged with 3,6-diphenyl-2- propyl-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione (0.5 g, 1.51 mmol), cesium carbonate (1.0 g, 3.0 mmol), 10-bromo-1-decanol (0.5 g, 2.1 mmol) and dimethylacetamide (40 ml). The reaction mixture was stirred at 60°C (mantle temperature) under argon and monitored by

TLC (DCM as eluent). After 6 hours the reaction mixture was cooled down to –10°C, and ice-cold hydrochloric acid (12M, 5 ml) was added dropwise. The precipitate formed was extracted with chloroform (80 ml) and appled onto silica gel. Chromatography with 50% ethyl ether in hexane to neat ether gave, after removal of solvents, 490 mg of brown oil.

The oil crystallized upon sonication with cold ether, was recrystallized from ether- chloroform to give 370 mg (49%) of 35 as bright yellow crystals. M.p. 105°C. 1H δ: 7.8

(m, 4H), 7.5 (m, 6H), 3.75 (m, 4H), 3.6 (t, 2H), 1.6-1.5 (m, 6H), 1.3-1.2 (m, 12H), 0.9 (t,

3H). 13C δ: 162.7, 162.63, 148.5, 131.1, 128.9, 128.7, 128.6, 128.2, 109.75, 109.71, 63.0,

43.3, 41.8, 32.7, 29.39, 29.32, 29.25, 28.91, 26.6, 25.7, 22.7, 11.1. UV-Vis λmax: 467,

490. Fluorescence λmax: 530, 561sh. ΦF=0.76.

276

3,6-diphenyl-2,5-bis(10-hydroxy-1-decyl)pyrrolo[3,4-c]pyrrole-1,4-dione 85.

O O

HNHN HO (CH2)10 NN (CH2)10 OH

O O 1 85

A 200 ml round bottom flask with a stirbar was charged with DPP 1 (1.0 g, 3.47 mmol), cesium carbonate (3.4 g, 10.4 mmol), 10-bromo-1-decanol (1.8 g, 7.6 mmol) and dimethylacetamide (70 ml). The reaction mixture was stirred at 120°C (mantle temperature) under argon and monitored by TLC (DCM:EtOAc = 1:1). After 6 hours the reaction mixture was cooled down to –10°C, and ice-cold hydrochloric acid (12M, 7 ml) was added dropwise. The precipitate formed was extracted with chloroform (80 ml), washed with water (2×50 ml), dried with MgSO4 and applied onto silica gel.

Chromatography with 50% ethyl acetate in dichloromethane gave 700 mg (34%) of dark brown oil, which was crystallized from 1-propanol to give 125 mg (6%) of orange crystals. The mother liquors after recrystallization contain only the target (by TLC) but fail to deposit second crop of crystals. M.p. 143°C. 1H δ: 7.8 (dd, 2H), 7.5 (m, 3H), 3.8 (t,

2H), 3.6 (t, 2H), 1.6-1.5 (m, 4H), 1.3-1.2 (m, 12H). 13C δ: 162.6, 148.4, 130.9, 128.8,

128.7, 127.6, 109.9, 63.0, 41.8, 32.8, 29.3, 29.2, 28.9, 26.6, 25.6. UV-Vis λmax: 475.

Fluorescence λmax: 525, 566sh. ΦF=0.87.

277

3,6-diphenyl-2-(10-hydroxy-1-decyl)-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione.

O O

HNHN HO (CH2)10 NNH

O O

3,6-diphenyl-2-(10-hydroxy-1-decyl)-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione was prepared similar to 85 starting from DPP (1.0 g, 3.47 mmol) and 10-bromo-1- decanol (1.8 g, 7.5 mmol). Chromatography with a gradient of 0-100% ethyl acetate : dichloromethane, gave 0.87 g of brown oil. The oil was crystallized from 1-propanol to

1 give 0.7 g (34%) of mono-(HOC10H21)DPP as orange crystals. M.p. 166°C. H δ: 7.8 (dd,

2H), 7.5 (m, 3H), 3.8 (t, 2H), 3.8 (t, 1H), 3.6 (t, 2H), 1.6–1.5 (m, 2H), 1.4–1.2 (m, 14H).

13C δ: 162.6, 148.4, 130.9, 128.8, 128.7, 128.0, 109.9, 63.0, 41.8, 32.8, 29.3, 29.2, 29.1,

28.9, 28.7, 26.6, 25.6. UV-Vis λmax: 468, 493. Fluorescence λmax: 523, 563sh. ΦF=0.86.

2-(6-iodohexyl)-3a,4,7,7a-tetrahydro-4,7-epoxy-1H-isoindole-1,3(2H)-dione 86.

CAS RN [874998-65-9]

O O

O NH O N I O O 120 86

A 100 ml round-bottom flask with a stirring bar was charged with the adduct of furan and maleimide (1.65 g, 0.01 mol), 1,6-diiodohexane (3.7 g, 0.011 mol), potassium

278

carbonate (1.4 g, 0.11 mol), and 35 ml of acetone. The flask was topped with a reflux condenser and the reaction mixture was stirred at 40°C for 12 hours. The course of the reaction was monitored by TLC with 1:1 eluent of hexane and ethyl acetate. The solvent was evaporated; the residue dissolved in a minimum amount of chloroform and chromatographed with 3:1 to 1:1 mixture of hexane and ethyl acetate. Recrystallization from ethyl acetate yields 1.0 g (26.6%) of white crystalline material, m.p. 71.5°C (DSC,

1 10°/min). H NMR (CDCl3), δ: 6.52 (s, 1H); 5.25 (s, 1H); 3.46 (t, 2H, J=6.6Hz); 3.17 (t,

2H, J=6.6Hz); 2.84 (s, 2H); 1.8 (m, 2H); 1.56 (m, 2H); 1.45-1.26 (m, 4H). 13C NMR

(CDCl3), δ: 176.4, 136.7, 81.1, 47.6, 38.9, 33.4, 30.1, 27.5, 25.7, 7.1.

3,6-Diphenyl-2,5-bis(6-[3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2-yl]- hexyl)pyrrolo[3,4-c]pyrrole-1,4-dione 87.

O N O O N O O HNHN + O N O N O O I O N O 1 86 87 O

A 200 ml round bottom flask fitted with a stirbar, air condenser and nitrogen bubbler was charged with DPP 1 (2.88 g, 10 mmol), potassium tert-butoxide (2.5 g, 22 mmol), dimethylacetamide (80 ml), and heated at 80°C for 2 hrs. The resulting mixture was cooled to room temperature and a solution of 2-(6-iodohexyl)-3a,4,7,7a-tetrahydro-

4,7-epoxy-1H-isoindole-1,3(2H)-dione419 (7.54 g, 20 mmol) in dimethylacetamide (10

279

ml) was added at once and the reaction mixture was stirred at room temperature for 12 hrs, then at 100°C for 12 hrs, and cooled down. The solvent was evaporated to 50 ml, hydrochloric acid (70 ml 2M) was added, the resulting precipitate was filtered off, air- dried (5.11 g), dissolved in chloroform (150 ml), filtered, applied onto silica gel and chromatographed with gradual elution from hexane – ethyl acetate (1:1) to neat ethyl acetate. Evaporation of solvents afforded 0.59 g (7%) of Fu-MI-(CH2)6-DPP as red crystals. 1H δ: 8.32 (m, 1H), 7.81 (m, 2H), 7.51-7.58 (m, 4 H), 6.49 (m, 2H), 5.23 (m,

1H), 3.81 (t, 2H), 3.41 (t, 2H), 2.81 (s, 1H), 1.64-1.47 (m, 8H). 13C δ: 176.4, 136.7,

132.3, 131.4, 129.4, 129.1, 128.9, 128.0, 81.0, 47.5, 42.1, 38.9, 29.5, 27.5, 26.4, 26.2.

3,6-Diphenyl-2,5-bis(6-[2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl]hexyl)pyrrolo[3,4- c]pyrrole-1,4-dione 88.

O O

O N N O N N O O O

O O N O O N N O N 87 88 O O

A 100 ml round bottom flask fitted with a stirbar, air condenser and nitrogen bubbler was charged with Fu-MI-(CH2)6-DPP (265 mg, 0.345 mmol), xylenes (mixture of isomers, 70 ml) and refluxed for 24 hrs. The solvent was evaporated and the residue was chromatographed with gradual elution from neat chloroform to 5% ethyl acetate in chloroform. Evaporation of solvents gave orange glass (194 mg, 87%). Recrystallization

280

from dimethylacetamide (4 ml) gave 33 mg (14%) of orange crystals. 1H δ: 7.79 (dd,

2H), 7.54 (dd, 2H), 7.18 (m, 1 H), 6.66 (s, 2H), 3.74 (t, 2H), 3.44 (t, 2H), 1.53-1.39 (m,

8H). 13C δ: 171.0, 163.2, 148.1, 134.2, 131.3, 129.1, 128.8, 128.6, 128.2, 41.7, 37.7, 29.5,

27.5, 26.4, 26.2. ES-MS: 647.2 (M+1), 527.1. UV-Vis (CH2Cl2) λmax, nm (lg ε): 231

(3.98), 264 (3.79), 464 (3.31). Fluorescence λmax: 525, 565sh. ΦF=0.67.

3,6-Diphenyl-2-(6-[3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2-yl]hexyl)-

5-(3-hydroxysulfonylpropyl)-pyrrolo[3,4-c]pyrrole-1,4-dione 89.

O 1 O O O N O + S O N O HNHN O 2 O O I N + O N O 1 86 89 SO3H O

A 100 ml round bottom flask fitted with a stirbar, air condenser and nitrogen bubbler was charged with DPP 1 (1.44 g, 5 mmol), potassium tert-butoxide (1.2 g, 10.5 mmol), dimethylacetamide (50 ml), and heated at 80°C for 12 hrs. The resulting mixture was cooled to room temperature and a solution of 1,3-propanesultone (0.61 g, 5 mmol) in dimethylacetamide (20 ml) was added dropwise. After the addition was complete, the reaction mixture was stirred at room temperature for 1 hr, and then at 140°C for 4 hrs, and cooled back to room temperature. A solution of 2-(6-iodohexyl)-3a,4,7,7a- tetrahydro-4,7-epoxy-1H-isoindole-1,3(2H)-dione (1.88 g, 5 mmol) in dimethylacetamide

(20 ml) was added at once and the reaction mixture was stirred at room temperature for

281

12 hrs, then at 120°C for 4 hrs, and cooled down. The solvent was evaporated to 50 ml, hydrochloric acid (70 ml 2M) was added, the resulting precipitate was filtered off, air- dried, dissolved in a mixture of chloroform and methanol (100 ml), applied onto silica gel and chromatographed with gradual elution from hexane – ethyl acetate (1:1) to neat ethyl acetate. Evaporation of solvents afforded 500 mg (15%) of 41 as red crystals. M.p.

165°C. 1H δ: 10.27 (s, 1H), 8.3 (m, 1H), 8.13 (m, 3H), 7.8 (m, 1H), 7.4-7.6 (m, 8 H), 6.49

(m, 1H), 5.24 (m, 1H), 3.81 (t, 2H), 3.41 (t, 2H), 2.82 (s, 1H), 1.65-1.14 (m, 12H). 13C δ:

176.5, 171.3, 164.2, 149.2, 144.8, 140.5, 136.7, 133.8, 132.3, 131.6, 130.3, 129.4, 129.2,

128.9, 128.6, 128.0, 81.0, 47.5, 42.1, 38.9, 32.1, 29.8, 27.5, 26.2, 26.1, 22.9, 14.3. UV-

Vis λmax: 470, 489. Fluorescence λmax: 524, 565sh.

3,6-Diphenyl-2,5-bis(4-perfluorobutylsulfonylphenyl)-pyrrolo[3,4-c]pyrrole-

1,4-dione 91.

O O

HNHN C4F9–SO2 NN SO2–C4F9

O O 1 91

A 100 ml round bottom flask with a stirbar was charged with DPP 1 (720 mg,

2.5 mmol), copper (II) oxide (795 mg, 10 mmol), 1-bromo-4-(perfluorobutylsulfonyl)- benzene (2.81 g, 7.0 mmol), and dimethylformamide (15 ml). The reaction mixture was stirred and refluxed for three days, cooled down, poured into water (100 ml). The

282

precipitate formed was filtered off and treated with boiling ethyl acetate (5×70 ml).

Combined hot ethyl acetate solutions were filtered from insoluble matter, solvent evaporated, and the residue was recrystallized from 1-propanol to give 786 mg (32%) of

43 as orange crystals. M.p. 262°C. 1H δ: 8.05 (d, 2H), 7.57-7.55 (m, 2H), 7.52-7.47 (m,

3H), 7.41-7.36 (m, 2H). 13C δ: 142.8, 132.6, 132.1, 130.8, 129.8, 129.1, 128.3, 126.4.

UV-Vis λmax: 271, 475. Fluorescence λmax: 515, 555sh. ΦF=0.91.

3,6-Diphenyl-2-(4-perfluorobutylsulfonylphenyl)-5(H)pyrrolo[3,4-c]pyrrole-

1,4-dione 92. The residue, insoluble in ethyl acetate was dissolved in hot dimethylformamide, filtered off while hot and after cooling gave 297 mg (18%) of 92 as orange crystals. M.p. 320°C. 1H (DMSO) δ: 11.48 (s, 1H), 8.46 (dd, 2H), 8.02 (d, 2H),

7.55-7.45 (m, 7H), 7.37-7.29 (m, 3H). 13C δ: 162.7, 160.0, 148.5, 143.5, 142.1, 132.3,

131.4, 130.6, 128.9, 128.7, 128.6, 128.3, 128.1, 127.2, 126.8. UV-Vis λmax: 461, 485,

490. Fluorescence λmax: 513, 554sh. ΦF=0.93.

3,6-Diphenyl-2,5-bis(3,5-bis[trifluoromethyl]phenyl)-pyrrolo[3,4-c]pyrrole-

1,4-dione 93 was prepared similar to 91, starting from DPP 1 (2.88 g, 10 mmol) and 1- bromo-3,5-bis(trifluoromethyl)benzene (14.6 g, 50 mmol). Yield: 2.66 g (37%). M.p.

298°C. 1H δ: 7.85 (m, 2H), 7.66 (m, 4H), 7.57 (m, 2H), 7.55 (m, 3H), 7.50 (m, 2H), 7.47

(m, 1H), 7.43 (m, 2H), 7.41 (m, 2H), 7.38 (t, 1H). UV-Vis λmax: 251, 315, 475.

Fluorescence λmax: 520, 552sh. ΦF=0.95.

283

3,6-Diphenyl-2,5-bis(4-fluorophenyl)-pyrrolo[3,4-c]pyrrole-1,4-dione 94.

O O

HNHN FFNN

O O 1 94

A 50 ml pear-shaped flask with a stir bar and a reflux condenser was charged with DPP 1 (576 mg, 2 mmol), 1-bromo-4-fluorobenzene (1.05 g, 6 mmol), copper (I) oxide (1.15 g, 8 mmol), and dimethylformamide (30 ml). The reaction mixture was stirred and refluxed for 8 days. After the reaction was complete by TLC (CHCl3), it was cooled, diluted with chloroform (70 ml), and applied onto silica gel. Flash- chromatography on silica gel with CH2Cl2 as eluent afforded 115 mg (12%) of bright yellow crystals. M.p. 344°C. 1H δ: 7.56-7.53 (m, 1H), 7.46-7.37 (m, 2H), 7.32-7.28 (m,

2H). 13C not available due to low solubility.

3,6-Diphenyl-2-(4-fluorophenyl)-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione 95.

O O

HNHN F NNH

O O 1 95

A 100 ml pear-shaped flask with a stir bar and a reflux condenser was charged with 3,6-diphenyl-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione (2.88 g, 10 mmol), 1-iodo-

284

4-fluorobenzene (9.0 g, 40 mmol), copper (I) oxide (6.0 g, 42 mmol), and dimethylformamide (60 ml). The reaction mixture was stirred and refluxed for 24 hours, cooled down, diluted with chloroform (70 ml), filtered through #50 filter and applied onto silica gel. Flash-chromatography with hot chloroform as eluent afforded, after evaporation of solvent, 1.8 g (37%) of 48 as bright yellow crystals. M.p. 404°C. 1H δ:

10.4 (s, 1H), 7.64-7.56 (m, 4H), 7.36-7.31 (m, 4H), 7.23-7.07 (m, 6H).

2,5-Bis-(4-nitrophenyl)-3,6-diphenyl-2,5-dihydropyrrolo[3,4-c]pyrrole-1,4-dione 96.

O O

HNHN O2NNONN 2

O O 1 96

A 100 ml round bottom flask with a stirbar was charged with DPP 1 (1.44 g, 5 mmol), potassium tert-butoxide, and dimethylacetamide (70 ml). The reaction mixture was heated at 80°C for 30 min and the tert-butanol formed was removed under reduced pressure. To the cooled reaction mixture, 4-fluoro-1-nitrobenzene (3.0 g, 21 mmol) was added and the reaction mixture was stirred for 48 hrs at 80°C and then 96 hrs at 140°C.

The reaction mixture was cooled down, poured into water (150 ml) and filtered off. The solid on the filter was air dried and recrystallized from dimethylacetamide (15 ml) to give

320 mg (12%) of 96 as orange crystals. M.p. 200°C. 1H (300 MHz) δ: 8.27 (d, 4H), 8.06

285

(d, 1H), 7.16 (d, 4H), 6.57 (d, 1H). 13C (75 MHz) δ: 160.8, 154.4, 144.3, 132.2, 129.0,

128.5, 127.9, 124.8, 124.4, 119.5, 110.3. UV-Vis λmax: 268, 470.

3,6-diphenyl-2-(2,4-dinitrophenyl)-5-propylpyrrolo[3,4-c]pyrrole-1,4-dione 97.

O O

Pr NNH PrNN NO2

O O 33 97

A 100 ml round bottom flask with a stir bar was charged with 3,6-diphenyl-2- propyl-5-hydropyrrolo[3,4-c]pyrrole-1,4-dione 21 (0.3 g, 0.91 mmol), cesium carbonate

(0.75 g, 2.3 mmol), 1-fluoro-2,4-dinitrobenzene (0.37 g, 2 mmol), and dimethylacetamide

(45 ml). The reaction mixture was stirred at 80°C and monitored by TLC (eluent neat chloroform). After 8 hrs the mixture was cooled down, diluted with cold hydrochloric acid (3 M, 50 ml), extracted with chloroform (3×50 ml) and applied onto silica gel.

Chromatography with 60 to 100% dichloromethane in hexane gave, after evaporation of solvents and recrystallization from 1-propanol : chloroform (5 ml) 113 mg (25%) of orange crystals. M.p. 234°C dec. ES-MS: 497 (M++1). 1H δ: 8.9 (d, 1H, J=2.5 Hz), 8.4

(dd, 1H, J1=8.8 Hz, J2=2.5 Hz), 7.8 (m, 2H), 7.6 (m, 2H), 7.5 (m, 3H), 7.4 (m, 3H), 7.3

(M, 1H), 3.8 (t, 2H), 1.7 (m, 2H), 0.9 (t, 3H). 13C δ: 162.7, 159.5, 151.9, 146.0, 145.6,

143.8, 135.0, 132.1, 131.8, 131.2, 129.1, 129.0, 128.9, 127.3, 126.7, 121.1, 113.9, 107.5,

43.9, 22.7, 11.2. UV-Vis λmax (lg ε): 263 (4.45), 298 (4.31), 466 (4.26).

286

Basic hydrolysis of DPP 1.

H O OOH COOH N O

HOOC O N O H

An autoclave was charged with DPP (4.35 g), sodium dodecylsulfate (0.1 g, as a dispersinf agent), water (200 ml), potassium hydroxide (10 g) and ethanol (3 ml). The reaction mixture was stirred in an autoclave at 150°C for 12 hrs and cooled down. The colorless homogeneous solution formed was acidified with hydrochloric acid and the precipitate formed was filtered off, washed with water and air-dried to give 1.25 g of gray solid. The crude product was chromatographed with ethyl acetate and recrystallized from

1-propanol to give 1.2 g of white crystals, which were identified as benzoic acid. M.p.

121.5°C. 1H NMR (acetone-d6) δ: 8.1 (dd, 2H), 7.6 (td, 1H), 7.5 (t, 2H). 13C NMR

(acetone-d6) δ: 168.7, 134.0, 131.2, 130.6, 129.4.

6-[(3-carboxypropanoyl)amino]caproic acid.

O O O OH O O + H N 2 OH NH OH O O

A 500 ml round bottom flask with a stir bar was charged with ω-aminocaproic acid (13.1 g, 0.1 mol), maleic anhydride (9.8 g, 0.1 mol), and acetic acid (400 ml). The reaction mixture was stirred at room temperature for 24 hrs, the precipitate formed was

287

filtered off on a Büchner funnel, washed with water (200 ml), and air-dried to give 22.3

(97%) of maleamic acid 101. M.p. 170 °C. Lit.420 m.p. 171–173 °C.

6-(maleimid-2-yl)caproic (6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)caproic) acid 101.

O O O OH O N NH OH OH O O

A 500 ml round bottom flask with a stir bar was charged with . maleamic acid

101 (12.3 g, 0.05 mol), acetic anhydride (50 ml), and sodium acetate (2.5 g, 0.03 mol).

The reaction mixture was stirred at 90 °C for 2 hrs, cooled, and poured onto ice (100 g).

The aqueous solution was extracted with ether (3×50 ml) and the extracts were

impregnated onto silica gel. Chromatography with hexane – ethyl acetate (1:2), followed

by recrystallization from isopropanol–water, gave (49%) of ω-maleimidylcaproic acid as

420 1 white crystals. M.p. 88–89.5 °C. Lit. m.p. 88–91 °C. H NMR (CDCl3) δ: 11.5 (s, br,

13 1H), 6.7 (s, 2H), 3.5 (t, 2H), 2.3 (t, 2H), 1.6 (m, 6H), 1.3 (m, 2H). C NMR (CDCl3) δ:

180.0, 170.2, 135.1, 38.9, 34.7, 28.0, 26.0, 24.0. IR (neat solid, HATR) ν, cm–1: 3429,

3313, 3051, 2946, 2869, 1708, 1624, 1570, 1512. ES-MS: 210.15 (M+). Analysis

(found/calc): C 56.67/56.86, H 6.10/6.20, N 7.50/6.63.

1,6-Diiodohexane

OH I HO I

288

A one liter three neck round bottom flask was charged with polyphosphoric acid

(H3PO4·½P2O5, 150 g), o-phosphoric acid (85% aq., 200 g), potassium iodide (342 g,

2.06 mol), and 1,6-hexanediol (118 g, 1 mol). The flask was fitted with a mechanical stirrer, reflux condenser, and a nitrogen/thermometer adapter and placed into a heating mantle. The reaction mixture was stirred at 120-130°C for five hours under nitrogen while dense oil separates at the bottom of the flask. The reaction mixture was cooled to room temperature, water (400 ml) and ether (300 ml) were added and the contents transferred to extraction funnel. The separated ether layer was washed with water (200 ml), dried with MgSO4 and ether evaporated on rotavap. The dark oily residue (297 g,

88%) was mixed with copper powder (2 g) and vacuum distilled (100-110°C at 0.5-0.3

20 mm) to give 271 g (80%) of clear product. nD 1.583. Notes: (1) Mixture of o-phosphoric and polyphosphoric acids must be allowed to cool down to room temperature before KI addition to avoid excessive loss of HI and I2 evolution, if exposed to air. (2) Vaccum distillation w/o copper powder produces a dark brown distillate.

1-(6-Iodohexyl)maleimide 102

N I O

A 250 ml round bottom flask with a stir bar was charged with 1-(6- hydroxyhexyl)maleimide 103 (1.9 g, 0.01 mol), chloroform (20 ml), and carbon disulfide

(20 ml). To that stirred mixture, a solution of diphosphorus tetraiodide (1.42 g, 2.5 mmol)

289

in carbon disulfide (100 ml) was added at once. After 30 min of stirring, solid potassium carbonate (5.0 g), followed by sat. aq. solution of potassium carbonate (30 ml) were added. The layers were separated and the aqueous layer was extracted with ether (3×50 ml). The combined organic layers were dried with MgSO4, solvents evaporated, and the oily residue was filtered through a short silica pad, washing it with hexane – ethyl acetate

(1:2). The filtrate, after removal of the solvents, gave 2.1 g (68 %) of 1-(6- iodohexyl)maleimide as colorless oil. The product is unstable and should be used

1 immediately or may be stored under argon in freezer for a few days. H NMR (CDCl3) δ:

13 6.6 (s, 2H), 3.4 (t, 2H), 3.2 (t, 2H), 1.5 (m, 8H). C NMR (CDCl3) δ: 175.6, 136.4, 47.6,

33.3, 30.0, 27.5, 25.5, 7.0. The same compound was also prepared by an alternative procedure from maleimide silver salt MI-Ag and 1,6-diiodohexane.

1-(6-Hydroxyhexyl)maleimide 103.

N OH O

1-(6-hydroxyhexyl)maleimide was prepared similar to 1-(6-iodohexyl)maleimide, starting from maleimide silver salt MI-Ag and 6-bromo-1-hexanol. 1H NMR

(CDCl3) δ: 6.7 (s, 2H), 3.5 (t, 2H), 3.4 (t, 2H), 1.5 (pentet, 2H), 1.4 (pentet, 2H), 1.3 (m,

13 4H). C NMR (CDCl3) δ: 175.4, 135.6, 61.8, 46.5, 38.6, 33.3, 28.0, 27.4, 26.5.

290

5-(diethylamino)-2-nitrosophenol 104.

NO 104 Et NOH 2 Et2NOH

A one-liter beaker, equipped with a mechanical stirrer, was charged with 3-

(N,N-diethylamino)phenol (50.0 g, 0.3 mol), ice (100 g), and hydrochloric acid (aq.,

12M, 130 ml). The beaker was immersed into an ice bath and the mixture was stirred until a homogeneous solution was formed. To that stirred solution, a solution of sodium nitrite (22.0 g, 0.32 mol) in water (200 ml), precooled to 5 °C, was added with vigorous stirring at such a rate that temperature of the reaction mixture did not rise above 5 °C.

After the addition was complete, the reaction mixture was stirred for additional 6 hrs, and filtered off on a Büchner funnel. The solid on filter was washed with cold water (2×100 ml), ethanol (2×100 ml, precooled to –20 °C) and vacuum dried at room temperature to give 68.2 g (97%) of 5-(diethylamino)-2-nitrosophenol as hydrochloride salt. The product decomposes above 50 °C and its recrystallization is thus not recommended. The product had the same Rf = 0.67 (EtOAc) as the authentic commercial sample from TCI America.

9-Diethylamino-2-hydroxy-5H-benzo[a]phenoxazin-5-one (Nile Red Phenol) 105.

OH OH

NO N + Et NOH 2 OH Et2NOO 104 105

291

A 500 ml round bottom flask with a stir bar was charged with 5-(diethylamino)-

2-nitrosophenol hydrochloride (9.2 g, 0.04 mol), 1,6-dihydroxynaphthalene (6.0 g, 0.375 mol), and DMF (200 ml). The flask was topped with a reflux condensor and the reaction mixture was stirred at reflux for 6 hrs. The reaction progress was monitored by TLC

(hexane : ethyl acetate = 1:2, target’s Rf = 0.48) and at least eight colored products can be distinguished. After the reaction was complete, DMF was removed on a rotovap with an oil pump, the residue was dissolved in methanol and impregnated onto silica gel.

Repeated chromatographies, performed on a dry-packed columns (Note 1) with hexane : ethyl acetate = 1:1 (target’s Rf = 0.25) gave crude Nile Red Phenol, which was further purified by several recrystallizations from 1-propanol to give 5.4 g (43%) of pure material. IR (neat solid, HATR) ν, cm–1: 3375, 2964, 1645, 1590, 1561. UV-Vis (MeOH)

λmax (lg ε): 210 (4.37), 265 (4.44), 326 (4.76), 547 (4.60). Fluorescence λmax = 632 nm.

1 H NMR (DMSO-d6) δ: 10.4 (s, br, 1H), 7.96 (d, 1H, J = 8.7 Hz), 7.87 (d, 1H, J = 2.4

Hz), 7.56 (d, 1H, J = 9.0 Hz), 7.08 (dd, 1H, J1 = 8.6 Hz, J2 = 2.5 Hz), 6.78 (dd, 1H, J1 =

9.0 Hz, J2 = 2.5 Hz), 6.61 (d, 1H, J = 2.5 Hz), 6.14 (s, 1H), 3.48 (q, 4H, J = 7.0 Hz), 1.15

13 (t, 6H, J = 7 Hz). C NMR (DMSO-d6) δ: 181.6, 160.6, 151.5, 150.6, 146.3, 138.6,

133.7, 130.8, 127.5, 123.8, 118.3, 109.9, 108.2, 107.9, 104.1, 96.0, 44.4, 12.4.

292

9-Diethylamino-2-(6-hydroxyhexyloxy)-5H-benzo[a]phenoxazin-5-one 106.

HO OH O

N N

Et2NOO Et2NOO 105 106

A 125 ml round bottom flask with a stir bar was charged with Nile Red Phenol

105 (0.5 g, 1.5 mmol), 6-bromo-1-hexanol (0.8 g, 4.4 mmol), potassium carbonate (0.45 g, 3.3 mmol), potassium iodide (50 mg), and DMF (50 ml). The reaction mixture was heated at 80 °C for 14 hrs and monitored by TLC (hexane : ethyl acetate = 1:2). After the reaction was complete, DMF was evaporated on a rotovap with an oil pump, the residue was dissolved in ethyl acetate (50 ml), filtered from insolubles, and impregnated onto silica gel. Chromatography in hexane : ethyl acetate with gradual elution from = 1:1 to

1:2, followed by recrystallization from iso-octane/1-propanol, gave 0.5 g (71%) of 9- diethylamino-2-(6-hydroxyhexyloxy)-5H-benzo[a]phenoxazin-5-one as dark red crystals.

1 Analysis (found/calc): C 68.21/71.87, H 6.93/6.96, N 6.17/6.45. H NMR (CDCl3) δ:

8.21 (d, 1H, J = 8.7 Hz), 8.04 (d, 1H, J = 2.4 Hz), 7.60 (d, 1H, J = 9.0 Hz), 7.16 (dd, 1H,

J1 = 8.6 Hz, J2 = 2.5 Hz), 6.65 (dd, 1H, J1 = 9.0 Hz, J2 = 2.5 Hz), 6.46 (d, 1H, J = 2.5 Hz),

6.30 (s, 1H), 4.17 (t, 2H, J = 6.3 Hz), 4.1 (t, 2H, J = 6.6 Hz), 3.47 (q, 4H, J = 7.2 Hz),

13 1.26 (t, 6H, J = 7.2 Hz). C NMR (DMSO-d6) δ: 183.4, 161.9, 152.2, 150.9, 147.0,

140.1, 134.2, 131.2, 127.9, 125.8, 124.9, 118.4, 109.7, 106.7, 105.4, 96.4, 68.4, 64.7,

45.3, 29.3, 28.8, 26.0, 21.2, 12.8.

293

N-(Hydroxymethyl)maleimide 107.

O O

NH + CH2O N OH O O

To a suspension of maleimide (9.8 g, 0.1 mol) in formaldehyde (37% aq., 8.1 ml) at 30°C was added NaOH (5% aq., 0.3 ml). Within 10 min all maleimide had dissolved and a mildly exothermic reaction ensued, rising the temperature to 35 °C.

