Synthesis, Study of Self-Assembly, and Trivalent Lanthanide Metal Ion Recognition Characteristics of Amphiphilic Acylpyrazolones and Amphiphilic Acylisoxazolones

Western Michigan University ScholarWorks at WMU

Dissertations Graduate College

12-2007

Venkat Reddy Guduru Western Michigan University

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Recommended Citation Guduru, Venkat Reddy, "Synthesis, Study of Self-Assembly, and Trivalent Lanthanide Metal Ion Recognition Characteristics of Amphiphilic Acylpyrazolones and Amphiphilic Acylisoxazolones" (2007). Dissertations. 868. https://scholarworks.wmich.edu/dissertations/868

This Dissertation-Open Access is brought to you for free and open access by the Graduate College at ScholarWorks at WMU. It has been accepted for inclusion in Dissertations by an authorized administrator of ScholarWorks at WMU. For more information, please contact [email protected]. SYNTHESIS, STUDY OF SELF-ASSEMBLY, AND TRIVALENT LANTHANIDE METAL ION RECOGNITION CHARACTERISTICS OF AMPHIPHILIC ACYLPYRAZOLONES AND AMPHIPHILIC ACYLISOXAZOLONES

Venkat Reddy Guduru

A Dissertation Submitted to the Faculty of The Graduate College in partial fulfillment of the requirements for the Degree of Doctor of Philosophy Department of Chemistry

Western Michigan University Kalamazoo, Michigan December 2007

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. SYNTHESIS, STUDY OF SELF-ASSEMBLY, AND TRIVALENT LANTHANIDE METAL ION RECOGNITION CHARACTERISTICS OF AMPHIPHILIC ACYLPYRAZOLONES AND AMPHIPHILIC ACYLISOXAZOLONES

Venkat Reddy Guduru, Ph.D.

Western Michigan University, 2007

This research project focused on the separation of trivalent lanthanide metal ions

by HPLC using amphiphilic acylpyrazolones and amphiphilic acylisoxazolones. The

central hypothesis of our research project is that the nanoscale self-assembly nature of

amphiphilic ligands (chelating lingads) can influence their metal ion recognition and

separation. To test the central hypothesis, we have synthesized a family of novel

amphiphilic ligands and employed them as model systems for the separation of trivalent

lanthanide metal ions. Several novel intermediates such as 4-acylated, iV-acylated, O-

acylated pyrazolones and isoxazolones, have been efficiently synthesized and fully

characterized.

CH3 CH3 /CH3 N =< Ph-Ny' ------y s " ^ ' Yy Y ' ^vopeg 0 V OH 0 - , —* ^" OHY 0 Pyrazolone 4-Haloacylpyrazolone Amphiphilic acylpyrazolone Q ■■ ^ — °rW , 0 OH 0 OH 0

Isoxazolone 4-Haloacylisoxazolone Amphiphilic acylisoxazolone

(R = Methyl, Pheny; R' = Valeroyl, Benzoyl; x = 750, 516, 913)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The trivalent lanthanide metal ion recognition efficacies of these amphiphilic

ligands have been demonstrated through baseline separations of a mixture of light,

middle, and heavy lanthanide metal ions by employing them in the aqueous mobile phase

of high performance liquid chromatography (HPLC) with Ci 8 silica gel as the stationary

phase. The complex separation mechanism is influenced by the chemical structure of 4-

acylated amphiphilic ligands and their spontaneous self-assembly in the aqueous phase as

well as on the stationary phase. Transmission electron microscopy (TEM) images of

these amphiphilic ligands in the aqueous phase in the absence and presence of metal ions

demonstrated several nanoscale structures such as spherical, dendritic, and linear (nano

rods, nano-tubes, nano-fibers) structures.

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Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ACKNOWLEDGEMENTS

I wish to profoundly thank my advisor, Dr. Subra Muralidharan, for accepting me

as his student and giving me the opportunity to do research in his laboratory, under his

guidance. I am greatly indebted to him for the freedom extended to me in every aspect of

my work. This enabled me to work in an area that interested me, and effectively partition

my time between my work and my family. Discussions with him were always thought

provoking and exceedingly helpful. I shall never forget my advisor’s support when I was

under stress after I finished my masters. In the same spirit, I want to thank the other

members of my doctoral committee, Dr. Ekkehard Sinn, Dr. Sherine Obare, Dr. Yirong

Mo, Dr. Muralidhar K. Ghantasala for their help and suggestions. I also wish to thank Dr.

John Miller, graduate student advisor, for his valuable academic guidance towards my

degree. I am sincerely thankful to the Office of Basic Energy Sciences, Department of

Energy for their financial support to continue this project and the Department of

Chemistry, WMU for providing me with the opportunity to pursue my graduate studies.

I am extremely grateful to Dr. Hengli Ma for her help in HPLC work, and also

wish to thank Dr. Srividhya Narayanan for her helpful guidance in my Ph.D. candidacy

exam. I would also like to thank our research group for their congeniality. I thank Dr.

Raymond Sung for his help and support in operating various departmental instruments. I

am especially thankful to Dr. Robert Eversole for his help in providing me with an

opportunity to get trained on transmission electron microscope.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Acknowledgements-Continued

I am also thankful to Annie Dobs and Pam McCortney for their prompt help, not

only in all administration related matters, but also in copying manuscripts and office

supplies.

I am very much thankful to my wife Chandana for being an understanding friend

and excellent wife without whose company living would be meaningless. I am grateful to

my parents and younger brother for their encouragement and emotional support

throughout my stay in the United States. I wish to thank my cousin Dr. Srinivas Reddy G.

for his professional and emotional help extended throughout the duration of my work.

Last but not least, I also wish to thank my friends for their guidance, encouragement and

constant support towards my Ph.D. degree.

Venkat Reddy Guduru

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Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. TABLE OF CONTENTS

ACKNOWLEDGEMENTS...... ii

LIST OF TABLES...... x

LIST OF FIGURES...... xiv

LIST OF SCHEMES...... xviii

CHAPTER

I. INTRODUCTION...... 1

1.1. Lanthanides and the importance of their separation ...... 1

1.2. Recent developments in the field of lanthanide metal ion separation ...... 3

1.3. Central hypothesis ...... 5

II. SYNTHESIS OF 4-HALO ACYL-3 -METHYL-1 -PHENYL-5- PYRAZOLONES AND THEIR AMPHIPHILIC DERIVATIVES 6

2.1. Introduction ...... 6

2.2. Chemical properties of acylpyrazolones ...... 6

2.3. Synthetic approach, results and discussion ...... 10

2.4. X-ray crystallographic analysis of acylpyrazolone ligands 13

2.5. X-ray diffraction data and their treatment ...... 16

2.6. Synthesis of amphiphilic acylpyrazolone ligands ...... 19

2.7. Experimental methods ...... 20

2.7.1. General...... 20

2.7.2. Instrumentation and characterization of compounds... 20

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CHAPTER

2.7.3. 0-(Chlorovaleroyl)-3-methyl-l-phenylpyrazol-5- one (0 2 ) ...... 2 1

2.7.4. 4-(Chlorovaleroyl)-3-methyl-1 -phenylpyrazolin-5- ol (C2)...... 22

2.7.5. 0-(Bromovaleroyl)-3-methyl-l-phenylpyrazol-5- one (04) ...... 23

2.7.6. 4-(Bromovaleroyl)-3 -methyl-1 -phenylpyrazolin-5- ol (C4) ...... 24

2.7.7. 3-Methyl-l-phenyl-0-valeroylpyrazol-5-one (0 5 ) ..... 25

2.7.8. 3-Methyl-1-phenyl-4-valeroylpyrazolin-5-ol (C5) ...... 26

2.7.9. 0(4-Chloromethyl)benzoyl-3-methyl-1 - phenylpyrazol-5-one (0 3 )...... 27

2.7.10. 4-(4-Chloromethyl)benzoyl-3-methyl-1- phenylpyrazolin-5-ol (C3)...... 28

2.7.11. 3-Methyl-l-phenyl-0-tolueoylpyrazol-5-one (06)... 29

2.7.12. 3-Methyl-1-phenyl-4-tolueoylpyrazolin-5-ol (C6).... 30

2.7.13. General procedure for the synthesis of amphiphilic acylpyrazolones ...... 31

2.8. X-ray studies ...... 31

2.8.1. Collection of X-ray diffraction data ...... 31

2.8.2. Structure solution and refinement ...... 32

III. SYNTHESIS OF 4-HALO AC YL-5-IS OXAZOLONE LIGANDS AND THEIR AMPHIPHILIC DERIVATIVES ...... 33

3.1. Introduction ...... 33

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CHAPTER

3.2. Chemical properties of isoxazolone ...... 34

3.3. Synthetic approach, results and discussion ...... 36

3.4. X-ray crystallographic analysis of acylisoxazolone ligands... 41

3.5. X-ray diffraction data and their treatment ...... 43

3.6. Synthesis of amphiphilic acylisoxazolone ligands ...... 45

3.7. Experimental methods ...... 46

3.7.1. General...... 46

3.7.2. 3-Methyl-5-isoxazolone ( 8 )...... 47

3.7.3. /V-Chlorovaleroyl-3-methyl-5-isoxazolone (9)...... 47

3.7.4. 4-Chlorovaleroyl-3-methyl-5-isoxazolone (10)...... 48

3.7.5. 3-Methyl-4-(tetrahydropyran-2-ylidene)-5- isoxazolone (11) ...... 49

3.7.6. 7V-(p-Chloromethyl)benzoyl-3-methyl-5- isoxazolone (12) & 0-(p-Chloromethyl)benzoyl-3- methyl-5-isoxazolone (13)...... 49

3.7.7. 4-(/>Chloromethyl)benzoyl-3 -methyl-5- isoxazolone (15) ...... 51

3.7.8. 4-(/>Chloromethyl)benzoyl-3-phenyl-5- isoxazolone (17), iV-(p-Chloromethyl)benzoyl-3- phenyl-5-isoxazolone (18), & 0-(p-Chloromethyl) benzoyl-3-phenyl-5-isoxazolone (19)...... 52

3.7.9. A-Chlorovaleroyl-3-phenyl-5-isoxazolone (20) & 0-Chlorovaleroyl-3-phenyl-5-isoxazolone (21)...... 53

3.7.10. 4-Chlorovaleroyl-3-phenyl-5-isoxazolone (22)...... 54

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CHAPTER

3.7.11. General procedure for the synthesis of amphiphilic acylisoxazolones ...... 55

IV. LANTHANIDE METAL IONS SEPARATION BY HPLC METHOD USING AMPHIPHILIC ACYLPYRAZOLONES...... 56

4.1. Introduction ...... 56

4.2. Instrumentation and experimental methods ...... 56

4.2.1. HPLC system ...... 56

4.2.2. pH meter ...... 57

4.2.3. Transmission electron microscope (TEM) ...... 57

4.2.4. HPLC column packing ...... 57

4.2.5. Reagents ...... 59

4.2.6. Procedure ...... 59

4.3. HPLC separation results and discussion ...... 60

4.4. HPLC separation chromatograms by HMPVP-750 and results...... 64

4.5. HPLC separation chromatograms by HMPVP-516 and results...... 6 6

4.6. HPLC separation chromatograms by HMPVP-913 and results...... 69

4.7. HPLC separation chromatograms by HBMPP-750 and results...... 71

4.8. HPLC separation chromatogram by HBMPP-516 and results...... 75

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CHAPTER

4.9. HPLC separation chromatogram by HBMPP-913 and results...... 75

4.10. Self-assembly behavior of amphiphilic acylpyrazolones in the aqueous phase ...... 77

4.11. Possible mechanism of lanthanide metal ion recognition and separation by amphiphilic ligands ...... 90

4.12. Analysis of separation results through logD vs. log[HL] and logD vs. pH plots of all amphiphilic ligands ...... 95

4.12.1. LogD vs. log[HL] of HMPVP-750 at pH 2.68 ...... 98

4.12.2. LogD vs. pH of HMPVP-750 at 1 xlO'5M ...... 100

4.12.3. LogD vs. log[HL] of HMPVP-516 at pH 2.90...... 104

4.12.4. LogD vs. pH of HMPVP-516 at 5 xlO'6M ...... 106

4.12.5. LogD vs. log[HL] of HMPVP-913 at pH 2.84 ...... 109

4.12.6. LogD vs. pH of HMPVP-913 at 1 xl0' 5M...... 112

4.12.7. LogD vs. log[HL] of HBMPP-750 at pH 2.40 ...... 115

4.12.8. LogD vs. pH of HBMPP-750 at 5 xlO'6M ...... 117

4.13. Discussion of separation mechanism ...... 120

V. LANTHANIDE METAL IONS SEPARATION BY HPLC METHOD USING AMPHIPHILIC ACYLISOXAZOLONES...... 121

5.1. Introduction ...... 121

5.2. Instrumentation and experimental methods...... 121

5.3. Results and discussion ...... 122

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CHAPTER

5.4. HPLC separation chromatograms by HBPIS-750, HBPIS-516, HBPIS-913, HPVIS-750, and HPVIS-516 ligands ...... 123

5.5. HPLC separation chromatograms by HMVIS-750 and results...... 126

5.6. HPLC separation chromatogram by HMVIS-516 and results ...... 128

5.7. HPLC separation chromatograms by HMVIS-913 and results...... 130

5.8. LogD vs. log[HL] of HMVIS-516 at pH 2.19...... 132

5.9. LogD vs. pH of HMVIS-516 at 5 x 10'5M ...... 135

5.10. Self-assembly behavior of amphiphilic acylisoxazolone ligands ...... 136

VI. SUMMARY AND FUTURE DIRECTIONS...... 141

6.1. Summary ...... 141

6.2. Future directions ...... 144

REFERENCES...... 146

APPENDICES...... 160

A. lR NMR, 13C NMR, IR, and LC/MS spectral data of acylpyrazolones ...... 161

B. *H NMR, 13C NMR, IR, and LC/MS spectral data of acylisoxazolones ...... 180

C. LC/MS spectral data of PEGs, amphiphilic acylpyrazolones, and amphiphilic acylisoxazolones ...... 196

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LIST OF TABLES

2.1. Acylation of pyrazolone with different acid chlorides and reaction conditions ...... 11

2.2. Acyl group transfer from O to C on the pyrazolone ring ...... 13

2.3. Bond lengths and bond angles of enol fragment ...... 15

2.4. Crystallographic data and structure refinement summary for compounds C2, C3, C5 and 0 3 ...... 16

2.5. Bond distances (A) for compounds C2, C3, C5, and 0 3 ...... 17

2.6. Bond angles (deg) for compounds C2, C3, C5, and 0 3 ...... 18

3.1. Crystallographic data and structure refinement summary for compounds 9 , 10, and 11...... 43

3.2. Bond distances (A) for compounds 9 , 10, and 11...... 44

3.3. Bond angles (deg) for compounds 9 , 10, and 11...... 44

4.1. Amphiphilic valeroylpyrazolone concentrations vs. pH ...... 93

4.2. Amphiphilic benzoylpyrazolone concentrations vs. pH ...... 93

4.3. D values for each lanthanide metal ion at pH 2.68 as a function of ligand concentration ...... 98

4.4. LogD values for each lanthanide metal ion at pH 2.68 as a function of ligand concentration ...... 98

4.5. Selectivity (a) for adjacent lanthanide metal ions...... 99

4.6. LogKex values for each metal ion as a function of ligand concentration 100

4.7. D values for each lanthanide metal ion at different pH ...... 100

4.8. LogD values for each lanthanide metal ion at different pH ...... 101

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4.9. Selectivity (a) for adjacent lanthanide metal ions ...... 101