Separation of the product began promptly. After 2.5 hrs at room temperature the reaction mixture was filtered to yield 9.6 g (75%) of product m.p. 100–102°C. Once recrystallized

421 1 from EtOAc gave m.p. 103-104°C. Lit. m.p. 104–106°C. H NMR (DMSO-d6) δ: 7.0

(s, 2H), 6.23 (t, 1H, J = 7.1 Hz), 4.79 (d, 2H, J = 7.1 Hz).

N-(2-hydroxyethyl)maleimide.

O O

H N N NH + 2 OH OH O O

A half a liter round bottom flask with a stir bar was charged with maleic anhydride (20 g, 0.20 mol), benzene (240 ml), and 2-aminoethanol (13 g, 0.21 mol). The flask was fitted with a Dean-Stark trap and the reaction mixture was refluxed for 2 hrs while separating water in the trap. After the reaction has been finished (no cloudy condensate in the trap), benzene was evaporated on rotavap and the yellow oil (28 g) was recrystallized from methanol (200 ml) to give 3.2 g (11%) of clear oil. 1H NMR (DMSO-

294

13 d6) δ: 7.01 (s, 2H), 6.23 (t, 1H), 4.79 (d, 2H), 3.47 (m, 2H). C NMR (DMSO-d6) δ:

171.01, 134.40, 57.87, 39.88. The yield is comparable to that in the literature.422

Diphosphorus tetraiodide P2I4. P4(α) + 4 I2 = 2 P2I4.

Commercial carbon tetrachloride was purified (Note 1) from traces of sulfur by shaking it with metallic mercury, drying with CaH2 (no alkali metals can be used!), and then distilling on a rotovap. The water-cooled condensor was replaced with a finger condensor, filled with dry ice-acetone mixture and distillation was done under argon.

White phosphorus (20.0 g, 0.161 mol of P4) was washed with acetone, ether, dried under argon, weighed into an argon-flushed flask with purified carbon disulfide

(100 ml), and filtered through an Acrodisk micron filter under argon pressure (do not use syringe!); some additional CS2 was used to wash the filter. Iodine (164.0 g, 0.645 mol) was dissolved in purified carbon disulfide (800 ml) with help of sonication. Solubility of

I2 in CS2 at 25 °C is 197 g I2 per 1000 g CS2, but it safer to use excess of CS2 to avoid crystallization.

A two-liter, three-neck ChemGlass reaction vessel, equipped with a mechanical stirrer, an efficient reflux condensor, and a dropping funnel, was flushed with argon and charged with white phosphorus (20.0 g, 0.161 mol of P4) solution in carbon disulfide

(100 ml). Iodine (164.0 g, 0.645 mol) solution in purified carbon disulfide (800 ml) was added from the addition funnel during 30 min period of time, with stirring. An exothermic reaction starts almost immediately. After the addition was complete, the reaction mixture was stirred for 24 hrs, and cannula transferred under argon into a two-

295

liter pear-shaped flask. The solvent was evaporated under argon on a rotovap (Note 2) until signs of P2I4 crystallization started to appear. The solution was then removed from the rotovap and left for crystallyzation – first, at room temperature and then in a fridge.

The precipitate was filtered off on a Büchner funnel and recrystallized once more from clean CS2 to give 79.8 g of crop 1. The combined mother liquors were evaporated, left to crystallize, and the precipitate was recrystallized from fresh CS2 (Note 3) to give 62.9 g of crop 2. The combined yield was 142.7 g (78%) of P2I4 as red-orange crystals. M.p.

126.8 °C. Lit.423 m.p. 124.5 °C. The material should be stored in a fridge under argon

(sensitive to moisture, but not air).

Note 1. If unpurified CS2 has been used, the m.p. of resulting P2I4 was 110 °C.

Note 2. The recovered CS2 was mixed with Ph3P to remove iodine, dried with CaH2, and redistilled. Note 3. Heat the solution very gently, for P2I4 decomposes very easily to PI3 and β-P4. For the same reason, m.p. depends on the heating rate: the higher the rate, the higher recorded m.p. value.

N-(Iodomethyl)maleimide 108.

O O OH I N N

O 107 O 108

N-(Iodomethyl)maleimide was prepared similar to 1-(6-iodohexyl)maleimide

1 102 from 107 and P2I4 in 37% yield as yellowish liquid. H NMR (CDCl3) δ: 6.8 (s, 1H),

296

13 5.3 (s, 1H). C NMR (CDCl3) δ: 175.5, 136.5, 10.5. The compound may be stored under argon in freezer for a week. Readily decomposes on open air and at room temperature.

Maleimid-2-yl-acetic acid (6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)acetic acid) 109.

O O O O OH O O H N N CH2—COOH + 2 OH NH OH O O O 109

Maleimid-2-yl-acetic acid was prepared similar to 6-(maleimid-2-yl)caproic acid

101, starting from glycine and maleic anhydride, in 44% overall yield. M.p. 110 °C (from

424 1 13 water, lit. m.p. 111–113.5 °C). H NMR (D2O) δ: 6.7 (s, 2H), 4.2 (s, 2H). C NMR

(D2O) δ: 172.5, 171.7, 130.5, 39.4,

Silver salt of maleimide MI–Ag.

O O

NH N Ag

O O MI MI-Ag

A one liter Erlenmeyer flask with a stir bar was charged with a solution of maleimide (7.1 g, 0.073 mol) in anhydrous ethanol (300 ml) and a solution of silver nitrate (12.4 g, 0.073 mol) in DMSO (50 ml). To that mixture, a solution of sodium hydroxide (aq., 0.4 M, 182.5 ml, 2.92 g NaOH) was added dropwise during 2 hrs at vigorous stirring. The precipitate formed was filtered off on a Büchner funnel, washed

297

with ethanol (100 ml), water (250 ml), methanol (200 ml), acetone (200 ml). The product was air-dried, followed by vacuum drying in a desiccator over P2O5 at 20 mm Hg to give

14.0 g (94%) of MI-Ag.

6-(Maleimid-2-yl)caproyl chloride (6-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)hexanoyl chloride) 111.

O O O O N N OH Cl O O

A 125 ml round bottom flask with a stir bar was charged with thionyl chloride

(45 ml) and the flask was cooled in a dry ice – acetone bath to –10 °C. To the cooled thionyl chloride, 6-(maleimid-2-yl)caproic acid 101 (2.0 g, 9.5 mmol) was added at stirring, followed by DMF (1 drop). The reaction mixture was allowed to warm up and then was heated for 3 hrs at 80 °C. Thionyl chloride was evaporated on a rotovap and its traces were removed by azeotropic distillation with benzene (2×80 ml). The residue was dissolved in dichloromethane (25 ml) and filtered through a thin pad of silica gel on a

Büchner funnel to give, after solvent removal, 2.0 g (92%) of colorless liquid, which was directly utilized in acylation.

N-(4-hydroxyphenyl)maleimide (1-(4-hydroxyphenyl)-1H-pyrrole-2,5-dione) 112 was prepared similar to 6-(maleimid-2-yl)caproic acid 101, starting from 4-aminophenol and maleic anhydride and substituting acetic anhydride for acetyl chloride in the

298

dehydration step, in overall yield of 68%. M.p. 185 °C (lit. m.p. 154425…195426). 1H

13 NMR (DMSO-d6) δ: 9.9 (br, 1H), 7.10 (d, 2H), 7.05 (d, 2H), 6.8 (s, 2H). C NMR

(DMSO-d6) δ: 171.2, 157.9, 135.4, 129.3, 123.3, 116.3.

N-(6-iodohexyl)maleimide (1-(6-iodohexyl)-1H-pyrrole-2,5-dione) 113.

N-(6-iodohexyl)maleimide was prepared similar to 1-(6-iodohexyl)maleimide

102, starting from maleimide silver salt MI-Ag and 1,6-diiodohexane, in 96% yield. 1H

13 NMR (CDCl3) δ: 6.6 (s, 2H), 3.4 (t, 2H), 3.2 (t, 2H), 1.5 (m, 8H). C NMR (CDCl3) δ:

175.6, 136.4, 47.6, 33.3, 30.0, 27.5, 25.5, 7.0.

9-Diethylamino-2-methoxy-5H-benzo[a]phenoxazin-5-one 115.

OH OMe

N N

Et NOO Et2NOO 2 115 105

A 250 ml round bottom flask with a stir bar was charged with Nile Red Phenol

(0.5 g, 1.5 mmol), iodomethane (0.25 g, 1.76 mmol), potassium carbonate (0.96 g, 7 mmol), and DMF (70 ml). The reaction mixture was stirred at 80 °C for 3 hrs and monitored by TLC (ethyl acetate – hexane 2:1, product Rf = 0.57). After the reaction was complete, the reaction mixture was cooled down and applied directly to a silica gel column. Chromatography with ethyl acetate – hexane (1:1) gave, after evaporation of solvents and recrystallization from 1-propanol, 0.43 g (77%) of dark red crystals. IR (neat

299

–1 1 solid, HATR) ν, cm : 2964, 1645, 1590, 1561. H NMR (CDCl3) δ: 8.2 (d, 1H, J = 9.0

Hz), 8.0 (d, 1H, J = 2.7 Hz), 7.56 (d, 1H, J = 9.0 Hz), 7.15 (dd, 1H, J1 = 9.0 Hz, J2 = 2.7

Hz), 6.61 (dd, 1H, J1 = 9.0 Hz, J2 = 2.7 Hz), 6.4 (d, 1H, J = 2.7 Hz), 6.27 (s, 1H), 4.0 (s,

13 3H), 3.45 (q, 4H, J = 7.2 Hz), 1.2 (t, 6H, J = 7.2 Hz). C NMR (DMSO-d6) δ: 183.2,

162.3, 152.1, 150.8 146.9, 140.1, 134.1, 131.1, 127.8, 125.8, 124.7, 118.1, 109.5, 105.9,

105.3, 96.4, 55.7, 45.0, 12.7.

9-Diethylamino-2-(2-hydroxyethyloxy)-5H-benzo[a]phenoxazin-5-one 116.

OH OH O

N N

Et NOO Et2NOO 2 116 105

9-Diethylamino-2-(2-hydroxyethyloxy)-5H-benzo[a]phenoxazin-5-one was prepared similar to NR-OMe 115, starting from Nile Red Phenol (2.0 g, 6 mmol) and substituting iodomethane for ethylene carbonate (1.6 g, 18 mmol), in 76% yield. 1H NMR

(CDCl3) δ: 8.2 (d, 1H), 8.0 (d, 1H), 7.56 (d, 1H), 7.15 (dd, 1H), 6.61 (dd, 1H), 6.4 (d,

13 1H), 6.27 (s, 1H), 4.2 (t, 2H), 4.0 (t, 2H), 3.45 (q, 4H), 1.2 (t, 6H). C NMR (DMSO-d6)

δ: 183.2, 162.3, 152.1, 150.8 146.9, 140.1, 134.1, 131.1, 127.8, 125.8, 124.7, 118.1,

109.5, 105.9, 105.3, 96.4, 55.7, 45.0, 12.7.

300

9-Diethylamino-2-(6-iodohexyloxy)-5H-benzo[a]phenoxazin-5-one 117.

I OH O

N N

Et2NOO Et2NOO 105 117

A 50 ml pear-shaped flask was charged with a stirring bar, Nile Red Phenol (9- diethylamino-2-hydroxy-5H-benzo[a]phenoxazin-5-one 105, 66 mg, 0.2 mmol), 1,6- diiodohexane (134 mg, 0.4 mmol), potassium hydroxide (55 mg, 0.4 mmol), and dimethylacetamide (10 ml). The reaction mixture was stirred at 90°C for one hour (most of the starting material was consumed during first several minutes, as monitored by TLC with 1:1 eluent of hexane and ethyl acetate). The solvent was evaporated and the residue was chromatographed on silica gel with 1:1 mixture of hexanes and ethyl acetate. Yield

74 mg (79%) of dark ruby red material, which is used directly in next step. 1H NMR

(CDCl3), δ: 8.16 (d, 1H, J=9Hz); 7.97 (d, 1H, J=2.4Hz); 7.56 (d, 1H, J=9Hz); 7.13 (dd,

1H, J1=9Hz, J2=2.4Hz); 6.68 (dd, 1H, J1=9Hz, J2=2.4Hz); 6.43 (d, 1H, J=2.4Hz); 6.37 (s,

1H); 4.15 (t, 2H, J=6.6Hz); 3.47 (q, 4H, J=7.2Hz); 3.19 (t, 2H, J=6.6Hz); 1.84 (m, 2H);

13 1.54 (m, 2H); 1.42 (m, 2H); 1.25 (t, 6H, J=7.2Hz). C NMR (CDCl3), δ: 182.2, 162.0,

152.1, 151.3, 147.2, 139.5, 139.4, 134.2, 131.5, 127.8, 125.8, 125.2, 118.5, 110.7, 106.7,

104.8, 96.7, 68.4, 45.6, 33.6, 29.9 (double intensity), 29.6, 12.8, 7.0.

301

9-Diethylamino-2-(caproyloxy)-5H-benzo[a]phenoxazin-5-one 118.

OH O

N N

Et2NOO Et2NOO 105 118

A 50 ml round bottom flask with a stir bar was charged with Nile Red Phenol

(33 mg, 0.1 mmol), anhydrous DMF (5 ml), and capryloyl chloride (23 mg, 0.15 mmol).

To that stirred mixture, potassium carbonate (powdered, 30 mg, 0.2 mmol) was added at once: no change on TLC (hexane – ethyl acetate 1:1) during one hour at room temperature and then one hour at 60 °C. To the reaction mixture, DBU (1,8-

Diazabicyclo[5.4.0]undec-7-ene, 33 mg, 0.2 mmol) was added at once and stirring continued at 60 °C for 30 min: a less polar spot on TLC wiith Rf = 0.61 appeared. The reaction mixture was cooled down, poured into water (100 ml) and extracted with ethyl acetate (10 ml). The extract was washed with HCl (2M, 5 ml), K2CO3 (aq., 10 ml), dried with MgSO4, and filtered through a thin silca gel pad on a Büchner funnel. Evaporation

1 of solvents allowed 27 mg (61%) of dark red material. H NMR (CDCl3) δ: 8.2 (d, 1H),

8.0 (d, 1H), 7.56 (d, 1H), 7.15 (dd, 1H), 6.61 (dd, 1H), 6.4 (d, 1H), 6.27 (s, 1H), 3.45 (q,

4H), 2.5 (t, 2H), 1.8 (pentet, 2H), 1.7 (pentet, 2H), 1.4 (pentet, 2H), 1.2 (t, 6H), 0.9 (t,

13 3H). C NMR (CDCl3) δ: 183.2, 170.8, 162.3, 152.1, 150.8 146.9, 140.1, 134.1, 131.1,

127.8, 125.8, 124.7, 118.1, 109.5, 105.9, 105.3, 96.4, 45.0, 34.5, 31.0, 24.5, 22.1, 14.7,

12.7.

302

Exo-cis-7-oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic anhydride Fu-MA 119.

O H O

O + O O O

O H O

Maleic anhydride (98 g, 1 mol) was placed in a 2 liter Erlenmeyer flask, dissolved in 900 ml of ether, furan (100 ml, 1.34 mol) added, the flask was sealed with rubber stopper and aluminum foil and left undisturbed for a week. Crystals formed were filtered and air-dried giving 152.5 g (91.8%) of the exo-cis Diels-Alder adduct.

Recrystallization of the direct product is not recommended since fine crystals give too violent reaction with ammonia. M.p. 116.5°C with decomposition. Analysis

(Found/Calc): C 57.59/57.84; H 3.71/3.64.

Exo-cis-7-oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboximide Fu-MI 120.

(10-oxa-4-azatricyclo[5.2.1.02,6]dec-8-ene-3,5-dione)

H O H O

O O O NH

H O H O

(a) A one liter pear-shaped round bottom flask was charged with exo-cis-7- oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic anhydride (150 g, 0.9 mol), 150 ml of ice- cold water and a stirring bar. Aqueous ammonia (450 ml of 29% solution, 3.7 mol) was added dropwise to the stirred suspension, then the flask was equipped with a reflux condenser, immersed in an oil bath with temperature 130-140°C and refluxed for 45 min

(at the end of this time period the temperature of the reaction mixture rises to

303

101…103°C). Longer reaction time and higher bath temperature cause decomposition of the product and reduce the yield. After cooling most of the solvent was rotoevaporated and 95% ethanol was added in such amount that all solid dissolved at boiling. The solution was cooled slowly, precipitated crystals filtered off, washed with 85% ethanol, then absolute ethanol and vacuum dried. Concentration of the mother liquors allowed additional crop of the crystals, combined yield is 140.6 g (94%). M.p. 157.5°C. Analysis

(Found/Calc): 57.50/58.18; H 4.33/4.27; N 8.43/8.48. Rf = 0.34 (hexane : ethyl acetate =

1:1, visualized with KMnO4).

Fu-MI [42074-03-3] 120.

O H O

O + NH O NH 120

O H O

Maleimide (7.4 g, 76 mmol) was placed in a 250 ml round bottom flask, dissolved in 150 ml of ether, furan (8.5 ml, 116 mmol) was added, the flask was covered with aluminum foil and left undisturbed for 24 hrs. Crystals formed were filtered and air- dried to give 11.8 g (94%) of the Diels-Alder adduct. M.p. 168.3°C (lit.427 m.p. 162 °C for exo isomer, 126–128°C for endo isomer, and 126–128°C for mixture thereof.) IR

(cm–1): 3028, 3012, 2970, 1738, 1726, 1365, 1353, 1287, 1229, 1216, 1205, 1142, 1090,

1018, 927, 896, 854, 821, 791, 733, 688, 633, 582. Analysis (Found/Calc): 58.22/58.18;

H 4.29/4.27; N 8.64/8.48. Rf = 0.34 (hexane : ethyl acetate = 1:1, visualized with

KMnO4).

304

N-(6-iodohexyl)-exo-cis-7-oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboximide 121.

(4-(6-iodohexyl)-10-oxa-4-azatricyclo[5.2.1.02,6]dec-8-ene-3,5-dione)

CAS RN [874998-65-9]

H O H O

O NH O N I H O H O 120 121

Exo-cis-7-oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboximide (10.0 g, 0.06 mol) was dissolved in 220 ml of acetone, then 1,6-diiodohexane (23.7 g, 0.07 mol) and potassium carbonate (16.6 g, 0.12 mol) were added and the mixture was refluxed for 12 hours upon which time the color gradually changes to deep red. Acetone was evaporated and 50 ml of chloroform added, the solution was filtered off of insoluble inorganic salts, applied to a dry-packed silica gel column (ID 40 mm, l=30 cm) and eluted with hexane : ethyl acetate mixture = 3:1 , then 1:1 composition. The former elutes out 1,6- diiodohexane, and the latter elutes the product. One may avoid chromatography and increase the yield if the solid obtained after evaporation of the chloroform filtrate is thoroughly washed with cold (–20°C) hexane and then recrystallized from hexane – chloroform mixture. Yield 6.2 g (27%). M.p. 71.5°C (DSC, 5°C/min). Analysis

(Found/Calc): C 44.84/44.82; H 4.83/4.84; N 3.78/3.73. Rf = 0.47 (hexane : ethyl acetate

= 1:1). 1H NMR, δ: 6.52 (s, 2H); 5.25 (s, 2H); 3.46 (t, 2H, 3J=6 Hz); 3.17 (t, 2H, 3J=6

Hz); 2.84 (s, 2H); 1.8 (pentet, 2H, 3J=6 Hz); 1.54 (pentet, 2H, 3J=6 Hz); 1.38, 1.27 (m,

4H, 3J=6 Hz). 13C NMR, δ: 176.4; 136.7; 81.1; 47.6; 38.9; 33.4; 30.1; 27.5; 25.7; 7.1.

305

4-[6-(9-Diethylamino-5-oxo-5H-benzo[a]phenoxazin-2-yloxy)-hexyl]-10-oxa-4-azatricyclo[5.2.1.02,6]dec-8-ene-3,5-dione [874998-66-0] 122.

H O

O N O H O N

Et2NOO 122

Diels-Alder protected Nile Red maleimide 122 was prepared by two methods.

(a) A 125 ml round-bottom flask with a stirring bar was charged with Nile Red phenol (9- diethylamino-2-hydroxy-5H-benzo[a]phenoxazin-5-one, 370 mg, 1.1 mmol), 2-(6- iodohexyl)-3a,4,7,7a-tetrahydro-4,7-epoxy-1H-isoindole-1,3(2H)-dione (500 mg, 1.33 mmol), potassium carbonate (400 mg, 2.9 mmol), and dimethylformamide (3 ml). The reaction mixture was stirred at 65°C for 12 hours and then chromatographed on silica gel with 1:1 mixture of hexane and ethyl acetate. After evaporation of the solvent, the residue was washed with distilled hexane (10 ml × 2), and dried in vacuum, Yield 418 mg (65%).

1 H NMR (CDCl3), δ: 8.18 (d, 1H, J=9Hz); 8.01 (d, 1H, J=2.4Hz); 7.57 (d, 1H, J=9Hz);

7.14 (dd, 1H, J1=9Hz, J2=2.4Hz); 6.63 (dd, 1H, J1=9Hz, J2=2.4Hz); 6.50 (d, 1H,

J=0.9Hz); 6.42 (s, 1H), 6.27 (d, 1H, J=0.9Hz); 5.26 (s, 2H), 4.14 (t, 2H, J=6.6Hz); 3.53-

3.41 (m, 6H, overlap of N1–H (q) and N2–H (t)); 2.88 (s, 2H); 1.85 (m, 2H); 1.75-1.52

13 (m, 4H); 1.25 (t, 6H, J=6.6Hz). C NMR (CDCl3), δ: 183.3, 176.3, 161.8, 152.1, 150.7,

146.6, 140.1, 136.5 (double intensity), 134.1, 131.1, 127.7, 125.6, 124.7, 118.3, 109.5,

106.7, 105.3, 96.3, 80.9, 68.2, 49.9, 47.4, 38.9, 25.6, 26.4, 27.5, 29.0, 12.7.

306

(b) Same as (a) except that 9-Diethylamino-2-(6-iodohexyloxy)-5H-benzo[a]phenoxazin-

5-one (74 mg, 0.156 mmol) and furan-maleimide adduct (33 mg, 0.2 mmol) were used as

reactants. Yield 55 mg (61%).

1-[6-(9-Diethylamino-5-oxo-5H-benzo[a]phenoxazin-2-yloxy)-hexyl]-pyrrole-2,5-dione.

[874998-67-1] 123 (Nile Red maleimide).

H O O

O N N O O H O O N N

Et2NOO Et2NOO 122 123

A one liter pear-shaped flask with a stirring bar was charged with Diels-Alder

protected Nile Red maleimide (4-[6-(9-Diethylamino-5-oxo-5H-benzo[a]phenoxazin-2-

yloxy)-hexyl]-10-oxa-4-aza-tricyclo[5.2.1.02,6]dec-8-ene-3,5-dione, 4.0 g, 6.87 mmol),

dichloromethane (100 ml), toluene (250 ml), and topped with a 40 cm Vigreaux column.

The flask was heated on a 140°C heating mantle with gentle reflux for 12 hours, allowing

of gradual evaporation of dichloromethane and evolved furan. The solvent was

evaporated, the residue dissolved in dichloromethane and chromatographed on silica gel

with a 1:1 mixture of hexane and ethyl acetate. Yield 2.26 g (64%) of dark red material,

which could be converted to a crystalline form by very slow evaporation of

dichloromethane solution thereof. Analysis (Found/Calc): C 69.26/70.16, H 6.20/6.08, N

1 8.10/8.18. H NMR (CDCl3), δ: 8.20 (d, 1H, J=9Hz); 8.01 (d, 1H, J=2.4Hz); 7.58 (d, 1H,

307

J=9Hz); 7.16 (dd, 1H, J1=9Hz, J2=2.4Hz); 6.69 (s, 2H); 6.65 (dd, 1H, J1=9Hz, J2=2.4Hz);

6.42 (d, 1H, J=0.9Hz); 6.28 (s, 1H); 4.15 (t, 2H, J=6.6Hz); 3.55 (t, 2H, J=9Hz); 3.45 (q,

4H, J=9Hz); 1.85 (m, 2H); 1.68-1.56 (m, 4H); 1.44 (m, 2H); 1.25 (t, 6H, J=6.6Hz). 13C

NMR (CDCl3), δ: 183.3, 170.9, 161.8, 152.1, 150.7, 146.9, 140.2, 134.1 (double intensity), 131.1, 127.7, 125.6, 124.7, 118.3, 109.5, 106.7, 105.3, 96.4, 68.2, 45.1, 37.8,

29.1, 28.5, 26.5, 25.7, 12.7.

[4-[4-Diethylamino-2-(6-{3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2- yl}hexyloxy)phenylvinyl]-3-cyano-5,5-dimethylfuran-2(5H)-ylidene]malonodinitrile.

NC NC NC NC CN CN O O O N N O NO OH + O NO O

O I

A 100 ml round bottom flask with a stir bar was charged with 4-(4- diethylamino-2-hydroxy-vinyl)-DCDHF (200 mg, 0.535 mmol), 2-(6-iodohexyl)-

3a,4,7,7a-tetrahydro-4,7-epoxy-1H-isoindole-1,3(2H)-dione (Fu-MI-C6-I, 220 mg, 0.586 mmol), potassium carbonate (720 mg, 5.2 mmol), and 3-pentanone (50 ml). The mixture was stirred at room temperature for 12 hrs (no change on TLC) and then at ~80°C for 12 hrs. After the reaction was complete by TLC (eluent: EtOAc neat) the solvent was evaporated on rotavap, dissolved in dichloromethane (50 ml), applied onto silica gel (70 g) and chromatographed with hexane : ethyl acetate = 1:1 to allow, after solvent

308

1 evaporation, 43 mg (13%) of dark blue solid. Rf=0.42 (neat EtOAc). H NMR (CDCl3) δ:

7.90 (d, 1H, J=16.2 Hz), 7.46 (d, 1H, J=9 Hz), 6.85 (d, 1H, J = 16 Hz), 6.50 (s, 2H), 6.34

(d, 1H, J = 9 Hz), 6.00 (s, 1H), 5.25 (s, 2H), 4.0 (t, 2H), 3.60 (m, 4H), 3.46 (m, 2H), 3.17

13 (m, 2H), 2.82 (s, 6H), 1.71 (s, 6H), 1.56-1.32 (m, 8H), 1.06 (t, 6H). C NMR (CDCl3) δ:

180.8, 179.2, 176.7, 176.6, 136.7, 106.2, 93.8, 68.3, 62.76, 62.73, 47.5, 46.0, 45.3, 38.9,

32.6, 27.6, 26.3, 25.3, 12.94.

4-Diethylamino-2-(6-[3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2-yl]- hexyl)benzaldehyde.

O ONO N N O OH O

A 200 ml round bottom flask with a stir bar was charged with 4- diethylaminosalicylaldehyde (10.3 g, 53.3 mmol), 1-iodo-6-[3a,4,7,7a-tetrahydro-4,7- epoxy-1,3-dioxo-1H-isoindol-2-yl]hexane (Fu-MI-C6-I, 20 g, 53.3 mmol), potassium carbonate (10 g), and acetone (120 ml). The reaction mixture was stirred at reflux for 3 days, filtered, the solid on filter washed with acetone (50 ml), and the combined acetone filtrates were impregnated onto silica gel. Chromatography with hexane:ethyl acetate from 3:1 to neat ethyl acetate allowed, after solvent evaporation, 18 g (77%) of greenish

1 yellow oil. ES-MS: 441.1 (M+1) and 373.2 (M+1–Fu). H NMR (CDCl3) δ: 10.11 (s,

1H), 7.65 (d, 1H), 6.50 (s, 2H), 6.23 (dd, 1H), 5.97 (d, 1H), 5.22 (s, 2H), 3.97 (t, 2H),

3.45 (t, 2H), 3.38 (q, 4H), 2.80 (s, 2H), 1.78 (pentet, 2H), 1.56 (m, 2H), 1.47 (m, 2H),

309

1.33 (m, 2H), 1.18 (t, 6H). 13C NMR δ: 187.0, 176.4, 163.9, 153.8, 136.5, 130.0, 114.2,

104.2, 93.1, 80.9, 67.8, 47.3, 44.7, 38.7, 28.9, 27.4, 26.2, 25.6, 12.6.

[4-[4-Diethylamino-2-(6-{3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2- yl}hexyloxy)phenylvinyl]-3-cyano-5,5-dimethylfuran-2(5H)-ylidene]malonodinitrile.

O NC NC ONO CN N O O O N O NC NC NO + O CN O O

A 200 ml round bottom flask with a stir bar was charged with 4-diethylamino-2-

(6-[3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2-yl]hexyl)benzaldehyde (AS-

3-04, 3.7 g, 8.4 mmol), 2-dicyanomethylene-3-cyano-4,5,5-trimethylfuran (1.6 g, 8.0 mmol), pyridine (21 g), and acetic acid (50 mg, 6 drops). The reaction mixture was stirred at 60°C for 12 hrs, cooled and the solvent was removed by rotary evaporation. The residue chromatographed on silica gel with gradient of hexane : ethyl acetate = 1:1 through neat ethyl acetate. Fractions containing pure product were combined and

1 concentrated to give 4.7 g (90%) of dark blue crystals. M.p. 110°C. H NMR (CDCl3) δ:

7.96 (d, 1H, J=16 Hz), 7.50 (d, 1H, J=9 Hz), 6.87 (d, 1H, J=16 Hz), 6.53 (s, 2H), 6.37

(dd, 1H, J1=9 Hz, J2=2 Hz), 6.05 (d, 1H, J=2 Hz), 5.26 (s, 2H), 4.04 (t, 2H, J=6 Hz), 3.50

(m, 6H), 1.89 (m, 2H), 1.74 (s, 6H), 1.66-1.36 (m, 6H), 1.27 (t, 6H, J=7 Hz). 13C NMR

(CDCl3) δ: 176.7, 176.4, 175.1, 162.3, 153.9, 144.1, 136.5, 113.3, 112.5, 112.2, 111.9,

310

108.2, 106.2, 96.3, 93.7, 80.97, 68.2, 47.4, 45.2, 38.7, 28.9, 27.50, 27.0, 26.2, 25.8, 12.8.

–1 ES-MS: 554.2 (M+H–C4H4O). IR (HATR, neat, ν, cm ): 2984, 2940, 2871, 2220 (m),

1710 (s), 1615, 1583, 1550, 1513 (s, br), 1251 (s), 1186 (s, br), 1073, 1011, 960, 873,

–1 811, 694, 651, 596. IR (HATR, CHCl3 solution, ν, cm ): 1710, 1517, 1419, 1361, 1259,

1088, 909, 643, 527.

3-cyano-2-dicyanomethylene-4-(2-[2-{6-hexyloxy}-4-{N,N-diethylamino}phenyl]vinyl)-5,5-dimethyl-2,5-dihydrofuran 124.

NC NC NC NC CN CN O O O O N N NO N O O O O

A 250 ml round bottom flask with a stir bar was charged with [4-[4-

Diethylamino-2-(6-{3a,4,7,7a-tetrahydro-4,7-epoxy-1,3-dioxo-1H-isoindol-2-yl}hexyloxy)phenylvinyl]-3-cyano-5,5-dimethylfuran-2(5H)-ylidene]malonodinitrile (4.7 g, 7.56 mmol) and xylene (mixture of isomers, 70 ml). The reaction mixture was refluxed for 2 hrs, at which time the starting material was absent by TLC analysis. The reaction was cooled and the solvent was removed by rotary evaporation. The residue was chromatographed on silica gel with a gradient of hexane : ethyl acetate = 1:1 through neat ethyl acetate to give 2.0 g (47%) of dark blue crystals, which was recrystallized from 1-

1 propanol to give 1.76 g (41 %) of pure product. H NMR (CDCl3) δ: 8.00 (d, 1H, J=16

Hz), 7.50 (d, 1H, J=9 Hz), 6.84 (d, 1H, J=16 Hz), 6.7 (s, 2H), 6.4 (dd, 1H, J1=9 Hz, J2=2

311

Hz), 6.05 (d, 1H, J=2 Hz), 4.05 (t, 2H, J=6 Hz), 3.50 (m, 6H), 1.89 (m, 2H), 1.74 (s, 6H),

13 1.66-1.36 (m, 6H), 1.27 (t, 6H, J=7 Hz). C NMR (CDCl3) δ: 176.8, 175.0, 170.9, 162.2,

154.0, 144.1, 134.1, 113.4, 112.6, 112.2, 112.0, 108.0, 106.3, 96.3, 93.7, 68.2, 45.2, 37.6,

28.9, 28.5, 27.0, 26.4, 25.8, 12.8. ES-MS: 554.2 (M+H). UV-Vis (CHCl3) λmax: 590 nm.

M.p. 174.8°C. IR (HATR, neat solid, ν, cm–1): 2972, 2934, 2870, 2220 (s), 1705 (s),

1617, 1581, 1553, 1511 (s, br), 1377, 1348, 1313, 1235 (s, br), 1182 (s, br), 1119, 1077,

1006, 964, 872, 822 (s), 709, 692, 653.

4,5-Dimethoxyphthalaldehyde.

O O CHO Cl Cl O O CHO

A 200 ml round bottom flask with a stir bar was charged with 4,5-dimethoxy-

1,2-bis(chloromethyl)benzene (2.35 g, 0.01 mol), tetra-n-butylammonium periodate (9.16 g, 0.02 mol), dioxane (30 ml), fitted with a reflux condenser and refluxed for 29 hrs. The mixture was cooled down, poured into water (200 ml), filtered, air-dried (7.43 g), and flash-chromatographed on silica (50 g) with gradual elution from hexane-ethyl acetate

(4:1) to ethyl acetate. Solvent evaporation and recrystallization of the second chromatographic fraction from ethanol allowed 0.8 g (41%) of 4,5- dimethoxyphthalaldehyde, identical to an authentic sample by TLC (ethyl acetate) and

NMR.

312

2,3,6,7-tetramethoxy-9,10-dihydroanthracene 129.

O H O O + H O O O O

A one liter Erlenmeyer flask equipped with a mechanical stirrer was charged with sulfuric acid (70%, 850 g) and veratrole (50 g, 0.36 mol). To that vigorously stirred mixture formaldehyde (37% aq., 70 g) was added dropwise during 30 min and stirring was continued for 2 additional hrs. The reaction mixture was poured onto crashed ice

(500 g), filtered, washed with water (500 ml), air-dried, and recrystallized from a mixture of 1-propanol (450 g) and chloroform (350 g). The crystals were washed with cold 1- propanol (3×50 ml) and air-dried, 42.2 g (78%). M.p. 232°C (lit.428 mp. 230°C). 1H NMR

(CDCl3) δ: 6.83 (s, 2H), 4.76 (d, 1H, J=13.5 Hz), 3.84 (s, 6H), 3.54 (d, 1H, J=13.5 Hz).

13 C NMR (CDCl3) δ: 147.8, 131.9, 113.2, 56.2, 36.6.

1,1,1-Trichloro-2,2-bis(3,4-dimethoxyphenyl)ethane.

CCl3 MeO CCl3 MeO OMe + HOHO MeO MeO OMe

A 200 ml round bottom flask with a stirring bar was charged with chloral hydrate (50.0 g, 0.3 mol), acetic acid (100 ml), sulfuric acid (50 ml), and heated to ~40

°C. To that stirred mixture, veratrole (37.6 g, 0.2 mol) was added from a syringe pump at

3.4 ml/hr. After the addition was complete (which takes ca. 11 hrs), the reaction mixture was stirred at 40 °C for additional 24 hrs, and poured onto crushed ice (150 g). The

313

precipitate formed was filtered off on a Büchner funnel, air-dried (51.0 g, 83%) and recrystallized from 1-propanol to give 31.0 g (51%) of white crystals. M.p. 117°C (lit.429 m.p. 115–116 °C). 1H NMR δ: 7.2 (dd, 2H), 7.1 (d, 2H), 6.9 (d, 2H), 4.9 (s, 1H), 3.9

(2×s, 12H). 13C NMR δ: 149.8, 149.5, 131.6, 123.2, 115.4, 112.4, 102.7, 70.7, 56.6, 56.3.

2,3,6,7-tetramethoxy-9,10-anthraquinone 130.

CCl 3 CHCl2 O MeO OMe MeO OMe

MeO OMe MeO OMe CCl3 O

A 200 ml round bottom flask with a stirring bar was charged with 1,1,1-

Trichloro-2,2-bis(3,4-dimethoxyphenyl)ethane (20.0 g, 5 mmol), chloral hydrate (18.0 g,

0.11 mol), and acetic acid (62 ml). To that stirred mixture, sulfuric acid (120 ml) was added from an addition funnel during 4 hrs. After the addition was complete, the reaction mixture was stirred for additional 3 hrs, and poured onto crushed ice (400 g). The precipitate formed was filtered off on a Büchner funnel, air-dried (41 g) and dissolved in potassium hydroxide solution (aq., 10%, 400 ml). To that stirred mixture, a solution of potassium permanganate (28 g, 0.177 mmol) in water (120 ml) was added dropwise from an addition funnel. After the addition was complete, the reaction mixture was heated to

80 °C and stirred at that temperature for 30 min. The solution was cooled and filtered on a Büchner funnel. The solid on filter was recrystallized from DMAc to give 1.1 g (67%) of 2,3,6,7-tetramethoxy-9,10-anthraquinone as yellow crystals. M.p. 348 °C (lit.430 345–

314

1 13 346 °C). H NMR (CF3COOD) δ: 7.8 (s, 4H), 4.1 (s, 12H). C NMR (CF3COOD) δ:

187.0, 156.1, 130.8, 111.6 (108.5*), 57.8 (56.6*). *Value in CDCl3.

2-Bromo-4,5-dimethoxybenzaldehyde 137.

O CHO O CHO 137 O O Br

A one-liter two-neck round bottom flask equipped with a mechanical stirrer and an addition funnel, was charged with 4,5-dimethoxybenzaldehyde (83.2 g, 0.5 mol) and chloroform (500 ml). To the stirred reaction mixture bromine (160.0 g, 1 mol) was added dropwise from the addition funnel during ca. 2 hrs. After ~¼ of all bromine had been added, the reaction mixture became warm and a precipitate began to deposit and an increase in power input to the mechanical stirrer was necessary. After all bromine had been added, the reaction mixture was stirred for additional two hours, cooled in frige to –

20 °C and filtered off on a Büchner funnel. The solid on filter was suspended in water (1 liter) and potassium carbonate (sat. aq., ~70 ml) was carefully added until basic reaction to pH paper, followed by sodium sulfite solution (sat. aq., ~120 ml) until bromine color disappeared. The decolorized suspension was filtered off on a Büchner funnel, washed with water (3×200 ml), air-dried, and recrystallized from acetic acid to give 192.0 g

(52%) of 2-bromo-4,5-dimethoxybenzaldehyde as white crystals. M.p. 150 °C (lit.431 m.p.

1 148–150 °C). H NMR (CDCl3) δ: 10.15 (s, 1H), 7.4 (s, 1H), 7.0 (s, 1H), 3.93 (s, 3H),

13 3.90 (s, 3H). C NMR (CDCl3) δ: 190.8, 154.5, 148.9, 126.5, 120.4, 115.4, 110.4, 56.5,

56.2.

315

2-Bromo-4,5-dimethoxybenzaldehyde dimethyl acetal 138.

O O CHO O O

O Br O Br 137 138

A 250 ml round bottom flask with a stir bar was charged with 2-Bromo-4,5- dimethoxybenzaldehyde (24.5 g, 0.1 mol), trimethoxymetane (31.2 g, 0.3 mol), anhydrous methanol (150 ml), and Dowex 50W-X8 ion exchange resin (10.0 g). The flask was topped with a Vigreaux column (Note 1) and the reaction mixture was refluxed for 20 hrs and monitored by TLC (CH2Cl2 neat). After the reaction was complete, the reaction mixture was filtered off on a Büchner funnel, the filtrate was evaporated from solvents and the oily residue was vacuum distilled, collecting fraction b.p. 105–120 °C at

1 0.05 mm Hg: 16.51 g (58%) of colorless oil. H NMR (CDCl3) δ: 7.1 (s, 1H), 7.0 (s, 1H),

13 5.5 (s, 1H), 3.91 (s, 3H), 3.86 (s, 3H), 3.4 (s, 6H). C NMR (CDCl3) δ: 149.8, 148.5,

129.3, 115.4, 113.2, 110.7, 103.3, 56.27, 56.17, 54.1.

2,3,6,7-tetrakis(pentyloxy)-9,10-dihydroanthracene 143.

O O O H11C5 H11C5 C5H11 H C H C C H 11 5O 11 5O O 5 11

A 250 ml Erlenmeyer flask with a stirring bar was charged with sulfuric acid

(70%, 50 ml) and 1,2-dipentyloxybenzene (5 g, 20 mmol). To that vigorously stirred mixture formaldehyde (37% aq., 4 g) was added dropwise during 10 min and stirring was

316

continued for 2 additional hrs, while a viscous oil separated out. The oily product was extracted with dichloromethane (2×50 ml), applied onto silica gel and chromatographed with hexane – ethyl acetate (gradual elution from 8:1 to 4:1). Evaporation of solvent afforded 2.4 g (45%) of colorless opalescent oil, which solidified in two days to slightly yellow crystals. M.p. 51°C.

2,3,6,7-tetrahydroxy-9,10-dihydroanthracene 146.

O O HO OH

OOHOHO

A 500 ml round bottom flask with a stir bar was charged with 2,3,6,7- tetramethoxy-9,10-dihydroxyanthracene (1.0 g, 3.33 mmol), chloroform (19 g) and cooled in an ice bath for 20 min. To the cooled stirred solution, boron tribromide (6.6 g,

26 mmol) was added dropwise from a syringe during 30 min. The reaction mixture was allowed to warm up to room temperature during 12 hrs and then methanol (20 ml) was added dropwise followed by water (50 ml) all at once. The white precipitate formed was dissolved in ethanol (150 ml), filtered off, evaporated, air- and vacuum dried: 0.6 g

(74%). M.p. 161.8°C. 1H (DMSO) δ: 11.36 (s, 4H), 9.17 (d, 2H), 7.90 (d, 2H). 13C

(DMSO) δ: 143.3, 130.8, 116.7, 34.9.

9,10-Dimethy1-2,3,6,7-tetramethoxyanthracene 147.

317

O O O + H O O O O

A half liter round bottom flask with a stir bar was charged with veratrole (13.8 g,

0.1 mol), acetic acid (30 ml), and acetaldehyde (12 ml, 0.21 mol). To this mixture, cooled in an ice bath, was added sulfuric acid (20 ml) dropwise during 20 min. The ice bath was removed and the reaction mixture was stirred for 6 hrs at room temperature. The deep-red mixture was poured onto crushed ice (300 g), extracted with chloroform (2×100 ml) and chromatographed on silica (150 g) with dichloromethane. Solvent evaporation afforded

1.2 g (7.4%) of off-white crystals. M.p. 328°C (DSC, 10°C/min). Lit. m.p. >340°C,432

433 1 13 323.5°C. H NMR (CDCl3) δ: 7.40 (s, 2H), 4.08 (s, 6H), 2.94 (s, 3H). C NMR

(CDCl3) δ: 148.8, 125.9, 124.01, 102.7, 55.8, 14.9.

2,3,6,7-Tetramethoxy-9,10-dipentylanthracene 148.

C5H11 O O O O + H11C5 H O OO C5H11

A 200 ml round bottom flask with a stirring bar was charged with veratrole (5.0 g, 36 mmol) and hexanal (5.0 g, 50 mmol). To the stirred reaction mixture methanesulfonic acid (15 g, 0.15 mol) was added dropwise during 20 min period. After the addition was complete, the reaction mixture was stirred at room temperature for 2 hrs, poured onto crushed ice (150 g), and extracted with chloroform. Chromatography with

318

gradual elution from neat hexane to 5% EtOAc in hexane, followed by solvent evaporation and recrystallization from 1-propanol, allowed 1.3 g (16%) of light yellow crystals. M.p. 182°C. 1H NMR δ: 7.4 (s, 2H), 4.1 (s, 6H), 3.35 (t, 2H), 1.8 (pentet, 2H),

1.6-1.5 (m, 4H), 0.9 (t, 3H). 13C NMR δ: 149.0, 129.7, 125.5, 102.7, 55.8, 32.7, 30.1,

28.9, 22.8, 14.4. UV-Vis (CHCl3) λmax, nm: 277, 357, 373, 392. (Cf. anthracene: 256,

342, 360, 379).

2,3,6-Tripentyloxy-7-hydroxyanthracene.

C5H11 O O OH H11C5 O H11C5 + H11C5 H C H H11C5 C5H11 11 5O O O C5H11

A 100 ml round bottom flask with a stirring bar was charged with 1,2- dipentyloxybenzene (2.0 g, 8 mmol) and hexanal (1.8 g, 18 mmol). To the stirred reaction mixture methanesulfonic acid (10 g, 104 mmol) was added dropwise over 30 min. After the addition was complete, the reaction mixture was stirred for 2 hrs at room temperature, poured onto crushed iced (100 g) and extracted with chloroform (100 ml).

Chromatography with gradual elution from neat hexane to 3% EtOAc in hexane allowed

1 0.8 g (20%) of the title compound as light yellow oil. H NMR (CDCl3) δ: 6.75 (m, 2H),

13 3.98 (t, 2H), 1.90-1.80 (m, 6H), 1.5 (m, 6H), 0.90 (m, 6H). C NMR (CDCl3) δ:149.1,

147.7, 138.7, 120.2, 114.4, 114.0, 69.51, 69.48, 36.4, 32.2, 30.0, 29.7, 28.5, 28.0, 22.8,

22.7, 14.3.

319

2,3,6,7-Tetrapentyloxy-9,10-dipentylanthracene 149.

C5H11 O O O O H11C5 H C C5H11 + H11C5 11 5 H11C5 H H C C H O 11 5O O 5 11 C5H11

A 100 ml round bottom flask with a stirring bar was charged with 1,2- dipentyloxybenzene (5.0 g, 20 mmol), methanesulfonic acid (20 g, 208 mmol), and cooled in an ice bath. To the stirred reaction mixture hexanal (2.1 g, 21 mmol) was added dropwise over 10 min. After the addition was complete, the reaction mixture was stirred for 2 hrs at room temperature, chloroform (20 ml) was added, stirring continued for 12 hrs and then water (100 ml) was added. The mixture was extracted with chloroform

(3×50 ml) and chromatographed with gradual elution from neat hexane to 1-2% EtOAc in hexane to allow 6.3 g (95%) of light yellow oil. Recrystallized from pentane in dry ice:

1 white crystals, m.p. 43°C. H NMR (CDCl3) δ: 6.8 (s, 2H), 3.98 (t, 4H), 3.76 (t, 2H),

13 1.90-1.80 (m, 9H), 1.5 (m, 12H), 0.90 (m, 9H). C NMR (CDCl3) δ: 149.1, 147.6, 138.7,

120.2, 114.3, 113.9, 69.5, 69.48, 50.5, 36.3, 32.1, 29.3, 28.5, 22.7, 14.2.

1,2,3,4-Tetrapentylnaphthalene.

C5H11 I C5H11 + H11C5 C5H11

I C5H11 C5H11

Ten ml round bottom flask with a stir bar was charged with 1,2-diiodobenzene

(131 μl, 1 mmol), 6-dodecyne (1.06 g, 6.3 mmol, 6×), silver acetate (347 mg, 2.07 mmol),

320

and anhydrous toluene (6.25 g). The mixture was refluxed under argon for 20 min, cooled below b.p. and palladium acetate (11 mg, 49 μmol, 5 mol%) was added at once. The reflux was continued for 12 more hours, until the peak of diiodobenzene almost disappeared on GC-MS. Conversion (by GC-MS) >90%. EI-MS: 409.36 (37), 408.41

(100), 295.24 (39), 282.24 (57), 281.21 (44), 239.28 (34), 225.26 (38), 211.19 (26),

183.20 (29), 169.19 (29). The solvent was evaporated on rotovap, and the residue was chromatographed on argenated silica gel (Note 1) to allow 392 mg (93%) of yellow oil.

1H NMR: 7.98-7.95 (m, 2H), 7.41-7.37 (m, 2H), 2.50 (t, 4H), 2.36 (t, 4H), 1.46-1.15 (m,

24H), 0.93 (t, 12H).

Note 1. Argentation of silica gel. Silver nitrate (14 g) was dissolved with sonication in methanol (550 ml). To this solution silica gel (300 g) was added and solvent evaporated on rotovap. The free-flowing solid was dried at 110°C in the dark for three hrs.

1,2,3,4,5,6,7,8-Octapentylanthracene 152.

C5H11 C5H11 I I H11C5 C5H11 + H11C5 C5H11 I I H11C5 C5H11 C5H11 C5H11

A 200 ml round bottom flaskwith a stir-bar was charged with charged with

1,2,4,5-tetraiodobenzene (5.82 g, 0.01 mmol), 6-dodecyne (10 g, 0.06 mol, 6×), silver acetate (7.0 g, 0.042 mol), and anhydrous, degassed toluene (60 ml). The mixture was refluxed under argon for 20 min, cooled below b.p. and palladium acetate (112 mg, 0.5

321

mmol, 5 mol%) was added at once and reflux was continued for 24 more hours. The reaction mixture was cooled down, filtered, washed with ether (2×50 ml), the solvent from combined washings was evaporated and the residue was chromatographed with neat hexane to allow 3.7 g (50%) of yellow oil, part of which crystallizes on standing to yellow crystals. M.p. 87°C (no LC phases). 1H NMR: 8.68 (s, 1H), 3.17 (t, 4H), 2.79 (t,

4H), 1.66-1.45 (m, 28H), 0.99 (t, 12H). 13C NMR: 135.9, 133.7, 129.1, 119.4, 33.2, 33.0,

31.5, 31.3, 30.6, 29.7, 23.0, 22.7, 14.4, 14.3. ES-MS: 739.9 (M+), 661.3, 590, 488.4,

424.4.

1,2,3,4,5,6,7,8-Octaheptylanthracene 154.

C7H15 C7H15 H15C7 C7H15

H15C7 C7H15 C7H15 C7H15

A 250 ml round bottom flask with a stir-bar was charged with 9,10- dihydroanthracene (1.8 g, 0.01 mol) and 1-bromoheptane (20 ml). To that stirred solution aluminum chloride (8 g, 0.07 mol) was added in small portions over 30 min period and stirring continued under air exposure at room temperature for 24 hrs. The reaction mixture was poured onto crushed ice (300 g) and extracted with ether (3×150 ml). The combined extracts were evaporated and the residue was chromatographed on argenated silica with neat hexane to afford 7.7 g (80%) of 154 as opalescent oil. 1H NMR: 8.68 (s,

1H), 3.17 (t, 4H), 2.79 (t, 4H), 1.66-1.25 (m, 36H), 0.89 (t, 12H). 13C NMR: 135.9, 133.7,

129.1, 119.4, 33.2, 33.0, 31.5, 31.3, 30.6, 29.7, 23.0, 22.7, 15.6, 15.3, 14.4, 14.3

322

1,2,3,4,5,6,7,8-Octa(decyl)anthracene.

C10H21 C10H21 H21C10 C10H21

H21C10 C10H21 C10H21 C10H21

A 500 ml round bottom flask with a stir-bar was charged with 9,10- dihydroanthracene (5.4 g, 0.03 mol), aluminum chloride (32 g, 0.24 mol), and heptane

(70 ml). To that stirred mixture, 1-bromodecane (55 ml, 0.26 mol) was added dropwise from an addition funnel over 2 hr period and stirring continued under air exposure at room temperature for 24 hrs. The reaction mixture was poured onto crushed ice (300 g) and extracted with ether (3×150 ml). The combined extracts were evaporated and the residue was chromatographed on silica gel with neat distilled hexane to afford, after

1 solvent evaporation, 21.2 g (54%) of pale yellow oil. Tg –15°C. H NMR (CDCl3) δ:

7.18 (s, 1H), 2.5-2.4 (m, 8H), 1.9-0.9 (m, 77H). UV-Vis (CHCl3) λmax: 263 nm.

2,3-Anthracenedicarbaldehyde.434

OCH3 CHO CHO + O CHO CHO OCH3

A 100 ml round bottom flask was charged with phthalic aldehyde (10 g, 0.074 mol), 2,5-dimethoxytetrahydrofuran (20 g, 0.15 mol), acetic acid (7.5 ml), water (7.5 ml), piperidine (10 drops), and was heated under reflux for 24 hrs. The reaction mixture was

323

cooled down, the dark red-brown precipitate was filtered and washed with water and methanol: 3.48 g (20%). The crude material was sublimed at 180°C and 0.1 mm Hg to

1 allow 2.84 g of lemon yellow crystals, 2.84 g (16%). H NMR (CDCl3) δ: 10.65 (s, 1H),

13 8.64 (s, 1H), 8.62 (s, 1H), 8.03 (dd, 1H), 7.64 (dd, 1H). C NMR (CDCl3) δ: 192.6,

136.6, 132.2, 131.1, 130.2, 129.7, 128.8, 127.9

2,3-Diazatetracene.

CHO N N CHO

A 200 ml round bottom flask was charged with 2,3-anthracenedicarbaldehyde

(1.0 g, 4.2 mmol), ethanol (120 ml), hydrazine (3 ml), and refluxed for an hour. The reaction mixture was cooled down, filtered, and the precipitate was air-dried, 0.6 g

1 (70%). Dec. 287°C. H NMR (CDCl3) δ: 9.59 (s, 1H), 8.84 (s, 1H), 8.77 (s, 1H), 8.11

13 (dd, 1H), 7.58 (dd, 1H). C NMR (CDCl3) δ: 152.2, 133.2, 132.4, 128.5, 128.2, 127.8,

127.2, 122.3.

4,5-Dimethoxyphthalic aldehyde 158.

O CHO O CHO

O Br O CHO 137 158

An oven-dried one-liter two-neck round bottom flask with a stir bar was charged

TriMEDA (17.3 g, 0.169 mol) and THP (150 ml). The necks were fitted with a

324

thermomter/argon adapter and septum. The mixture was cooled under argon in a dry ice – acetone bath to –40 °C and a solution of n-butyl lithium (5M in hexanes, 30 ml) was added dropwise via a syringe with vigorous stirring, while the temperature rose to –

10 °C. After the addition of n-BuLi was complete, the reaction mixture was stirred at –20

°C for 20 min and cooled down to –40 °C, at which temperature a solution of 2-bromo-

4,5-dimethoxybenzaldehyde (30.0 g, 0.122 mol) in THP (450 ml) was added via a syringe pump. Additional THP (100 ml) used to wash the syringe, was added to the reaction mixture at once. After the addition of bromoaldehyde was complete, the reaction mixture was stirred at –20 °C for 20 min and cooled down to –40 °C. A solution of n- butyl lithium (5M in hexanes, 29 ml) was added dropwise via a syringe with vigorous stirring, while the temperature rose to –20 °C. After the addition of n-BuLi was complete, the reaction mixture was stirred at –20 °C for 20 min and anhydrous DMF (20 ml) was added at once. The cooling bath was removed and the reaction mixture was allowed to warm up to room temperature.

The solvents were removed to ~1/4 of initial volume on a rotovap, the solution was cooled to 0 °C and hydrochloric acid (aq., 3M, ~350 ml) was added slowly at vigorous stirring. The brown precipitate formed was filtered off on a Büchner funnel, washed with water (3×200 ml), ice-cold ethanol (3×100 ml), and air-dried to give 19.0 g

(80%) of 4,5-dimethoxyphthalic aldehyde as brownish-gray crystals. The crude material was recrystallized from ethanol (aq. az., 30 ml/g) to give 10.0 g (42%) of white crystals.

435 1 M.p. 170 °C (lit. m.p. 168–169 °C). H NMR (CDCl3) δ: 10.6 (s, 1H); 7.5 (s, 1H); 4.00

13 (s, 3H); C NMR (CDCl3) δ: 190.1, 153.2, 131.0, 111.6, 56.5.

325

2,3-Dimethoxytetracene-6,11-dione 159.

OH O O CHO O + O CHO O 158 OH 155 159 O

A 500 ml round bottom flask with a stir bar was charged with a solution of 4,5- dimethoxyphthalic aldehyde (3.88 g, 0.02 mol) in a hot mixture of ethanol (150 ml) and

THF (150 ml) and a solution of 1,4-dihydroxynaphthalene (3.20 g, 0.02 mol) in hot ethanol (50 ml). To that stirred mixture, sodium hydroxide solution (aq., 10%, 5 ml) was added at once and the reaction mixture was stirred for 30 min. The yellow precipitate formed was filtered off on a Büchner funnel, washed with hot ethanol (200 ml), acetone

(100 ml), and air-dried to give 5.1 g (80%) of 2,3-dimethoxytetracene-6,11-dione as

436 1 bright orange crystals. M.p. 323 °C (lit. m.p. 328–329 °C). H NMR (CDCl3) δ: 8.7 (s,

13 2H), 8.4 (m, 2H), 7,8 (m, 2H), 7,3 (s, 2H), 4.1 (s, 6H). C NMR (CDCl3) δ: 183.1, 152.3,

134.5, 133.9, 131.8, 128.6, 127.4, 127.3, 107.9, 56.2.

2,3-Dihydroxytetracene-6,11-dione 160.

O O O HO

O HO 159 O 160 O

A 500 ml round bottom flask was charged with pyridine hydrochloride (60.0 g,

0.17 mol) and lithium iodide (10 g, 0.075 mol), and the flask was heated under nitrogen

326

at 150 °C for 12 hrs. To thus dried molten salts, 2,3-dimethoxytetracene-6,11-dione (8.3 g, 0.026 mol) was added at once (the color of the starting material solution in the melt is deep orange) and the heating was continued at 200 °C for 30 min (color gradually changes to brown-green). After the reaction was complete (TLC, CHCl3 neat), the reaction mixture was cooled down, water (150 ml) was added and the flask was shaken until the bulb of solid salts has dissolved. Hydrochloric acid (aq., 12 M, 5 ml) was added and the formed suspension was filtered off on a Büchner funnel. The solid on filter was washed with water (50 ml) and air-dried to give 7.3 g (96%) of 2,3-dihydroxytetracene-

1 6,11-dione as green-brown powder. H NMR (acetone-d6) δ: 8.6 (s, 2H), 8.3 (dd, 2H), 7.9

13 (dd, 2H), 7.2 (s, 2H). C NMR (CDCl3) δ: 185.2, 152.4, 138.6, 134.7, 131.8, 128.4,

127.4 (double intensity), 108.9.

2,3-Didecyloxytetracene-6,11-dione 161.

H C O 21 10 O HO O

HO O 160O 161 O H21C10

A 250 ml round bottom flask with a stir bar was charged with 2,3- dihydroxytetracene-6,11-dione (4.0 g, 13.8 mmol), 1-bromodecane (12.2 g, 55 mmol), potassium carbonate (7.5 g, 55 mmol), potassium iodide (75 mg), and NMP (120 ml).

The reaction mixture was stirred at 80 °C for 12 hrs and monitored by TLC (neat CHCl3).

After the reaction was complete, it was cooled down and poured into an ice-water

(250 ml) with vigorous stirring. The precipitate was filtered off on a Büchner funnel,

327

washed with water (200 ml), ethanol (20 ml), hot ethanol (30 ml) and air-dried. The crude product was dissolved in boiling chloroform (2×100 ml) and filtered from insoluble matters. The filtrate was reduced in volume to ~75 ml and impregnated onto silica gel.

Chromatography with neat chloroform allowed, after evaporation of solvent, 1.32 g

(17%) of 2,3-didecyloxytetracene-6,11-dione as bright orange gel. Recrystallization proved to be difficult, for the material is a strong gelator in most solvents (CHCl3, PrOH,

1 AcOH), and for this reason a m.p. was not measured. H NMR (CDCl3) δ: 8.6 (s, 2H), 8.3

(dd, 2H), 7.8 (dd, 2H), 7.2 (s, 2H), 4.1 (t, 4H), 1.9 (pentet, 4H), 1.5–1.3 (m, 28H), 0.9 (t,

13 6H). C NMR (CDCl3) δ: 185.1, 152.4, 138.6, 134.7, 131.9, 128.4, 127.4, 108.9, 69.2,

32.2, 29.9, 29.8, 29.7, 29.6, 29.2, 26.3, 22.9, 14.4.

2,3-Didecyloxytetracene 162.

H C 21 10 O H21C10 O O

O O H C 161 O 162 21 10 H21C10

A 250 ml round bottom flask with a stir bar was charged with anhydrous cyclohexanol (distilled from sodium, 20 ml), aluminum (1.0 g, 37 mmol), and mercry (II) chloride (22 mg, 0.08 mmol). The flask was topped with a reflux condensor, flushed with nitrogen, and carefully heated over flame of a Bunsen burner until hydrogen evolution started. At that point the flask was removed from the flame and reaction went spontaneously. After the hydrogen evolution ceased, carbon tetrachloride (0.5 ml) was added, followed by 2,3-didecyloxytetracene-6,11-dione (1.32 g, 2.3 mmol). The reaction

328

mixture was heated at 120 °C and monitored by TLC (dichloromethane : hexanes = 1:1).

After the reaction was complete (~3 hrs), the reaction mixture was cooled to ~70 °C and the following reagents were added dropwise in this order: ethanol (95% aq., 40 ml), acetic acid (4 ml), water (15 ml), hydrochloric acid (aq., 2M, 5 ml). The precipitate formed was filtered off on a Büchner funnel, washed with water (70 ml), ethanol (2×20 ml), and air-dried to give 1.1 g (88%) of 2,3-didecyloxytetracene as off-orange crystals.

The crude product was chromatographed on a 150 mm Biotage column with hexanes : dichloromethane (gradient elution from 2:1 to 1:1) to give, after evaporation of solvents,

140 mg (11%) of pure 2,3-didecyloxytetracene as orange crystals. M.p. (EtOH) 130–

1 132 °C (microscope, hot stage). UV-Vis (CHCl3) λmax: 240, 304, 393. H NMR (CDCl3)

δ: 8.8 (s, 2H), 8.1 (dd, 2H), 7.7 (s, 2H), 7.6 (dd, 2H), 7.3 (s, 2H), 4.2 (t, 4H), 1.9 (pentet,

13 4H), 1.5 (pentet, 4H), 1.3 (m, 24H), 0.9 (t, 6H). C NMR (CDCl3) δ: 154.2, 135.2, 130.3,

130.2, 129.5, 129.4, 129.3, 109.7, 69.6, 32.1, 29.8 (two peaks), 29.6 (double intensity),

29.2, 26.2, 22.9, 14.3.

Pentacene 163.