4.10. Slopes for each lanthanide metal ion ...... 103

4.11. LogKex values for each lanthanide metal ion at different pH ...... 103

4.12. D values as a function of ligand concentration at pH 2.90 ...... 104

4.13. LogD values as a function of ligand concentration at pH 2.90 ...... 104

4.14. Selectivity (a) for adjacent metal ions at different ligand concentration.... 106

4.15. LogKex values for each metal ion as a function of ligand concentration 106

4.16. D values for each lanthanide metal ion at different pH ...... 107

4.17. LogD values for each lanthanide metal ion at different pH ...... 107

4.18. Selectivity (a) for adjacent lanthanide metal ions at different pH ...... 107

4.19. Slopes for each metal ion as a function of concentration and pH ...... 109

4.20. LogKex values for each lanthanide metal ion as a function of pH ...... 109

4.21. D values for each lanthanide metal ion at different ligand concentration.... 110

4.22. LogD values for each lanthanide metal ion at different ligand concentration ...... 1 1 0

4.23. Selectivity (a) for adjacent lanthanide metal ions at different concentration ...... 1 1 0

4.24. LogKex values as a function of ligand concentration ...... 112

4.25. D values for each lanthanide metal ion at different pH ...... 112

4.26. LogD values for each lanthanide metal ion at different pH ...... 113

4.27. Selectivity (a) for adjacent lanthanide metal ions at different pH ...... 114

4.28. Slopes for each lanthanide metal ion ...... 114

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4.29. LogKex values for each lanthanide metal ion as a function of pH ...... 114

4.30. D values for each lanthanide metal ion at different ligand concentration.... 115

4.31. LogD values for each lanthanide metal ion at different ligand concentration ...... 115

4.32. Selectivity (a) for adjacent metal ions at different ligand concentration 116

4.33. LogKex values for each lanthanide metal ion as a function of concentration ...... 117

4.34. D values for each lanthanide metal ion at different pH ...... 117

4.35. LogD values for each lanthanide metal ion at different pH ...... 118

4.36. Selectivity («) for adjacent metal ions at different pH ...... 119

4.37. Slopes for each lanthanide metal ion ...... 119

4.38. LogKex values for each lanthanide metal ion at different pH ...... 119

5.1. D values at different ligand concentration ...... 127

5.2. Selectivity (a) for adjacent lanthanide metal ions at different ligand concentration ...... 127

5.3. D values at different ligand concentrations at pH 2.19 ...... 133

5.4. LogD values as a function of ligand concentration at pH 2.19 ...... 133

5.5. Selectivity (ot) for adjacent lanthanide metal ions at different ligand concentration ...... 133

5.6. LogKex values for each metal ion as a function of ligand concentration 134

5.7. D values for each lanthanide metal ion as a function of pH ...... 135

5.8. LogD values for each lanthanide metal ion as a function of pH ...... 135

5.9. Selectivity (a) for adjacent lanthanide metal ions at different pH ...... 135

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5.10. Slopes for each lanthanide metal ion ...... 136

5.11. LogKex values for each lanthanide metal ion at different pH ...... 136

xiii

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LIST OF FIGURES

2.1. Tautomeric forms of 4-acyl-3-methyl-1 -phenyl-5-pyrazolone ...... 9

2.2. X-ray crystal structures of C-acylated and O-acylated pyrazolones 14

2.3. Chemical structures of amphiphilic acylpyrazolone ligands ...... 20

3.1. Possible tautomeric structures of isoxazolone ...... 34

3.2. Possible tautomeric forms of 4-acylisoxazolone and its schematic representation of metal complex formation ...... 35

3.3. Crystal structures of N-, 4-chlorovaleroyl-3-methyl-5-isoxazolones (9,10) and cyclised-3-methyl-5-isoxazolone (11) ...... 42

3.4. Amphiphilic acylisoxazolone ligands ...... 46

4.1. Chemical structure of arsinazo III indicator ...... 57

4.2. Structures of a family of amphiphilic acylpyrazolone ligands ...... 60

4.3. Surfactants that are used for the preparation of amphiphilic ligands 61

4.4. Chemical structure of HMPVP-750 and HBMPP-750 ...... 61

4.5. Parent acylpyrazolones (C2 and C3) and their amphiphilic derivatives (HMPVP-PEGX and HBMPP-PEGX)...... 63

4.6. HPLC separation chromatograms by HMPVP-750 ligand with different pH and concentrations ...... 64

4.7. HPLC separation chromatograms by HMPVP-516 with different conditions ...... 67

4.8. HPLC separation chromatograms by HMPVP-913 with different conditions ...... 69

4.9. HPLC separation chromatograms by HBMPP-750 with different conditions ...... 72

4.10. HPLC separation chromatogram by HBMPP-516 ...... 75

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4.11. HPLC separation chromatogram by HBMPP-913 ...... 76

4.12. TEM images of valeroylpyrazolone-PEG 75o (HMPVP-750) ...... 78

4.13. TEM images of valeroylpyrazolone-PEG 5i6 (HMPVP-516)...... 78

4.14. TEM images of valeroylpyrazolone-PEGgn (HMPVP-913) ...... 78

4.15. TEM images of benzoylpyrazolone-PEG 7so (HBMPP-750) ...... 78

4.16. TEM images of benzoylpyrazolone-PEG 5i6 (HBMPP-516)...... 79

4.17. TEM images of benzoylpyrazolone-PEGgn (HBMPP-913) ...... 79

4.18. Schematic representation of ligand self-assemblies in aqueous phase 80

4.19. TEM images of HMPVP-750 with Mg +2 metal ion (12 hr solution) 83

4.20. TEM images of HMPVP-750 with Mg +2 metal ion (one week solution)... 83

4.21. TEM images of HMPVP-516 with Mg +2 metal ion (12 hr solution) 84

4.22. TEM images of HMPVP-516 with Mg +2 metal ion (one week solution)... 84

4.23. TEM images of HMPVP-913 with Mg +2 metal ion (12 hr solution) 85

4.24. TEM images of HMPVP-913 with Mg +2 metal ion (one week solution)... 85

4.25. TEM images of HBMPP-750 with Mg +2 metal ion (12 hr solution) 8 6

4.26. TEM images of HBMPP-750 with Mg +2 metal ion (one week solution)... 8 6

4.27. TEM images of HBMPP-516 with Mg +2 metal ion (12 hr solution) 8 6

4.28. TEM images of HBMPP-516 with Mg +2 metal ion (one week solution).... 87

4.29. TEM images of HBMPP-913 with Mg +2 metal ion (12 hr solution) 87

4.30. TEM images of HBMPP-913 with Mg +2 metal ion (one week solution)... 87

4.31. TEM images of valeroylpyrazolone-PEG 75o (HMPVP-750) with Sm+3.... 8 8

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4.32. TEM images of valeroylpyrazolone-PEG 5i6 (HMPVP-516) with Sm+3.... 8 8

4.33. TEM images of valeroylpyrazolone-PEGgn (HMPVP-913) with Sm +3 8 8

4.34. TEM images of HBMPP-750 with Sm +3 metal ion ...... 89

4.35. TEM images of HBMPP-516 with Sm +3 metal ion (one week solution).... 89

4.36. TEM images of HBMPP-913 with Sm +3 metal ion ...... 89

4.37. TEM images of HBMPP-913 with Sm +3 metal ion (one week solution).... 90

4.38. Schematic representation of metal ion complexation and separation mechanism ...... 91

4.39. Equilibration of the stationary phase (Cis column) with HMPVP-750 96

4.40. Real run time vs. retention time plots of each lanthanide metal ion 97

4.41. LogD vs. log[HL] plots of each lanthanide metal ion ...... 99

4.42. LogD vs. pH plots of each lanthanide metal ion ...... 102

4.43. LogD vs. log[HL] plots of each lanthanide metal ion ...... 105

4.44. LogD vs. pH plots of each lanthanide metal ion ...... 108

4.45. LogD vs. log[HL] plots of each lanthanide metal ion ...... I l l

4.46. LogD vs. pH plots of each lanthanide metal ion ...... 113

4.47. LogD vs. log[HL] plots of each lanthanide metal ion ...... 116

4.48. LogD vs. pH plots of each lanthanide metal ion ...... 118

5.1. Chemical structures of synthesized amphiphilic acylisoxazolone ligands 122

5.2. HPLC separation chromatogram by HBPIS-750 (5 x 10' 6M)...... 123

5.3. HPLC separation chromatogram by HBPIS-516 (1 x 10‘ 6M)...... 124

5.4. HPLC separation chromatogram by HBPIS-916 (1 x 10" 6M)...... 124

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5.5. HPLC separation chromatogram by HPVIS-750 (2 x 10' 5M)...... 125

5.6. HPLC separation chromatogram by HPVIS-516 (5 x 10' 6M)...... 125

5.7. HPLC separation chromatogram by HMVIS-750 (2 x 10' 5M)...... 126

5.8. HPLC separation chromatogram by HMVIS-750 (4 xlO‘ 5M)...... 127

5.9. HPLC separation chromatogram by HMVIS-516 (5 x 10' 5M)...... 128

5.10. HPLC separation chromatogram for six metal ions by HMVIS-516 ...... 129

5.11. HPLC separation chromatogram by HMVIS-516 (1 x 10' 5M)...... 129

5.12. HPLC separation chromatogram by HMVIS-516 (2 x 10' 5M)...... 130

5.13. Separation chromatograms for five metal ions by HMVIS-913 ...... 131

5.14. LogD vs. log[HL] plots of each lanthanide metal ion ...... 134

5.15. TEM images of benzoylphenylisoxazolone-PEG 75o (HBPIS-750) ...... 137

5.16. TEM images of benzoylphenylisoxazolone-PEG 5i6 (HBPIS-516)...... 137

5.17. TEM images of benzoylphenylisoxazolone-PEGgB (HBPIS-913) ...... 137

5.18. TEM images of phenylvaleroylisoxazolone-PEG 75o (HPVIS-750) ...... 138

5.19. TEM images of phenylvaleroylisoxazolone-PEG 5i6 (HPVIS-516)...... 138

5.20. TEM images of phenylvaleroylisoxazolone-PEGgn (HPVIS-913) ...... 138

5.21. TEM images of methylvaleroylisoxazolone-PEG75o (MVIS-750) ...... 139

5.22. TEM images of methylvaleroylisoxazolone-PEG 5i6 (MVIS-516)...... 139

5.23. TEM images of methylvaleroylisoxazolone-PEGg 13 (MVIS-913) ...... 139

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LIST OF SCHEMES

1.1. Schematic representation of central hypothesis of the research...... 5

2.1. Mechanism of acylation on carbon (4th position) and oxygen ...... 8

2.2. Synthetic approach for 4-acylpyrazolones (C-acylpyrazolones) ...... 10

2.3. Synthesis of C-acylpyrazolone from O-acylpyrazolone ...... 12

2.4. Synthesis of amphiphilic acylpyrazolone ligands ...... 19

3.1. Synthesis of 3-methyl-5-isoxazolone ( 8 ) ...... 36

3.2. Synthesis of N- and 4-chlorovaleroyl-3-methyl-5-isoxazolones (9 and 10).. 37

3.3. Synthesis of N-, O-, and 4-(p-chloromethyl)benzoyl-3- methyl-5-isoxazolones (12,13, and 15) ...... 38

3.4. Synthesis of N-, O-, and 4-(o-chloromethyl)benzoyl-5-isoxazolones (18,19, and 17)...... 39

3.5. Synthesis ofTV-, 0-, and 4-chlorovaleroyl-3-phenyl-5-isoxazolones (20, 21, and 22)...... 40

3.6. Synthesis of amphiphilic acylisoxazolone ligands ...... 45

xvm

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CHAPTER I

INTRODUCTION

1.1. Lanthanides and the importance of their separation

Lanthanides belong to the 4/-Block series and are placed in the sixth period of the

periodic table. Lanthanides are also known as Lanthanones and the first inner transition

elements. These are the elements in which the 4/orbitals (antepenultimate shell) are

progressively filled by the differentiating electron, and this includes elements from

Cerium (Cesg) to Lutetium (L U 7 1 ). The general electronic configuration of these elements

is os 2 (n-l)d0' 1 (n-2)/1'14. The name lanthanide has been derived from lanthanum (La)

element which is the prototype of lanthanides. ’ 1 2

It is important to note that these elements are placed outside the body of the

periodic table due to their remarkable similarities among the chemical properties of the

lanthanides. These elements differ in the number of/-electrons which do not take part in

chemical bonding. The similarities in properties, in turn is due to the similar electronic

configuration of the outermost shell and can also be attributed to the lanthanide

contraction which is a steady decrease in the radii of metal/metal ions as the series moves

across from La to Lu. The cause of this contraction is the imperfect shielding (screening)

of 4/-electrons from the increasing nuclear charge. The poor shielding of /electrons is

due to the part of their wave function (shape of orbital ) . 1 As atomic number increases, the

nuclear charge increases by unity at each step, while no comparable increase in the

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. mutual shielding effect of 4/-electrons occurs. This causes a contraction in the size of the

4Asubshell. Consequently the atomic and ionic size go on decreasing as we move from

La to Lu; thus the ionic radius changes from 1.06A (La+3) to 0.85A (Lu+3) . 1,2

As these elements have very close chemical properties due to lanthanide

contraction, the separation of lanthanides is still a problem. In spite of several reported T ft procedures such as physical, pyrochemical and chemical ' for the separation and

purification of these metals, it has been a formidable challenge for the analytical chemists

to develop new separation methods to separate lanthanides as a group from actinides as

well as to separate individual lanthanides from each other from nuclear waste in fewer

stages. Recognition of metal ions is an important factor for a variety of applications such

as recovery of metal ions and environmental remediation. In spite of sixty five years of

research that dates back to the Manhattan Project during World War II, separation of

lanthanide metal ions with high selectivity continues to be a challenge. Recycling of

lanthanides from nuclear wastes and disposing the nuclear wastes in proper way is very

important.