164 O 163

Preparation of aluminum cyclohexanolate stock solution. Commercial

20 cyclohexanol (Acros, 98%, nD = 1.4625) was found to contain 1…2% of water and was purified by distillation from sodium under normal pressure, employing a 15 cm Vigreaux

329

column, thermally insulated with an asbestos tape, wrapped around. The forerun boiling below 160 °C (50 ml from a liter of commercial cyclohexanol) was discarded and the fraction boiling 160…161 °C was collected and stored over powdered437 3Å molecular sieves. A one-liter three-neck round bottom flask with a stirbar was charged with cyclohexanol (500 ml), aluminum turnings (25 g, 0.925 mol), mercury (II) chloride

(0.625 g, 2.3 mmol), and carbon tetrachloride (12.5 ml). The flask was topped with a reflux condensor, thermometer, and nitrogen inlet and the reaction mixture was heated under nitrogen on an IR hot plate to ~160 °C until hydrogen evolution started. The temperature was lowered to ~120 °C or as necessary to keep the hydrogen evolution vigorous, but not out of control (intermittent cooling of the flask by immersing it into an ice-water bath was used to control the reaction rate). After the hydrogen evolution had ceased (~3 hrs) and all aluminum had dissolved, the solution of aluminum cyclohexanolate was cooled down, and another portion of carbon tetrachloride (7 ml) was added.

To the warm (60…80 °C) solution of aluminum cyclohexanolate in cyclohexanol obtained above, pentacene-6,13-dione (25.0 g, 0.08 mol) was added at once under nitrogen and the reaction mixture was refluxed for 4 hrs. The precipitate of pentacene (insoluble in cyclohexane) started to appear after 2 hrs of reflux. The reaction mixture was cooled down, acetic acid (500 ml), followed by methanol (1250 ml) were added and stirred for an hour. The suspension was then filtered off on a Büchner funnel, washed with methanol (300 ml), and air-dried to give 12.8 g (57%) of crude pentacene.

Triple sublimation in vacuum (p = 0.1 mm Hg) gave 7.4 (33%) of purified pentacene.

330

Subsequent sublimations did not increase the purity of pentacene (determined as residue after sublimation by TGA). Analysis (found/calculated): C 94.34/94.93, H 5.66/5.07.

Pentacene-6,13-dione 164.

O O CHO + CHO O 164 O

o-Phthalic aldehyde (10.0 g, 0.075 mol) was dissolved in ethanol (95.6%, aq.,

250 ml). 1,4-Hexanedione was dissolved in ethanol (95.6%, aq., 150 ml). The solutions were mixed into a one-liter Erlenmeyer flask and stirred with a large stir bar. To that mixture, potassium hydroxide (aq., 5%, 7.5 ml) was added at once. Marsh-green suspension was formed immediately. The reaction mixture was heated at 50 °C for an hour, during which time the color changed to yellow-brown. The hot reaction mixture was filtered off on a Büchner funnel, washed with hot ethanol (300 ml), methanol (200 ml), and air-dried to give 10.1 g (87%) of pentacene-6,13-dione as yellow crystals. The crude material may be recrystallized from DMAc (80 ml per gram) to give orange-yellow crystals, m.p. 390 °C or sublimed to give yellow needles, m.p. 389 °C. Lit.256 m.p. 377–

394 °C.

331

2,5-Dibenzoylterephthalic acid and 2,4-Dibenzoylisophthalic acid [165].438

O O HOOC O O COOH

+ O O + COOH OO O O COOH

A 600 ml autoclave fitted with a mechanical stirrer and a thermocouple was charged with pyromellitic dianhydride (25 g, 0.115 mol), benzene (200 ml, 2.25 mol), and anhydrous aluminum chloride (60 g, 0.45 mol). The autoclave was closed and heated with stirring for 4 hours at 100°C (internal temperature). After cooling, the contents were poured into a 2 l beaker with ice (500 g), and hydrochloric acid (37% aq., 50 ml), and stirred for 15 min until the dark color disappeared. The precipitate formed was filtered, washed with hydrochloric acid (2M, 200 ml), water (300 ml), and air-dried resulting in brown-gray powder (48 g). The crude product was dissolved in hot solution of potassium hydroxide (22 g, 0.4 mol) in water (200 ml), filtered, and the residue on filter was treated with additional solution of potassium hydroxide (7 g, 0.125 mol) in water (70 ml), and filtered. The combined filtrates were cooled to ~5°C and slowly acidified with hydrochloric acid (2M, 350 ml). The white precipitate formed was filtered, washed with water (300 ml), and air-dried to allow 17 g (40%) of white powder, which was used in subsequent reaction.

332

Pentacene-5,7,12,14-tetraone 166.

O COOH

O O HOOC O + O O OO

HOOC COOH

A 250 ml Erlenmeyer flask with a stir bar was charged with a mixture of two isomeric dibenzoylphthalic acids (AS-3-25a, 16.8 g, 0.045 mol), sulfuric acid (98%, 135 ml), the neck of the flask was sealed with Parafilm, and the reaction mixture was heated at 100-120°C for four hours, being occasionally stirred with a teflon spatula. After cooling, the reaction mixture was poured onto ice (400 g), filtered on fiber glass filter, washed with sodium bicarbonate (sat.aq., 100 ml), hot water (300 ml), and hot ethanol

(200 ml). The brown-gray residue on the filter was then suspended in boiling ethanol

(2×100 ml), filtered, and air-dried to give 14.0 g (92%) of the pentacene-5,7,12,14- tetraone as a greenish-yellow powder. M.p. 407°C (lit.438 m.p. 408°C).

1,4-Anthracenequinone 167.

O OH O

O OH 167 O

A two-liter two-neck round bottom flask equipped with a mechanical stirrer and a reflux condensor, was charged with quinizarine (Note 1, 57.0 g, 0.2375 mol),

333

anhydrous methanol (1000 ml). While stirring, the flask was cooled in an acetone — dry ice bath to ca. –30 °C and sodium borohydride (37.8 g, 1 mol) was added during half an hour from a powder-addition funnel, placed atop of the reflux condensor. When the addition has been complete, the cooling bath was removed, the reaction mixture was warmed up to room temperature and heated on a heating mantle to reflux for 24 hrs. The reaction mixture was cooled and most of methanol was removed on a rotovap. To the residue after evaporation water (700 ml) was added and the resulting solution was filtered

(Note 2). To the filtrate hydrochloric acid (aq., 3 M, 400 ml) was added dropwise at stirring, causing at pH~7 an abrupt precipitation of thick yellow mass. The yellow precipitate was filtered on a Büchner funnel, washed with water (300 ml), air- and vacuum-dried to give 46.4 g (94%) of 1,4-anthracenequinone as yellow crystals. The crude material is of sufficient purity, but may be recrystallized from BuOAc (800 ml per

30 g) with 72% recovery. M.p. 224 °C (lit. m.p. 204–206,439 225 °C440). 1H NMR δ: 8.6

(s, 2H), 8.0 (dd, 2H, J = 3.3 Hz), 7.7 (dd, 2H, J = 3.3 Hz), 7.1 (s, 2H). 13C NMR δ: 184.7,

140.0, 134.8, 130.2, 129.6, 128.9, 128.4.

Note 1. Commercial quinizarine from Acros was recrystallized from acetic acid

(14 g AcOH per g). Note 2. The filtration of the water solution before acidification is essential to get pure product.

334

2,3-Dihydro-1,4-anthracenequinone 168.

O O

167 O 168 O

A 250 ml Erlenmeyer flask was charged with 1,4-anthracenequinone (3.0 g, 14.4 mmol), trifluoroacetic acid (85 g), and zinc dust (3.78 g, 5.8 mmol). The reaction mixture was sonicated for 5 min, and filtered. The filtrate was poured into water (300 ml) and the precipitate formed was filtered off on a Büchner funnel. The crude product (2.8 g, 92%) was washed with water (200 ml), cold ethanol (30 ml), dichloromethane (100 ml), and recrystallized from ethanol (aq. az., 90 ml) — chloroform (10 ml) mixture to give 2.6 g

(86%) of 2,3-dihydro-1,4-anthracenequinone as crystals of marsh-green color. M.p.

227 °C (lit.262c m.p. 175 °C). 1H NMR δ: 8.6 (s, 2H), 8.0 (dd, 2H, J = 3.3 Hz), 7.7 (dd,

2H, J = 3.3 Hz), 7.1 (s, 2H), 2.16* (s, 4H). 13C NMR δ: 199.7*, 184.6, 140.0, 134.8,

130.2, 129.6, 128.8, 128.3, 21.7*.

*quickly disappears due to isomerization into 1,4-dihydroxyanthracene

1-Fluoropentacene 169.

F 173 O F 169

1-Fluoropentacene was prepared similar to pentacene 163, starting from 1- fluoropentacene-6,13-dione 173 (2.78 g, 8.5 mmol). Crude material (1.1 g, 43%) was

335

sublimed: two times covered with degreased iron filings and two times neat to give 110 mg (4%) of dark blue crystals. M.p. 289 °C. UV-Vis (benzene, degased) λmax: 279, 291,

303, 322, 349. IR (neat solid, HATR) ν, cm–1: 3014, 1443, 1371, 1306, 1230, 1220, 814,

734. Analysis (found/calc.): C 88.19/89.17, H 4.85/4.42.

2-Fluoropentacene 170.

2-Fluoropentacene was prepared by Kihong Park similar to 1-fluoropentacene. The crude material received was sublimed in vacuum (p = 0.15 mm Hg) at 180–210 °C. Sublimes in inert atmosphere without decomposition, TGA peak (2nd derivative maximum) at 385 °C.

Analysis (found/calc.): C 87.89/89.17, H 4.94/4.42.

3-Fluorophthalaldehyde 172.

CHO OH OH CHO F F 179 172

A 250 ml round bottom flask with a stir bar was charged with oxalyl chloride (5 ml, 55 mmol) and dichloromethane (120 ml). The flask was topped with an addition funnel and cooled in a dry ice – acetone bath to –40 °C under argon. From the addition funnel, a solution of DMSO (7 ml, 90.5 mmol) in dichloromethane (20 ml) was added dropwise during 10 min. The reaction mixture was stirred for 15 min and 1,2- bis(hydroxymethyl)-3-fluorobenzene (3.0 g, 19.2 mmol) in dichloromethane (10 ml) was added dropwise from the same addition funnel during 5 min. The reaction mixture was

336

stirred for additional 30 min and triethylamine (30 ml, 0.2 mol) was added at once. The reaction mixture was allowed to warm up to room temperature, poured onto ice (300 g) and extracted with ether (3×150 ml). The combined extracts were dried with MgSO4, solvents evaporated on rotovap, and the oily residue was chromatographed with hexane – ethyl acetate (7:3) to give, after removal of solvents, 2.1 g (72%) of 3-

1 fluorophthalaldehyde as colorless oil. H NMR (CDCl3) δ: 10.6 (s, 1H), 10.5 (s, 1H), 8.0

(m, 1H), 7.8 (m, 1H), 7.7 (m, 1H).

1-Fluoropentacene-6,13-dione 173.

O O CH2Br + CH2Br 167 O F 177 F 173 O

A 250 ml round bottom flask with a stir bar was charged with 1,4-anthraquinone

167 (4.16 g, 0.02 mol), 1,2-bis(bromomethyl)-3-fluorobenzene 177 (10.0 g, 0.036 mol), sodium iodide (20 g.0.13 mol), and DMF (150 ml). The reaction mixture was heated at

70 °C for 24 hrs (dark brown tar forms), cooled down, poured into water (450 ml). The precipitate formed was filtered on a Büchner funnel, washed with water (300 ml), and air- driedto give 7.3 g of dark-brown solid. The crude product was sublimed in vacuum twice ti give 2.6 g (35%) of 1-fluoropentacene-6,13-dione as brownish crystals. The NMR characterization was not performed due to low solubility.

337

1-Fluoro-2,3-dimethylbenzene 176.

176

NH2 F

A four-liter beaker, equipped with a mechanical stirrer, was charged with water

(500 ml), and o-xylidine (2,3-dimthylaniline, 200.0 g, 1.65 mol). The beaker was immersed into a dry ice – acetone cooling bath (bath temperature –10 °C) and with stirring, ice water mixture (1500 g) was added. When the temperature in the beaker had reached 10 °C, a solution of hydrochloric acid (aq., 12M, 176 ml) in water (200 ml) was added at once. When the temperature had lowered to 0–5 °C after HCl addition, a precooled to 5 °C solution of sodium nitrite (125.3 g, 1.8 mol) in water (300 ml) was added with vigorous stirring at such a rate that temperature did not rise above 5 °C. After the addition was complete, the reaction mixture was stirred for additional 15 min and fluoroboric acid solution (aq., 39%, 380 g, 1.7 mol), precooled to 5 °C was added in 20-

30 ml portions, simultaneously increasing the power input for the mechanical stirrer.

After the addition of HBF4 was complete, the suspension was stirred for 10 additional min, and filtered off on a Büchner funnel. The solid on filter was washed with cold water

(200 ml), methanol (200 ml, precooled to –30 °C), and ether (300 ml, precooled to –30

°C). After suction drying on the filter, the solid was transferred into a one-liter pear- shaped flask and additionally dried on the rotovap at 30–35 °C and 20 mm Hg, followed by 40–45 °C at 0.1 mm Hg. After the drying was complete, the flask was removed from the rotovap, topped with a long reflux condensor, and heated with a heating gun on one side until local decomposition of the diazonium salt started. Heating was continued for 20

338

min until all solid had decomposed and nitrogen evolution had ceased. The liquid resulting from decomposition was distilled twice at normal pressure, collecting fraction

441 1 with b.p. 144–147 °C. Lit. b.p. 146–148 °C. H NMR (CDCl3) δ: 6.9–7.0 (m, 3H), 2.2

13 (s, br, 3H), 2.1 (d, 3H, JH–F = 2 Hz). C NMR (CDCl3) δ: 161.9 (d, JC–F = 240 Hz), 139.0

(d, JC–F = 5 Hz), 126.7 (d, JC–F = 10 Hz), 125.0 (d, JC–F = 3 Hz), 123.6 (d, JC–F = 17 Hz),

+ 112.9 (d, JC–F = 22), 19.5 (d, JC–F = 3 Hz), 10.8 (d, JC–F = 7 Hz). EI-MS: 124 (M ).

1,2-bis(bromomethyl)-3-fluorobenzene 177

CH2Br

CH2Br F F 176 177

A two-liter round bottom flask with a stir bar was charged with N- bromosuccinimide (127.2 g, 0.707 mol, Note 1), anhydrous carbon tetrachloride (500 ml), 1-fluoro-2,3-dimethylbenzene (39.9 g, 0.321 mol), and benzoyl peroxide (1.1 g, 4.5 mmol). The flask was topped with a reflux condensor, and the reaction mixture was heated to reflux on a heating mantle and irradiated with a 450 W medium pressure Hg

Hanovia lamp. After approximately 15 min of irradiation, a vigorous reaction was initiated. Reflux and irradiation was continued for an additional hour. The reaction mixture was cooled down, filtered from succinimide on a Büchner funnel and the filtrate was rotovaped. The oily residue after evaporation was vacuum distilled, collecting fraction b.p. 100–130 °C at 0.5 mm Hg: 81.5 g (91%), which partially solidified on

442 1 standing: m.p. 38–39 °C (lit. m.p. 41–42 °C). H NMR (CDCl3) δ: 7.5–6.9 (m, 3H), 4.8

339

13 (d, 2H, JH–F = 2 Hz), 4.6 (s, 2H). C NMR (CDCl3) δ: 161.9 (d, JC–F = 240 Hz), 139.0 (d,

JC–F = 5 Hz), 131.0 (d, JC–F = 10 Hz), 127.0 (d, JC–F = 3 Hz), 125.0 (d, JC–F = 17 Hz),

116.0 (d, JC–F = 22 Hz), 29.0 (d, JC–F = 3 Hz), 21.0 (d, JC–F = 7 Hz). EI-MS: 282.9 (55),

281.0 (100), 279,3 (55), 202.1 (22), 200.1(21), 122.3 (28), 121.3 (51), 102.2 (14), 101.2

(18), 75.4 (20), 74.3 (8).

Note 1. When exactly two equivalents of NBS have been used, a mixture of mono- and dibromo- products in 2:1 ratio was separated (determined by GC-MS).

Increasing the NBS amount to four equivalents, however, gave an 85:5 mixture of dibromo- and tribromo- isomers only (determined by GC-MS). Only traces (on GC-MS) of o-fluoro-tetrabromo-o-xylene 178 have been detected when dibromo- compound 176 was subjected to radical bromination with 4 equivalents of bromine.

1,2-Bis(hydroxymethyl)-3-fluorobenzene 179.

CH Br 2 OH OH CH2Br F 177 F 179

A 2000 ml round bottom flask with a stir bar was charged with 1,2- bis(bromomethyl)-3-fluorobenzene (87.2 g, 0.31 mol), potassium carbonate (87.0 g, 0.63 mol), water (1000 ml), and tetrabutylammonium tetrafluoroborate (5.0 g). The reaction mixture was stirred at 80 °C for 8 hrs, cooled down, and extracted with ether (5×200 ml).

The combined extracts were dried with MgSO4, and ether evaporated to give 38.0 g

1 (79%) of 1,2-bis(hydroxymethyl)-3-fluorobenzene as colorless liquid. H NMR (CDCl3)

340

13 δ: 7.8 (d, 1H, JH–F = 8 Hz), 7.4 (m, 1H), 7.1 (s, 2H), 7.0 (m, 1H), 4.7 (m, 2H). C NMR

(CDCl3) δ: 160 (d), 143.8, 131.0, 126.3, 119.8, 116.5 (d), 61.3, 37.0. The diol becomes pink if left to open air for more than a day.

2,3,9,10-Tetrahexyloxypentacene-6,13-dione 185.

O O

HO OH H13C6O OC6H13

HO OH H13C6O OC6H13 187 O 185 O

2,3,9,10-Tetrahexyloxypentacene-6,13-dione was prepared similar to 2,3- didecyloxytetracene-6,11-dione 161, starting from 2,3,9,10-tetrahydroxypentacene-6,13-

1 dione in 4% yield. M.p. 189.5–194.5 °C (microscope, hot stage). H NMR (CDCl3) δ: 8.6

(s, 1H), 7.3 (s, 1H), 4.2 (t, 2H), 1.9 (pentet, 2H), 1.6 (pentet, 2H), 1.4 (m, 4H), 0.9 (t, 3H).

13 C NMR (CDCl3) δ: 183.4, 152.4, 131.9, 129.5, 127.4, 108.9, 69.2, 31.8, 29.1, 25.9,

22.8, 14.2.

2,3,9,10-Tetrahydroxypentacene-6,13-dione 187.

O O O O HO OH

O O HO OH 188 O 187 O

2,3,9,10-Tetrahydroxypentacene-6,13-dione was prepared similar to 2,3- dihydroxytetracene-6,11-dione 160, starting from 2,3,9,10-tetramethoxypentacene-6,13- dione in 90% yield. M.p. 409 °C (dec.). TGA decomposition maximum at 440 °C. IR

341

(solid, HATR) ν, cm–1: 3220, 3150, 3050, 1670, 1640, 1630, 1567, 1270, 1256, 1190,

1150, 1138, 1100, 951, 770, 711.

2,3,9,10-Tetramethoxypentacene-6,13-dione 188.

O O O CHO O O + O CHO O O O 158 O 188

4,5-Dimethoxyphthalic aldehyde (6.6 g, 0.034 mol) was dissolved in boiling ethanol (95.6%, aq., 400 ml). 1,4-Hexanedione (2.0 g, 0.018 mol) was dissolved in warm ethanol (95.6%, aq., 20 ml). The solutions were mixed into a 500 ml Erlenmeyer flask and stirred with a large stir bar. To that mixture, potassium hydroxide (aq., 10%, 0.8 ml) was added at once. Marsh-green suspension was formed immediately. The reaction mixture was heated at 70 °C for an hour, during which time the color changed to yellow- brown. The hot reaction mixture was filtered off on a Büchner funnel, washed with hot ethanol (300 ml) and air-dried to give 6.7 g (93%) of 2,3,9,10-tetramethoxy pentacene-

314 1 6,13-dione as yellow crystals. M.p. 400 °C (lit. m.p. >320 °C). H NMR (CDCl3) δ: 8.7

(s, 4H), 7.3 (s, 4H), 4.1 (s, 12 H). 13C NMR (solid-state) δ: 183.0, 150.5, 128.2, 107.7,

55.2. IR (neat, HATR) ν, cm–1: 1670, 1620, 1590, 1510, 1480, 1435, 1390, 1250, 1210,

1160.

342

2,3,9,10-Tetramethoxypentacene 189.

O O O O O

O O O O 188 O 189

2,3,9,10-Tetramethoxypentacene was prepared similar to pentacene from

2,3,9,10-Tetramethoxypentacene-6,13-dione 188 (4.0 g, 9.3 mmol) and aluminum cyclohexanolate solution in cyclohexanol (100 ml) to give 2.4 g (65%) of 189 as dark ruby red crystals. M.p. 413 °C (subl.). 13C NMR (solid-state) δ: 154.6, 132.6, 128.1,

108.7, 59.8, 58.2 (two peaks for methoxy groups are probably due to their magnetic nonequivalence in crystalline lattice).

349 HgO/SiO2 Mercury tetrafluoroborate on silica gel.

2 HBF4 + HgO + SiO2 = Hg(BF4)2–SiO2 + H2O

A one liter pear-shaped flask was charged with hydrofluoroboric acid (50% aq. solution, 175.6 g, 1 mol) and yellow mercury (II) oxide (107 g, 0.495 mol) was added at once. The flask was swirled until clear solution was formed and silica gel (107 g) was added. The slurry was evaporated on rotavap (80°C, 0.2 mm Hg) till constant mass.

+ – Bis(pyridine) iodonium tetrafluoroborate Py2I B F4 .

+ – 2 Py + I2 + Hg(BF4)2 / SiO2 + CH2Cl2 = Py2I BF4

A one liter Erlenmeyer flask was charged with mercury tetrafluoroborate impregnated on silica gel (50 wt%, 215 g, 0.5 mol), dichloromethane (900 ml), pyridine

343

(96 ml), and large stir bar. To vigorously stirred reaction mixture, iodine (127 g, 0.5 mol) was added in portions (10 g) and stirring was continued for two hours. Reaction mixture warmed up (~40°C) and changed color to yellow as the reaction progressed. The solvent was evaporated to 200 ml and ether (300 ml) was added to precipitate yellow bis(pyridine) iodonium tetrafluoroborate. The precipitate was filtered, washed with ether

(200 ml) and air dried to yield 146.8 g (79%). M.p. 153°C (lit.350 149-151°C).

Iodopentamethylbenzene 194.

A one liter two neck round bottom flask with a stir bar was charged with pentamethylbenzene (44.35 g, 0.3 mol), acetic acid (225 g), and iodine (34.3 g, 0.135 mol). The flask was fitted with a reflux condensor, thermometer, and an addition funnel containing periodic acid dihydrate (44.1 g, 0.193 mol) in water (60 ml). This solution was added dropwise to the stirred reaction mixture during 20 min. After the addition was complete, the reaction mixture was brought to 110°C and refluxed for 4 hrs. When the reaction was complete, the reaction mixture was cooled down to room temperature, diluted with water (500 ml), filtered off and the precipitate was washed with water (200 ml), cold ethanol (2 × 50 ml), and air-dried to give 63 g (76%) of crude product. The crude product was recrystallized twice from 1-propanol (250 ml) to give 38 g (46%) of product. M.p. 132°C (lit. m.p. 127-135°C). EI-MS: 274.03 (100), 147.13 (80), 128.07

344

1 (10), 119.11 (38), 91.05 (39), 77.11 (19), 63.03 (10). H NMR (CDCl3) δ:2.51 (s, 6H),

2.27 (s, 6H), 2.18 (s, 3H). 13C NMR: 137.0, 135.1, 133.4, 109.3, 28.4, 18.7, 17.1.

1,2,4,5-Tetraiodo-p-xylene 195.

I I

A one liter three neck round bottom flask was charged with sulfuric (500 ml) and periodic (26 g, 0.114 mol) acids, fitted with a mechanical stirrer and a thermometer and stirred until clear solution was formed (~30 min). The flask was cooled then in dry ice – acetone bath (–15°C) to –5°C and finely ground iodine (87 g, 0.343 mol) was added at once. After a short induction period (~10 min) the temperature raised to +10°C and more dry ice was added to the bath to keep the temperature of the reaction mixture below

0°C. To the reaction mixture cooled to –5°C, p-xylene (21.5 g, 0.2 mol) was added dropwise from an addition funnel during 20 min. After the addition was complete, the cooling bath was replaced with a heating mantle and reaction was stirred at 80°C for three days, while monitored by GC-MS. Upon completion (GC peak of triiodo-p-xylene disappeared), the reaction mixture was cooled down, poured onto crushed ice (2 kg), filtered, the precipitate on filter washed with water (500 ml), re-suspended in water (1000 ml), filtered, re-suspended in sodium bicarbonate (10% aq., 500 ml), filtered, washed with sodium bisulfite (10% aq., 200 ml), water (500 ml), re-suspended and sonicated in ethanol (500 ml), filtered, and air-dried to yield crude dark-brown product 108.2 g (88%).

345

The crude material was sublimed (3×, 230°C, 0.015 mm Hg) to yield 45.4 g (37%) of white crystals. M.p. 245.5°C. (Lit.443 m.p. 245-248°C). EI-MS: 609.69 (87), 482.70 (39),

483.78 (20.5), 355.83 (74), 229.01 (60.5), 102.06 (100), 103.05 (17), 91.08 (2), 74.02 (9),

75.04 (6).

1,2,4,5-Tetraiodobenzene 196.

I I I I I + I I I I I

0°C. To the reaction mixture cooled to –5°C, benzene (15.6 g, 0.2 mol) was added dropwise from an addition funnel during 20 min. After the addition was complete, the cooling bath was removed and reaction was stirred at room temperature for three days, while monitored by GC-MS. Upon completion (GC peak of triiodobenzene disappeared), the reaction mixture was poured onto crushed ice (1 kg), filtered, the precipitate on filter washed with water (500 ml), sodium bisulfite (10% aq., 200 ml), and air-dried. The crude brown-gray product was washed with hot BuOAc (500 ml) to remove most of the dark

346

color and recrystallized from di(ethylene glycol) monoethyl ether (Note 1) to give 67.8 g

(58%) of tetraiodobenzene 196 as yellowish crystals. The residue insoluble in di(ethylene glycol) monoethyl ether was recrystallized from nitrobenzene to give 9.9 g of yellow crystals of tetraiodobenzene and 2.7 g of orange powder of hexaiodobenzene443 (m.p.

423°C dec 430°C), insoluble in hot nitrobenzene, but recrystallizable from boiling nitrobenzene. Total yield of tetraiodobenzene was 77.7 g (73%). M.p. 252°C. (Lit.444 m.p.

249-254°C). EI-MS: 581.75 (M+, 81), 455.79 (100), 329.90 (15), 74.00 (30).

Note 1. Di(ethylene glycol) monoethyl ether (one liter) was brought to boiling and added to the crude tetraiodobenzene, boiled with stirring for ten minutes and rapidly filtered. The filtrate was allowed to cool down to room temperature, filtered from the formed crystals of tetraiodobenzene, and returned for recrystallization. This operation was repeated three times. The recrystallized material may be further purified by sublimation (two times, 150-210°C, 0.05 mm Hg) to leave behind 4.7 g of hexaiodobenzene and then repeatedly recrystallized from dimethoxyethane (50 g of solvent per gram of material) until material with a pure white color was obtained.

2,6-Diiodo-6-methylaniline 198.

I I NH2 NH2

A 300 ml round bottom flask with a stir bar was charged with p-toluidine (2.56 g, 0.02 mol), iodine (13 g, 0.051 mol), ethanol (150 ml), and water (70 ml). The mixture

347

was stirred for three hours at room temperature and then for 24 hrs at 80°C. After this time the reaction mixture was cooled, diluted with water (100 ml) and sodium bisulfite solution (10% aq., 50 ml), filtered off and the filtrate washed with water (200 ml) and cold ethanol (50 ml) to give the crude product 5.3 g (62%) as a black powder. The crude product was recrystallized from 1-propanol to give 2.22 g (25%) of brown crystals. M.p.

124°C.

N-Acetyl-4-methylaniline.

NH2 NH O

A 500 ml round bottom flask with a stir bar was charged with p-toluidine (35.5 g, 0.331 mol) and acetic acid (100 ml). The flask was topped with an addition funnel, filled with acetic anhydride (40 g, 0.392 mol), which was added to the reaction mixture dropwise during 30 min. After the addition was complete, ethanol (100 ml) and water (50 ml) were added and the resulting mixture was recrystallized to afford 31.4 g (63.5%) of

N-Acetyl-4-methylaniline. M.p. 152.2°C (lit. m.p. 148-155°C). EI-MS: 149.08 (23),

107.07 (70), 106.10 (100), 79.09 (10), 77.1 (24).

348

N-Acetyl-2-iodo-4-methylaniline 199.

I NH NH

O O

A 200 ml round bottom flask with a stir bar was charged with N-acetyl-4- methylaniline (5.3 g, 0.035 mol), sodium hydrocarbonate (6.5 g, 0.077 mol), dichloromethane (200 ml), methanol (10 ml), water (3 ml), and iodine monochloride

(11.9 g, 0.079 mol). The reaction mixture was monitored on TLC (hexane : ethyl acetate

= 1:1) and by GC-MS as it was stirred at room temperature for 12 hrs, then refluxed for

12 hrs (s.m. still present on TLC), then dichloromethane was removed on rotavap and replaced with 1,2-dichloroethane (200 g) and pyridine (20 g) and refluxed for another 12 hrs. The reaction mixture was cooled down, filtered, washed with NaHSO3 (10% aq.,

2×100 ml), HCl (10% aq., 2×200 ml) and an aliquot injected into GC-MS: 63% conversion to N-acetyl-2-iodo-4-methylaniline, EI-MS: 274.96 (14), 233.01 (32), 148.11

(100), 106.1 (60), 77.07 (39).

3,4,5-Triiodotoluene 201.

I I I I I NH2

A 125 ml Erlenmeyer flask with a stir bar was charged with 2,6-diiodo-6- methylaniline (1.38 g, 3.84 mmol), sulfuric acid (40 g) and cooled down in an ice-acetone

349

bath until the amine completely dissolves. To the cooled reaction mixture an ice-cold solution of sodium nitrite (0.3 g, 4.3 mmol) in water (2 ml) was added dropwise at vigorous stirring and left for diazotization for 30 min. Then an ice-cold solution of potassium iodide (0.76 g, 4.57 mmol) in water (10 ml) was poured into the reaction mixture at once, resulting in gas evolution. When the gas evolution ceased, the flask was removed from the ice-acetone bath, allowed to warm up to room temperature, stirred for

2 hrs, and poured onto crushed ice (50 g). The black precipitate was filtered off, washed with water (100 ml) and cold ethanol (50 ml) and sublimed twice (110°C, 1 mm Hg) to give 1.6 g (88%). The sublimed product was recrystallized from 1-propanol (50 ml) to give 1.2 g (66%). M.p. 119.5°C. EI-MS: 469.81 (100), 342.92 (22), 216.02 (18), 89.11

(28), 63.01 (35). 1H NMR: 7.69 (s, 2H), 2.17 (s, 3H). 13C NMR: 141.5, 139.8, 116.9,

106.9, 20.0.

1,4-Diiodonaphthalene 202.

A two-liter three-neck round bottom flask with a stir bar was charged with 1,4- dibromonaphthalene (50.0 g, 0.175 mol) and anhydrous ether (1000 ml). The flask was fitted with a thermometer, nitrogen inlet, and a septum. The reaction mixture was cooled to –20 °C in a dry ice – acetone bath and tert-BuLi (393 g, 595 ml of 1.7 M solution in hexane, 0.88 mol, 5 eq.) was added dropwise during a course of 40 min via a cannula from a bottle, tared on top-load balances. After addition was complete, the reaction mixture was allowed to warm up to room temperature, stirred at that temperature for 20 min, and cooled back to –20 °C. To the cooled reaction mixture, iodine (152.0 g, 0.6 mol,

350

3.3 eq.) was added in 5-7 g portions via a powder funnel under a nitrogen counterflow at such a rate that the temperature did not rise above –15 °C. After addition was complete, the reaction mixture was allowed to warm up to room temperature, stirred at that temperature for 1 hour and quenched with Na2S2O3 (aq., 10%, 200 ml).

The solution was transferred into a separatory funnel and the organic layer was washed in this order with: Na2S2O3 (aq., 10%, 200 ml), water (3×200 ml), hydrochloric acid (aq., 2M, 2×200 ml), water (3×200 ml), treated with freshly activated carbon (2×70 g), and dried with MgSO4. The solvents were evaporated on a rotovap and the residue crystallized to give 28.0 g (40%) of 1,4-diiodonaphthalene as canary yellow crystals.

M.p. 108.5 °C (lit.373 m.p. 110–111 °C). 1H NMR δ: 8.05 (dd, 2H, J = 3.3 Hz), 7.8 (s,

2H), 7.6 (dd, 2H, J = 3.3 Hz). 13C NMR δ: 138.3, 134.9, 133.2, 128.8, 100.9.

The ultimate purification was performed via a series of column chromatographies with doubly distilled solvents (hexane – ether 8:2 and hexane – dichloromethane 8:2), treatment of cyclohexane solution with freshly activated charcoal and neutral alumina, and recrystallization from residue-free 1-propanol. Before the final recrystallization, a saturated solution in chloroform was micron-filtered, chloroform evaporated, and the material was recrystallized from micron-filtered 1-propanol.

2,3-Diaminonaphthalene.

OH NH2

351

A 100 ml autoclave was charged with a stir bar, 2,3-dihydroxynaphthalene (7.7 g, 48 mmol), ammonium sulfite monohydrate (9.2 g, 68 mmol), and ammonium hydroxide (29% aq. sln, 80 ml). The reaction mixture was heated in a mantle filled with iron filings to 250°C (mantle temperature) until pressure reached 520 psi (35 bar, takes ~

1.5 hr) and then heating continued for 12 hours. The cooled reaction mixture was poured into water (200 ml), filtered, washed with water (2×200 ml), and air dried to allow 6.4 g

(84%) of yellow crystals of 2,3-diaminonaphthalene. M.p. 193°C (lit. 190-199°C). The crude material was recrystallized from 1-propanol (170 ml) to yield 4.8 g (63%) of tan- yellow crystals. EI-MS: 159.18 (M+1, 12), 158.14 (M+, 100), 141.11 (5), 140.14 (6),

131.12 (17), 130.12 (62), 114.11 (7), 103.10 (16), 77.05 (12).

2,3,6,7,10,11-Hexakis(pentyloxy)triphenylene 205.

OC5H11 OC5H11

OC5H11 H11C5O

OC5H11 OC5H11

Two-neck 250 ml round bottom flask with a stir bar was charged with 1,2- bis(pentyloxy)benzene (F.W. 250.38, 17.0 g, 67.9 mmol) and anhydrous dichloromethane

(150 ml). The flask was fitted with a thermometer/nitrogen inlet and vanadium oxychloride (27.1 g, 156 mmol) was added dropwise from an addition funnel at such a rate that the temperature of the reaction mixture does not rise above 30-34°C (addition takes approximately 20 min, the color changes to bluish-green). The reaction mixture was

352

stirred for additional 20 min and anhydrous methanol (100 ml) followed by water (250 ml) was added at once. The mixture was extracted with dichloromethane (4×200 ml) and subjected to two consecutive chromatographic separations with hexane:dichloromethane

= 1:1 (first) and 9:1 (second) to afford, after solvent evaporation, 14.4 g (85.4%) of grayish crystals.

The crude product (14.4 g, ~19.3 mmol) was re-alkylated with 1-bromopentane

(F.W. 151.05, 5.8 g, 38 mmol), potassium carbonate (6.0 g, 43 mmol), potassium iodide

(0.1 g, 0.6 mmol), and 2-butanone (150 ml) at 90°C for 12 hrs. The reaction mixture was cooled to room temperature, filtered, solvent evaporated on a rotavap and the residue dissolved in boiling cyclohexane. The warm solution was treated with charcoal (2×15 g), filtered, and applied onto silica gel (50 g). The impregnated material was placed on top of a 1500 ml flash chromatography column, filled with silica gel (25 cm of column height), was eluted with neat hexane (3×500 ml) to remove excess of 1-bromopentane and left under suction for 12 hrs. Then elution was resumed with dichloromethane (1 to 10%) in hexane. First ~10% (pink) of the HAT-5 fraction was discarded, the rest of the fraction was collected, solvents evaporated on a rotavap to allow 8.3 g (49%) of white crystals.

The purified material was recrystallized from 1-propanol/chloroform (residue-free), dissolved in dichloromethane (residue-free), the solution was filtered through 0.45μ syringe filter, solvent evaporated and the residue was recrystallized from cyclohexane (10 ml/g, with charcoal and alumina) to provide 6.04 g (35%) of snow-white crystals, which do not change color when exposed to air and ambient light for two weeks.

353

1-Nitro-2,3,6,7,10,11-hexakis(hexyloxy)triphenyelene MN-HAT-5.

OC H 5 11 OC5H11 OC H 5 11 OC5H11

H11C5O H C O 11 5 O + H C O N - 11 5 H11C5O O OC H 5 11 OC5H11 OC H 5 11 OC5H11

A 50 ml round bottom flask with a stir bar was charged with 2,3,6,7,10,11- hexakis(hexyloxy)triphenyelene (HAT-5, 740 mg, 0.99 mmol), dichloromethane (25 ml), and cooled under nitrogen to –25°C in a dry ice-acetone bath. To this mixture a precooled to –25°C solution of nitric acid (aq., 65.8%, d=1.39227, n=1.40318, 95.1 mg, 0.99 mmol) in nitromethane (2 ml) was added at once with vigorous stirring. The cooling bath was removed and the reaction mixture was allowed to warm up to room temperature. At –

5°C the clear reaction mixture changed color to deep green and then slowly (within 20 min) turned to marsh yellow-green. The rection mixture was allowed to stir for one hour at room temperature, and then applied onto silica gel. Chromatography on silica gel with

1 to 3% ether in distilled hexane allowed, after removal of solvents, 700 mg (89%) of a yellow wax, >95% pure by HPLC (Zorbax NH2 = 3-aminopropyldiethoxysilane ID 4.6 mm; l= 250 mm; 5 μm; eluent i-OtH:CHCl3=70:30 at 1 ml/min, Rt=3.6 min). Hot-stage

13 microscopy: Colh–I 137.3-137.9°C (lit. 141.4°C). C NMR (CDCl3) δ: 150.0, 149.5,

148.7, 148.5, 148.4, 143.3, 139.9, 126.3, 124.3, 123.8, 121.4, 118.6, 113.5, 106.4, 105.6,

75.1, 69.0, 68.9, 68.6, 29.9, 29.3, 29.15, 29.11, 28.54, 28.51, 28.1, 22.78, 22.76, 22.66,

14.2, 14.12, 14.11.

354

1,5,9-Trinitro-2,3,6,7,10,11-hexakis(hexyloxy)triphenyelene TN-HAT-5.

OC H - 5 11 O OC5H11 + OC5H11 N OC H O 5 11 H11C5O H C O 11 5 O + H C O N - 11 5 H11C5O O + OC H - N 5 11 O O OC5H11 OC H 5 11 OC5H11

A 100 ml round bottom flask with a stir bar was charged with 2,3,6,7,10,11- hexakis(hexyloxy)triphenyelene (HAT-5, 4.13 g, 5.5 mmol), dichloromethane (50 ml), and cooled under nitrogen to –10°C in a dry ice-acetone bath. To this mixture an impregnation of nitric acid (aq., 65.8%, d=1.39227, n=1.40318, 2.66 g, 27.7 mmol) on silica gel (15 g) was added at once with vigorous stirring. The cooling bath was removed and the reaction mixture was allowed to warm up to room temperature and stir for 48 hours at room temperature. The solvent was evaporated and the resulting brown impregnation was was placed at the top of a silica gel column and chromatographed with

1 to 7% dichloromethane in distilled hexane. The first bright yellow fraction gave, after removal of solvents, 0.6 g (12%) of a yellow oil, >80% pure by HPLC (Zorbax NH2 = 3- aminopropyldiethoxysilane ID 4.6 mm; l= 250 mm; 5 μm; eluent i-octane:CHCl3=70:30

+ at 1 ml/min, Rt=3.0 min). APCI-MS: 879 (M ). UV-Vis (CH2Cl2) λmax (lg ε): 283 (4.87).

1 H NMR (CDCl3) δ: 7.55 (s, 1H), 4.25 (t, 2H), 4.11 (t, 2H), 1.92 (pentet, 2H), 1.80

13 (pentet, 2H), 1.6-1.4 (m, 8H), 1.02-0.96 (m, 6H). C NMR (CDCl3) δ: 151.9, 143.9,

141.6, 123.0, 114.4, 108.3, 75.4, 69.3, 29.7, 28.7, 28.3, 27.9, 22.4, 14.0, 13.97. Cr–Colh

33.9-34.3°C (microscope hot stage, 0.5°C/min). Colh–I 142°C (DSC, 10°C/min).

CONCLUSIONS

1. We have prepared a series of fluorescent 3,6-diphenyl-2,5-dihydropyrrolo[3,4- c]pyrrole-1,4-dione dyes, useful for the field of single-molecule spectroscopy (SMS) and have applied several of them to the detection and characterization of single biological molecules. The dyes have been shown to posses high quantum yields and useful (up to 84 nm) Stoke’s shifts. Photobleaching measurements have been conducted for several of the synthesized dyes and revealed that the anticipated structure–photostability does not hold true in all instances. Maleimide, hydroxy, halogen and some other functional groups have been incorporated into the DPP bicyclic structure. N,N′-Diarylated DPPs have been prepared by two new methods.

2. A cysteine-specific fluorescent probe has been synthesized and applied to testing of the local polarity and conformational changes in a single-cysteine mutant of GroEL chaperonin of E. coli. The methodology to introduce a maleimide moiety, learned during the synthesis, has been utilized to synthesize other maleimide-containing fluorophores.

3. A series of iodinated aromatic compounds has been prepared and purified to study charge transport in organic molecular crystals. Several iodination approaches have been explored and compared to attain the highest possible purity of the final compounds.

355 356

4. Several polyalkyl- and polyalkoxy- acenes (anthracenes, tetracenes) and some important key intermediates en route to polyalkoxypentacenes have been prepared. None of the synthesized compounds exhibited mesogenic properties.

5. A reliable, highly effective purification protocol has been elaborated for purification of hexapentyloxytriphenylene and its analogs. New trinitro-HAT-5 has been synthesized and characterized as a discotic liquid crystal in the temperature range 34…140 °C.

REFERENCES

1 Chandra, B. P.; Kalia, V; Datt, S. C. Crystalloluminescence: a new tool to determine the critical size of a crystal nucleus. J. Phys. D: Appl. Phys. 1985, 18, L189–L193. 2 Moerner, W. E.; Kador, L. Phys. Rev. Lett. 1989, 62, 2535–2538. 3 Moerner, W. E. A Dozen Years of Single-Molecule Spectroscopy in Physics, Chemistry, and Biophysics. J. Phys. Chem. B 2002, 106, 910–927. 4 a) Itano, W. M.; Bergquist, J. C.; Wineland, D. J. Science 1987, 237, 612; b) Dehmelt, H; Paul W., Ramsey, N. F. Rev. Mod. Phys. 1990, 62, 525. 5 Jabłoński, A. Über den Mechanisms des Photolumineszenz von Farbstoffphosphoren. Z. Phys. 1935, 94, 38–46. 6 Moerner, W. E.; Fromm, D. P. Methods of Single-Molecule Fluorescence Spectroscopy and Microscopy. Rev. Sci. Instrum. 2003, 74, 3597–3619. 7 Haw Y.; Guobin L.; Pallop K.; Tai-Man L.; Ivan R.; Sergio C.; Luying X.; X. Sunney, X. Protein Conformational Dynamics Probed by Single-Molecule Electron Transfer. Science 2003, 302, 262–266. 8 Tinnefeld, P.; Sauer, M. Branching Out of Single-Molecule Fluorescence Spectroscopy: Challenges for Chemistry and Influence on Biology Angew. Chem. Int. Ed. (Eng.) 2005, 44 (18), 2642–2671. 9 Arnaud, C. H. The $1,000 Genome Chem. Eng. News 2005, 83 (51), 60–61. 10 Förster, T. Ann. Phys. 1948, 2, 55–75. 11 Tuschl. T.; Gohlke, C.; Jovin, T. M.; Westhof, E.; Eckstein, F. Science 1994, 266, 785– 789.

357 358

12 Rigler, R.; Mets, Ü.; Widengren, J.; Kask, P. Eur. Biophys. J. Biophys. Lett. 1993, 22, 169–175. 13 Cohen, A. E.; Moerner, W. E. Method for trapping and manipulating nanoscale objects in solution. Appl. Phys. Lett. 2005, 86 (9), 0931091–0931093. 14 Denk, W.; Strickler, J. H.; Webb, W. W. Science 1990, 248, 73–76. 15 Willets, K. A.; Nishimura, S. Y.; Schuck, P. J.; Twieg, R. J.; Moerner, W. E. Nonlinear Optical Chromophores as Nanoscale Emitters for Single-Molecule Spectroscopy. Accounts of Chemical Research 2005, 38 (7), 549–556. 16 Schuck, P. J.; Willets, K. A.; Fromm, D. P.; Twieg, R. J.; Moerner, W. E.. A novel fluorophore for two-photon-excited single-molecule fluorescence. Chemical Physics 2005, 318 (1–2), 7–11. 17 Heinze, K. G.; Rarbach, M.; Jahnz, M.; Schwille, P. Biophys. J. 2002, 83, 1671–1681. 18 Moerner, W. E.; Ambrose, W. P. Phys. Rev. Lett. 1991, 66, 1376. 19 Ormo, M., et al., Science 1996, 273, 1392-5. 20 Xue, Q.; Yeung, E. S. Nature 1995, 373, 681. 21 Craig, D. B.; Arriaga, E. A.; Wong, J. C. Y.; Lu, H.; Dovichi, N. J. J. Am. Chem. Soc. 1996, 118, 5245. 22 Lu, H. P.; Xun, L.; Xie, X. S. Science 1998, 282, 1877. 23 Tsien, R. Y. Rev. Biochem. 1998, 67, 509. 24 Single Molecule Optical Detection, Imaging, and Spectroscopy, Ed. T. Basche, W. E. Moerner, M. Orrit, and U. P. Wild, Verlag Chemie, Munich, 1997. 25 Seisenberger, G.; Ried, M. U.; Endreß, T.; Büning, H.; Hallek, M.; Bräuchle, C. Science 2001, 294, 1929–1932. 26 Nano Lett. 2006, 6, 45. 27 Kapanidis, A. N.; Weiss, S. Fluorescent probes and bioconjugation chemistries for single-molecule fluorescence analysis of biomolecules. J. Chem. Phys. 2002, 117, 10953- 10964.

359

28 Weiss, S. Fluorescence Spectroscopy of Single Biomolecules. Science 1999, 283, 1676-1683. 29 Wilkinson, F.; McGarvey, D. J.; Olea, O.F. J.Phys.Chem. 1994, 98, 3762. 30 Hubner, C. G.; Renn, A.; Renge, I.; Wild, U.P. J. Chem. Phys. 2001, 115, 9619. 31 Single-Molecule Optical Detection, Imaging and Spectroscopy Basché, T.; Moerner, W. E.; Orrit, M; Wild, U. P. Eds.; VCH Verlaggesellschaft: 1996; p 10. 32 Zarrin , F.; Dovichi, N. J. Anal. Chem. 1985, 57, 2690–2692. 33 Enderlein, J. Recent advances in single molecule fluorescence spectroscopy. Reviews in Fluorescence. Kluwer Academic Publishers/Plenum Publishing: New York, NY, 2004, pp 121–163. 34 Miller, L. W.; Cornish, V. W. Selective Chemical Labeling of Proteins in Living Cells. Current Opinion in Chemical Biology 2005, 9, 56–61. 35 Stokes, G. G. On the Change of Refrangibility of Light. Phil. Tans. R. Soc. London 1852, 142, 463–562. 36 Bruches, M.; Moronne, M.; Gin, P.; Weiss, S.; Alivisatos, A. P. Science 1998, 281, 2013–2016. 37 Handbook of Fluorescent Probes and Research Products, Ninth Edition – http://probes.invitrogen.com/handbook/ 38 Bruchez, M. P.; Moronne, M.; Gin, P.; Weiss, S.; Alivisatos, A.P. Science, 1998, 281, 2013. 39 Michalet, X.; Pinaud, F.; Lacoste, T. D.; Dahan, M., Bruchez, M.P.; Alivisatos, A.P.; Weiss, S. Single Molecules 2001, 2, 261. 40 J. Phys. Chem. B 2005, 109, 24220. 41 Lakowicz, J. R. Principles of Fluorescence Spectroscopy 2nd ed. Kluwer Academic Publishers/Plenum Publishing: New York, NY, 1999 42 Farnum, D. G.; Mehta, G.; Moore, G. G. I.; Siegal, F. P. Attempted Reformatskii reaction of benzonitrile. 1,4-Dioxo-3,6-diphenylpyrrolo[3,4-c]pyrrole, a lactam analog of pentalene. Tetrahedron Lett. 1974, 29, 2549–52.

360

43 a) Jost M.; Iqbal A.; Rochat, A. C. 1,4-Diketopyrrolopyrrole Pigments. U.S. Patent 4,791,204, 1988; b) Wooden, G.; Guy de Weck; Wallquist O. Diketopyrrolopyrroles and Pigments Thereof. U.S. Patent 5,424,452, 1995; c) Lamatsch B.; Wallquist O.; Scloeder I. Yellow Diketopyrrolopyrrole Pigments. U.S. Patent 5,672,716, 1997; d) Hendi, S. B. 1,4- Diketo-3,6-diarylpyrrolo[3,4]cpyrrole Pigment Derivatives. U.S. Patent 5,786,487, 1998; e) Wallquist O.; Lamatsch B.; Ruch T. Blue Diketopyrrolopyrrole Pigments. U.S. Patent 5,817,832, 1998. 44 Srivatsavoy, V. J. P.; Eschle, M.; Moser, J.-E.; Grätzel, M. Excited State Electron and Energy Transfer of a Highly Fluorescent Heterocyclic Dye: a Laser Flash Photolysis Study of 2,5-Dimethyl-3,6-diphenylpyrrolo[3,4-c]pyrrole-1,4-dione. J. Chem. Soc. Chem. Commun. 1995, 3, 303–304. 45 Fukuda, M.; Kodama, K.; Yamamoto, H.; Mito, K. Solid State Laser with Newly Synthesized Pigment. Dyes and Pigments 2002, 53, 67–72. 46 Potrawa, T.; Langhals, H. Fluorescent dyes with large Stokes shifts – soluble dihydropyrrolopyrrolediones. Chem. Ber. 1987, 120 (7), 1075–8. 47 Langhals, H.; Grundei, T.; Potrawa, T.; Polborn, K. Highly photostable organic fluorescent pigments – a simple synthesis of N-arylpyrrolopyrrolediones (DPP). Liebigs Ann. Chem. 1996, 5, 679–682. 48 Zambounis, J. S.; Hao, Z.; Iqbal, A. Latent pigments activated by heat. Nature (London) 1997, 388 (6638), 131–132. 49 Hall-Goulle, V. Soluble Chromophores Containing Solubilizing Groups Which Can Be Easily Removed. U.S. Patent 6,063,924, 2000. 50 Hall-Goulle, V., Bize, A. Soluble Chromophores Having Improved Solubilizing Groups U.S. Patent 6,274,728, 2001. 51 Chan, W. K.; Chen, Y.; Peng, Z.; Yu, L. Rational Designs of Multifunctional Polymers. J. Am. Chem. Soc. 1993, 115, 11735–11743.

361

52 Beyerlein, T.; Tieke, B. New photoluminescent conjugated polymers with 1,4-dioxo- 3,6-diphenylpyrrolo[3,4-c]pyrrole (DPP) and 1,4-phenylene units in the main chain. Macromol. Rapid Commun. 2000, 21 (4), 182–189. 53 Heim, I.; Tieke, B.; Lenz, R. et al. Novel Diketopyrrolopyrrole Polymers. Int. Patent WO 2005/049695 A1, 2005. 54 Yu, L.; Chan, W. K.; Peng, Z.; Gharavi, A. Multifunctional Polymers Exhibiting Photorefractive Effects. Acc. Chem. Res. 1996, 29, 13–21. 55 Rochat, A. C.; Wallquist, O.; Iqbal, A.; Mizuguchi, J. Process for the Preparation of Aminated Diketobis (aryl or heteroaryl)pyrrolopyrroles and the Use Thereof as Photoconductive Substances. U.S. Patent 5,973,146, 1999. 56 Mizuguchi, J.; Iqbal, A.; Giller, G. Preparation of Electrochromic Diketopyrroles for Electrochromic Display Devices. Ger. Offen. DE 4435211 A1, 1995-04-27, 1995. 57 Mizuguchi, J. Pyrrolopyrrole pigments and their electronic applications. Shikizai Kyokaishi 1997, 70 (6), 393–403. 58 Heim, I.; Tieke, B.; Lenz, R.; Schmidhalter, B.; Rabindranath, A. R.; Dueggeli, M. Novel diketopyrrolopyrrole polymers for electronic devices. Int. Patent WO 2005/049695, 2005. 59 Hofkens, J.; Verheijen, W.; Shukla, R.; Dehaen, W.; De Schryver, F. C. Detection of a Single Dendrimer Macromolecule with a Fluorescent Dihydropyrrolopyrroledione (DPP) Core Embedded in a Thin Polystyrene Polymer Film. Macromolecules 1998, 31 (14), 4493–4497. 60 Smet, M.; Metten, B.; Dehaen, W. Construction of rod-like diketopyrrolopyrrole oligomers with well-defined length. Tetrahedron Lett. 2001, 42 (37), 6527–6530. 61 Hao, Z.; Iqbal, A.; Tebaldi, N.; Praefke, K. Liquid crystalline diketopyrrolopyrroles. U.S. Patent 5,969,154, 1999. 62 Praefcke, K.; Jachmann, M.; Blunk, D.; Horn, M. Novel family of liquid crystals based on a known biheterocyclic pigment material: mesomorphic derivatives of 2,5- dihydropyrrolo[3,4-c]pyrrole-1,4-dione. Liq. Cryst. 1998, 24 (1), 153–156.

362

63 Wendy V.; Johan H.; Bert M.; Jo V.; Ramesh S.; Mario S.; Wim D.; Yves E.; Frans De Schryver The Photo Physical Properties of Dendrimers Containing 1,4-Dioxo-3,6- Diphenylpyrrolo[3,4-c]pyrrole (DPP) as a Core. Macromol. Chem. Phys. 2005, 206, 25– 32. 64 Eggeling, C.; Widengren, J.; Rigler, R.; Seidel, C. A. M. Photostability of fluorescent dyes for single-molecule spectroscopy: Mechanisms and experimental methods for estimating photobleaching in aqueous solution. In Applied Fluorescence in Chemistry, Biology and Medicine; Rettig, W., Strehmel, B., Schrader, S., Seifert, H., Eds.; Springer- Verlag: New York, 1999; pp 193–240. 65 Hofkens, J.; Verheijen, W.; Shukla, R.; Dehaen, W.; De Schryver, F. C. Detection of a Single Dendrimer Macromolecule with a Fluorescent Dihydropyrrolopyrroledione (DPP) Core Embedded in a Thin Polystyrene Polymer Film. Macromolecules 1998, 31 (14), 4493–4497. 66 Iqbal, A.; Jost, M.; Kirchmayr, R.; Pfenninger, J.; Rochat, A.; Wallquist, O. Synthesis and properties of 1,4-diketopyrrolo[3,4-c]pyrroles. Bull. Soc. Chim. Belg. 1988, 97 (8–9), 615–43. 67 Lenz, R.; Wallquist, O. DPP Chemistry – Continuous Innovation. Surface Coatings International, Part B: Coatings Transactions, 2002, 85 (B1), 19–26. 68 Hao, Z.; Iqbal, A. Some Aspects of Organic Pigments Chem. Soc. Rev. 1997, 26, 203– 213. 69 Iqbal A.; Cassar, L. Process for Dyeing High-Molecular Organic Material, and Novel Polycyclic Pigments U.S. Patent 4,415,685, 1983. 70 Iqbal A.; Cassar, L. 1,4-Diketopyrrolo[3,4-c]pyrroles. U.S. Patent 4,490,542, 1984. 71 Rochat, A.; Cassar, L.; Iqbal, A. Preparation of Pyrrolo[3,4-c]pyrroles. U.S. Patent 4,579,949, 1986. 72 Kaul, B. Solvent-free prodn. and use of pyrrolo[3,4-c]pyrroledione pigments. Int. Patent WO 2004076457, 2004. 73 Sparke, J. M.; Watson, K.D. J. Chem. Soc. Perkin Trans. 1 1976, 5.

363

74 Jost, M.; Iqbal, A.; Rochat, A. C. N-Substituted 1,4-Diketopyrrolo[3,4-c]pyrroles. U.S. Patent 4,585,878, 1986. 75 Jost, M.; Iqbal, A.; Rochat, A. C. Compositions Pigmented with N-Substituted 1,4- Diketopyrrolo[3,4-c]pyrroles. U.S. Patent 4,666,455 , 1987 76 Pfenninger, J.; Iqbal, A.; Rochat, A. C. Pyrrolo[3,4-c]-pyrroles and process for their preparation. Eur. Patent EP0184982, 1986. 77 Pfenninger, J.; Iqbal, A.; Rochat, A. C. Process for the Preparation of Pyrrolo[3,4-c]- pyrroles. U.S. Patent 4,778,899, 1988. 78 Pfenninger, J.; Iqbal, A.; Rochat, A. C.; Wallquist, O. Process for the Preparation of Pyrrolo[3,4-c]-pyrroles and Novel Pyrrolo[3,4-c]-pyrroles. U.S. Patent 4,659,775, 1987. 79 STN substructure search in CA/CAplus/CASREACT databases, performed on February, 7, 2006. 80 MDL Crossfire Commander substructure search in Beilstein database (ver. 2005/03). 81 Morton, Colin J. H.; Gilmour, Ryan; Smith, David M.; Lightfoot, Philip; Slawin, Alexandra M. Z.; MacLean, Elizabeth J. Synthetic studies related to diketopyrrolopyrrole (DPP) pigments. Part 1: The search for alkenyl-DPPs. Unsaturated nitriles in standard DPP syntheses: a novel cyclopenta[c]pyrrolone chromophore. Tetrahedron 2002, 58 (27), 5547–5565. 82 Closs, F.; Gompper, R. 2,5-Diazapentalene. Angew. Chem. 1987, 99 (6), 564–567; Angew. Chem., Int. Ed. Engl. 1987, 26 (6), 552–3. 83 Knorr, L.; Scheidt, M. Ber. Dtsch. Chem. Ges. 1894, 27, 1167–1168. 84 a) Pfleger, R.; Reinhardt, F. Chem. Ber. 1957, 90, 2404–2411; b) Wu, A.; Zhao, Y.; Chen, N.; Pan, X. A Modification of the Knorr Oxidative Coupling Method for Preparation of 1,4-Diketones. Synth. Commun. 1997, 331; c) J. Chem. Soc. Perkin 1, 1980, 2670; d) Pelter, A.; Ward, R. S.; Watson, D. J.; Jack, I. R. Synthesis and NMR Spectra of 2,6- and 2,4-Diaryl-3,7-dioxabicyclo[3.3.0]octanes J. Chem. Soc. Perkin 1, 1982, 183–190; e) Synth. Commun. 1997, 2087; f) Tetr. Lett. 1978, 1509; g) J. Am.

364

Chem. Soc., 1964, 2392; h) Thorp, L.; Brunskill, E. R. J. Am. Chem. Soc., 1915, 37, 1258. 85 a) Knorr, L.; Scmidt, J. Liebigs Ann. Chem. 1896, 293, 111; b) Lohaus, H. Liebigs Ann. Chem. 1934, 509, 130. 86 a) Bromme, E.; Claisen, L. Chem. Ber. 1888, 21, 1134; b) Scmidt, P. F. Chem. Ber. 1895, 28, 1206; c) Keglević, D.; Malnar, M.; Tomljenović, T. Arkiv Chemi, 1954, 26, 67. 87 Rubin, M. B.; Bargurie, M.; Kaftory, M.; Kosti, S. Synthesis and reactions of 1,6- diaryl-2,5-bis(diazo)-1,3,4,6-tetraoxohexanes J. Chem. Soc., Perkin Trans. 1 1980 (12), 2670–2677. 88 Jost, M.; Iqbal, A.; Rochat, A. C. N-Substituted 1,4-Diketo-3,6-diarylpyrolo[3,4- c]pyrroles. U.S. Patent 4,585,878, 1986. 89 Jost, M.; Iqbal, A.; Rochat, A. C. Compositions Pigmented with N-Substituted 1,4- Diketo-3,6-diarylpyrolo[3,4-c]pyrroles. U.S. Patent 4,666,455, 1987. 90 Zambounis, J. S.; Hao, Z.; Iqbal, A. Pyrrolo[3,4-c]pyrroles. U.S. Patent 5,484,943, 1996 91 Zambounis, J. S.; Hao, Z.; Iqbal, A. Pyrrolo[3,4-c]pyrrole synthesis. U.S. Patent 5,616,725, 1997. 92 Hall-Goulle, V. Soluble chromophores containing solubilising groups which can be easily removed U.S. Patent 6,063,924, 2000. 93 Hall-Goulle, V.; Bize, A. Soluble chromophores having improved solubilizing groups. U.S. Patent 6,274,728, 2001. 94 Mizuguchi, J. A pigment precursor based on 1,4-diketo-3,6-diphenyl-pyrrolo-[3,4-c]- pyrrole and its regeneration into the pigment. Journal of Imaging Science and Technology 2005, 49 (1), 35–40. 95 Hendi, S. B. 1,4-Diketo-3,6-diarylpyrolo[3,4-c]pyrroles. U.S. Patent 5,785,750, 1998. 96 Hendi, S. B. Process for Preparing Diketopyrrolopyrrole Derivatives. U.S. Patent 5,840,907, 1998.