Efficient separation and recovery of metal ions from industrial wastes as well as

raw materials are gaining more importance because of the increasing demand for high

purity products and also due to environmental concerns. The lanthanide metals have been

used in many fields2, 7’ 8 since long back. The following are the uses of lanthanides. (1)

The cerium alloy ( Mischmetals) with steel is sufficiently pyrophoric and hence used in

cigarette lighters, toys and flame throwing tanks. (2) For removing oxygen and sulfur

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. from other metals. (3) Cerium-magnesium alloys are very brittle and so are used in flash

light powders. (4) Useful as catalytic converters means CeC >2 is used as ceramic support

for the platinum metal components of the catalyst, and used as oxygen reservoir due to

the ability to exist as Ce 2C>3 under reduction conditions. Oxides of cerium (Ce 0 2 ) and

thorium (TI 1O2) are used in gas mantles. (5) Lanthanide salts have recently been used in

lasers. Neodymium oxide dissolved in selenium oxychloride serves as a powerful liquid

laser. (6 ) Since Cerium absorbs both heat and UV light, cerium glass is used in glare

reducing spectacles. (7) Cerium salts are used as catalysts in hydrogenation,

dehydrogenation, oxidation, and cracking of petroleum. They are also used in qualitative

analysis as well as for dyeing cotton. ( 8 ) Europium phosphors are used in CRT TV

screens and Nd-YAG lasers. Luminescent lanthanide complexes (Eu +3 and Tb+3) have a

wide variety of photonic applications, such as planar waveguide amplifiers, plastic lasers,

light emitting diodes and luminescent probes .9' 12 Since lanthanides have potential

applications their recycling from wastes and separation of lanthanide metal ions is

important.

1.2. Recent developments in the field of lanthanide metal ion separation

As the properties of metal ions are determined by their size and charge, the

lanthanide ions with typically tripositive charge and almost identical sizes, resemble each

other very much in chemical properties which cause the separation of lanthanide metals a

difficult task. However, the separation may be affected by one or another or combination

of below mentioned various methods based on the slight differences in their solubilities,

complex formation, basic properties and hydration. These differences in their properties

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. in turn are due to very slight size differences of their trivalent ions. Available methods 2 ,6

for the separation of lanthanides: (1) Fractional crystallization method, (2) Fractional

precipitation, (3) Fractional thermal decomposition of salts, (4) Ion exchange method, (5)

Oxidation reduction method, ( 6 ) Solvent extraction method, (7) Membrane-based and

supercritical fluid processes, and ( 8 ) Chromatographic methods. However, these methods

have their own advantages and disadvantages. They use large volume of environmentally

harmful organic solvents, have a slow extraction speed, and a low concentration rate . 13

Though there are several separation procedures available in the field, more efficient multi

stage methods are necessary for complete separation of adjacent lanthanides.

A survey of the literature showed that the extraction behavior of metal ions using

/?-diketones, 4-acyl-5-pyrazolones, and 4-acyl-5-isoxazolones has been studied

extensively using solvent (liquid-liquid) extraction methods 14' 31 and centrifugal partition

chromatography .5 ,3 2 ,33 There are no reports found that use HPLC for the separation of

lanthanide metal ions, and all of the ligands those were used in the solvent extraction for

effective separation cannot be dissolved in water and most of them are effective only at

very low pH values which is not suitable for silica support. We have employed these

ligands (pyrazolones and isoxazolones) and converted them into amphiphilic in behavior,

to use in high performance liquid chromatographic method (HPLC) for the first time in

the field of science. In our research, we have focused on a family of 4-

haloacylpyrazolone and 4-haloacylisoxazolone ligands that can be converted into

amphiphilic in nature that are soluble in aqueous phase, which can eventually provide

efficient separation of tri valent lanthanides in the pH range 2.0 - 3.0.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 1.3. Central hypothesis

Nanoscale structures often have unique properties that are different from

individual atoms, molecules, and bulk structures. Our hypothesis is that the amphiphilic

chelating ligands, which form organized nanoscale molecular self-assemblies, are capable

of providing excellent selectivity and separation for closely related metal ions, such as

tri valent lanthanide metal ions. To substantiate our hypothesis we have employed

amphiphilic acypyrazolones and amphiphilic acylisoxazolones, which spontaneously

form self-assembled nanoscale structures in the aqueous phase, as chelating ligands for

the separation of lanthanide metal ions. This type of approach for the recognition of

trivalent lanthanide metal ions is the first approach in the science field.

R /CH3 CH, N =< N = \ -N, Ph- Ph- ° A r' Cl Cl 0 0 OH 0 OH 0 Isoxazolone 4-haloacylisoxazolone 4-haloacylpyrazolone Pyazolone (R = Me, Ph) R /CH3 N = f Ph- W' "OPEG "OPEG 0 OH O O OH r Amphiphilic acylisoxazolone Amphiphilic acylpyrazolone (R’ = valeroyl, benzoyl) (R' = valeroyl, benzoyl)

Lanthanide metal ligand complex on HPLC column

Separation of Lanthanides by HPLC

Scheme 1.1. Schematic representation of central hypothesis of the research

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CHAPTER II

SYNTHESIS OF 4-HALO AC YL-3 -METHYL-1 -PHENYL-5-PYRAZOLONES AND THEIR AMPHIPHILIC DERIVATIVES

2.1. Introduction

The 4-acyl-5-pyrazolones, a kind of yS-diketones or enols, have several

O JA 2*7 advantages. ’ ' These acylpyrazolones act as strong acids. They can extract metal ions

from acidic media at a lower pH, because of their small dissociation constant . 14 Various

kinds of derivatives can be easily synthesized, and obvious substituent effects were

observed in the extraction of lanthanide metal ions . 14 Acylpyrazolones are an important

2^ 21 10 and interesting class of transition metal and lanthanide metal ion chelating ligands ’ ’

40 and continue to attract significant attention as a result of good chelating agents,

eventually better lanthanide metal ion separation ligands. Physico-chemical constants of

these acylpyrazolone ligands such as acid dissociation constants, distribution coefficients

etc can be easily varied within a wide range by proper substitution in the molecules.

Hence this class of compounds seems to serve well as a basis for a closer study of the

influence of the physical and chemical properties of a chelating agent on its extractive

and chelating behavour 41

2.2. Chemical properties of acylpyrazolones

First part of our research program in metal ion recognition was focused on the

design, synthesis, and fundamental characterization of amphiphilic ligands that are

soluble in aqueous and nonaqueous media (polar organic solvents). Acylpyrazolones

have long attracted attention in solvent extraction studies for a various families of metal

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ions such as transition metals, trivalent lanthanides etc . 14,32’ 3 3 ,3 5 ,38-40 The 4-haloacyl-3-

methyl-l-phenylpyrazol-5-ones are analogous to /Tdiketones and their enols, offer

several advantages including their relatively low pKa values (3.0-4.0) and the flexibility

in introducing various substituents to gain a fundamental understanding of their metal ion

recognition efficacies . 14 The introduction of substituents can influence their pKa values,

distribution ratios between organic and aqueous phases, and stability constants of their

complexes with metal ions which are important parameters in a traditional solvent

extraction system. 14 ’ 39 ’ 40

These ligands have been particularly attractive for the extraction and separation of

lanthanide metal ions due to' their low pKa values and the ability to modify their

structures to optimize the distribution and stability constants .8 ,3 2 ,3 3 ,3 8 ,41-44 The impetus

for our research stems from designing ligands that can be employed in aqueous solvents

for the recognition of metal ions to obtain chelating systems that can provide efficient

separation of metal ion mixtures through HPLC. The water soluble ligands are especially

important towards developing “green” separation methods. To this end we have designed,

synthesized, and characterized 4-haloacyl-3-methyl-l-phenylpyrazol-5-ol (halo = chloro,

bromo) compounds and is to the best of our knowledge the first report of such

compounds. The halogen end group on the alkyl and aryl substituents in the 4 position

has been further reacted with Na salts of polyethylene glycol methyl ethers (PEG) to

obtain acylpyrazolones with hydrophilic side chains and in turn they are amphiphilic in

nature. These amphiphilic ligands spontaneously self-assemble in the aqueous phase to

yield a variety of nanostructures and exhibit excellent selectivity for trivalent lanthanide

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. metal ions when employed in the aqueous mobile phase in HPLC separations in which

Cig silica gel column was used as stationary phase. The self-assembling and separation

results are discussed in Chapter 4.

While synthesizing the desired 4-haloacylpyrazolone compounds, the interesting

acid chloride chemistry with pyrazolones has been observed. The initial significant

component of our studies was the demonstration that the O-acyl (oxygen on 5th carbon)

product was formed initially in varying yields depending on the acid chloride and

reaction conditions employed which then could be quantitatively converted to the C-acyl

(4th carbon) pyrazolone ligand through acyl group transfer. Two reactive sites (4th carbon fVi and oxygen on 5 carbon) are available on the pyrazolone ring due to its resonance

structures that lead to the formation of C-acylation and O-acylation. The mechanism is

shown in scheme 2 .1 .

c h 3 ,c h 3 N = / / \ P h " Ns ^ Ph T H 0 O 0 © Pyrazolone (1) acid chloride acid chloride

,c h 3

C-acylation O O-acylation

Scheme 2.1. Mechanism of acylation on carbon (4th position) and oxygen

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The acyl group transfer, analogous to the Fries rearrangement, was catalyzed by a

strong base such as calcium hydroxide. The acyl group transfer has been previously

observed by Jensen 45 who reported only one O-acylated compound which was not

completely characterized. We have modified Jensen’s procedure and synthesized both O-

acyl and C-acyl compounds and characterized them by NMR ( 1H, 13C{'H}), mass

spectrometry, elemental analysis, and X-ray crystallography. The 4-acylpyrazolones (C-

acylated pyrazolone) with terminal haloacyl groups were reacted with sodium salts of

polyethylene glycol methyl ethers with varying number of oxyethylene groups to obtain

the amphiphilic acylpyrazolones that are soluble in aqueous and polar nonaqueous media.

The amphiphilic acylpyrazolones were primarily characterized by LC/MS to determine

their purity and molecular mass.

The 4-haloacyl-3-methyl-l-phenylpyrazol-5-ol could exist in four tautomeric

forms (Figure 2.1) where R represents a terminal halo alkyl or aryl group 46 Resonance,

inductive, and steric effects, due to the substituents in 1, 3, and 4 positions of the

pyrazolone ring, influence the extent of keto-enol tautomerization and the position of the

hydroxyl group .47

Figure 2.1. Tautomeric forms of 4-acyl-3-methyl-1-phenyl-5-pyrazolone

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. We have determined from *H NMR in CDCI 3, and X-ray crystallographic studies

that 4-haloacyl-3-methyl-l-phenylpyrazol-5-ol compounds exist predominantly in the

enol form with the OH group on the pyrazolone ring and X-ray crystallographic

structures are shown in section 2.4.

2.3. Synthetic approach, results and discussion

The synthetic approach for 4-haloacyl-3-methyl-l-phenylpyrazol-5-ol ligands

employing commercially available 3-methyl-l-phenyl-2-pyrazolin-5-one (1), is shown in

scheme 2.2. These 4-haloacylpyrazlones exist in both diketone and enol forms and they

can extract most metal ions at a lower pH due to their small dissociation constants.

Therefore, it is very likely that the introduction of electron withdrawing substituents and

bulky groups into the 4-acyl group of the pyrazolone would increase the acidity of the

final ligand and also increase their solubilities in the organic solvents, and the metal ions

might still be extracted at a lower pH, with a larger distribution ratio.

,CH3 N = / / \ R + P IT 1^

O acid chloride C-acylpyrazolone 1 O O-acylpyrazolone

Scheme 2.2. Synthetic approach for 4-acylpyrazolones (C-acylpyrazolones)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Hence, valeroyl (electron donating group) and benzoyl (electron withdrawing

group) functional groups have been chosen as side chains on the pyrazolone ring. Various

reaction conditions such as the nature of base, solvent, temperature and reaction time

(Table 2.1) have been investigated. The optimum reaction conditions for the formation of

O-acyl ated and C-acylated products and the conversion of C-acylated to C-acylated

products were determined. In this regard we have modified Jensen’s procedure who

employed dioxane under reflux conditions with Ca(OH )2 as the base .45 The C-acylated

(4-acylated) and O-acylated products were separated by crystallization. Yields of the

desired 4-acylated pyrazolone, depended on the acid chloride used in the reaction and

reaction conditions such as base (Et 3N, Ca(OH)2), time and temperature. Reaction

conditions and % yields are listed in table 2 .1 for the different acid chlorides studied.

Table 2.1. Acylation of pyrazolone with different acid chlorides and reaction conditions

R Base Temp Time C-Acylpyrazolone O-Acylpyrazolone yield (%) yield (%)

2 Ca(OH) 2 0°C 18 hr 1 0 % 80% (02) 70°C 6 hr 80% (C2) 5% Et3N rt 5 hr 0 % 80%

3 Ca(OH) 2 0°C 2 hr 1 0 % 75% (03) 80°C 18 hr 84% (C3) 1 0 % Et3N rt 1 hr 0 % 80%

4 Et3N rt 1 hr 0 % 90% (04)

5 Et3N rt 1 hr 0 % 85% (05)

6 Ca(OH) 2 80°C 36 hr 50% 0 % Et3N rt 1 hr 0 % 85% (06) Note: The C-acylated and C-acylated products are denoted by C and C associated with the acid chloride number.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The products were identified by NMR ( 1H, 13C{1H}), mass spectrometry,

elemental analysis, and X-ray crystallography (in cases where single crystals could be

obtained). The formation of the O-acylated and C-acylated pyrazolones was confirmed by

'fl NMR spectra. Proton NMR spectrum of O-acylated pyrazolone shows a singlet peak

at 5 6.28 ppm for the proton in position 4 of the pyrazolone ring. This peak is absent for

the C-acylated pyrazolone. In the 'H NMR spectrum of 4-acylpyrazolone (C-acylated

product) a peak corresponding to the enolic -OH could not be observed. However, the

absence of a peak at 8 3.2 ppm, corresponding to the methylene proton at the fourth

position of the pyrazolone ring, and X-ray crystallographic structures confirms the

existence of 4-acylpyrazolones in the enolic form with -OH group in the pyrazolone ring.

The mass spectra for chloromethyl pyrazolones exhibited molecular ion peaks differing

by m/z values of 2 due to 35C1 and 37C1 isotopes.