365

97 Hendi, S. B. Process for Preparing Diketopyrrolopyrrole Derivatives. U.S. Patent 5,786,487, 1998. 98 Hendi, S. B. U.S. Process for Preparing Diketopyrrolopyrrole Derivatives. Patent 5,919,945, 1999. 99 Buckles, R. E.; Hausman, E. A.; Wheeler, N. G. The Action of Bromine Vapor on Solid Aromatic Compounds J. Am. Chem. Soc. 1950, 72, 2494. 100 Buckles, R. E.; Wheeler, N. G. 4,4′-Dibromobiphenyl. Org. Synth. Coll. Vol. 4, 256. 101 Wallquist, O.; Iqbal, A.; Pfenninger, J.; Rochat, A. Process for the Preparation of Brominated Pyrrolo[3,4-c]pyrroles and Mixtures Thereof. U.S. Patent 4,810,802, 1989. 102 Rochat, A. C.; Iqbal, A.; Ettingen; Jeanneret, R.; Mizuguchi, J. Dithioketo Pyrrolopyrroles U.S. Patent 4,760,151, 1988. 103 Rochat, A. C.; Iqbal, A.; Wallquist, O. Monoketopyrrolopyrrole. U.S. Patent 5,017,706, 1991. 104 Zambounis, J. S.; Hao, Z.; Iqbal, A. Pyrrolo[3,4-cipyrroles containing cyanimino groups. U.S. Patent 5,527,922, 1996. 105 Wallquist, O.; Lamatsch, B.; Ruch, T. Blue diketopyrrolopyrrole pigments. U.S. Patent 5,817,832, 1998. 106 Rochat, A. C.; Wallquist, O.; Iqbal, A.; Mizuguchi, J. Process for the preparation of aminated diketobis (aryl or heteroaryl)pyrrolopyrroles and the use thereof as photoconductive substances U.S. Patent 5,973,146, 1999. 107 Lamatsch, B.; Wallquist, O. Process for the preparation of diketopyrrolopyrrolecarboxylic acids and their esters and amides. U.S. Patent 5,874,588, 1999. 108 Mizuguchi, J.; Grubenmann, A.; Gary W.; Rihs, J. Structures of 3,6- Diphenylpyrrolo[3,4-clpyrrole-l,4-dione and 2,5-Dimethyl-3,6-diphenylpyrrolo[3,4- c]pyrrole-l,4-dione. Acta Cryst. 1992, B48, 696–700. 109 Bäbler, F. Process for Preparing 1,4-Diketo-3,6-diphenylpyrrolo-[3,4-c]-pyrrole. U.S. Patent 5,347,014, 1994.

366

110 Bäbler, F. Process for the Preparation of Diaryldiketopyrrolopyrrole Pigments. U.S. Patent 5,565,578, 1996. 111 Surber, W.; Iqbal, A.; Stern, C. Process for the preparation of pyrrolo[3,4-c]pyrroles. U.S. Patent 4,931,566, 1990. 112 Mizuguchi, J.; Rihs, G. Electronic spectra of 1,4-diketo-3,6-diphenyl-pyrrolo-[3,4-c]- pyrrole in the solid state. Ber. Bunsen-Gess. Chem. 1992, 96 (4), 597–606. 113 Mizuguchi, J.; Wooden, G. A large bathochromic shift from the solution to the solid state in 1,4-diketo-3,6-diphenyl-pyrrolo-[3,4-c]-pyrrole. Ber. Bunsen-Gess. Chem. 1991, 95 (10), 1264–1274. 114 Riggs, R. L.; Morton, C. J. H.; Slawin, A. M. Z.; Smith, D. M.; Westwood, N. J.; Austen, W. S. D.; Stuart, K. E. Synthetic studies related to diketopyrrolopyrrole (DPP) pigments. Part 3: Synthesis of tri- and tetra-aryl DPPs. Tetrahedron, 2005, 61, 11230– 11243. 115 Brooker, L. G. S.; Van Lare, E. J. Color and constitution of organic dyes. Kirk-Othmer Encyclopedia of Chemical Technology, 2-nd Ed., 1964, 5, 763–88. 116 Dewar, M. J. S. The electronic theory of organic chemistry. Clarendon Press: Oxford, 1949. 117 Dewar, M. J. S.; Dougherty, R. C. The Molecular Orbital Theory of Organic Chemistry. McGraw-Hill: New York, 1969. 118 Frei, U.; Kirchmayr, R. Process for the preparation of tert-alkyl esters. U.S. Patent 4,904,814, 1990. 119 Wang, E. C. Tetrahedron Lett. 1998, 39, 4047–4050. 120 Wallquist, O. Cosmetic formulations comprising diketodiphenyl pyrrolopyrrole pigments. Int. Patent WO 2005039513, 2005. 121 Fukuda, M. Kodama, K.; Yamamoto, H.; Mito, K. Evaluation of new organic pigments as laser-active media for a solid-state dye laser. Dyes and Pigments 2004, 63 (2), 115–125

367

122 Enokida, T.; Tamano, M. Organic electroluminescent device and pyrrolo[3,4-c]pyrrol- based electron-transporting material for it. Patent JP 09003448, 1997. 123 Waldron, W. R.; Franklin, R. C. Process for preparing aromatic sulfonic esters of branched chain aliphatic alcohols. U.S. Patent 2,728,788, 1953. 124 Zambounis, J.; Bize, A. Process for producing N-methylated organic pigments. Int. Patent WO 9608537, 1996. 125 Kitao, T.; Yoshida, O.; Kaieda, O.; Shimoyama, F.. Pyrrolopyrrole derivatives as petroleum product identifying agents and method of adding the agents. Patent JP 02216457, 1990. 126 Goldfinger, M. B.; Crawford, K. B.; Swager, T. M. Directed Electrophilic Cyclizations: Efficient Methodology for the Synthesis of Fused Polycyclic Aromatics J. Am. Chem. Soc. 1997, 119, 4578. 127 Mallory, F. B.; Mallory, C. W.; Butler, K. E.; Lewis, M. B.; Xia, A. Q.; Luzik, E. D., Jr.; Fredenburgh, L. E.; Ramanjulu, M. M.; Van, Q. N.; Francl, M. M.; Freed, D. A.; Wray, C. C.; Hann, C.; Nerz-Stormes, M.; Carroll, P. J.; Chirlian, L. E. Nuclear Spin- Spin Coupling via Nonbonded Interactions. 8.1 The Distance Dependence of Through- Space Fluorine-Fluorine Coupling J. Am. Chem. Soc. 2000, 122, 4108. 128 Jeffery, T. Palladium-catalysed Vinylation of Organic Halides under Solid-Liquid Phase Transfer Conditions. J. Chem. Soc. Chem. Commun. 1984, 1287. 129 (a) Mignani J. Tetrahedron Lett. 1990, 31 (33), 4743. (b) Littke A. F. J. Am. Chem. Soc. 2000, 122 (17), 4020. 130 a) Momose, T.; Uchida, S.; Imanishi, T. Octahydro-7(1H)-quinolones. VII. A stereoselective synthesis of cis,trans-decahydro-3H,8H-benzo[i,j]quinolizine-3,8-dione. Chem. Pharm. Bull. 1979, 27 (1), 215–21. b) Scammells, P. J.; Baker, S. P.; Belardinelli, L.; Olsson, R. A. Substituted 1,3-Dipropylxanthines as Irreversible Antagonists of A1 Adenosine Receptors. J. Med. Chem. 1994, 37 (17), 2704–12. 131 Lu, Z.; Twieg, R. J. Tet. Lett. 2005, 46, 2997–3001.

368

132 Greene T.; Wuts, P. G. M. Protective Groups in Organic Synthesis. 3-d Ed. Wiley and Sons: New York, 1999; p 173–178. 133 Herzig, J.; Nudelman, A.; Gottlieb, H. E.; Fischer, B. Studies in Sugar Chemistry. 2. A Simple Method for O-Deacylation of Polyacylated Sugars. J. Org. Chem. 1986, 51, 727– 730. 134 Copola, G. M. N-Arylation of Isatins. A Direct Route to N-Arylisatioc Anhydrides. J. Heterocycl. Chem. 1987, 24, 1249–1251. 135 Smith, M. B.; March, J. Aromatic Nucluophilic Substitution. (Chapter 13) In: March’s Advanced Organic Chemistry. Reactions, Mechanisms, and Structure. Wiley and Sons: New York, 2001; pp 850–893. 136 Weber, G. Polarization of the Fluorescence of Macromolecules. Biochem J. 1952, 51, 155, 13136. 137 Bergstroem, F.; Mikhalyov, I.; Haeggloef, P.; Wortmann, R.; Ny, T.; Johansson, L. B. A. Dimers of dipyrrometheneboron difluoride (BODIPY) with light spectroscopic applications in chemistry and biology. J. Am. Chem. Soc. 2002, 124 (2), 196–204. 138 Amat-Guerri, F.; Liras, M.; Carrascoso, M. L.; Sastre, R. Methacrylate-tethered analogs of the laser dye PM567-synthesis, copolymerization with methyl methacrylate and photostability of the copolymers. Photochem. Photobiol. 2003, 77 (6), 577–584 139 Haugland, R. P. Handbook of Fluorescent Probes and Research Products. 9-th Ed. p 3. http://probes.invitrogen.com/handbook/figures/0667.html 140 Nietzki, R.; Becker, V. Oxazine Dyes. Ber. Dtsch. Chem. Ges. 1908, 40, 3397–3400. 141 Kehrmann, F.; Grillet, E.; Borgeaud, P. New syntheses of oxazine dyes. Helvetica Chimica Acta 1926, 9, 866–880. 142 Du, H.; Fuh, R.-C. A.; Li, J.; Corkan, L. A.; Lindsey, J. S. Photochem CAD: a computer-aided design and research tool in photochemistry. Photochemistry and Photobiology 1998, 68 (2), 141–142. 143 Davis, M. M.; Hetzer, H. B. Titrimetric and equilibrium studies using indicators related to Nile Blue A. Anal. Chem. 1966, 38, 451–461.

369

144 Sahyun, M. R. V. Total luminescence spectroscopy in a reverse micellar system. J. Phys. Chem. 1998, 92, 6028–6032. 145 Sackett, D. L.; Wolff, J. Nile red as a polarity-sensitive fluorescent probe of hydrophobic protein surfaces. Anal. Biochem. 1987, 167, 228–234, 146 Golini, C. M.; Williams, B. W.; Foresman, J. B. Further Solvatochromic, Thermochromic, and Theoretical Studies on Nile Red. J. Fluoresc. 1998, 8 (4), 395–403. 147 Briggs, M. S. J.; Bruce, I.; Miller, J. N.; Moody, C. J.; Simmonds, A. C.; Swann, E. Synthesis of functionalized fluorescent dyes and their coupling to amines and amino acids. J. Chem. Soc., Perkin Trans. 1. 1997, (7), 1051–1058. 148 Moerner, W. E.; Twieg, R. J.; Kline, D. W.; He, M. Fluorophore Compounds and Their Use in Biological Systems. U.S. Patent Appl. 20050009109, 2005. 149 Wright, D.; Gubler, U.; Roh, Y.; Moerner, W. E.; He, M.; Twieg, R. J. High- performance photorefractive polymer composite with 2-dicyanomethylen-3-cyano-2,5- dihydrofuran chromophore. Applied Physics Letters 2001, 79 (26), 4274–4276. 150 Willets, K. A.; Ostroverkhova, O.; He, M.; Twieg, R. J.; Moerner, W. E. Novel fluorophores for single-molecule imaging. J. Am. Chem. Soc. 2003, 125 (5), 1174–1175. 151 Willets, K. A.; Callis, P. R.; Moerner, W. E. Experimental and Theoretical Investigations of Environmentally Sensitive Single-Molecule Fluorophores. J Phys. Chem. B 2004, 108 (29), 10465–10473. 152 Willets, K. A.; Twieg, R. J.; Moerner, W. E. Single-Molecule Magic. SPIE’s OE Magazine of Photonics Technologies and Applications 2004, 4 (6), 13–15. 153 (a) Chem. Pharm. Bull. 1977, 25, 1289. (b) Anal. Biochem. 1977, 79, 83. (c) Chem. Pharm. Bull. 1977, 25, 1678. 154 J. Hystochem. Cytochem. 1993, 41, 1413. 155 Haugland, R. P. Handbook of Fluorescent Probes and Research Products. 9-th Ed. p 82. http://probes.invitrogen.com/handbook/sections/0201.html 156 Long, J.; Wang, Y.-M.; Matsuura, T.; Meng, J.-B. J. Heterocycl. Chem. 1999, 36 (4), 895–900. (Condensation of 1,3-dihydroxy-2-carbethoxy-naphthalene [BRN 2217406]

370

with N,N-bis-(2-hydroxyethyl)-4-nitrosoaniline · HCl [BRN 3737456] in ethanol to give 6-carbethoxy-9-bis(2-hydroxyethyl)amino-5H-benzo[a]phenoxazin-5-one [BRN 8364710] in 60% yield.) 157 Bris, M.-T. L. J. Heterocycl. Chem. 1989, 26, 429–433. (Preparation of 9-nitroso- 2,3,6,7-tetrahydro-1H,5H-pyrido[3,2,1-ij]quinolin-8-ol [BRN 4457827] in 60% yield.) 158 (a) J. Heterocycl. Chem. 1985, 1275. (b) Org, Synth. 1943, Coll. Vol. 2, 223. http://www.orgsyn.org/orgsyn/prep.asp?rxntypeid=182&prep=CV2P0223 159 (a) J. Org. Chem. 1961, 26, 19. (b) J. Org. Chem. 1972, 37, 392–393. 160 Lauwers Tetrahedron Lett. 1979, 1801–1804. 161 (a) Germann, F. J. Am. Chem. Soc. 1927, 49, 307–312. (from α-P4 and I2) (b) Tetrahedron Lett. 1979, 1801–1804. (c) Newkome, G. J. Chem. Soc. Chem. Commun.

1975, 885. (from PCl3 and KI in Et2O) 162 J. Med. Chem. 1996, 39, 1700. 163 J. Chem. Res. Synopsis 1998, 272–273. 164 Choi, D. H.; Song, S. Jahng, W. S. Optimal synthetic design of second-order nonlinear optical material with good temporal stability. Mol. Cryst. Liq. Cryst. Sci. Technol. Sect. A. 1996, 280, 17. 165 Zentz, F.; Valla, A.; Guilou, R. L.; Labia, R.; Mathot, A.-G.; Sirot, D. Synthesis and antimicrobial activities of N-substituted imides. Il Farmaco 2002, 57, 421–426. 166 Schwartz, A. L.; Lerner, L.M. J. Org. Chem. 1974, 39, 21–23. 167 (a) Bergson, J. J. Am. Chem. Soc. 1954, 76, 2835–2836. (b) Bergson, J. J. Am. Chem. Soc. 1954, 76, 4060–4069. (c) Bergson, J.; Swidler, R. J. Am. Chem. Soc. 1953, 75, 1721–1726. 168 Walker, M. A. A High Yielding Synthesis of N-Alkylated Maleimides Using a Novel Modification of the Mitsunobu Reaction. J. Org. Chem. 1995, 60, 5352–5355 169 Tetrahedron Lett. 1994, 35 (5), 665–668.

371

170 Aponte, M. A.; Butler, G. B. Copolymers Containing Alternating Sequences of Nucleic Acid-Base Pairs. I. Monomer Synthesis. J. Polym. Sci., Polym. Chem. Ed. 1984, 22, 2841–2858. 171 Narita, M.; Teramoto, T.; Okawara, M. Bull. Chem. Soc. Jpn. 1971, 44, 1084. 172 Rickborn, B. Organic Reactions 1998, 52, 1–393. 173 Prill, E. D. Maleimide and N-substituted derivatives. U.S. Patent 2,524,136, 1950. 174 Clevenger, R. C.; Turnbull, K. D. Synthesis of N-Alkylated Maleimides. Synth. Commun. 2000, 30 (8), 1379–1388. 175 Kwart, H.; Burchuk, I. J. Am. Chem. Soc. 1952, 74, 3094–3097. 176 Kayser M. Can. J. Chem. 1982, 60, 1199–1206. (NMR and m.p.) 177 Zawadowski, T.; Grabowska, M. Roczniki Chemii 1973, 47, 189–191. 178 Zawadowski, T. Acta Poloniae Pharmaceutica 1995, 52, 125–128. 179 Performed with Dr. Zhikuan Lu. 180 Simon, J.; André, J.-J. Molecular Semiconductors. Springer: Berlin, 1985. 181 Chemical Encyclopedia. Eds. Knuniants, I. L.; Zefirof, N. S. Big Russian Encyclopedia: Moscow; Vol. 4, 55 182 Kittel, C. Introduction into Solid State Physics 8 Ed. 2005, 208. 183 Fritzsche, H. J. J. Phys. Chem. Solids 1958, 6, 69. 184 Seeger, K. Semiconductor Physics. 9-th Ed. Springer: Berlin, 2004. 185 The term “organic semiconductor” was used for the first time in 1948: (a) Vartanyan, A. T. Zhur. Phys. Khim. 1948, 22, 769. (b) Eley, D. D. Nature 1948, 162, 819. 186 Heilmeier, G.H.; Warfield, G.; Harrison, S. E. J. Appl. Phys. 1963, 34, 2278. 187 Duke, C.; Schein, L. Phys. Today, 1980, 33, 42. 188 Simon, J.; Bassoul, P. Molecular Semiconductors: Properties and Applications. In: Design of Molecular Materials, Chapter 6; Wiley and Sons: New York, 2000. 189 Schein L.B., Brown D.W. Mobilities In Organic Molecular Crystals. Mol. Cryst. Liq. Cryst., 1982, 87, 1–12.

372

190 Karl, N. Defect Control in Semiconductors, Ed. Sumino, K. Elsevier Scientific: Amsterdam, 1990; p 1725. 191 Warta, W.; Stehle, R.; Karl, N. Appl. Phys. A 1985, 36, 163. 192 Friedel, G. The mesomorphic states of matter. Ann. Phys. 1922, 18, 273–474. 193 Goodby, J. W.; Gray, G. W. Guide to the Nomenclature and Classification of Liquid Crystals. In: Handbook of Liquid Crystals, Vol. 1, Chapter 2; Eds. Demus, D.; Goodby, J. W.; Gray, G. W.; Spiess, H.-W.; Vill, V. Wiley-VCH: Weinheim, 1998. 194 Lehmann, O. Liquid Crystals. J. Pysique 1910, 7, 713–35. 195 Friedel, G.; Grandjean, F. The Anistropic Liquids of Lehmann. Bull. Soc. Franc. Min. 1911, 33, 192–239. 196 Collings, P.J.; Hird, M. Introduction to Liquid Crystals. Taylor & Francis: Bristol, PA, 1997. ISBN 0748406433. 197 de Gennes, P. G.; Prost, J. The Physics of Liquid Crystals. Clarendon Press: Oxford, 1993. ISBN: 0198520247. 198 Wegewijs, B. R.; Siebbeles, L. D. A.; Boden, N.; Bushby, R. J.; Movaghar, B.; Lozman, O. R.; Liu, Q.; Pecchia, A.; Mason, L. A. Charge-carrier mobilities in binary mixtures of discotic triphenylene derivatives as a function of temperature. Phys. Rev. B 2002, 65 (24), 245112/1–245112/8. 199 Bushby, R. J.; Lozman, O. R. Photoconducting liquid crystals. Curr. Opin. Solid State Materials Sci. 2003, 6 (6), 569–578. 200 van de Craats, A. M.; Warman, J. M.; Fechtenkötter, A.; Brand, J. D.; Harbison, M. A.; Müllen, K. Adv. Mater., 1999, 11, 1469. 201 Lever, L.; Kelsall, R.; Bushby, R. A band transport model for highly ordered discotic mesophases. J. Comput. Electronics 2005, 4 (1/2), 101–104. 202 Lever, L. J.; Kelsall, R. W.; Bushby, R. J. Band transport model for discotic liquid crystals. Phys. Rev. B 2005, 72 (3), 035130/1–035130/11. 203 Bushby, R. J.; Boden, O. R. Curr. Opin. Colloid Interface Sci. 2002, 7, 343. 204 Meier, H. Organic Semiconductors. Chemie-Verlag: Weinheim, 1974.

373

205 Burland, D. M. Phys. Rev. Lett. 1974, 33, 833. 206 Sakagami, S.; Nakamizo, M. Mesomorphic properties of 2-(4-n- alkoxybenzylideneamino)anthracenes. Bull. Chem. Soc. Jpn. 1977, 50 (4), 1009–1010. 207 Mery, S.; Haristoy, D.; Nicoud, J.-F.; Guillon, D.; Monobe, H.; Shimizu, Y. Liquid crystals containing a 2,6-disubstituted anthracene core — mesomorphism, charge transport and photochemical properties. J. Mater. Chem. 2003, 13 (7), 1622–1630. 208 Gimenez, R.; Pinol, M.; Serrano, J. L. Luminescent Liquid Crystals Derived from 9,10-Bis(Phenylethynyl)anthracene. Chem. Mater. 2004, 16 (7), 1377–1383. 209 Hanna, J.; Kogo, K.; Kafuku, K. Fluorescent liquid crystalline charge transfer materials. Eur. Pat. Appl. 1999, 67 pp. EP 915144 A1. 210 Substituted anthracenes for liquid crystal, light-emitting or semiconducting materials and devices. Goulding, Mark John; Thompson, Marcus; Duffy, Warren; Heeney, Martin; Mcculloch, Iain. (Merck Patent GmbH, Germany). PCT Int. Appl. (2005), 70 pp. WO 2004-EP7089 211 Norvez, S.; Tournilhac, F.-G.; Bassoul, P.; Herson, P. Mesomorphism and Polar Distortion in 1,4,5,8-Tetrasubstituted Anthraquinones and Anthracenes. Chem. Mater. 2001, 13, 2552–2561. 212 Norvez, S. J. Org. Chem. 1993, 2414–2418. 213 Billard, J.; Dubois, J. C.; Vaucher, C.; Levelut, A. M. Structures of the two discophases of rufigallol hexa-n-octanoate. Mol. Cryst. Liq. Cryst. 1981, 66 (1-4), 435– 442. 214 Carfagna, C.; Roviello, A.; Sirigu, A. Disk-like mesogens: synthesis and characterization of a series of rufigallol hexa-n-alkanoates. Mol. Cryst. Liq. Cryst. 1985, 122 (1-4), 151–160. 215 Carfagna, C.; Iannelli, P.; Roviello, A.; Sirigu, A. Discotic mesomorphism of rufigallol hexa-n-alkoxylates. Liq. Cryst. 1987, 2 (5), 611–616. 216 Billard, J.; Luz, Z.; Poupko, R.; Zimmermann, H. The mesophases of octa- alkanoyloxy-9,10-anthraquinone. Liq. Cryst. 1994, 16 (2), 333–342.

374

217 Bender, D.; Muellen, K.. Novel alkylanthracenes synthesis, reductive alkylation, and reductive polymerization. Chem. Ber. 1988, 121 (6), 1187–1197. 218 Pozzo, J.-L.; Clavier, G. M.; Colomes, M.; Bouas-Laurent, H. Different Synthetic Routes towards Efficient Organogelators: 2,3-Substituted Anthracenes. Tetrahedron 1997, 53 (18), 6377–6390. 219 Haworth, R. D.; Mavin, C. R. The Structure of Diisoeugenol. J. Chem. Soc. 1931, 1363–1366. 220 Robinson, G. M. A Reaction of Homopiperonyl and of Homoveratryl Alcohols. J. Chem. Soc. 1915, 267–276. 221 (a) Johnson, J. R.; Jobling, W. H.; Bodamer, G. W. J. Am. Chem. Soc. 1941, 63, 131– 135. (b) McDonald. E.; Suksamarn, A.; Wylie, R. D. J. Chem. Soc. Perkin Trans. 1 1979, 1893–1900. 222 (a) Harig, M.; Neumann, B. Eur. J. Org. Chem. 2004, 11, 2381–2397. (b) Leblois, D.; Danielle, P.; Piessard, S.; Baut, G. L.; Kumar, P.; Brion J.-D. Eur. J. Med. Chem. Chim. Ther. 1987, 22, 229. (c) Maekawa, N. Bull. Chem. Soc. Jpn. 1959, 32, 1311–1315. (d) Perkin J. Chem. Soc. 1902, 81, 1032. (e) Perkin, W. H.; Robinson G. M. J. Chem. Soc. 1910, 97, 1139. 223 Hanson, L. Acta Chem. Scand. 1989, 43, 304–306. 224 Mono-adduct of such reaction is known: Anderson, D. R.; Koch, T. H. J. Org. Chem. 1978, 43, 2726–2728. 225 Vanzetti, B. L.; Oliverio, A. Some derivatives of veratrole and of methylvanillin. II. 2,3,6,7-Tetramethoxyanthraquinone. Gazz. Chim. Ital. 1930, 60, 620–632. 226 Shklyaev, Y. V.; Nifontov, Y. V. Three-component synthesis of 3,4-dihydroiso- quinoline derivatives. Russ. Chem. Bull., Int. Ed. 2002, 51 (5), 844–849. 227 Miao, Q.; Nguyen, T.-Q.; Someya, T.; Blanchet, G. B.; Nuckolls, C.; J. Am. Chem. Soc. 2003, 125 (34), 10284–10287. 228 (a) Criegee, R. Oxidation with quadrivalent lead salts. Justus Liebigs Ann. Chem. 1930, 481, 263–286. (b) Criegee, R. Chem. Ber. 1931, 64B, 260–266.

375

229 Kitamura, M.; Shen, B.; Liu, Y.; Zheng, H.; Takahashi, T. Aromatization of Highly Alkyl-substituted Dihydroanthracenes Using n-BuLi/TMEDA/MeI. Chem. Lett. 2001, 646–647. 230 (a) Arcoleo, A.; Paternostro, M. P. Condensation of phenyl ethers with aliphatic aldehydes. XXI. Reaction of veratrole with monobromo- and tribromoacetaldehydes. Ann. Chim. (Rome) 1968, 58 (3), 290-7. (b) Arcoleo, A.; Natoli, M. C. Condensation of phenolic ethers with aliphatic aldehydes. XIX. Structure of a compound derived from the condensation of veratrole with acetaldehyde. Ann. Chim. (Rome) 1967, 57 (6), 716–722. 231 Chung, Y. J. Org. Chem. 1989, 54 (5), 1018–1032. 232 Boldt, P. Chem. Ber. 1967, 100, 1270–1280. 233 Arcoleo, A. Ann. Chim. (Rome) 1967, 57, 716–720. 234 Huang, W.; Zhou, X.; Kanno, K.-I.; Takahashi, T. Pd-Catalyzed Reactions of o- Diiodoarenes with Alkynes for Aromatic Ring Extension. Org. Lett. 2004, 6 (14), 2429– 2431. 235 Marks, V.; Gottlieb, H. E.; Melman, A.; Byk, G.; Cohen, S.; Biali, S. E. Polyethylated Aromatic Rings: Conformation and Rotational Barriers of 1,2,3,4,5,6,7,8- Octaethylanthracene, 1,2,3,4,6,7,8-Heptaethylfluorene, and 1,2,3,4,5,6,7,8- Octaethylfluorene. J. Org. Chem. 2001, 66 (20), 6711–6718. 236 Zeis, R.; Celine, C.; Siegrist, T.; Schlockermann, C.; Chi, X.; Kloc, C. Field Effect Studies on Rubrene and Impurities of Rubrene. Chem. Mater. 2006, 18, 244–248. 237 Podzorov, V.; Menard, E.; Borissov, A.; Kiryukhin, V.; Rogers, J. A.; Gershenson, M. E. Phys. Rev. Lett. 2004, 93, 086602. 238 (a) Laatsch, H. Liebigs Ann. Chem. 1980, 140. (b) Fieser, L. F.; Tishler, M.; Wendler,

N. L. Extensions of the Vitamin K1 Synthesis. J. Am. Chem. Soc. 1940, 62, 2861–2866./ 239 Wilds, A. L. Reduction with Aluminum Alkoxides. Organic Reactions 1944, 178– 223. 240 Hinshaw, J. C. Convenient synthesis of 2,3,6,7-tetramethylanthracene. Org. Prep. Proc. Int. 1972, 4 (5), 211–213.

376

241 Coffey, S.; Boyd, V. The reduction of anthraquinone and other polycyclic quinones with aluminum alkoxides (Meerwein-Pondorff reagent). J. Chem. Soc. 1954, 2468–2470. 242 Gaylord, N. G.; Stepan, V. Reduction of methylated and anthraquinones to methylated anthracenes with aluminum tris(cyclohexyl oxide). Coll. Czech. Chem. Commun. 1974, 39 (7), 1700–1710. 243 Afzali, A.; Dimitrakopoulos, C. D.; Breen, T.L. High-Performance, Solution- Processed Organic Thin Film Transistors from a Novel Pentacene Precursor. J. Am. Chem. Soc. 2002, 124, 8812–8813. 244 Nelson, S. F.; Lin, Y. Y.; Gundlach, D. J.; Jackson, T. N. Appl. Phys. Lett. 1998, 72, 1854. 245 Yamabe, T,; Tonala, K.; Oheki, K. Solid State Commun. 1982, 44, 823. 246 Kivelson, S.; Chapman, O. L. Phys. Rev. B. 1983, 28, 7236. 247 Aihara, J.-I. Reduced HOMO-LUMO Gap as an Index of Kinetic Stability for Polycyclic Aromatic Hydrocarbons. J. Phys. Chem. A 1999, 103, 7487–7495. 248 Clar, E. Polycyclic Hydrocarbons Academic Press: London, 1964; Vols. 1, 2. 249 Harvey, R. G. Polycyclic Aromatic Hydrocarbons. Wiley-VCH: New York, 1997. 250 Aihara, J.-I. Why are some polycyclic aromatic hydrocarbons extremely reactive? Phys. Chem. Chem. Phys. 1999, 1, 3193–3197. 251 Marschalk, C. Researches in the acene series (anthracene and its linear benzologs). Bull. Soc. Chim. Fr. 1954, 877–892. 252 Schleyer, P. von R.; Manoharan, M.; Jiao, H.; Stahl, F. The Acenes: Is There a Relationship between Aromatic Stabilization and Reactivity? Org. Lett. 2001, 3 (23), 3643–3646. 253 Berg, O.; Chronister, E. L.; Yamashita, T.; Scott, G. W.; Sweet, R. M.; Calabrese, J. s- Dipentacene: Structure, Spectroscopy, and Temperature- and Pressure-Dependent Photochemistry. J. Phys. Chem. A 1999, 103, 2451–2459. 254 Clar, E. The Aromatic Sextet. Wiley: London, 1972. 255 Herwig, P. T.; Müllen, K. Adv. Mater. 1999, 11 (6), 480–483.

377

256 Ried, W.; Anthöfer, F. Einfache Synthese für Pentacen-6,13-chinon. Angew. Chem. 1953, 601. 257 Mills, W. H.; Mills, M. J. Chem. Soc.1912, 2194. 258 (a) Brukner, V.; Karczag (Wilhelms), A.; Körmendy, K.; Mészáros, M.; Tomasz, J. Einfache und Ausgiebige Synthese des Pentacens. Acta Chim. Acad. Sci. Hung. 1960, 443–448. (b) Brukner, V.; Karczag (Wilhelms), A.; Körmendy, K.; Mészáros, M.; Tomasz, J. Einfache Synthese des Pentacens. Tetrahedron Lett. 1960, (1), 5–6. (c) Brukner, V.; Tomasz, J. Weitere Vereinfachung der Pentacensynthese. Acta Chim. Acad. Sci. Hung. 1961, 405–408. 259 Bigelow, L. A.; Reynolds, H. H. Quinizarin. Org. Syn. Coll. Vol. 1, 476. 260 Marchand, A. P.; Reddy, G. M. Mild and Highly Selective Ultrasound-Promoted Zinc/Acetic Acid Reduction of C=C Bonds in α,β-Unsaturated γ-Dicarbonyl Compounds. Synthesis 1991, 198–200. 261 Blaszczak, L. C.; McMurry, J. E. Reduction of Enedicarbonyl Compounds with Titanous Ion. J. Org. Chem. 1974, 39 (2), 258. 262 (a) Serpaud, B.; Lepage, Y. Condensation of o-dialdehydes with 1,4-diphenols and γ- diketones. Bull. Soc. Chim. Fr. 1977, (5-6, Pt. 2), 539–542. (b) Lepage, L.; Lepage, Y. Synthesis of o-dibenzoyl compounds by unstable diene trapping. C. R. Seances Acad. Sci. C (Paris) 1975, 280 (12), 847–849. (c) Verine, A.; Lepage, Y. Synthesis of polycyclic quinones by carbanions. Bull. Soc. Chim. Fr. 1973, (3, Pt. 2), 1154–1159. (d) Lepage, Y.; Verine, A. Synthesis of polycyclic quinones through carbanions. C. R. Seances Acad. Sci. C (Paris) 1972, 274 (17), 1534–1536. 263 Roberson, L.B.; Kowalik, J.; Tolbert, L.M.; Kloc, C.; Zeis, R.; Chi, X.; Fleming, R.; Wilkins, C. Pentacene Disproportionation during Sublimation for Field-Effect Transistors. J. Am. Chem. Soc. 2005, 127, 3069–3075. 264 Campbell, R. B.; Robertson, J. Monteath; Trotter, J. The crystal structure of hexacene, and a revision of the crystallographic data for tetracene and pentacene. Acta Cryst. 1962, 15, 289–290.

378

265 Holmes, D.; Kumaraswamy, S.; Matzger, A. J.; Vollhardt, K. P. C. On the nature of nonplanarity in the [N]phenylenes. Chem. Eur. J. 1999, 11, 3399–3412. 266 Swartz, C. R.; Parkin, S. R.; Bullock, J. E.; Anthony, J. E.; Mayer, A. C.; Malliaras, G. G. Synthesis and Characterization of Electron-Deficient Pentacenes. Org. Lett. 2005, 7 (15), 3163–3166. 267 (a) Cava, M. P. J. Org. Chem. 1980, 516. (b) Cava, M. P.; Deana, A. A.; Muth, K. Condensed Cyclobutane Aromatic Compounds. VIII. The Mechanism of Formation of 1,2-Dibromobenzocyclobutene; A New Diels-Alder Synthesis. J. Am. Chem. Soc. 1959, 81, 6458–6460. (c) Cava, M. P. J. Am. Chem. Soc. 1958, 6149. (d) Cava, M. P. J. Am. Chem. Soc. 1957, 1701. 268 (a) Martín, N.; Seoane, C.; Hanack, M. Org. Prep. Proced. Int. 1991, 23, 237. 269 Segura, J. L.; Martín, N. o-Quinodimethanes: Efficient Intermediates in Organic Synthesis. Chem. Rev. 1999, 99, 3199–3246. 270 Cambridge Crystallographic Data Centre Publication CCDC-114447, ccdc.cam.ac.uk. 271 Rice, J. E. J. Org. Chem. 1988, 1775–1779. 272 Flood, D. T. Fluorobenzene. Org. Syn. Coll. Vol. 2, 295. 273 Stephenson, E. F. M. o-Xylylene Dibromide. Org. Syn. Coll. Vol. 4, 984. 274 Smith, M. B.; March J. March’s Advanced Organic Chemistry. Fifth Ed. Wiley and Sons: New York, 2001. pp 911–914. 275 Wenner, W. Bis(bromomethyl) compounds. J. Org. Chem. 1952, 523–528. 276 Smith, M. B. Organic Synthesis. 2-nd Ed. McGraw Hill: New York, 2002. pp 204– 206. 277 Farooq, O. Oxidation of aromatic 1,2-dimethanols by activated dimethyl sulfoxide. Synthesis 1994, (10), 1035–1036. 278 Jarvis, W. F.; Dittmer, D.C. J. Org. Chem. 1988, 53, 5750–5756. 279 Hoey, M. D.; Dittmer, D.C. J. Org. Chem. 1991, 56, 1947–1948.

379

280 Performed in collaboration with Dr. Kihong Park, Electronic Materials and Devices Research Center, KIST, P.O. Box 131, Cheongryang, Seoul 130-650, Korea. +82-2-958- 5292. 281 Smith, J. G.; Dibble, P. W.; Sandborn, R. E.; J. Org. Chem. 1986, 51 (20), 3762–3768. 282 (a) Reetz, M. T.; Toellner, K.; Tetrahedron Lett. 1995, 36 (52), 9461–9464. (b) Mikami, K.; Ohmura, H.; Org. Lett. 2002, 4 (20), 3355–3358. 283 LiqCryst Database, ver.4.3. 284 Allen, C. F. H.; Bell, A. Action of Grignard Reagents on Certain Pentacenequinones, 6,13-Diphenylpentacene. J. Am. Chem. Soc. 1942, 64, 1253–1260. 285 Weygand, F.; Kinkel, K. G.; Tietjen, D. The preparation of aromatic o-dialdehydes from o-dialcohols via the cyclic selenous acid esters. Chem. Ber. 1950, 83 (4), 394–399. 286 Bhattacharjee, D.; Popp, F. D. The oxidation of a series of phthalyl alcohols. J. Heterocycl. Chem. 1980, 17 (2), 315–320. 287 Meziane, M. A. A.; Royer, S.; Bazureau, J. P. A practical one-pot synthesis of ethyl isoquinoline-3-carboxylate by domino reactions: a potential entry to constrained nonproteogenic amino acid derivatives. Tetrahedron Lett. 2001, 42 (6), 1017–1020. 288 Harig, M.; Neumann, B.; Stammler, H.-G.; Kuck, D. Eur. J. Org. Chem. 2004, 11, 2381–2397. 289 Meziane, M. A. A. A.; Bazureau, J. P. Molecules 2002, 7 (2), 252–263. 290 Sato,T. J. Chem. Soc. Perkin Trans. 1 1973; 891–895. 291 Abou-Teim, O.; Jansen, R. B.; McOmie, J. F. W.; Perry, D. H. J. Chem. Soc. Perkin Trans. 1 1980, 1841–1846. 292 Snieckus, V. directed Ortho Metallation. Tertiary Amide and O-Carbamate Directors in Synthetic Strategies for Polysubstituted Armatics. Chem. Rev. 1990, 90 (6), 879–933. 293 Narasimhan, N. S.; Mali, R. S. Heteroatom Directed Aromatic Lithiation Reactions for the Synthesis of Condensed Heterocyclic Compounds. In: Topics in Curr. Chem. Springer-Verlag: Berlin, 1987, 138, 63–147.

380

294 Narasimhan, N. S.; Mali, R. S. Synthesis of Heterocyclic Compounds Involving Aromatic Lithiation Reactions in the Key Step. Synthesis 1983, 957–986. 295 Meier, H.; Fetten, M. A new synthetic route to tribenzo[a,e,i][12]annulenes. Tetrahedron Lett. 2000, 41, 1535–1538. 296 Gschwend, H. W.; Rodriguez, H. R. Heteroatom-facilitated lithiations. Organic Reactions 1979, 26, 1–360. 297 Winkle, M. R.; Ronald, R. C. Regioselective metalation reactions of some substituted (methoxymethoxy)arenes. J. Org. Chem. 1982, 47 (11), 2101–2108. 298 Ziegler, F. E.; Fowler, K. W. Substitution reactions of specifically ortho-metalated piperonal cyclohexylimine. J. Org. Chem. 1976, 41 (9), 1564–1566. 299 Comins, D. L.; Brown, J. D. Ortho metalation directed by a-amino alkoxides. J. Org. Chem. 1984, 49 (6), 1078–1083. 300 Napolitano, E.; Giannone, E.; Fiaschi, R.; Marsili, A. Influence of alkoxyalkyl substituents in the regioselective lithiation of the benzene ring. J. Org. Chem. 1983, 48 (21), 3653–3657. 301 Plaumann, H. P.; Keay, B. A.; Rodrigo, R. The regiospecific lithiation of aromatic acetals. Tetrahedron Lett. 1979, 51, 4921–4924. 302 Wakefield, B. J. The Chemistry of Organolithium Compounds. Pergamon: New York, 1974; 335 pp. 303 De Costa, B. R.; Radesca, L.; Di Paolo, L.; Bowen, W. D. Synthesis, characterization, and biological evaluation of a novel class of N-(arylethyl)-N-alkyl-2-(1- pyrrolidinyl)ethylamines: structural requirements and binding affinity at the s receptor. J. Med. Chem. 1992, 35 (1), 38–47. 304 (a) Fornasier, R.; Scrimin, P.; Tecilla, P.; Tonellato, U. Bolaform and classical cationic metallomicelles as catalysts of the cleavage of p-nitrophenyl picolinate. J. Am. Chem. Soc. 1989, 111 (1), 224–229. (b) Hall, L. A. R.; Stephens, V. C.; Burckhalter, J. H. 2-Dimethylaminoethyl chloride hydrochloride. Org Syn. 1951, 31, 37–39 (Coll. Vol. 4, 333).

381

305 Einhorn, J.; Luche, J. L. Ultrasound in organic synthesis. 10. Selective ortho-lithiation of the Bouveault reaction intermediate. Tetrahedron Lett. 1986, 27 (16), 1793–1796. 306 Posner, G. H.; Canella, K. A. Phenoxide-directed ortho lithiation. J. Am. Chem. Soc. 1985, 107 (8), 2571–2573. 307 Lieberman, S. V. Connor, R. p-Nitrobenzaldehyde. Org. Syn. 1943, Coll. Vol. 2, p.441. 308 Bauer, V. J.; Duffy, B. J.; Hoffman, D.; Klioze, S. S.; Kosley, R. W., Jr.; McFadden, A. R.; Martin, L. L.; Ong, H. H.; Geyer, H. M., III. Synthesis of spiro[isobenzofuran- 1(3H),4'-piperidines] as potential central nervous system agents. J. Med. Chem. 1976, 19 (11), 1315–1324. 309 Hartford, W. H.; Darrin, M. The chemistry of chromyl compounds. Chem. Rev. 1958, 2, 1–61. 310 Ataei, S. M. A convenient method for one step oxidative decarboxylation by KMnO4 in non aqueous media. J. Chem. Res. Synopses 2000, (3), 148–149. 311 by Dr. Lionel Sanguinet. 312 Wood, J. H.; Perry, M. A.; Tung, C. C. Bis-chloromethylation of aromatic compounds. J. Am. Chem. Soc. 1950, 72, 2989–2991. 313 Lee, W. Y.; Park, C. H.; Kim, E. H. Orthocyclophanes. 4. Functionalization of p1n]Orthocyclophanes on the Aromatic Rings. J. Org. Chem. 1994, 59, 4495–4500. 314 Martin, N.; Behnisch, R.; Hanack, M. Syntheses and electrochemical properties of tetracyano-p-quinodimethane derivatives containing fused aromatic rings. J. Org. Chem. 1989, 54 (11), 2563–2568. 315 Payne, M. M.; Delcamp, J. H.; Parkin, S. R.; Anthony, J. E. Robust, Soluble Pentacene Ethers. Org. Lett. 2004, 6 (10), 1609–1612. 316 Clar, E.; John, Fr. Polynuclear aromatic hydrocarbons and their derivatives. VII. A new class of deeply colored radical hydrocarbons and the supposed pentacene of E. Philippi. Chem. Ber. 1930, 63B, 2967–2977.

382

317 Sparfel, D.; Gobert, F.; Rigaudy, J. Thermal transformations of meso-acenic photooxides. VI. Pentacenic photooxides. Tetrahedron 1980, 36 (15), 2225–2235. 318 (a) Uno, H.; Yamashita, Y.; Kikuchi, M.; Watanabe, H.; Yamada, H.; Okujima, T.; Ogawa, T.; Ono, N. Photo precursor for pentacene. Tetrahedron Lett. 2005, 46 (12), 1981–1983. (b) Yamada, H.; Yamashita, Y.; Kikuchi, M.; Watanabe, H.; Okujima, T.; Uno, H.; Ogawa, T.; Ohara, K.; Ono, N. Photochemical synthesis of pentacene and its derivatives. Chem. Eur. J. 2005, 11 (21), 6212–6220. 319 Afzali, A.; Dimitrakopoulos, C. D.; Breen, T. L. High-Performance, Solution- Processed Organic Thin Film Transistors from a Novel Pentacene Precursor. J. Am. Chem. Soc. 2002, 124 (30), 8812–8813. 320 Afzali, A.; Kagan, C. R.; Traub, G. P. N-sulfinylcarbamate-pentacene adduct: A novel pentacene precursor soluble in alcohols. Synthetic Metals 2005, 155 (3), 490–494. 321 Mott, N. F.; Davis, E. A. Electronic Processes in Non-Crystalline Materials, 2nd Ed., Clarendon Press: Oxford, 1979. 322 Holstein, T. Studies of Polaron Motion. Annals of Physics 2000, 281 (1/2), 725–773. 323 D. Emin; A.M. Kriman Transient small-polaron hopping motion. Phys. Rev. B, 1986, 34, 7278. 324 Anthony, J. E.; Eaton, D. L.; Parkin, S. R. A Road Map to Stable, Soluble, Easily Crystallizable Pentacene Derivatives. Org. Lett. 2002, 4 (1), 15–18. 325 Emsley, J. The Elements. 2nd Ed. Clarendon Press: Oxford, 1991. 326 Periodic Table v. 2.5. http://nautilus.fis.uc.pt/st2.5/scenes-e/elem/e05392.html 327 Schwartz, L.; Melvin; I.; Henry G.; Hornig, J. F.; Photoconductivity in p- diiodobenzene. Mol. Cryst. 1967, 2 (4), 379–384. 328 Ellman, B. Ab-Initio Study of the Electronic Structure of the Crystalline High- Mobility Organic Semiconductor 1,4-Diiodobenzene. Submitted for publication. 329 Ellman, B.; Nene, S.; Semyonov, A. N.; Twieg, R. J.. High Mobility, Low Dispersion Hole Transport in 1,4-Diiodobenzene. Adv. Mater. 2006, 18, accepted for publication.

383

330 Suzuki, H.; Kondo, A.; Ogawa, T. Preparation of Aromatic Iodides from Bromides via the Reverse Halogen Exchange. Chem. Lett. 1985, 411–412. 331 Suzuki, H.; Kondo, A.; Inouye, M.; Ogawa, T. An alternative synthetic method for polycyclic aromatic iodides. Synthesis 1986, 121–122. 332 Vonk, W. F. M.; Louw, R. Vapor phase chemistry of arenes. V. Vapor phase iodination of benzene derivatives. Formation of aryl iodides from bromides based on halogen exchange with iodobenzene. Rec. Trav. Chim. Pays-Bas 1977, 96 (2), 59–60. 333 Abraham, M. H.; Grellier, P. L.; Priscilla L. Heterolytic cleavage of main group metal-carbon bonds. In: The Chemistry of Metal–Carbon Bond, Eds. Hartley, F. R.; Patai, S. 5 vols. Wiley: New York, 1984–1990; Vol. 2, 1985, p 72. 334 Taylor, E. C.; Kienzle, F.; McKillop, A. 2-Iodo-p-xylene. Org. Syn. 1976, 55, 70–73; Coll. Vol. 6, 709. 335 Takagi, K.; Hayama, N.; Inokawa, S. The in Situ-generated Nickel(0)-catalyzed Reaction of Aryl Halides with Potassium Iodide and Zinc Powder. Bull. Chem. Soc. Jpn. 1980, 53 (12), 3691–3695. 336 Gan, Z.; Roy, R. Sialoside clusters as potential ligands for siglecs (sialoadhesins). Can. J. Chem. 2002, 80 (8), 908–916. (1,3,5-Triiodobenzene from 1,3,5-tribromobenzene via KI/Ni/DMF halogen exchange reaction.) 337 Larock, R. C. Organomercury Compounds in Organic Synthesis. Springer: New York, 1985. 338 Merkushev, E. B. Advances in the synthesis of iodoaromatic compounds. Synthesis 1988, 12, 923–937. 339 Chaikovski, V. K.; Jharlova, T. S.; Filimonov, V.D.; Saryucheva, T.A. Superactive iodination reagent on a base of iodine chloride and silver sulfate. Synthesis 1999, 748. 340 (a) Miller, L. L.; Kujawa, E. P.; Campbell, C. B. Iodination with Electrolytically Generated Iodine(I). J. Am. Chem. Soc. 1970, 92 (9), 2821–2825. (b) Miller, L. L.; Watkins, B. F. Scope and Mechanism of Aromatic Iodination with Electrochemically Generated Iodine(I). J. Am. Chem. Soc. 1976, 98 (6), 1515–1519.

384

341 Lines, R.; Parker, V. D. Electrophilic Aromatic Substitution by Positive Iodine Species. Iodination of Deactivated Aromatic Compounds. Acta Chem. Scand. B 1980, 34 (1), 47–51. 342 Slezak, F. B.; Bluestone, H.; Magee, T. A.; Wotiz, J. H. Preparation of substituted glycolurils and their N-chlorinated derivatives. J. Org. Chem. 1962, 27, 2181–2183. 343 Nematollahi, J.; Ketcham, R. Imidazoimidazoles. I. Reaction of ureas with glyoxal. Tetrahydroimidazo[4,5-d]imidazole-2,5-diones. J. Org. Chem. 1963, 28 (9), 2378–2380. 344 Slezak, F. B.; Hirsch, A.; Rosen, I. Halogenation of glycoluril and diureidopentane. J. Org. Chem. 1960, 25, 660–661. 345 Yagovkin, A. Yu.; Bakibaev, A. A.; Bystritskii, E. I. Successful synthesis of 2,4,6,8- tetraiodo-2,4,6,8-tetraazabicyclo[3.3.0]octane-3,7-dione. Khimiya Geterotsiklicheskikh Soedinenii 1995, (12), 1695–1696. 346 Chaikovski, V. K.; Filimonov, V. D.; Yagovkin, A. Yu.; Ogorodnikov, V. D. 2,4,6,8- Tetraiodo-2,4,6,8-tetraazabicyclo[3.3.0]octane-3,7-dione as a mild and convenient reagent for iodination of aromatic compounds. Russ. Chem. Bull. 2001, 50 (12), 2411– 2415. 347 Chaikovski, V. K.; Filimonov, V. D.; Yagovkin, A. Y.; Kharlova, T. S. 2,4,6,8- Tetraiodoglycoluril in sulfuric acid as a new powerful reagent for iodination of deactivated arenes. Tetrahedron Lett. 2000, 41 (47), 9101–9104. 348 Kajigaeshi, S.; Kakinami, T.; Yamasaki, H.; Fujisaki, S.; Kondo, M.; Okamoto, T. Iodination of Phenols by Use of Benzyltrimethylammonium Dichloroiodate (1–). Chem. Lett. 1987, 2109–2112. 349 Barluenga, J.; Campos, P. J.; Gonzalez, J. M.; Asensio, G. A simple and general route to aryl iodides from arenes. J. Chem. Soc. Perkin Trans. 1 1984, (11), 2623–2624. 350 Barluenga, J.; Rodriguez, M. A.; Campos, P. J. Electrophilic additions of positive iodine to alkynes through an iodonium mechanism. J. Org. Chem. 1990, 55 (10), 3104– 3106.

385

351 Barluenga, J.; Gonzalez, J. M.; Garcia-Martin, M. A.; Campos, P. J.; Asensio, G. Acid-mediated reaction of bis(pyridine)iodonium(I) tetrafluoroborate with aromatic compounds. A selective and general iodination method. J. Org. Chem. 1993, 58 (8), 2058–2060. 352 Merkushev, E. B.; Sedov, A. M.; Simakhina, N. D. Simple method for the synthesis of iodomethylbenzenes. Zhurnal Organicheskoi Khimii 1978, 14 (5), 1115–1116. 353 Britton, D.; Gleason, W. B. Diiododurene: four centrosymmetric molecules in general positions. Acta Cryst. C 2003, 59 (8), o439–o442. 354 Suzuki, H.; Nakamura, K.; Goto, R. The direct iodination of polyalkylbenzenes bearing bulky groups. Bull. Chem. Soc. Jpn. 1966, 39 (1), 128–131. 355 Deacon, G. B.; Farquharson, G. J. Permercurated arenes. III. Syntheses of periodoarenes and perchloroarenes by iodo- and chloro-demercuration of some permercurated arenes. Aust. J. Chem. 1977, 30 (8), 1701–1713. 356 Bugueno-Hoffmann, R. Spectrochim. Acta A 1992, 48 (4), 509–517. 357 Suzuki, H; Goto, R. A convenient synthesis of certain aromatic polyiodo compounds. Bull. Chem. Soc. Jpn. 1963, 36, 389–391. 358 Mattern, D. L. Direct Aromatic Periodination. J. Org. Chem. 1984, 49, 3051–3053. 359 Barluenga, J.; Gonzalez, J. M.; Garcia-Martin, M. A.; Campos, P. J. Polyiodination on benzene at room temperature: a regioselective synthesis of derivatives. Tetrahedron Lett. 1993, 34 (24), 3893–3896. 360 (a) Khotsyanova, T. L.; Smirnova, V. I. Crystal and molecular structures of hexaiodobenzene. Kristallografiya 1968, 13 (5), 787–790. (b) Steer, R J.; Watkins, S. F.; Woodward, P. Crystal and molecular structure of hexaiodobenzene. J. Chem. Soc. C 1970, (2), 403–408. 361 Nakayama, A.; Fujihisa, H.; Takemura, K.; Aoki, K.; Carlon, R. P. Structural study on pressure-induced metallization of C6I6. Synth. Metals 2001, 120 (1-3), 767–768.

386

362 Iwasaki, E.; Shimizu, K.; Amaya, K.; Nakayama, A.; Aoki, K.; Carlon, R. P. Metallization and superconductivity in hexaiodobenzene under high pressure. Synth. Metals 2001, 120 (1-3), 1003-1004. 363 Nakayama, A.; Fujihisa, H.; Aoki, K. Crystal and molecular structures of C6I6 under high pressure. Koatsuryoku no Kagaku to Gijutsu 2000, 10 (3), 214–220. 364 Tateyama, Y.; Ohno, T. First-principles study on molecular dissociation under metallization pressure in aromatic monomolecular crystals with iodine atoms. J. Phys. Condensed Matter 2002, 14 (44), 10429–10432. 365 (a) Satyamurthy, N.; Barrio, J. R. Cation exchange resin (hydrogen form) assisted decomposition of 1-aryl-3,3-dialkyltriazenes. A mild and efficient method for the synthesis of aryl iodides. J. Org. Chem. 1983, 48 (23), 4394. (b) Barrio, J. R.; Satyamurthy, N.; Ku, H.; Phelps, M. E. The acid decomposition of 1-aryl-3,3- dialkyltriazenes. Mechanistic changes as a function of aromatic substitution, nucleophile strength, and solvent. J. Chem. Soc. Chem. Commun. 1983, (8), 443–444. (c) Foster, N. I.; Heindel, N. D.; Burns, H. D.; Muhr, W. Aryl iodides from anilines via triazene intermediates. Synthesis 1980, (7), 572–573. 366 Wheeler, H. L.; Liddle, L. M. Researches on Halogen Amino Acids. Iodine Derivatives of o-Toluidine. 3-Iodoaminobenzoic Acids. American Chemical Journal 1910, 42, 498–505. 367 Gore, R.; Suzuki, H. Jacobsen reaction of polyhalobenzenes and monoaikylpoly- halobenzenes. Nippon Kagaku Zasshi 1963, 84 (2), 167–173. 368 Liu, Y. X.; Knobler, C. B.; Trueblood, K. N.; Helgeson, R. 3,4,5-Triiodotoluene,

C7H5I3. Acta Cryst. C 1985, 41 (6), 959–961. 369 Friedman, L.; Logullo, F. M. Synthesis of o-dihalobenzenes from benzenediazonium- 2-carboxylate. Angew. Chem. 1965, 5 (77), 217–218. 370 Janczewski, M.; Prajer, L. Synthesis and properties of naphthalenesulfonic acids. Replacement of the sulfonic acid group with halogens. Roczniki Chemii 1954, 28, 681– 682.

387

371 Pan, Y.; Peng, Z. A convinient synthetic approach to 1,5-diiodonaphthalene derivatives. Tetrahedron Lett. 2000, 41, 4537–4540. 372 Cakmak, O.; Demitras, I.; Balaydin, H. T. Selective bromination of 1- bromonaphthalene: efficient synthesis of bromonaphthalene derivatives. Tetrahedron 2002, 58, 5603–5609. 373 Kajanus, J.; van Berlekom, S. B.; Albinsson, B.; Mårtensson, J. Synthesis of Bis(phenylethynyl)arylene-Linked Diporphyrins Designed for Studies of Intramolecular Energy Transfer. Synthesis 1999, (7), 1155–1162. 374 Novak, I.; Jiang, H.; Kovac, B. Intramolecular Interactions in Diiodonaphthalenes. J. Phys. Chem. A 2003, 107 (4), 480–484. 375 Hellberg, J.; Allared, F.; Pelcman, M. Facile synthesis of 2,3-diiodonaphthalene and 2-bromo-3-iodonaphthalene. Synth. Commun. 2003, 33 (15), 2751–2756. 376 Seeboth, H. Bucherer reaction and the preparative use of its intermediate products. Angew. Chem., Int. Ed. Eng. 1967, 6 (4), 307–317. 377 Boden, N.; Bushby, R. J.; Clements, J. Mechanism of quasi-one-dimensional electronic conductivity in discotic liquid crystals. J. Chem. Phys. 1993, 98 (7), 5920– 5931. 378 (a) Adam, D.; Haarer, D.; Closs, F.; Frey, T.; Funhoff, D.; Siemensmeyer, K.; Schuhmacher, P.; Ringsdorf, H. Discotic liquid crystals — A new class of fast photoconductors. Ber. Bunsen-Ges. 1993, 97 (10), 1366–1370. (b) Adam, D.; Closs, F.; Frey, T.; Funhoff, D.; Haarer, D.; Ringsdorf, H.; Schuhmacher, P.; Siemensmeyer, K. Transient photoconductivity in a discotic liquid crystal. Phys. Rev. Lett. 1993, 70 (4), 457–460. 379 Marquardt, F. H. 2,3,6,7,10,11-Hexamethoxytriphenylene. J. Chem. Soc. 1965, 1517– 1518. 380 Matheson, I. M.; Musgrave, O. C.; Webster, C. J. Oxidation of veratrole by quinones. Chem. Commun. 1965, (13), 278–279.

388

381 Boden, N.; Borner, R. C.; Bushby, R. J.; Cammidge, A. N.; Jesudason, M. V. The synthesis of triphenylene-based discotic mesogens: new and improved routes. Liq. Cryst. 1993, 15 (6), 851–858. 382 Cooke, G.; Sage, V.; Richomme, T. Synthesis of Hexa-alkyloxytriphenylenes Using

FeCl3 Supported on Alumina. Synth. Commun. 1999, 29 (10), 1767–1771. 383 (a) Kumar, S.; Manickam, M. Oxidative trimerization of o-dialkoxybenzenes to hexaalkoxytriphenylenes: molibdenum (V) chloride as novel reagent. Chem. Commun.

1997, 1615–1616. (b) Waldvogel, S. R. The Reaction Pattern of the MoCl5-Mediated Oxidative Aryl-aryl Coupling. Synlett 2002, (4), 622–624. 384 Kumar, S.; Varshney, S. K. Vanadium oxychloride, a novel reagent for the oxidative trimerization of o-dialkoxybenzenes to hexaalkoxytriphenylenes. Liq. Cryst. 1999, 26 (12), 1841–1843. 385 Kumar, S.; Varshney, S. K. Synthesis of Triphenylene and Dibenzopyrene Derivatives: Vanadium Oxychloride a Novel Reagent. Synthesis 2001, (2), 305–311. 386 Chapuzet, J.-M.; Simonet-Gueguen, N. A versatile anodic source of triphenylenes possessing at least one ionophoric site. Tetrahedron Lett. 1991, 32 (50), 7405–7408. 387 Chapuzet, J.-M.; Simonet, J. The anodic trimerization of aromatic orthodiethers: new development. Tetrahedron 1991, 47 (4-5), 791–798. 388 Dietrich, M.; Heinze, J. On the determination of redox potentials of highly reactive aromatic mono- and multications. J. Am. Chem. Soc. 1990, 112 (13), 5142–5145. 389 Bechgaard, K.; Parker, V. D. Mono-, di-, and trications of hexamethoxytriphenylene. Novel anodic trimerization. J. Am. Chem. Soc. 1972, 94 (13), 4749–4750. 390 Ronlan, A.; Aalstad, B.; Parker, V. D. Unsymmetrical anodic coupling of veratrole with various anisole derivatives. Products and mechanisms. Acta Chem. Scand. B 1982, 36 (5), 317–325. 391 Le Berre, V.; Angely, L.; Simonet-Gueguen, N.; Simonet, J. Anodic trimerization. A facile one-step synthesis of tris(15-crown-5)triphenylene. J. Chem. Soc. Chem. Commun. 1987, (13), 984–986.

389

392 Private communication from Dr. Owen Roger Lozman , currently at “Avecia Inkjet Limited”, UK. 393 Bushby, R. J.; Boden, N.; Kilner, C. A.; Lozman, O. R.; Lu, Z.; Liu, Q.; Thornton- Pett, M. A. Helical geometry and liquid crystalline properties of 2,3,6,7,10,11- hexaalkoxy-1-nitrotriphenylenes. J. Mater. Chem. 2003, 13 (3), 470–474. 394 Kumar, S.; Manickam, M.; Varshney, S. K.; Rao, D. S. S.; Prasad, S. K. Novel heptasubstituted discotic liquid crystals. J. Mater. Chem. 2000, 10, 2483–2489. 395 Demas, J. N.; Crosby, G. A. The Measurement of Photoluminescence Quantum Yields. A Review. J. Phys. Chem. 1971, 75 (8), 991–1024. 396 Parker, C. A.; Rees, W. T. Correction of fluorescence spectra and measurement of fluorescence quantum efficiency. Analyst (London) 1960, 85, 587–600. 397 Optical Radiation Measurements. Volume 3. Measurement of Photoluminescence. Ed. Mielentz, K. D. 398 Standards in Fluorescence Spectroscopy. Ed. Miller, J. N. QD459.S72x 399 Eaton, D. F. Reference Materials for Fluorescent Measurement. Pure and Appl. Chem. 1988, 60 (7), 1107–1114. 400 Madge, D.; Wong, R.; Seybold, P. G. Fluorescence Quantum Yields and Their Relation to Lifetimes of Rhodamine 6G and Fluorescein in Nine Solvents: Improved Absolute Standards for Quantum Yields. Photochem. Photobiol. 2002, 75 (4), 327–334. 401 Karstens, T.; Kobs, K. Rhodamine B and Rhodamine 101 as Reference Substances for Fluorescence Quantum Yield Measurement. J.Phys.Chem. 1980, 84, 1871–1872. 402 Langhals, H.; Karolin, J.; Johansson, L. B-Å. Spectroscopic properties of new and convinient standards for measuring fluorescence quantum yields. J. Chem. Soc., Faraday Trans. 1998, 94, 2919–2922. 403 Peterman, E. J. G.; Brasselet, S.; M erner, W. E. The Fluorescence Dynamics of Single Molecules of Green Fluorescent Protein. J. Phys. Chem. A 1999, 103, 10553- 10560. 404 Dr. Katherine A. Wilets, private communication.