All O-acylated pyrazolones could be converted to the C-acylated pyrazolones (Scheme

2.3) employing Ca(OH )2 as acyl transfer agent at different reaction time in high yields

(Table 2.2).

dry THF, 80°C

u C-acylpyrazolone O-acylpyrazolone

Scheme 2.3. Synthesis of C-acylpyrazolone from O-acylpyrazolone

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 2.2. Acyl group transfer from O to C on the pyrazolone ring

R Time Yield

2 1 hr 8 8 % (C2)

3 5 hr 60% (C3)

4 4 hr 85% (C4)

5 6 hr 97% (C5)

6 6 hr 60% (C6 ) All reactions were carried out under Argon.

2.4. X-ray crystallographic analysis of acvlpyrazolone ligands

Recrystallizing of compounds C2, C3, C5 and 03 from THF/H 20 , CH2CI2, THF,

and hexane gave suitable crystals for structure determination by X-ray analysis. Single

crystal X-ray structural analysis of acylpyrazolones was performed as shown in section

2 .8 in order to understand their molecular structures in terms of bond angles, bond

lengths and the position of the OH group in the ligands C2, C3, C5 and 03. The X-ray

structures are shown in figure 2.2. The structures indicate that the pyrazolones exist in the

enol form with the OH group on the pyrazolone ring.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Compound C5 Compound C2

C !5a , C15

N2a C !3a C l 4a C3a C13 C14

N1a C2a C4a C8a C12a C6a G12 C5a C7a C1a C10a C10 0 1 a C !1a C11

Compovmd 0 3 Compound C3

fC 3

Figure 2.2. X-ray crystal structures of C-acylated and C-acylated pyrazolones

The crystal structure of the O-acylated product (03) for reaction of 3-methyl-1-

phenyl-2-pyrazolin-5-one (1) with />-chloromethylbenzoyl chloride (3) was also obtained

as shown in figure 2.2. The bond distances and bond angles for the enol fragment of

pyrazolones C2, C3, and C5 are compared (Table 2.3) and complete crystallographic

data for all compounds is given in tables 2.4, 2.5, and 2.6. These structural parameters are

most relevant for metal ion complexation and as seen from table 2.3, the values are very

similar for the acylpyrazolones with aliphatic and aromatic acyl groups.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 2.3. Bond lengths and bond angles of enol fragment

ch3

N-— / =3

nr \C2

r 1 1 ° \ H

Bond lengths (A) Bond C2 C3 C5 Ci-Oi 1.329(2) 1.325(10) 1.327(15)

C4-0 2 1.249(2) 1.258(11) 1.238(15)

C 1-C2 1.388(2) 1.404(11) 1.392(18)

C2-C4 1.445(2) 1.439(13) 1.450(17)

Bond angles (degrees) Ci-C2-Oi 127.14(16) 127.56(8) 130.56(11)

C2-C4 - 0 2 118.58(16) 118.52(8) 118.40(11)

crc2-c3 103.94(15) 103.86(7) 104.34(11)

C 1-C2-C4 120.34(16) 118.64(8) 121.05(11)

c3-c2-c4 135.73(16) 137.41(7) 134.59(12)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 2.5. X-rav diffraction data and their treatment

The collection of diffraction data and their treatment are given in the following

tables 2.4, 2.5, and 2.6. Type of the crystal and space group for each compound are

displayed in table 2.4.

Table 2.4. Crystallographic data and structure refinement summary for compounds C2, C3, C5 and 03

Parameters Compound C2 Compound C3 Compound C5 Compound 03

empirical formula c 15h 17cin 2o 2 C18H15C1N20 2 c 15h 18n 2o 2 c 18h 15cin 2o 2 fw 292.76 326.78 258.32 326.78 cryst habit colorless plate Pale yellow prism colorless prism colorless plate cryst dimens (mm) 0.28 x 0.21 x 0.02 0.40 x 0.20 x 0.20 0.30x0.12x0.08 0.40 x 0.30 x 0.08 cryst syst monoclinic monoclinic monoclinic monoclinic space group P lln CUc Pl\!n P2i/c a (A) 11.1005(5) 15.8055(4) 7.7972(5) 16.5050(3) b(A) 10.8112(5) 13.1260(4) 16.6266(10) 12.9905(2) c ( A) 12.2243(5) 14.9386(4) 21.5007(13) 7.37330(10) a (deg) 90 90 90 90 A (deg) 102.154(2) 98.0713(15) 99.412(3) 98.0390(8) 7 (deg) 90 90 90 90 V(A3) 1385.82(11) 3068.51(15) 2749.8(3) 1565.36(4) z 4 8 8 4 D (Mg m'3) 1.403 1.415 1.248 1.387 ju (mm'1) 0.28 0.26 0.08 0.25 cGoF 1.69 1.80 1.45 1.92 T (K) 110 110 110 110 R“/wRi 0.039/0.048 0.055/0.059 0.059/0.053 0.046/0.049 “RF = Sum(Fo-Fc)/Sum(Fo). Rw = Sqrt[Sum(w(Fo-Fc)**2)/Sum(wFo**2)]. cGoF = Sqrt[Sum(w(Fo- Fc)**2)/(No. of reflns - No. of params.)]

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 2.5. Bond distances (A ) for compounds C2, C3, C5, and 03

Compd Distance (A) distance (A) distance (A)

C2 Ol-Cl 1.329(2) C2-C3 1.429(2) C8-C11 1.8096(19) 02-C4 1.249(2) C2-C4 1.445(2) C9-C10 1.395(2) N1-N2 1.3991(20) C3-C15 1.501(3) C9-C14 1.391(3) Nl-C l 1.346(2) C4-C5 1.499(3) C10-C11 1.385(3) N1-C9 1.422(2) C5-C6 1.520(3) C11-C12 1.382(3) N2-C3 1.313(2) C6-C7 1.531(3) C12-C13 1.391(3) C1-C2 1.388(2) C7-C8 1.508(3) C13-C14 1.384(3)

C3 cn-cir 0.291(6) N1-N2 1.3940(10) C5-C10 1.3949(13) Cll-Cll" 1.290(18) Nl-C l 1.3405(12) C6-C7 1.3901(12) C ll-C ll 1.8207(18) N1-C12 1.4234(11) C7-C8 1.3882(13) cir-cir 1.004(18) N2-C3 1.3216(11) C8-C9 1.3926(13) Cll'-Cll 1.671(6) C1-C2 1.4048(11) C8-C11 1.5059(12) C9-C10 1.3910(13) C16-C17 1.3899(14) C12-C13 1.3957(13) cii"-cii"a 0.985(13) C2-C3 1.4336(13) C12-C17 1.3886(14) C11"-C11 2.65(3) C2-C4 1.4394(13) C13-C14 1.3971(14) Ol-Cl 1.3252(10) C3-C18 1.4961(12) C14-C15 1.3806(17) Ol-Hl 0.910(16) C4-C5 1.4818(12) C15-C16 1.3890(17) 02-C4 1.2586(11) C5-C6 1.3960(13)

C5 Ola-Cla 1.3277(15) C7a-C8a 1.5196(20) Clb-C2b 1.3907(18) Ola-Hla 0.96(2) C9a-C10a 1.3888(19) C2b-C3b 1.4276(18) 02a-C4a 1.2387(15) C9a-C14a 1.3914(18) C2b-C4b 1.4465(18) Nla-N2a 1.3969(14) ClOa-Clla 1.3870(19) C3b-C15b 1,4888(19) Nla-Cla 1.3494(16) Clla-C12a 1.383(2) C4b-C5b 1.5008(19) Nla-C9a 1.4265(16) C12a-C13a 1.383(2) C5b-C6b 1.5197(19) N2a-C3a 1.3239(17) C13a-C14a 1.3815(20) C6b-C7b 1.5200(20) Cla-C2a 1.3928(18) Olb-Clb 1.3214(16) C7b-C8b 1.518(2) C2a-C3a 1.4257(17) Olb-Hlb 0.92(2) C9b-C10b 1.3929(18) C2a-C4a 1.4504(17) 02b-C4b 1.2457(16) C9b-C14b 1.3886(18) C3a-C15 1.4919(19) Nlb-N2b 1.4019(14) ClOb-Cllb 1.3861(20) C4a-C5a 1.5037(18) Nlb-Clb 1.3513(16) Cllb-C12b 1.384(2) C5a-C6a 1.5176(18) Nlb-C9b 1.4232(17) C12b-C13b 1.387(2) C6a-C7a 1.5218(19) N2b-C3b 1.3178(17) C13b-C14b 1.3837(20)

03 N1-N2 1.3735(14) C3-C18 1.4999(18 C ll-C ll 1.8097(13) Nl-C l 1.3557(16) C4-C5 1.4777(17) C12-C13 1.3902(17) N1-C12 1.4239(15) C5-C6 1.3958(17) C12-C17 1.3909(17) N2-C3 1.3296(16) C5-C10 1.3944(17) C13-C14 1.3835(19) C l-01 1.3786(14) C6-C7 1.3778(18) C14-C15 1.3904(20) C1-C2 1.3709(17) C7-C8 1.3957(17) C15-C16 1.3879(19) 01-C4 1.3684(14) C8-C9 1.3959(18) C16-C17 1.3910(18) 02-C4 1.1975(15) C8-C11 1.4929(18) C2-C3 1.4118(18) C9-C10 1.3852(18)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 2.6. Bond angles (deg) for compounds C2, C3, C5, and 03

Compd Angle (deg) Angle (deg) angle (deg)

C 2 N2-N1-C1 109.92(13) N2-C3-C15 118.96(15) C10-C9-C14 120.05(16) N2-N1-C9 118.64(13) C2-C3-C15 129.55(16) C9-C10-C11 119.20(17) C1-N1-C9 131.44(15) 02-C4-C2 118.58(16) C10-C11-C12 121.23(17) N1-N2-C3 105.98(14) 02-C4-C5 121.35(16) Cl 1-C12-C13 119.17(17) 01-C1-N1 124.17(15) C2-C4-C5 120.07(16) C12-C13-C14 120.51(18) 01-C1-C2 127.14(16) C4-C5-C6 115.92(15) C9-C14-C13 119.83(17) N1-C1-C2 108.68(15) C5-C6-C7 109.87(15) C8-H8b-Cll 51.9(10) C1-C2-C3 103.94(15) C6-C7-C8 112.84(15) N1-C9-C14 118.17(15) C1-C2-C4 120.34(16) C7-C8-C11 110.03(13) N2-C3-C2 111.49(15) C3-C2-C4 135.73(16) N1-C9-C10 121.77(16)

C 3 Cll'-Cll-Cll" 9.9(14) N2-N1-C1 110.69(7) C2-C3-C18 130.72(8) cir-cn-cii 55.1(12) N2-N1-C12 118.92(7) 02-C4-C2 118.52(8) C11"-C11-C11 63.6(7) C1-N1-C12 130.34(7) 02-C4-C5 118.65(8) Cll-Cll'-Cll" 167.2(18) N1-N2-C3 106.12(7) C2-C4-C5 122.82(8) Cll-Cll'-Cll 116.7(13) Ol-Cl-Nl 124.22(7) C4-C5-C6 118.94(8) cir-cii'-cn 74.3(9) 01-C1-C2 127.56(8) C4-C5-C10 121.25(8) C11-C11"-C11’ 2.9(4) N1-C1-C2 108.22(7) C6-C5-C10 119.71(8) Cll-Cll"-Cll"a 124.6(10) C1-C2-C3 103.86(7) C5-C6-C7 119.78(8) C11-C11"-C11 73.6(7) C1-C2-C4 118.64(8) C6-C7-C8 120.72(8) Cir-Cll"-Cll"a 126.5(12) C3-C2-C4 137.41(7) C7-C8-C9 119.37(8) cii'-cir-cn 71.1(9) N2-C3-C2 111.09(7) C7-C8-C11 119.13(8) Cll"a-Cll"-Cl 1 155.4(9) N2-C3-C18 118.15(8) C9-C8-C11 121.47(8) C11-C11-C11' 8.2(2) C11"-C11-C8 122.9(4) C8-C9-C10 120.41(8) C11-C11-C11" 42.8(6) N1-C12-C13 120.34(8) C5-C10-C9 119.98(8) C11-C11-C8 111.08(8) N1-C12-C17 118.19(8) C13-C14-C15 120.86(10) Cll'-Cll-Cll" 34.6(6) C13-C12-C17 121.40(8) C14-C15-C16 120.03(9)

C 5 N2a-Nla-Cla 110.70(10) C6a-C7a-C8a 112.66(12) C3b-C2b-C4b 136.28(12) N2a-Nla-C9a 118.89(10) Nla-C9a-C10a 121.55(11) N2b-C3b-C2b 111.71(11) Cla-Nla-C9a 130.41(11) Nla-C9a-C14a 117.99(11) N2b-C3b-C15b 118.90(11) Nla-N2a-C3a 105.43(10) C10a-C9a-C14a 120.44(12) C2b-C3b-C15b 129.39(12) Ola-Cla-Nla 121.40(11) C9a-C10a-Clla 118.98(12) 02b-C4b-C2b 118.98(12) 01a-Cla-C2a 130.56(11) C10a-Clla-C12a 120.85(13) 02b-C4b-C5b 120.89(11) Nla-Cla-C2a 108.01(11) Clla-C12a-C13a 119.66(13) C2b-C4b-C5b 120.13(11) Cla-C2a-C3a 104.34(11) C12a-C13a-C14a 120.35(13) C4b-C5b-C6b 115.21(11) Cla-C2a-C4a 121.05(11) C9a-C14a-C13a 119.67(13) C5b-C6b-C7b 112.21(12) C3a-C2a-C4a 134.59(12) N2b-Nlb-Clb 110.06(10) C6b-C7b-C8b 112.82(13) N2a-C3a-C2a 111.52(11) N2b-Nlb-C9b 119.44(10) Nlb-C9b-C10b 121.58(11) N2a-C3a-C15a 118.48(12) Clb-Nlb-C9b 130.48(11) Nlb-C9b-C14b 117.81(11) C2a-C3a-C15a 129.97(12) Nlb-N2b-C3b 105.69(10) C10b-C9b-C14b 120.60(12) 02a-C4a-C2a 118.40(11) Olb-Clb-Nlb 123.50(12) C9b-C10b-Cllb • 118.86(13) 02a-C4a-C5a 121.00(11) 01b-Clb-C2b 128.01(12) C10b-Cllb-C12b 121.06(13) C2a-C4a-C5a 120.59(11) Nlb-Clb-C2b 108.48(11) Cl lb-C12b-C13b 119.34(13) C4a-C5a-C6a 114.72(11) Clb-C2b-C3b 104.05(11) C12b-C13b-C14b 120.58(13)