390

405 Cournoyer, M. E.; Dare, J. H. The use of alternatie solvent purification techniques. Chemical Health and Safety 2003, 10 (4), 15–18. 406 Vogel, A. I. J. Chem. Soc. 1948, 631. 407 Bowman, N. S. J. Am. Chem. Soc. 1965, 87, 4477–4481. 408 Azeotropic Data. Volume III. Advances in Chemistry Series. 1973, 116. QD1.A355no116 — includes all previous data from volumes 6 and 35. 409 Frei, U.; Kirchmayr, R. Process for the preparation of tert-alkyl esters. U.S. Patent 4,904,814, 1990 410 Chmyr, I. M.; Bukeikhanov, N. R.; Suvorov, B. V. Synthesis of p-fluorobenzonitrile by the oxidative ammonolysis of p-fluorotoluene on vanadium-titanium oxide catalysts. Izvestiya Akademii Nauk Kazakhskoi SSR, Seriya Khimicheskaya 1977, 27 (1), 40–41. 411 Herbst, R. M.; Wilson, K. R. Apparent acidic dissociation of some 5-aryltetrazoles. 1957, 22, 1142–1145. 412 Gillis, D. J. Org. Chem. 1971, 36, 518. 413 Ibata, T.; Isogami, Y.; Toyoda, Bull. Chem. Soc. Jpn. 1991, 64 (1), 42–49. 414 Beach, S. F.; Hepworth, J. D.; Sawyer, J.; Hallas, G.; Marsden, R. J. Chem. Soc. Perkin Trans. 2 1984; 2, 217–222. 415 Allbright J. Med. Chem. 1983, 26, 1406. 416 Barbero, M.; Degani, I.; Dughera, S.; Fochi, R. Synthesis 2003; 5, 742–750. 417 Hellerman J. Am. Chem. Soc. 1946, 68, 1890–1891. 418 (a) Soula, G. Tris(polyoxaalkyl)amines (trident), a new class of solid-liquid phase- transfer catalysts. J. Org. Chem. 1985, 50 (20), 3717–3721 (b) Loupy, A.; Philippon, N.; Pigeon, P.; Sansoulet, J.; Galons, H. Solid-liquid phase transfer catalysis without solvent: further improvement in SNAr reactions. Synthetic Communications 1990, 20 (18), 2855– 2864. 419 Kim, S. Y.; Semyonov, A. N.; Twieg, R. J.; Horwich, A. L.; Frydman, J.; Moerner, W. E. Probing the Sequence of Conformationally-Induced Polarity Changes in the Molecular

391

Chaperonin GroEL with Fluorescence Spectroscopy J. Phys. Chem. 2005, 109 (51), 24517-24525. 420 Kalgutkar, A. S. J. Med. Chem. 1996, 39, 1692–1703. 421 Tawney, P.O. J. Org. Chem. 1961, 26, 15–21. 422 (a) Beyer, U. Monatsch.Chem. 1997, 128, 91. (b) Zentz F. Il Farmaco 2002, 57, 421– 426. 423 Germann, F. J. Am. Chem. Soc. 1927, 49, 307–312 424 Krojidlo, M.; Barth, T.; Blaha, K.; Jost, K. Coll. Czech. Chem. Commun. 1976, 41 (7), 1954–1958. 425 Boucherle, A.; Carraz, G.; Revol, A. M.; Dodu, J. N-Substituted maleimides. Bull. Soc. Chim. Fr. 1960, 500–503. 426 Choi, D. H.; Song, S.; Jahng, W. S.; Kim, N. Mol. Cryst. Liq. Cryst. Sci. Technol. Sect. A 1996, 280, 17–26. 427 Zawadowski, T.; Grabowska, M.; Zbikowski, B. N-Substituted derivatives of exo-7- oxabicyclo[2.2.1]hept-5-ene-2,3-dicarboximide. Roczniki Chemii 1973, 47 (1), 189–191. 428 Robinson, J. J. Chem. Soc. 1915, 267–276. 429 Arcoleo, A.; Oliverio, A. Ann. Chim (Rome, Italy) 1957, 47, 415–432. 430 Arcoleo, A.; Natoli, M. C.; Marino, M. L. Condensation of phenol ethers with aliphatic aldehydes. XXX. Veratrole and isobutyraldehyde: isolation of a dibenzo[b,f]thiepin S,S-dioxide derivative. Ann. Chim (Rome, Italy) 1970, 60 (4), 323– 334. 431 Coltart, D. M.; Charlton, J. L. Can. J. Chem. 1996, 74, 88–94. 432 Chung, Y. J. Org. Chem. 1989, 54 (5), 1018-1032. 433 Boldt, P. Chem. Ber. 1967, 100, 1270-1280. 434 Lepage Y. Synthesis 1980, 689. 435 Bhattacharjee, D.; Popp, F. D. J. Heterocycl. Chem. 1980; 17, 315–320. 436 McOmie, J. F. W.; Perry, D. H. Synthesis 1973, 416–417.

392

437 Gordon, A. J.; Ford, R. A. The Chemist’s Companion: : a handbook of practical data, techniques, and references. Wiley: New York, 1972, 537 pp. 438 Mills, W.H.; Mills, M. J. Chem. Soc. 1912, 101, 2194. 439 Hua, D. H.; Tamura, M.; Huang, X.; Stephany, H. A.; Helfrich, B. A.; Perchellet, E. M.; Sperfslage, B. J.; Perchellet, J.-P.; Jiang, S.; Kyle, D. E.; Chiang, P. K.J. Org. Chem. 2002, 67 (9), 2907–2912. 440 Dufraisse, C.; Rigaudy, J.; Basselier, J. J.; Nguyen-Kim-Cuong. „Abnormal” fixation of O in three 1,4-dimethoxyanthracene photooxides, dissociable in the cold. Luminescence during the dissociation of the photooxides. Compt. Rend. 1965, 260 (19), 5031–5036. 441 Newman, M. S.; Wiseman, E. H. J. Org. Chem. 1961, 26, 3208–3211. 442 Rice, J. E.; Czech, A.; Hussain, N.; La Voie, E. J. J. Org. Chem. 1988, 53, 1775. 443 Mattern, C. J. Org. Chem. 1984, 49, 3051. 444 Jan, C. Can. J. Chem. 2002, 80 (8), 908.

Appendix A

Instructions to run a GC-MS experiment on the Thermo Electron Trace GC 2000/Polaris Q MS.

393 394

The compound being injected into the Trace GC must be proven (by usual GC,

TGA) to be volatile and contain NO non-volatile or thermally decomposable residues.

That is, NO reaction mixtures or crude products may be injected. Spec sensitivity is 10 picogram (!) of decafluorobenzophenone. Both volume and amount/concentration of the sample matter, for the injector liner cannot accept more vapor volume than equivalent to

3 μl of liquid. If you need to inject more than 3 μl, change the injector liner. By default,

the split liner is set up.

Method 1. Prepare Solution #1 by dissolving 1 mg in 10 ml. Take an aliquot of 1

ml and dilute to 10 ml of Solution #2. Take an aliquot of 1 ml of Solution #2 and dilute to

10 ml of Solution #3. Inject 1 μl of Solution #3. If you get no signal on GC-gram, inject 1

μl of Solution #2.

Method 2. Prepare Solution A by dissolving 1 mg in 10 ml. Take an aliquot of 1

μl and dilute to 1 ml of Solution B. Inject 1 μl of Solution B. If you get no signal on GC-

gram, take an aliquot of 10 μl of Solution A, dilute to 1 ml of Solution C and inject 1 μl

thereof.

Method 3. Prepare Solution Z by dissolving 1 μg (0.001 mg) in 1 ml. Inject 1 μl

of Solution Z.

Inject no more than 1–100 ng (nanogram) = 10–9 g = 10–6 mg

Inject no more than 1 μl (microliter) = 10–3 ml

1 ng/μl = 1 μg/ml = 1 mg/liter

100 ng/μl = 0.1 mg/ml = 1 mg/10 ml

395

Column max = 350°C ‘Alex-Stand-by.meth’:

Injector nominal = 225°C Injector = OFF

Transfer line nominal = 300°C Gas Saver Flow = 10 ml/min

MS Source nominal = 200°C

RUNNING GC-MS

1. Double click on Xcalibur. This opens the "Roadmap-Home page".

Xcalibur Icon.

2. Click on Instrument Setup icon in the Roadmap-Home Page.

This opens "Untitled-Instrument Setup". File Æ Open Æ Open your method file

(*.meth) from D:\Methods\Your name\

396

Xcalibur Home Page.

Instrument Setup.

397

3. If this is the first run of the day, check tuning. Otherwise, skip this step. To check tuning, click on Tune icon in the "Untitled-Instrument Setup". This opens the "tune window". Check the Foreline pressure at the bottom right in the status bar – it must be in the range of 40…65 mTorr.

Polaris Q MS Tuning.

3a. Click on Experiment in the menu bar of "tune window".

Click on Air Water in the drop down menu.

Click on Instrument in the menu bar of "tune window".

Click on System On in the drop down menu.

398

If the instrument is set for CI mode, click on “red flask” button to

switch on CI reagent gas (or from menu Instrument Æ CI Reagent Gas). A dialog

window appears where the check-box “Reagent Gas On” must be checked and the Flow

set to 1.5 ml/min (range: 0.5 to 2.5 ml/min).

CI Reagent Gas Control.

Click on On green button in the button bar of "tune window"

(or from menu Instrument Æ Filament/Dynode ON).

In EI mode, if the MS is working properly a spectrum with m/z= 14, 16, 18, 29

will appear. The water and oxygen peaks must NOT be the most abundant. (If you

observe high foreline pressure or you think the MS is not working properly and/or the air-

water spectrum does not look right, please let Mahinda Gangoda (2-3843 & 330-687-

4157) know about it.)

399

Click on OFF red button in the button bar of "tune window".

Close the "tune window".

Close "Untitled -Instrument setup" window. The "Roadmap-Home page" is

visible.

4a. If you do NOT use the autosampler.

Click on Sequence Setup icon in the "Roadmap-Home Page". This opens the

"Untitled[Open]-Sequence Setup" window.

Press Ctrl + O to open a sequence.

Double click on the name of the sequence belongs to your research group.

This will bring this sequence to the #1 row of the "Untitled[Open]-Sequence Setup".

Make necessary changes in #1 row. a) Double click File Name cell (Column #1) and in the dialog window type a file name to save your data to. The file name should be at D:\Data\Your_Name directory and end up with a *.raw extension. b) Double click on

Inst Method cell and select method file to run. c) Press Ctrl+S to save the sequence.

Unsaved sequence WILL give you an error message later on.

Click on green icon (7th from right) in the “Sequence Setup” to initialize GC.

A “Run Sequence” window opens. Click on OK in this window. Wait several seconds

until file names appear in the status window on the left and then

400

Sequence Window.

Click green icon (4th from right) in the “Sequence Setup” to initialize MS.

Check on the status window to see “GCQ/Polaris MS” is “Waiting for Contact Closure”

401

Select Rows to Run.

Then wait till the “TRACE GC 2000” turns (status window) to “Ready for Run”

(On GC key pad, the “READY TO INJECT” will turn green)

Inject only 1μL of 10-100 ng/μL (10-100 μg/ml) sample and simultaneously press

“START” (the blue round key on GC key pad) to start the run.

Click on “Real Time Plot View” (3rd icon from left) in the “Sequence Setup” window to

see the chromatogram (TIC).

The Real Time Plot View (Chromatogram) will appear after some delay time preset in the

method file (3-5 min). Wait till the run is completed (about 20 min.)

402

4b. If you DO use the autosampler.

Proceed same as 4a, but click on green icon “Run samples” (8th from right) in the

“Sequence Setup” and DO NOT click green icon “Start Analysis” (4th from right).

DATA PROCESSING

Go to “Roadmap-Home Page” by clicking on the 1st icon from left on the “Real Time

Plot” window.

Click on Qual Browser icon in “Roadmap-Home Page”

Press Ctrl + O to open the window “Open Raw File”

Double click and open your data file.

This will show the chromatogram (top) and a mass spectrum (bottom)

Click on a chromatographic peak while the mass spectral window is active (green pin on

upper right corner) to see the Mass Spectrum for a given peak.

403

Quantitative Browser Window.

STAND-BY MODE

From “Instrument setup” window open method “Alex-Stand-by.meth.

Click on the pictogram of Trace GC 2000 on the left.

Click on GCQ in the menu bar of “Instrument Setup”

This will open a drop down menu.

Click on Send Method to GC

Close Xcalibur software (Do not turn the computer off)

Appendix B

NMR EXPERIMENTS

INSTRUCTIONS and SUMMARY

404 405

Bruker Avance 400 MHz

033 SRL; Phone 2-2677 host: BH041304; MAC: 00-11-0A-00-79-B8; IP: 131.123.233.48

HP XW4100 P4 2.8 GHz Bus 533MHz L1/L2/L3 20/1024/0 kB RAM 512 MB DDR 333

Topspin 1.3 for Win requirements:

CPU > 1GHz; RAM > 512 Mb NTFS partition (for program and data)

Video RAM > 64 Mb (non-shared) NMRData 131.123.235.4:5505

Extra ethernet card D:\NMR\Data\Username\NMR\ExpNo\ProcNo

Run: Topspin 1.3 zz = zg + ftp start acquisition; ej / ij to eject / inject sample tube; tr trace FID to hard drive; rpar mg* read parameters from file; fp Fourier process FID; atma auto tune/wobble; apk auto phase correctiom; lockdisp show lock display; abs auto base line correction; lock lock the solvent; pp peak picking; gradshimau perform auto shimming; xwinplot = plot plot spectrum

GCOSY

Run 1H. Note rg = x value. Rpar mg-H-COSY. Enter manually rg x. atma zg xfb Adjust intensity and save intensity. Plot: chose 2D exp first, then 1D for projections.

HMQC

Run 1H. Note rg = x value. Rpar mg-C13-1H-HMQC. Enter manually rg x. atma (tunes

for both nuclei). ns = 8. zg xfb Adjust intensity and save intensity. If separate

independent 13C is available, may plot it as y projection.

406

Sample Preparation

The 5-mm tube should be filled with a deuterated solvent to a minimum depth of

5.0…5.5 cm (about 0.60…0.75 ml). Lesser depths will make shimming the magnet homogeneity difficult. Greater depths are O.K., except for variable temperature experiments. The amount (concentration) of sample required for a proton spectrum ranges from less than 1 mg/ml to about 20 mg/ml (mw=400). Too much sample can result in a loss of resolution or a distorted spectrum. This includes not just the sample of interest, but any proton source such as protonated buffers, residual protonated solvents, and water. About 5mg/ml is a sufficient maximum concentration for 1H. For 13C the

higher the concentration the better. The solution should be free from any solid, such as

undissolved solute, or dust. Filter the solution through a Pasteur pipette with a tiny cotton

plug, if necessary.

At the Instrument

The Bruker AMX 300 NMR instrument may be controlled and the data may be

processed via: 1) Spectrometer Control Module (SCM) – a keypad with a black rotating

knob on the right to the computer keyboard; 2) typing commands in the command line of uxnmr program; 3) clicking icon-buttons in Graphical User Interface (GUI) of uxnmr program. The mouse has three buttons thus there may be left, right, and middle clicks (L-

, R-, M-click). L-Click is default.

Press orange button, and then Lift on SCM – the spinner with a tube lifts up. Insert the spinner into the wood block. Insert your tube into the spinner all the way down.

Carefully remove the spinner from the wood block so that the tube retains its position in

407

the spinner. If the tube is inserted too deep into the spinner it will break inside the magnet when loaded. Place the spinner on top of NMR (air should be coming out!).

Press Lift on SCM – а the tube is lowered into the magnet. Press Spin and AutoLock.

Log your name in the sign-up sheet, login into the unix shell (get your group’s login and password from Mahinda) and type ‘uxnmr’ in the shell command prompt. Wait until

GUI is loaded. Type edc and in the appeared window enter your name in the NAME field, “1” for 1H, “2” for 13C in EXPNO, “1” for PROCNO and hit “Save”. Raw data –

FIDs – are identified at the experiment number (EXPNO) level. Do not save more than

two files as the disk space is limited. Lack of free disk space will preclude higher-

order experiments from running. Type rpar HCDCl3H13C to read parameters from

file HCDCl3H13C, then hit “copy all”. For 13C experiment type rpar and chose

MGZGDCCHC13 file from the list.

Shimming

If necessary, reset the shims to the standard best values by typing rsh test-test or rsh

, where is the appropriate shim file. L-Click the Lock dg icon in

GUI to bring the lock level to display. Optimize Z1 and Z2 as described below. Be sure

the Fine button is illuminated on SCM. The goal is to maximize the lock level by adjusting the shim values (LockGain is not a shim value; use LockGain to keep the lock display in the middle of the display range). Adjust the shim values slowly since there is a delay in the lock response. The procedure for shimming is as follows. Press Z1 and bring

the lock level as high as you can rotate the black knob on SCM. If the level goes off

scale, press LockGain and bring it back to screen. Repeat Z1 optimization until the lock

408

level reaches maximum and does not rise anymore with variation in Z1. Then press Z2 and repeat optimization rotating the knob very slowly as lock is very sensitive to Z2 variation. Return to Z1 and shim it again since these two values affect each other. Press

Standby to “lock” all the shim values from occasional change. Do not adjust Z3 or

Z4!! The proper value of Z4 requires hours of shimming.

Running experiment

Type rga (1H ONLY) to start receiver gain acquisition. After “rga:finished” has

appeared in the status line, type “rg” to see the value (1…8•103). Skip this command for

13C experiment. Type zg – this command clears all previous FIDs and starts scans. The

acquisition may be stopped at any time hereafter typing halt. It will stop acquisition and save all FIDs acquired hitherto to the disk.

Processing

During the acquisition, all FIDs are temporarily stored in the acquisition processor’s memory until ether halt or tr command is entered. Command tr traces all hitherto

collected FIDs from the acquisition processor’s memory to hard drive and do not

interrupt further acquisition. To see your spectrum during acquisition type tr followed by

fp (1H) or efp (13C) = em+ft+pk = exponential multiplication (em) followed by Fourier transformation (ft) followed by last phase correction (fk). Then apply Automatic phase correction (only once) by typing apk. To see any improvement in the signal since last tr command, type tr followed by fp again. After the acquisition has been stopped (halted) one need not to type tr, fp alone is sufficient.

409

The following commands are useful for 1D NMR: em - exponential multiplication on the FID, uses the parameter LB. This improves signal to noise at the expense of resolution. lb - this controls the degree of broadening added and affects your signal-to-noise. To see its effect, simply change its value and re-Fourier Transform with ef. gm - gaussian multiplication on the fid ft - Fourier transform. ef - combines em and ft. gf - combines gm and ft. pk - phase correct, applies the last phase correction to the spectrum. This process is useful when you have phased a preliminary spectrum, (with only a few scans) and wish to apply the same phase correction to the final spectrum. efp - combines em, ft , and pk. abs- automatic baseline correction apk – automatic phase correction

Expansions

The easiest way to do horizontal expansions on the screen is to click on the left mouse button which converts the pointer to a cursor. Position the cursor on the left edge of the region you want to expand and click the middle mouse button. Move the cursor to the right edge and click the middle mouse button again. Vertical expansion is performed via

L-click on ∧2 or M-click on ∨2 to increase or decrease the intensity correspondingly by a factor of 2.

410

Calibration

Click Calibrate. Move the cursor to the reference peak (usually TMS) and click the middle mouse button. Type in the chemical shift (sharp zero). L-Click Return.

Integration

Click Integrate. To define the integrals, convert the pointer to a cursor (click on left mouse button), and click the middle mouse button on either side of region to be integrated. To phase or reference an integral, they must first be selected. To select an integral, double-click inside it using the left mouse button. An * will appear next to the integral. To phase an integral, select it, place the pointer on slope or bias, and while depressing the left mouse button, move the mouse to phase it. A properly defined integral should extend beyond the apparent ends of the peak (if there is no other adjacent peak). A properly phased integral should be horizontal before and after the peak. Good integration requires careful attention. L-Click on Write+Return to save the integration.

Title

To add a title, type setti – this recalls vi, unix text editor. Use the following commands.

To begin type text, enter in append mode depressing “a”. To exit append mode, hit

“Esc”. To delete text, exit from append mode and press “x”. To finish: “Esc”,

Shift+”Z”+”Z”, then L-click on Clear icon. Other commands: dd – delete current line; J

– join lines; u – undo; i – insert mode; a – append mode; r – replace mode; Esc – back from mode; ZZ – store and exit; :q! – discard and exit; :w – store.

411

Plotting

L-click on Edplot icon – editing and printing window appears. Click “Fix-wind”, chose

IntegLables and PeaksLables to display. Click Sto+Plot to print. DO NOT use

Sto+Plots – this will not print anything for you until flplot command is entered.

Finishing

Remove the tube from the magnet: Press Spin (the green light on the key goes off), orange button, then Lift. Insert blank tube into the spinner and load the spinner with a blank tube into the magnet, do not spin. Type “exit” two times to exit the program and to log off from unix shell.

Commands Summary uxnmr Unix NMR program; uxnmr –r restores default parameters. edc Edits current file set rpar HCDCl3H13C Reads parameters from file HCDCl3H13C, chose MGZGDCCHC13 file for 13C

ii initializes interface

Lock dg Displays lock signal window

Auto lock Automatic lock should be on and steady green (not blinking).

Lock gain Press and rotate the black knob to bring the lock value, to the top of the screen rsh test-test Reads shim parameters from file.

Fine Should be “on”, meaning the knob changes the value slowly

Z1, Z2 Press “Z1” and bring the lock gain level as high as you can. If the level goes off

scale, press Lock gain and bring it back to screen. Repeat “Z1” optimization until the

412

level reaches maximum and does not rise anymore with variation in Z1. Then press “Z2”

and repeat optimization rotating the knob very slowly as lock is very sensitive to Z2 variation.

Stand-by Press to “lock” all the values from occasional change lb Line broadening, more lb increases sensitivity, but lowers resolution: 2 for 1H, 0 for

13C

rga (1H ONLY) Receiver gain acquisition. rg to see value (1…8•103). Skip this

command for 13C experiment.

zg Clears all previous FIDs and starts scans

tr Traces collected FIDs to hard drive

fp (1H) efp (13C) Exponential multiplication followed by Fourier transformation followed by phase correction apk Automatic phase correction (type only once) halt Terminates data acquisition and stores all FIDs to disk.

Integ Integrate the peaks and press “Write+Return”

Edplot Editing and printing window eda Edits acquisition parameters edg Edits graphics parameters edp Edits processing parameters edsp Set frequencies (also ased)

d1 1…10 for 1H, 1…5 for 13C

ns Number of scans to be completed

413

ds Number of dummy scans (to bring the spin system to an equilibrium) lock = AutoLock. Waits 2 min to find lock. ro = Spin. Waits 15 sec to reach set point. ro yes 15 yes will rotate with 15Hz and modulation to suppress solvent side bands. ej, ij – Eject and inject the sample. lo = LockPower (0–60 dB). 45 for CDCl3.

lopo Sets lo and magnetic field wrt SOLVENT parameter.

lg = LockGain (22–140 dB).

wsh, rsh, delsh, vish, lsh, setsh – Write, read, delete, views, prints, and displays a graph

of shim file.

tune –Auto shims using shim gradient file, editable with edtune .

vi commands:

dd – delete current line; J – join lines; u – undo; i – insert mode; a – append mode; r –

replace mode; Esc – back from mode; ZZ – store and exit; :q! – discard and exit; :w –

store;

In .profile: UXNMR_REQMSG=NO to suppress requests for printing.

In .uxnmrrc type those uxnmr commands one would like to execute at startup (one in a

line): param no; plunit ppm; pldigit 3

To insert a part of a spectrum into a plot: get_w12 Æ Sto+Plots Æ fplot (flush plot)

Edpul Edits and prints (“list”) specified pulse sequence file

Edcpul Edits current pulse sequence file (defined in PULPROG with eda)

1 u = 1 μs (millisecond, 10–6 sec); 1 ms = 1 microsec (10–3 sec); 1 s = 1 second

414

Lock_opt then →← to change grid.

COSY

Run usual 1H NMR in say u/user/alex/1/1. d1=1 s

Include all peaks in the sweep (expansion); include large TMS peak, but exclude small TMS peak.

Left click SW–SFO1 (accepts the display limits of your expansion for the sweep) and note the values of SW, O1, and rg. Change delay time to de 10 μs; check aq time. Run new spec with these parameters. cre 2 or re 2 rpar mgcosy-CDCl3 (for all solvents)

Change (enter) the noted parameters: SW, O1, and rg: rg = enter value

SW= enter value

1 SW = enter same value p1 p1 O1=enter value de de=10 aq d1=200 ns=16-32 d1 ds=4 (dummy scans to set the equilibrium in the spin system) expt – experiment time td – time domain size for 1D experiment

1 td – the number of cross spectra; the higher the value the higher 2D resolution

415

rser – 1st increment xfb – to see the spectrum (no tr) thres Click middle button and move mouse to change intensity of threshold

DefPlot: “y” to change intensities; levels<7; contours “y” edg Æ edproj1 Æ ed Æ change 1D filename; PF1CY – length of the most intense peak.

Save. edg Æ edproj2 Æ … (same)

Type plot to print. To expand: click Zoom strip → select the region: L-click, M-click2 →

DefPlot. in co ob – operate only in intensity

Limits: F1LO and F1HI (vertical); F2LO and F2HI (horizontal).

The horisontal resolution is always higher since td > 1 td.

After you are done, delete your 2D file!!! Data → Delete → Delete 2D data → chose your file and click “Execute”.

19F

Run usual 1H NMR, shim very well.

Remove (physically) 1H attenuators (4-2-6 dB) and place BNC connector directly to the

decoupler. Switch BNCs (together with 300 Low Pass Filter 0–31P Rejects 1H) on top of

the decoupler between 1H and 19F (X).

Tune the probe as follows. Find RED and BLACK probes in front of the decoupler

probe. For RED: push up, then turn CW until stops, then turn 21¼ turns CCW. For

BLACK: push up, then turn CW until stops, then turn 8¼ turns CCW.

416

Rpar mg-F19-CDCl3; rg=max is setup in the parameter file, so do NOT type rga.

However, if the FID signal is too strong (cut-off on top and bottom), manually reduce rg.

Set lb=1; de=10μsec. Run a spec with ns=1, use efp to transform and locate your signal.

Enlarge it and then click on SW-SFO1. Check the parameters above and collect spec

with large number of ns in the region of interest.

Return the 1H attenuators, switch back BNC connectors and tune the probes back to 1H:

For RED: push up, then turn CW until stops, then turn 9 ½ –9 ¾ turns CCW. For

BLACK: push up, then turn CW until stops, then turn 10 ½ turns CCW.

Running samples in unusual solvents

Put pure solvent in the tube. Call for Lock_dg. Press Field on SCM, look for resonance turning the knob, or set to a known value (905 for CF3COOD). Lock power ~35…50

depending on solvent saturation limit. Set the sweep width wide: sw 2e4 Hz. Each time

the sw changed, change de back to 10 μsec: de 10. Set the reference to zero: sr 0. Take a

scan (ns=1). Find solvent reference peak and calibrate it. Check TMS signal to

correspond. Change sweep to a practical width (check de!) and run sample in the new

solvent. Save parameter file if desired.

Appendix C

M. Braun Solvent Purification System (SPS)

User’s Instructions

417 418

A. Principles

The solvents are nitrogen-pushed through a column filled with either activated alumina or (for hydrocarbons) alumina and active (reduced) copper. In the latter case the solvent gets deoxygenated as well. The columns are color-coded: if the lower color band on the top of a cylinder is white – it’s alumina packed; if green – it’s copper packed. The active copper catalyst is compatible with hydrocarbons only and incompatible with THF,

CH2Cl2, and Et2O. The SPS does not purify solvents from non-volatile impurities and you

get the same grade of solvent as you put into the supply reservoir, yet anhydrous. Thus,

fill the supply reservoir with clean solvents only.

B. Operation

1. Start with a flask you intend to run a reaction in. Calculate the volume of

anhydrous solvent you will need. Fill the flask with acetone in that volume and make a

mark on the flask. Dry the flask in an oven or with a heat-gun. Cool it down under

nitrogen and stopper it with a matched stopper.

2. At SPS you’ll find it in Stand-by mode (Pic. 1), with nitrogen and solvent

supplies closed, vacuum pump operating, and solvent outlets capped and vacuumized.

Turn all SPS manifold valves to OFF positions (Pic. 2).

419

Pic. 1. Stand-by Mode.

Pic. 2. Manifold valves in OFF positions.

3. Check that all blue valves on all the connected solvent supply reservoirs are closed (Pic. 3). Open main nitrogen valve and check the pressure to be less than 5-7 psi

420

(Pic. 4). Open the blue valve on the solvent supply reservoirs you need to use (Pic. 3).

Double check that all manifold valves are closed (Pic. 2).

Pic. 3. Blue valves on all solvent supply reservoirs, but the one in use, are closed.

Pic. 4. Nitrogen pressure less than 5-7 psi.

421

For exemplary purposes, let’s now consider dispensing of dry THF.

While holding the storage cap by hand:

4. Turn the Main Manifold Valve to “Nitrogen” position.

5. Slowly turn THF’s Nitrogen valve to “Nitrogen/Vacuum” position. (Pic. 5).

Pic. 5. Removing the storage cap from THF’s tap.

422

6. The cap will be filled with nitrogen. Hold the white Teflon adapter with one hand and remove the cap with another hand. A stream of nitrogen should be heard flowing from the tap.

7. Place your flask onto the THF’s tap and hold it with your hand throughout the rest of operations.

8. Turn the Main Manifold Valve to “Vacuum” position. Wait for 15-30 seconds to evacuate your flask.

9. Turn the Main Manifold Valve to “Nitrogen” position for 2-3 seconds to refill the flask with nitrogen.

10. Repeat steps 8 and 9 three to five times.

11. Turn the Main Manifold Valve to “Vacuum” position. Wait for 30-50 seconds to completely evacuate your flask for the final time.

12. Turn THF’s Nitrogen valve to “Fill Collection Vessel” position. (Pic. 6).

423

Pic. 6. Turn THF’s Nitrogen valve to “Fill Collection Vessel” position.

13. Slowly turn THF’s Solvent valve to “To Collection Vessel” position (Pic. 7).

Fill the flask with solvent to the necessary volume or mark. Do not fill more than 2/3 of a flask.

424

Pic. 7. Fill the flask with solvent.

14. Turn THF’s Solvent valve to “OFF” position.

15. Turn the Main Manifold Valve to “Nitrogen” position.

16. Very slowly turn THF’s Nitrogen valve towards “Nitrogen/Vacuum” position.

(Pic. 8). Fill the flask with nitrogen. If you turn the Nitrogen valve too quickly or fully to

“Nitrogen/Vacuum” position, the blow or fast stream of nitrogen may blow the solvent out of the flask and/or the flask itself from the tap.

425

Pic. 8. Slowly filling the void of the flask with nitrogen.

17. With the nitrogen flowing into the flask, detach the flask from the tap and recap it with a stopper as quickly as possible. Turn THF’s Nitrogen valve completely to

“Nitrogen/Vacuum” position to flush residual solvent off.

18. Replace the storage cap on the THF’s tap.

19. Turn the Main Manifold Valve to “Vacuum” position.

20. Turn THF’s Nitrogen valve to “Nitrogen/Vacuum” position (Pic. 9).

426

Pic. 9. Recapping THF’s tap.

C. Stand-by mode

1. Close all blue valves on solvent supply reservoirs (Pic. 3).

2. Turn all “Nitrogen” valves to “Nitrogen/Vacuum” position (Pic. 1).

3. Turn the Main Manifold Valve to “Vacuum” position. (Pic. 1).

4. Close the main nitrogen valve.

The system now is in stand-by mode.