0 3 N2-N1-C1 110.46(10) 01-C4-02 122.71(11) C5-C10-C9 119.92(11) N2-N1-C12 119.62(10) 01-C4-C5 112.38(10) C8-C11-C11 109.88(8) C1-N1-C12 129.69(10) 02-C4-C5 124.92(11) N1-C12-C13 118.82(11) N1-N2-C3 104.82(10) C4-C5-C6 117.12(10) N1-C12-C17 120.52(10) N l-C l-O l 116.02(10) C4-C5-C10 123.14(11) C13-C12-C17 120.66(11) N1-C1-C2 108.77(11) C6-C5-C10 119.67(11) C12-C13-C14 119.74(12) 01-C1-C2 135.18(11) C5-C6-C7 120.23(11) C13-C14-C15 120.23(12) C1-01-C4 119.47(9) C6-C7-C8 120.44(12) C14-C15-C16 119.69(12) C1-C2-C3 103.65(11) C7-C8-C9 119.26(11) C15-C16-C17 120.65(12) N2-C3-C2 112.29(11) C7-C8-C11 120.15(11) C12-C17-C16 118.99(11) N2-C3-C18 120.14(12) C9-C8-C11 120.59(11) Cll-Hlla-Cll 50.4(7) C2-C3-C18 127.56(12) C8-C9-C10 120.43(11)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 2.6. Synthesis of amphiphilic acvlpvrazolone ligands

The -CH2CI group on the acyl group of the acylpyrazolones was reacted with the

sodium salt of monodisperse (PEG 516, PEG913 ) and polydisperse (PEG 750) polyethylene

glycol in dry THF under reflux conditions for about 12-14 hr (Scheme 2.4) by adopting a

modified procedure of Chiba et a l4&

dry THF .ONa .OH Na Me" Me" 12 hr, rt

mPEGx-OH mPEGx-ONa

/CH3 N / C h 3 -ONa dry THF Me" -PEGy Ph Cl Ph' -n' / y r t reflux, 24 hr OH 0 OH 0 4-acylpyrazolone amphiphilic acylpyrazolone ligand

x is the molecular weight of PEG, x = 516 (monodisperse), 913 (monodisperse), 750 (polydisperse), R is aliphatic side chain (valeryl group), and aromatic side chain (methyl benzene)

Scheme 2.4. Synthesis of amphiphilic acylpyrazolone ligands

Completion of the reaction was confirmed by LC/MS, TLC, and the formation of

NaCl. All amphiphilic acylpyrazolone ligands were red brown waxy solids, and were

characterized by LC/MS (Appendix C) which indicated the presence of a single

amphiphilic compound, when the monodisperse polyethylene glycol methyl esters were

employed and a mixture of amphiphilic compounds when polydisperse polyethylene

glycol methyl ester was employed. These amphiphilic acrypyrazolone ligands exhibited

high solubility in water as well as organic solvents and good thermal stability .49

Synthesized amphiphilic acylpyrazolone ligands with different PEG moieties are shown

in figure 2.3.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. OPEG

OPEG Ph' OH 0 OH 0 HMPVP-PEG HBMPP-PEG

x = 516 (monodisperse), 750 (polydisperse), 913 (monodisperse)

Figure 2.3. Chemical structures of amphiphilic acylpyrazolone ligands

2.7. Experimental methods

2.7.1. General

3-Methyl-1-phenyl-2-pyrazolin-5-one (1), all acid chlorides (2,3,4, 5, and 6 ), and

other reagents were analytically pure compounds obtained from Aldrich, and used

without further purification. The tetrahydrofuran (THF) solvent was distilled over Na

metal and benzophenone. Triethylamine (Et 3N) was dried over KOH and distilled over

CaH2 to obtain in anhydrous form .50 Mixture of products was chromatographically

separated with a silica column and crude products were purified by recrystallization. The

elaborated synthetic procedures of all compounds are provided in detail with the relevant

spectral data and the spectra are separately included in the appendix A.

2.7.2. Instrumentation and characterization of compounds

Melting points, reported in degree Celsius, were determined in open capillaries

using a Thomas-Hoover Unimelt instrument. 'H NMR and 13C NMR spectra were

recorded on a JEOL Eclipse 400 MHz NMR spectrometer. Chemical shifts were reported

using residual CDCI 3 (5 = 7.25), ( 8 = 77.0), and MeOD (8 = 49.0) as internal standards.

NMR spectral data were assigned according to the Pretsch text .51 IR spectra were

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. recorded on a Mattson-Satellite 3000 FTIR spectrophotometer. MS data were recorded

using a Shimadzu LC/MS-2010A spectrometer. Purification by column chromatography

was carried out with silica gel (32-63p). All reactions were monitored by normal phase

silica gel TLC plates (2:1 Hexane/ethyl acetate). All yields refer to isolated and highly

pure compounds.

O 2.7.3. 0-(ChlorovalerovlV3-methvl-l-phenvlpyrazol-5-one (021

The 3-methyl-l-phenyl-2-pyrazolin-5-one (1, 1.0 g, 5.7mmol) was taken into a

round bottom flask and flushed with argon gas and dissolved in dry THF (25 ml) and

anhydrous triethylamine (0.889 g, 1.0 ml, 7.0 mmol, 1.23 eq) was added and the reaction

mixture was stirred at room temperature for half an hour. 5-chlorovaleroyl chloride (2,

1.085 g, 0.899 ml, 7.0 mmol, 1.23 eq) was added to the reaction flask and stirring was

continued for 2 hours at room temperature. The reaction was monitored by TLC (2:1

hexane/ethyl acetate). After completion of the reaction, IN HC1 (25 ml) was added when

two layers separated. The aqueous layer was extracted with dichloromethane (3 x 30 ml).

The combined dichloromethane extract was washed with water (100 ml) and dried over

anhydrous MgSCL. Thick orange yellow oily compound (02) was obtained by

evaporating the organic solvent in a rotary evaporator and the crude product was purified

by column chromatography (silica gel, 2 :1 hexane/ethyl acetate) to obtain pure pale

yellow oily product (02, 80%). *H NMR (CDC13, 400 MHz) 5 7.50 (d, J = 8.08 Hz,

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. aromatic 2H), 7.42 (t, J = 7.86 Hz aromatic 2H), 7.29 (t, J = 7.32 Hz , aromatic 1H), 6.07

(s, 3H), 3.48 (t, J = 6.22 Hz, 2H), 2.53 (t, J = 6.96 Hz, 2H), 2.30 (s, 3H), 1.81-1.71 (m,

4H); 13C NMR (CDC13, 100 MHz) 6 168.6, 149.0, 144.4, 137.9, 129.2 (2C), 127.4, 123.4

(2C), 95.9, 44.3, 33.2, 31.5, 21.9, 14.7; IR (NaCl) 3054 cm ' 1 (m, aromatic C-H), 2962 cm'

1 (m, aliphatic C-H), 1780 cm ' 1 (C=0), 1265 cm' 1 (ester C-O), 756 cm' 1 (bs, C-Cl); MS

(ESI'): m/z = 291 (M+-l, 100%; 35C1), 293 (M+-l, 30%; 37C1).

,CH

Ph OH O 2.7.4. 4-(ChlorovalerovlV3-methyl-l-phenvlpvrazolin-5-ol (CD

Compound 0 2 (8.0 g, 27.39 mmol) was dissolved in dry THF (50 ml) and

anhydrous Ca(OH )2 (2.03 g, 27.39 mmol, 1 eq) was added to the reaction mixture. The

reaction mixture was refluxed at 80°C for 1 hr. It was monitored via TLC (2:1

hexane/ethyl acetate). After completion of the reaction, the mixture was allowed to cool

to room temperature, quenched with 2N HC1, and diluted with excess of 2N HC1 until

solid precipitate was formed. The mixture was left in the refrigerator for overnight to

obtain more solid. The precipitate was filtered and dried under vacuum and purified by

recrystallization from THF-H 2O to obtain completely pure product C2 (7.76 g, 8 8 %) mp:

96°C. 'H NMR ( C D C I 3 ,400 MHz) 8 7.80 (d, J = 8.04 Hz, aromatic 2H), 7.45 (t, J = 7.88

Hz aromatic 2H), 7.28 (t, J = 7.32 Hz , aromatic 1H), 5.17 (bs, OH), 3.59 (t, J = 5.86 Hz,

2H), 2.78 (t, J = 6.78 Hz, 2H), 1.92-1.89 (m, 4H); 13C NMR (CDC13, 100 MHz) 8 196.9,

160.4, 147.4, 137.3, 129.2 (2C), 126.7, 120.8 (2C), 103.8, 44.7, 38.3, 32.1, 21.7, 15.9; IR

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. (NaCl) 3435 cm' 1 (-0H), 3058 cm ' 1 (m, aromatic C-H), 2955 cm ' 1 (m, aliphatic C-H),

1625 cm' 1 (C=0), 1570 cm' 1 (enol C=C), 755 cm ' 1 (s, C-Cl); MS (ESI"): m/z = 291 (M+-l,

100%; 35C1 ), 293 (M+-l, 30%; 37C1); E.A Calculated for Q 5H 17CIN2O2: C, 60.61; H,

5.93; N, 9.42; Cl, 11.93; Found: C, 60.97; H, 5.70; N, 9.37; Cl, 11.76.

0 2.7.5. 0-(BromovalerovlV3-methyl-l-phenvlpvrazol-5-one (041

' The 3-methyl-l-phenyl-2-pyrazolin-5-one (1, 2.599 g, 14.9 mmol) was taken in a

round bottom flask, flushed with argon, and dissolved in dry THF (30 ml) and anhydrous

triethylamine (1.856 g, 2.55 ml, 18.4 mmol, 1.23 eq) was added and stirred at room

temperature for half an hour. 5-Bromovaleroyl chloride (4, 2.98 g, 2.0 ml, 16.4 mmol,

1 .1 2 eq) was added to the reaction flask and stirred for an hour at room temperature, and

reaction was monitored by TLC (2:1 hexane/ethyl acetate). After the completion of

reaction, it was quenched with IN HC1 (30 ml) when two layers separated. The aqueous

layer was extracted with dichloromethane (3x30 ml). The combined organic layers were

washed with water (3 x 30 ml) and dried over anhydrous MgSC> 4. The pale brown oily

compound was obtained by evaporating the organic solvent by rotary evaporator and was

purified by column chromatography (silica gel, 2 :1 hexane/ethyl acetate) to obtain pure

0 4 as a pale orange oil (4.95 g, 90%). lK NMR (CDC13, 400 MHz) 5 7.51 (d, J = 8.40

Hz, aromatic 2H), 7.43 (t, J= 7.86 Hz, aromatic 2H), 7.32 (t, J= 7.34 Hz, aromatic 1H),

3.36 (t, J = 6.22 Hz, 2H), 2.54 (t, J = 6.96 Hz, 2H), 2.31 (s, 3H), 1.87-1.78 (m, 4H); 13C

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. NMR (CDC13, 100 MHz) 6 168.5, 149.0, 144.3, 138.0, 129.2 (2C), 127.4, 123.4 (2C),

95.9, 33.1, 32.7, 31.6, 23.1, 14.6; IR (NaCl) 3053 cm ' 1 (m, aromatic C-H), 2985 cm ' 1 (m,

aliphatic C-H), 1781 cm ' 1 (C=0), 1265 cm' 1 (ester C-O), 754 cm' 1 (s, C-Br);MS (ESI'):

m/z = 335 (M+-l, 100%; 79 Br), 337 (M+-l, 99%; 81 Br).

,CH

OH 0 2.7.6. 4-(Bromovalerovli-3-methvl-1 -phenvlpvrazolin-5-ol (C41

Compound 04 (4.95 g, 14.6 mmol) was dissolved in dry THF (50 ml) and

anhydrous Ca(OH )2 (1.00 g, 14.6 mmol, 1 eq) was added to the reaction mixture. The

reaction was refluxed at 80°C for 4 hr. The reaction was monitored by TLC (2:1

hexane/ethyl acetate). After completion of the reaction, it was cooled to room

temperature, quenched with 2N HC1, and diluted with excess of 2N HC1 until a solid

precipitate formed. It was placed in the refrigerator overnight to obtain more solid. Then

precipitate (pale yellow solid, C4) was isolated by filtration, dried under vacuum and

obtained highly pure compound (4.18 g, 85%), mp: 104-105°C. 'H NMR (CDCI 3, 400

MHz) 8 7.81 (d, J = 7.68 Hz, aromatic 2H), 7.44 (t, J= 8.06 Hz, aromatic 2H), 7.28 (t, J =

7.5 Hz, aromatic 1H), 3.45 (t, J = 6.42 Hz, 2H), 2.77 (t, J = 7.16 Hz, 2H), 2.47 (s, 3H),

2.01-1.86 (m, 4H); 13C NMR (CDC13, 100 MHz) 5 196.9, 160.4, 147.4, 137.3, 129.2

(2C), 126.8, 120.8 (2C), 103.8, 38.2, 33.2, 32.2, 22.9, 15.9; IR (NaCl) 3432 cm ' 1 (b, -

OH), 3053 cm"1 (m, aromatic C-H), 2955 cm "1 (m, aliphatic C-H), 1625 cm ' 1 (conjugated

C=0), 1566 cm' 1 (enol C=C), 754 cm ' 1 (C-Br).; MS (ESI'): m/z = 335 (MM, 100%;

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 79 Br), 337 (M+-l, 99%; 81 Br); E.A.Calculated for Ci 5Hi7BrN20 2 : C, 53.43; H, 5.08; N,

8.31; Br, 23.70; Found: C, 53.56; H, 4.97; N, 8.15; Br, 21.30.

Ph^NN j^

° Y ~ ^ ' c h 3 2.7.7. 3-Methvl-l-phenvl-0-valerovlpvrazol-5-one (051

The 3-methyl-1-phenyl-2-pyrazolin-5-one (1, 2.0 g, 11.49 mmol) was taken in a

round bottom flask, flushed with argon, and dissolved in dry THF (50 ml). Anhydrous

triethylamine (1.43 g, 1.98 ml, 14.13 mmol, 1.23 eq) was added and the reaction mixture

was stirred at room temperature for half an hour. The valeroyl chloride (5, 1.40 g, 1.40

ml, 11.49 mmol, 1 eq) was added slowly to the reaction and mixture was stirred for an

hour at room temperature. The reaction was monitored by TLC (2:1 hexane/ethyl

acetate). After completion of the reaction it was quenched with IN HC1 (50 ml) when two

layers separated. The aqueous layer was extracted with dichloromethane (3 x 30 ml). The

combined organic layers were washed with water ( 1 0 0 ml) and dried over anhydrous

MgSCL. Thick orange yellow oily compound (05) was obtained by evaporating the

organic solvent by rotary evaporator, and was purified by column chromatography (silica

gel, 2:1 hexane/ethyl acetate) with high purity (3.38 g, 87%). lH NMR (CDCI 3, 400

MHz) 8 7.51 (d, J = 7.32 Hz, aromatic 2H), 7.40 (t, J = 7.88 Hz aromatic 2H), 7.28 (t, J =

7.5 Hz , aromatic 1H), 6.07 (s, 1H), 2.48 (t, J = 7.32 Hz, 2H), 2.31 (s, 3H), 1.62 (p, Ji =

7.32 Hz, J2 = 7.72 Hz, 2H), 1.32 (sextet, Ji - 7.32 Hz, J2 = 7.68 Hz, 2H), 0.88 (t, J = 7.32

Hz, 3H); 13C NMR (CDC13, 100 MHz) 8 169.1, 148.9, 144.5, 138.1, 129.1 (2C), 127.2,

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 123.3 (2C), 95.9, 33.8, 26.6, 22.1, 14.5, 13.7; IR (NaCl) 3054 cm ' 1 (aromatic C-H), 2934

cm' 1 (m, aliphatic C-H), 1781 cm ' 1 (C=0), 1265 cm' 1 (ester C=0); MS (ESI-): m/z = 257

(M+-l); 259 (M++l).

.CH

Ph CH OH O 2.7.8. 3-Methvl-l-phenvl-4-valerovlpvrazolin-5-ol (C51

Compound 0 5 (2.88 g, 8.55 mmol) was dissolved in dry THF (50 ml) and

anhydrous Ca(OH )2 (635 mg, 8.55 mmol, 1 eq) was added to the reaction mixture. It was

refluxed at 80°C for 6 hr. and monitored by TLC (2:1 hexane/ethyl acetate). After

completion of the reaction it was cooled to room temperature, quenched with 2N HC1,

and diluted with excess of 2N HC1 until a solid precipitate formed. The mixture was

placed in the refrigerator overnight to obtain more solid. The precipitated pink solid was

isolated by filtration and dried under vacuum to obtain pure product C5 ( 2.68 g, 97%),

mp: 60-61 °C (lit:62-63°C).46 lH NMR46 (CDC13, 400 MHz) 6 7.82 (d, J = 8.04 Hz,

aromatic 2H), 7.43 (t, J = 7.88 Hz aromatic 2H), 7.26 (t, J = 7.50 Hz , aromatic 1H), 2.73

(t, J = 7.52 Hz, 2H), 2.46 (s, 3H), 1.72 (p, ^ = 7.16 Hz, J 2 = 7.68 Hz, 2H), 1.43 (sextet, Ji

= 7.32 Hz, J2 = 7.72 Hz, 2H), 0.97 (t, J = 7.32 Hz, 3H); 13C NMR 14 (CDC13, 100 MHz) 5

197.4, 160.9, 147.5, 137.4, 129.2 (2C), 126.6, 120.7 (2C), 103.9, 38.7, 26.9, 22.6, 15.9,

14.0; IR (NaCl) 3440 cm"1 (-OH), 3068 cm "1 (m, aromatic C-H), 2952 cm ' 1 (s, aliphatic C-

H), 1627 cm' 1 (conjugated C=0), 1567 cm ' 1 (enol C=C) ; MS (ESI"): m/z = 257 (M M );

ESI+: 259 (M++1).

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0 2.7.9. 0-(4-Chloromethyltbenzovl-3-methyl-1 -phenylpyrazol-5-one (03)

The 3-methyl-l-phenyl-2-pyrazolin-5-one (1, 500 mg, 2.87 mmol) was taken in a

round bottom flask, flushed with argon, dissolved in dry THF (50 ml), anhydrous

triethylamine (357.61 mg, 492.60 pL, 3.53 mmol, 1.23 eq) was added and the reaction

mixture stirred at room temperature for half an hour. 4-Chloromethylbenzoyl chloride (3,

597 mg, 3.16 mmol, 1.1 eq) was added and stirred for an hour at room temperature and

the reaction was monitored by TLC (2:1 hexane/ethyl acetate). After completion of the

reaction it was quenched with IN HC1 (50 ml) and when two layers separated. The

aqueous layer was extracted with dichloromethane (3 x 50 ml). The combined all organic

layers were washed with water (3 x 30 ml) and dried over anhydrous MgSCL. The pale

brown solid was obtained by evaporating the organic solvent by rotary evaporator and the

crude product was purified by column chromatography (silica gel, 4:1 Hexane/ethyl

acetate) to obtain pure 0 3 as a white solid (80%), mp: 98°C. *H NMR (CDCI 3, 400 MHz)

8 8.06 (d, J = 8.08 Hz, aromatic 2H), 7.58 (d, J = 8.08 Hz aromatic 2H), 7.50 (d, J = 8.44

Hz , aromatic 2H), 7.43 (t, J = 7.88 Hz, 2H), 7.31 (t, J = 7.50 Hz, 1H), 6.28 (s, 1H), 4.61

(s, 1H), 2.37 (s, 1H); 13C NMR (CDC13, 100 MHz) 8 161.4, 149.2, 144.4, 143.9, 138.1,

131.1(2C), 129.2 (2C), 129.0 (2C), 127.9, 127.4, 123.3 (2C), 96.0, 45.2, 14.6; IR (NaCl)

3054 cm' 1 (aromatic C-H), 2987 cm "1 (aliphatic C-H), 1756 cm ' 1 (C=0), 1252 cm"1 (ester

C=0), 738 cm ' 1 (bs, C-Cl); MS (ESI'): m/z = 325 (M+-l, 100%; 35C1), 327 (M+-1, 33%;

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 37C1); E.A. Calculate4 for C 18 H 15CIN2O2: C, 66.16; H, 4.63; N, 8.57; Cl, 10.85; Found: C,

66.15; H, 4.90; N, 8.50; Cl, 11.10.

OH 0 2.7.10. 4-(4-Chloromethvlfbenzovl-3-methyl-1 -phenvlpvrazolin-5-ol (C3t

The 3-methyl-l-phenyl-5-pyrazolin-5-one (1,2.0 g, 11.49 mmol) was dissolved in

dry THF (50 ml), anhydrous Ca(OH )2 (850 mg, 11.49 mmol, 1 eq) was added to the

solution and the suspension was stirred at 35°C for 15 minutes. 4-(Chloromethyl) benzoyl

chloride (3, 2.17 g, 11.49 mmol, 1 eq) dissolved in dry THF was added slowly to the

reaction mixture and was refluxed at 80°C for 18 hr. The reaction was monitored by TLC

(2:1 hexane/ethyl acetate). After completion of the reaction, it was cooled to room

temperature, quenched with 2N HC1, and diluted with excess of 2N HC1 until a yellow

solid precipitate was formed. The reaction mixture was placed in the refrigerator

overnight to obtain more solid. The yellow precipitate separated by filtration and dried

under vacuum. The crude compound was dissolved in methyl tert-butyl ether (400 ml)

and ether solution was extracted with 0.5M NaHC03 solution (3 x 200 ml). Aqueous

layer was neutralized with 2N HC1 until yellow precipitate was formed and was filtered,

dried in vacuum oven for overnight to obtain highly pure C3 (3.16 g, 84%), mp: 143°C.

'H NMR (CDCI3, 400 MHz) 6 7.86 (d, J = 8.04 Hz, aromatic 2H), 7.65 (d, J = 8.08 Hz

aromatic 2H), 7.54 (d, J = 8.04 Hz aromatic 2H), 7.47 (t, J = 7.70 Hz , aromatic 2H), 7.31

(t, J - 6.96 Hz, aromatic, 1H), 4.65 (s, 2H), 2.11 (s, 3H); 13C NMR (CDCI 3, 100 MHz) 6

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 191.6,161.4,147.9,141.4,137.7,137.2,129.2 (2C), 128.6 (2C), 128.5 (2C), 126.9,120.9

(2C), 103.7, 45.5, 16.0; IR (NaCl) 3437 cm' 1 (-OH), 3065 cm' 1 (m, aromatic C-H), 2927

cm' 1 (m, aliphatic C-H), 1611 cm ' 1 (conjugated C=0), 1553 cm "1 (enol C=C), 764 cm "1

(C-Cl); MS (ESI'): m/z = 325 (M M , 100%; 35C1), 327 (M+-l, 30%; 37C1); E.A.

Calculated for C 18 H 15CIN2O2: C, 66.16; H, 4.63; N, 8.57; Cl, 10.85; Found: C, 66.18; H,

4.63; N, 8.52; Cl, 9.73.

.CH N =

Ph CH

2.7.11. 3-Methyl-l-phenvl-Otolueovlpvrazol-5-one (061 0

The 3-methyl-l-phenyl-2-pyrazolin-5-one (1, 500 mg, 2.87 mmol) was taken in a

round bottom flask, flushed with argon, dissolved in dry THF (25 ml), anhydrous

triethylamine (357.6 mg, 492.6 pL, 3.534 mmol, 1.23 eq) was added and stirred at room

temperature for half an hour. The /?-tolueoyl chloride ( 6 , 488.66 mg, 418.0 pL, 3.16

mmol, 1 .1 eq) was added to the reaction and stirred for an hour at room temperature.

Reaction was monitored by TLC (2:1 hexane/ethyl acetate). After completion of the

reaction, it was quenched with IN HC1 (25 ml) when two layers separated and the

aqueous layer was extracted with dichloromethane (3 x 20 ml). The combined organic

layers were washed with water (3 x 20 ml) and dried over anhydrous MgS(> 4. The pale

yellow color solid compound (06) was obtained by evaporating the organic solvent by

rotary evaporator and crude product was purified by column chromatography (silica gel,

2:1 hexane/ethyl acetate) to obtain pure 06 (713 mg, 85%), mp: 93°C. ]H NMR (C D C I 3 ,

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 400 MHz) 8 7.96 (d, J = 8.04 Hz, aromatic 2H), 7.59 (d, J = 7.32 Hz aromatic 2H), 7.42

(t, J = 7.88 Hz , aromatic 2H), 7.31-7.23 (m, aromatic, 3H), 6.25 (s, 1H), 2.42 (s, 3H),

2.37 (s, 3H); 13C NMR (CDC13, 100 MHz) 8 162.0, 149.2, 145.5, 138.2, 130.5 (2C),

130.2, 129.7 (2C), 129.2 (2C), 127.3, 125.0, 123.3, (2C), 96.0, 21.9, 14.6; IR (NaCl)

3060 cm"1 (aromatic C-H), 2976 cm ' 1 (aliphatic C-H), 1779 cm "1 (C=0), 1264 cm' 1 (ester

C=0), 746 cm"1 (s, C-Cl); MS (ESI"): m/z = 291 (M+-l); (ESI+): m/z = 293 (M+-l).

OH O 2.7.12. 3-Methyl-l-phenvl-4-tolueovlpvrazo1in-5-ol 1C61

Compound 06 (200 mg, 0.685 mmol) was dissolved in dry THF (15 ml),

anhydrous Ca(OH )2 (51 mg, 0.685 mmol, 1 .0 eq) was added to the solution and the

suspension was stirred at 80°C for 6 hours. The reaction was monitored by TLC (2:1

hexane/ethyl acetate). After completion of the reaction, the mixture was cooled to room

temperature, quenched with 2N HC1, and diluted with excess of 2N HC1 until yellow

precipitate was formed. The reaction mixture was placed in the refrigerator overnight to

obtain more solid. Then yellow precipitate was filtered and dried under vacuum to obtain

pure product C6 ( 122 mg, 61%) mp: 104-105°C (lit: 105-106°C).52 ’H NMR 52 (CDC13,

400 MHz) 8 7.87 (d, J = 7.68 Hz, aromatic 2H), 7.56 (d, J = 8.08 Hz aromatic 2H), 7.47

(t, J = 8.06 Hz , aromatic 2H), 7.31 (t, J = 8.08 Hz, aromatic, 1H), 7.31 (t, J = 8.08 Hz,

aromatic, 2H), 7.07 (d, J = 8.08 Hz, aromatic, 2H), 2.45 (s, 3H), 2.14 (s, 3H); 13C NMR

(CDC13, 100 MHz) 8 191.7, 161.8, 148.0, 142.8, 137.4, 134.7, 129.2 (2C), 129.2 (2C),

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 128.3 (2C), 126.7, 120.8 (2C), 103.6, 21.8, 16.1; IR (NaCl) 3431 cm ’1 (-OH), 3061 cm' 1

(m, aromatic C-H), 2925 cm ' 1 (m, aliphatic C-H), 1612 cm "1 (C=0), 1556 cm"1 (enol

C=C), 763 cm' 1 (s, C-Cl); MS (ESI'): m/z = 291 (M+-l); (E S f): m/z = 293 (M+-l).

OH 0 2.7.13. General procedure for the synthesis of amphiphilic acvlpyrazolones

Polyethylene glycol methyl ether (535 mg) was flushed with argon gas prior to

dissolve in dry THF (15 ml). Small pieces of Na metal was added to reaction flask and

stirred at room temperature for about 12 hr. The reaction mixture changed to light brown

color from colorless liquid. Excess of undissolved Na metal pieces were filtered and the

clear filtrate was mixed with 4-haloacylpyrazolone (l.leq) in dry THF. The reaction

mixture was refluxed at 80°C for about 24 hr, when some solid (NaCl) was formed. The

solid was removed by filtration and the filtrate was concentrated by evaporating the

excess THF to obtain red brown thick waxy product (amphiphilic acylpyrazolones).

2.8. X-ray studies

2.8.1. Collection of X-rav diffraction data

The sample was mounted on a nylon loop with a small amount of silicone grease.

All x-ray measurements were made on a Bruker-Nonius X 8 Apex2 CCD diffractometer

at a temperature of 11 OK. The unit cell dimensions were determined from a symmetry

constrained fit of 7355 reflections with 4.98° < 20 < 58.26°. The data collection strategy

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. was a number of co and (p scans which collected data up to 61.3° (20). The frame

C ’l integration was performed using SAINT+. The resulting raw data was scaled and

absorption corrected using a multi-scan averaging of symmetry equivalent data using

SADABS.54

2.8.2. Structure solution and refinement

The structure was solved by direct methods using SIR92.55 All non-hydrogen

atoms were obtained from the initial E-map. The hydrogen atoms were introduced at

idealized positions and were allowed to refine isotropically. The structural model was fit

to the data using full matrix least-squares based on F. The calculated structure factors

included corrections for anomalous dispersion from the usual tabulation. The structure

was refined, final tables and graphic plots were produced using the NRCVAX

crystallographic program suite .56 Additional information and other relevant literature

references can be found in the REPORT.OUT file and the reference section of the

Facility's Web page (http://www.xrav.ncsu.edul.

Crystallographic data for the structures (compound C2, C3, C5 and 0 3 ) has been

deposited with the Cambridge Crystallographic Data Centre as supplementary publication

numbers CCDC 644510, CCDC 644511, CCDC 644513, CCDC 644512 with

respectively. Copies of the data can be obtained, free of charge, on application to CCDC,

12 Union Road, Cambridge CB2 1EZ, UK [Fax: +44 1223 336033 or e-mail:

deposit@ccdc. cam. ac.uk].

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CHAPTER III

SYNTHESIS OF 4-HALOACYL-5-ISOXAZOLONE LIGANDS AND THEIR AMPHIPHILIC DERIVATIVES

3.1. Introduction

Since the amphiphilic acylpyrazolones are being promising chelating ligands and

provided good separation for the mixture of lanthanide metal ions, we proceeded further

on this research project using 4-acylisoxazolone ligands, which are similar to pyrazolone

ligands. 4-acylpyrazolones and 4-acylisoxazolones are extensively used in the metal ion

separation. This class of ligands is fascinating because of their common unique property

of extracting metal ions over a wide range of acidities with the proper choice of

substituents. Similar to acylpyrazolones, acylisoxazolones, a /?-diketones derived from 4-

acyl derivatives of isoxazolones, are also considered to be an important and interesting

alternative class of lanthanide metal ion chelating reagents. Due to their lower pKa value

17 7 S i a (~ 1 .0 - 2 .0 ), ’ ' acylisoxazolones have been reported to be an efficient extractants for

9 9 9Q < 7 metal ions from acidic media. ’ ’

The low pKa value of 4-acylisoxaozlones is because of the electron delocalization

by the isoxazolone group in the ring .26,29 The lower pKa value of these acylisoxazolones

varies due to the different substituents (electron withdrawing or donating groups) on the

isoxazolone ring. The lower pKa value for the enolic -OH group in acylisoxazolones

compared to acylpyrazolones is attributed to the higher electro negativity of the oxygen

atom compared to the nitrogen atom.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 3.2. Chemical properties of isoxazolone

Though the chemistry of 5-isoxazolones has been studied very well by earlier

chemists,58,59 it does not allow understanding of the structural assignments of the reaction

products because of the existence of various tautomers in this class of compounds .60

There are three tautomeric forms of 5-isoxazolones recognized and designated as CH-,

OH-, NH-forms as shown in figure 3.1 .61' 63 Generally, isoxazolones predominantly exist

in the CH-form in less polar solvents like chloroform, whereas in more polar solvent such

as dimethylsulfoxide in the OH-form .61,63 The OH- and NH-forms are predominant in the

solid state, probably due to the better hydrogen-bonded associations .64

NH-form CH-form OH-form

Figure 3.1. Possible tautomeric structures of isoxazolone

The chemistry of 5-isoxazolones is attractive as the active methylene carbon, 4th

position in the ring, represents the /?-carbon atom of cyclic enamines and is electron rich,

which is the preferred reaction site for many electrophiles. Alkylation and acylation of

isoxazol-5-ones have been reported in several cases .61 Normally, three attacking sites are

available on the ring and more than one product may be formed due to high

delocalization of electrons and formed stabilized resonance ring structure. Further studies

are still necessary to clarify the rules, which are governing the selectivity of these

complicated acylation reactions.58, 61 Usually, N-acylation seems to be preferred 61 and

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. matched with our results. In general, aliphatic acid anhydrides and chlorides react at

nitrogen, but aroyl halides provide major proportion of O-acylated products with minor

iV-acylated .65 The groups present at C-3 have great effect. Different reaction conditions

and presence of groups at C-3 may strongly influence the reaction .65

The structural chemistry of 4-acylisoxazolone ligands, similar to /?-diketones,

involves keto-enol tautomerism 19’ 23,25 includes four isomers as shown in figure 3.2. The

presence of an enolic OH peak at ~ 10-12 ppm of TMS in the 'H NMR in CDCI 3

confirms the existence of the enolic form. In several cases, 4-acylisoxazolones have been

shown to behave like a bidentate enol forming neutral metal complexes of the type shown

in figure 3.2. Our previous studies match well with this statement.

OH 0

Figure 3.2. Possible tautomeric forms of 4-acylisoxazolone and its schematic representation of metal complex formation

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 3.3. Synthetic approach, results and discussion

The parent 3-methyl-5-isoxazolone (8), and its 4-haloacylisoxazolones derivatives

have been synthesized. The amphiphililc acylisoxazolones with PEG moieties were

synthesized using similar procedure that was used for the synthesis of amphiphilic

acylpyrazolones to examine their metal ion recognition efficacies and study their

nanoscale self-assembled structural properties. The 3-methyl-5-isoxazolone (8) was

synthesized according to the most used method 66 that involves the long known

condensation reaction of /Nketoesters with hydroxylamine. According to Allan E.

Hydom’s procedure 66 (Scheme 3.1), the parent compound 8 was synthesized by using

ethylacetoacetate (7) and hydroxylamine. This compound has a lower shelf life, but when

it is in the chloroform solvent, it has longer shelf life. Methylisoxazolone (8) was

obtained in high yield and characterized by *H NMR, 13C NMR, IR and MS and

compared with* literature • data. f \ f’ i ( \ !

P H 3 0 0 75% Methanol N=\ + h o h 2n .h c i 4 hr at rt ^ v rr 7 78% (lit: 50%) 8

Scheme 3.1. Synthesis of 3-methyl-5-isoxazolone (8)

The same reaction conditions, which were discussed in Chapter II for the

acylation of pyrazolone, were employed to obtain 4-acylisoxazolone ligands. When the 3-

methyl-5-isoxazolone (8) was acylated in the presence of Ca(OH )2 in our initial trials, we

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. preferentially obtained A-acylated product (9, 3-methyl-A-chlorovaleroyl-5-isoxazolone)

according to Imo’s procedure 68 as shown in scheme 3.2.

.CH, Cl N= Ca(OH)2, dry THF ,CH + Cl rt, 30 min 0 8 93% 0

,CH3 /CH3 DMAP DMAP, dry C6H6 £ ^ dry C6H6, 80°C 40°C for 8 hr OH 0 O 50°C for 16 hr 17 hr, 78% 9 72%

Scheme 3.2. Synthesis of N- and 4-chlorovaleroyl-3-methyl-5-isoxazolones (9 and 10)

After synthesizing A-acylated compound 9, the chlorovaleroyl side chain was

transferred from nitrogen to carbon (4th position in the isoxazolone ring) according to the

procedure of Sato et al, 69 to obtain the desired 4-chlorovaleroyl-3-methyl-5-isoxazolone

(10). As mentioned in the literature, the reaction was carried out at higher temperature

(80°C) with 4 -(N, A-dimethyl)aminopyridine (DMAP) where the side chain turned into

cyclization and eventually pure 3-methyl-4-(tetrahydropyran-2-ylidene)-5-isoxazolone

(11) was obtained with high yield (Scheme 3.2). Direct C-acylation 69 using DMAP (dry

benzene, 0°C, rt, and 80°C) also provided cyclised product (11) instead of C-acylated

product (10). The transfer of the acyl side chain was not successful when Ca(OH )2 was

used as a base.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. To avoid the cyclisation in the side chain, reaction conditions were optimized.

When the temperature of the reaction was gradually decreased, the formation amount of

4-acylation was increased. When the reaction was carried out at 80°C, completely

cyclised product was observed, whereas the mixture of compounds ( 6 8 % of C-acylated

product and 5% of cyclised product) were obtained at lower temperature like 60°C to

65°C. Eventually 4-acylation occurred when the reaction was performed at 40 to 50°C

with 72% yield of pure desired product (10, Scheme 3.2).

As the substituents on the isoxazolone ring play a key role in metal ion complex

formation, another type of side chain that has electron withdrawing nature was introduced

on the 4 position of methylisoxazolone ( 8 ) using />-chloromethylbenzoyl chloride (3).

s e t The reaction was carried out with modified procedure of Imo’s procedure. It was

completed with in 30 minutes, but with A-(p-chloromethyl)benzoyl-3-methyl-5-

isoxazolone (12) and 0-(p-chloromethyl)benzoyl-3-methyl-5-isoxazolone (13) and 4-

chloromethylbenzoic acid (14, hydrolysis product of 4-chloromethylbenzoyl chloride) as

a side product (Scheme 3.3). CH; CH; N=- N = Cl + Ca(OH)2, dry THF + rt, 45 min 3 12 (80%) OH

Cl CH; DMAP, dry C6H6 ;N=

50-60°C for 16 hr OH 20% Scheme 3.3. Synthesis of N-, 0-, and 4-(p-chloromethyl)benzoyl-3-methyl-5- isoxazolones (12,13, and 15)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Direct C-acylation was also tried according to Sato’s procedure 69 (DMAP, 80°C,

dry benzene) with no success (starting material and hydrolysis product of acid chloride

were recovered). Finally, with the same above mentioned procedure 69 at lower

temperature (50-60°C), the acyl group was successfully transferred from nitrogen to

carbon (4 position of the isoxazolone ring) and desired product 15 was obtained with

low yield (Scheme 3.3).

The 3-phenyl-5-isoxazolone (16) is another starting material that was used in

amphiphilic ligand systems as it has a phenyl functional group (electron withdrawing

nature, which makes isoxazolone ligand more acidic) at the 3rd position in the

isoxazolone ring. Both electron withdrawing and donating side chains such as

chloromethylbenzoyl and valeroyl chloride have been introduced at the 4th position on the

phenylisoxazolone ring to obtain ligand structures similar to 4-acylpyrazolones. 3-

Phenyl-4-(p-chloromethyl)benzoyl-5-isoxazolone (17) was synthesized by reacting p-

chloromethylbenzoylchloride (3) with 3-phenyl-5-isoxazolone (16) according to a

modification of Imo’s procedure 68 (Scheme 3.4).

Cl Ca(OH)2, dry THF

rt, 30 min

Ph N = Cl Ph 0 . / = \ .Cl

0 OH + hc T ^ 0 14 17 (50%) 0 18 (25%) 1 9 (15%) Scheme 3.4. Synthesis of N-, O-, and 4-(/>chloromethyl)benzoyl-3-phenyl-5- isoxazolones (18,19, and 17)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The desired C-acylated product (17) was obtained along with the other side

products such as A-acylated (18), O-acylated (19), and chloromethylbenzoic acid (14)

compounds at room temperature, whereas at higher temperature like 75°C, after 7 hr

(longer reaction time) of the reaction time, only C-acylatedisoxazolone (17, 70%) and

compound (14) as a side product were obtained (Scheme 3.4)

The chlorovaleroyl side chain was introduced at the 4th position of 3-phenyl-5-

isoxazolone (16). The 4-acylation 68 of phenylisoxazolone (16) with 5-chlorovaleroyl

chloride (2) has been tried using Ca(OH )2 as a base to obtain 4-chloroveleroyl-3-phenyl-

5-isoxazolone (22). However, iV-acylated (20) and O-acylated (21) products were

obtained with this procedure. Direct 4-acylation 69 with DMAP as base, provided the

starting material (16) back and 5-chlorovaleroic acid (hydrolysis product of

chloroveleroyl chloride) as another side product. Then direct acylation reaction was split

into two step reaction such as the first step includes N-acylation, and the second step

includes acyl group transfer from nitrogen to carbon (4th position, scheme 3.5).

Ph Ph Ca(OH)2, dry THF Cl rt, 7 hr 0 0

16 21 (15%)

Cl Ph

50°C, 30 min 70%

Scheme 3.5. Synthesis of N-, O-, and 4-chlorovaleroyl-3-phenyl-5-isoxazolones (20,21, and 2 2 )

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. A7-Acylation was carried out using Ca(OH )2 at room temperature (Scheme 3.5)

along with O-acylatedisoxazolone (21) as minor side product. Both these compounds

were easily separated on silica gel column chromatography as well as recrystallization

from EtOH. After the pure jV-chloroveleroyl-3phenyl-5-isoxazlone (20) was obtained, the

following step of acyl group transfer was carried out using DMAP as base according to

Sato’s procedure 69 with a modification of the reaction conditions. The reaction was run at

lower temperature (50°C) for 30 minutes to avoid cyclization of the side chain. The yield

of the reaction was very high for the desired product 22.

3.4. X-ray crystallographic analysis of acylisoxazolone ligands

Nice single crystals were grown from THF/H 20, EtOH, and THF for N-

chlorovaleroyl-3-methyl-5-isoxazolone (9), 4-chlorovaleroyl-3-methyl-5-isoxazolone

(1 0 ), and the cyclised product (11) for structure determination by X-ray analysis. Single

crystal X-ray structural analysis of acylisoxazolones (9, 10) was performed as mentioned

in section 2 .8 in order to understand their molecular structures in terms of isomeric form,

bonding arrangement, and the position of the OH group. The crystal structures for

compounds 9, 10, and 11 are shown in figure 3.3. As it was reported in the earlier

literature,64 our results confirmed that the 4-acylisoxazolone (10) exists in the NH form in

its solid state.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Compound 9 Compound 10

Compound 11

Figure 3.3. Crystal structures of N-, 4-chlorovaleroyl-3-methyl-5-isoxazolones (9,10) and cyclised-3-methyl-5-isoxazolone (11)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 3.5. X-ray diffraction data and their treatment

The collection of diffraction data and their treatment are given in the following

tables 3.1, 3.2, and 3.3.

Table 3.1. Crystallographic data and structure refinement summary for compounds 9,10, and 11

Parameters Compound 9 Compound 10 Compound 11

empirical formula C9 H 12CINO3 c 9 h 12c i n o 3 C9 H h N 0 3 fw 217.65 217.65 181.19 cryst habit colorless prism pale yellow prism colorless plates cryst dimens (mm) 0.36 x 0.14 x 0.08 0.48x0.28x0.16 0.32 x 0.28 x 0.08 cryst syst monoclinic orthorhombic triclinic space group F lx!c Fix 2x 2x P-l a (A) 10.1838(3) 4.7294(10) 7.10270(10) b(A) 11.2810(3) 9.0118(2) 7.7387(2) c (A) 9.9180(3) 23.5920(6) 8.6393(2) a (deg) 90 90 72.8570(11) P (deg) 118.4832(15) 90 70.3820(11) y (deg) 90 90 74.2910(10) Z(A3) 1001.50(5) 1005.50(4) 419.571(16) Z 4 4 2 D (g.cm'3) 1.438 1.444 1.434 fi (cm'1) 0.36 0.36 0 .1 1 cGoF 1.29 1.71 2.06 T (K) 1 1 0 1 1 0 1 1 0 R7wR6 0.030/0.038 0.035/0.039 0.044/0.053 “RF = Sum(Fo-Fc)/Sum(Fo). ^Rw = Sqrt[Sum(w(Fo-Fc)**2)/Sum(wFo**2)]. cGoF = Sqrt[Sum(w(Fo- Fc)**2)/(No. of reflns - No. of params.)]

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 3.2. Bond distances (A ) for compounds 9,10, and 11

Compd distance (A) Distance (A) distance (A)

9 01-Cl 1.2031(12) N1-C5 1.3973(12) C6-C7 1.5215(13) 02-C1 1.4121(11) C5-03 1.2104(11) C7-C8 1.5291(13) 02-N1 1.4055(10) C5-C6 1.5076(13) C8-C9 1.5111(13) C1-C2 1.4323(14) C3-C4 1.4864(13) C9-C11 1.7998(10) C2-C3 1.3473(14) C3-N1 1.3784(12)

10 01-Cl 1.2254(13) C1-C2 1.4168(14) C6-C7 1.5198(15) 02-N1 1.3876(12) C2-C3 1.3960(14) C7-C8 1.5275(15) 02-C1 1.4054(12) C2-C5 1.4715(14) C8-C9 1.5047(15) 03-C5 1.2240(13) C3-C4 1.4838(16) C9-C11 1.8057(11) N1-C3 1.3187(14) C5-C6 1.5045(14)

11 01-Cl 1.2162(9) C2-C3 1.4423(9) C6-C7 1.5228(10) 02-N1 1.4421(8) C2-C5 1.3763(9) C7-C8 1.5201(11) 02-C1 1.3820(8) C3-C4 1.4887(10) C8-C9 1.5014(11) N1-C3 1.3003(9) C5-C6 1.4991(9) C9-03 1.4728(8) C1-C2 1.4512(9) C5-03 1.3238(8)

Table 3.3. Bond angles (deg) for compounds 9,10, and 11

Compd Angle (deg) Angle (deg) Angle (deg)

9 C1-02-N1 106.29(7) C2-C3-N1 108.54(8) N1-C5-C6 115.84(8) Ol-Cl-02 118.14(8) C4-C3-N1 121.80(8) 03-C5-C6 125.49(8) 01-C1-C2 135.39(9) 02-N1-C3 109.57(7) C5-C6-C7 111.23(8) 02-C1-C2 106.46(8) 02-N1-C5 117.43(7) C6-C7-C8 111.85(8) C1-C2-C3 109.09(8) C3-N1-C5 132.44(8) C7-C8-C9 109.90(8) C2-C3-C4 129.64(9) N1-C5-03 118.67(8) C8-C9-C11 111.29(7)

10 N1-02-C1 106.10(7) C3-C2-C5 126.95(9) C2-C5-C6 117.15(9) 02-N1-C3 111.08(8) N1-C3-C2 108.95(9) C5-C6-C7 114.16(9) Ol-Cl-02 117.12(9) N1-C3-C4 119.46(9) C6-C7-C8 110.91(9) 01-C1-C2 135.64(9) C2-C3-C4 131.59(9) C7-C8-C9 110.96(9) C1-C2-C3 106.59(9) 03-C5-C2 120.21(9) C8-C9-C11 111.49(8) C1-C2-C5 126.95(9) 03-C5-C6 122.63(9) 02-C1-C2 107.23(9)

11 N2-02-C1 109.90(5) N1-C3-C2 112.03(6) C5-C6-C7 114.19(6) 02-N1-C3 107.05(5) N1-C3-C4 119.00(6) C6-C7-C8 108.93(6) Ol-Cl-02 120.02(6) C2-C3-C4 128.97(6) C7-C8-C9 108.52(6) 01-C1-C2 133.89(6) C2-C5-C6 122.09(6) C8-C9-03 112.24(6) 02-C1-C2 106.10(6) C2-C5-03 116.05(6) C5-03-C9 118.98(12) C1-C2-C3 104.92(5) C6-C5-03 121.82(6) C3-C2-C5 122.11(5) C1-C2-C5 125.45(6)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 3.6. Synthesis of amphiphilic acylisoxazolone ligands

Amphiphilic acylisoxazolone ligands were synthesized according to modified

procedure of Chiba et aV s procedure that was used to synthesize amphiphilic

acylpyrazolone ligands. 4 -acylisoxazolones were reacted with different olyethylene

glycol moieties (PEG516, PEG750, PEG913) to obtain amphiphilic acylisoxazolones as

shown in scheme 3.6. PEG516 and PEG913 are completely monodisperse whereas PEG750

is polydisperse in nature.

+ Na

mPEGx-OH mPEGx-ONa

reflux, 24 hr OH O amphiphilic acylisoxazolone 4-acylisoxazone

x is the molecular weight of PEG, x = 516, 913 (monodisperse), 750 (polydisperse) R' is aliphatic side chain (valeryl group), and aromatic side chain (methyl benzene) R is methyl and phenyl group

Scheme 3.6. Synthesis of amphiphilic acylisoxazolone ligands

Completion of the reaction was confirmed by LC/MS, TLC ( 2:1 Hexane/ethyl

acetate), and the formation of NaCl salt at the bottom of the flask. All amphiphilic

acylisoxazolone ligands were yellow, orange, and pink orange red waxy solids, and were

characterized by LC/MS (Appendix C) which indicated the presence of a single

amphiphilic compound when the monodisperse polyethylene glycol methyl esters were

employed and a mixture of amphiphilic compounds when polydisperse polyethylene

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. glycol methyl ester was employed. These amphiphilic isoxazolone ligands exhibited high

solubility in water. The chemical structure of synthesized amphiphilic acylisoxazolone

ligands are shown in figure 3.4. Ph ,N=^ OPEGx

OH 0 OH 0 HMVIS-PEGX HPVIS-PEGx

Ph N = OPEG OPEG

OH 0 OH 0 HBMIS-PEGx HBPIS-PEGx

x = 516, 913 (monodisperse), 750 (polydisperse)

Figure 3.4. Amphiphilic acylisoxazolone ligands

3.7. Experimental methods

3.7.1. General

3-Phenyl-5-isoxazolone (16), acid chlorides (2, 3), and other reagents were

analytically pure compounds and purchased from Aldrich, and used without further

purification. All solvents and reagents were purified if necessary according to

Armarego’s procedure .50 Mixtures of products were chromatographically separated with

a silica column and crude products were purified by recrystallization. The elaborated

synthetic procedures of all compounds are provided in detail with the relevant spectral

data and the spectra are separately included in the appendix B. All pure acylisoxazolones

and their amphiphilic derivatives are characterized using same instrumentation (section

2.7.2) that was used for the characterization of acylpyrazolone compounds.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. .ch3

/ 3.7.2. 3-Methyl-5-isoxazolone (8) <^Nj|

This compound was prepared according to0 Allan E. Hydom et a /’s procedure. 66 A

mixture of ethylacetoacetate (7, 20.56 g, 20 ml, 0.158 mol) and hydroxylamine

hydrochloride (12 g, 0.174 mol, 1.1 eq) in 75% methanol (85 ml) was stirred at room

temperature for 4 hr. According to the TLC (2:1 hexane/ethyl acetate), after completion

of the reaction, all the solvent was evaporated by rotary evaporator and white needles like

compound (excess hydroxylamine) was filtered. The thick light yellow oily compound

was purified by column chromatography using ethyl acetate as an eluent to obtain pure

methylisoxazolone (8, 78%). 'H NMR (CDC13, 400 MHz) 8 3.39 (s, 2H), 2.12 (s, 3H);

13C NMR (CDC13, 100 MHz) 8 175.8, 164.2, 37.11, 14.8; IR (NaCl): 3582 cm '1 (-OH),

2955, 2930 cm'1 (m, aliphatic C-H), 1799 cm'1 (C=0); MS (ESI'): m/z = 98 (M M ),

(ESI+) 100 (M++l).

3.7.3. A-Chlorovalerovl-3-methvl-5-isoxazolone yy) q

Methylisoxazolone (8, 5.0 g, 0.05 mol) was dissolved in dry THF (80 ml), after it

was flushed with argon. Ca(OH )2 (3.73 g, 0.05 mol, 1 eq) was added into the reaction

mixture and was stirred for 10 minutes, then 5-chlorovaleroyl chloride (2, 8.61 g, 0.720

ml, 0.055 mol, 1.1 eq) was added drop wisely and the reaction was stirred at room

temperature for about 15 minutes. Completion of the reaction was monitored by TLC (2:1

hexane/ethyl acetate). After the completion of the reaction, stirring was stopped, excess

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. of undissolved Ca(OH )2 was filtered and solvent was evaporated from the filtrate to

obtain thick orange yellow oil, and the product 9 was purified by column chromatography

(1:1 Hexane/ethyl acetate) with 93% yield. After 10 days nice crystals were formed from

that thick yellow oil, and the crystals melt at 28-30°C. *H NMR (C D C I 3 , 400 MHz) 8

5.26 (s, 1H), 3.53 (t, J = 5.86 Hz, 2H), 2.74 (t, J = 6.78 Hz, 2H), 2.54 (s, 3H), 1.85-1.78

(m, 4H); 13C NMR (CDC13, 100 MHz) 8 167.5, 166.1, 158.8, 94.9, 44.4, 33.8, 31.7, 20.9,

15.6; MS (ESI'): m/z = 216 (M+-l, 100%; 35C1), 218 (M+-l, 30%; 37C1);

,CH3

3.7.4. 4-Chlorovalerovl-3-methvl-5-isoxazolone 6101 ^ q

This compound was prepared by a modified procedure of Sato’s method .69 N-

chlorovaleroyl-3-methyl-5-isoxazolone (9, 3.2 g, 14.8 mmol) was dissolved in anhydrous

benzene (60 ml), followed by DMAP (1.802 g, 14.8 mmol, 1 eq) under argon gas, and

reaction mixture was stirred at 40°C for 8 hr under argon, then temperature was increased

to 50°C and the reaction was stirred for 16 hr. Completion of the reaction was confirmed

by TLC (2:1 hexane/ethyl acetate). The reaction mixture was allowed to cool to room

temperature and was quenched with excess of DI water (200 ml), and aqueous layer was

washed with benzene twice (2 x 300 ml). An aqueous layer was acidified to pH 1 with 2N

HC1, and then was extracted with dichloromethane (2 x 300 ml), and this organic layer

was extracted with saturated Na2CC>3 solution (2 x 100 ml). The base solution was again

acidified with conc.HCl to pH 1, and was extracted with dichloromethane (3 x 100 ml),

and solvent was evaporated to obtain orange yellow solid. Crude compound was

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. recrystallized from ethyl acetate to obtain orange brown crystals (10, 2.3 g, 72%). m.p:

78-80°C; !H NMR (CDC13, 400 MHz) 5 10.33 (bs, NH), 3.58 (t, J = 6.04 Hz, 2H), 2.71 (t,

J = 7.26 Hz, 2H), 2.36 (s, 3H), 1.98-1.85 (m, 4H); 13C NMR (CDC13, 100 MHz) 5 193.2,

171.0, 161.7, 95.5, 44.1, 36.9, 32.0, 22.2, 12.2; MS (ESI'): m/z = 216 (M+-l, 100%; 35C1

), 218 (M M , 30%; 37C1).

.CH

3.7.5. 3-Methvl-4-(tetrahvdropvran-2-vlideneV5-isoxazolone (111

Reaction procedure to prepare this compound is same as above but at a higher

temperature (80°C) with 78% yield (11). m.p: 149-150°C; 'H NMR (CDC13, 400 MHz) 8

4.45 (t, J = 5.68 Hz, 2H), 3.24 (t, J = 6.98 Hz, 2H), 2.25 (s, 3H), 1.98 (p, Ji = 6.60 Hz, J 2

= 6.24 Hz, 2H), 1.88 (p, h = 6.60 Hz, J2 = 6.96 Hz, 2H); 13C NMR (CDC13, 100 MHz) 8

181.9, 172.8, 158.5, 98.6, 69.8, 24.2, 22.0, 15.9, 15.2; MS (E Sf): m/z = 182 (M*+l),

(ESIO 198 (M+-l, adduct with H 20 ).

,CH3 xCH3 N—rf N = o 3 o v ^

0 0 3.7.6. iV-fp-Chloromethvllbenzovl-3-methvl-5-isoxazolone (121 & Q-(p-Chloromethvllbenzovl-3-methvl-5-isoxazolone (131 ®

Methylisoxazolone (8, 500 mg, 5.05 mmol) was added to the round bottom flask, flushed

with argon gas and dissolved in dry THF (30 ml), then followed by Ca(OH ) 2 (373.73 mg,

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 5.05 mmol, 1 eq). /?-Chloromethylbenzoyl chloride (3, 1.05 g, 5.55 mmol, 1.1 eq) was

added slowly to the reaction mixture after it was stirred for 30 minutes at room

temperature. Consumption of starting material was confirmed by TLC (1:1 Hexane/ethyl

acetate). The reaction was completed within 15 minutes, then the reaction was stopped,

and excess of undissolved Ca(OH )2 was filtered from the reaction mixture. All the

solvent (THF) was evaporated from filtrate to obtain orange red yellow solid (crude yield

was 110%). Proton NMR showed mixture of compounds (TV-acylated + O-acylated +

chloromethyl benzoic acid). Products were separated by silica gel column

chromatography (2:1 hexane/ethyl acetate). A-(p-chloromethyl)benzoyl-3-

methylisoxazolone (12) was obtained as white powder (80%, mp: 80-82°C) and 'H NMR

(CDC13, 400 MHz) 8 7.90 (d, J = 8.44 Hz, aromatic 2H), 7.51 (d, J = 8.04 Hz, aromatic

2H), 5.41 (s, 1H), 4.62 (s, 2H), 2.70 (s, 3H); 13C NMR (CDCI3, 100 MHz) 6 166.3, 162.7,

160.3, 142.8, 130.8, 130.5 (2C), 128.6 (2C), 95.8, 45.2, 16.1; MS (ESI'): m/z = 250 (M+-

1, 100%; 35C1), 252 (M+-l, 30%; 37C1). 0-(p-chloromethyl)benzoyl-3-methylisoxazolone

(13) was obtained as a milky white glassy wool (17%, mp: 130-132°C). *H NMR (CDCI 3,

400 MHz) 8 8.18 (d, J - 8.04 Hz, aromatic 2H), 7.56 (d, J = 8.08 Hz, aromatic 2H), 6.05

(s, 1H), 4.64 (s, 2H), 2.32 (s, 3H); 13C NMR (CDC13, 100 MHz) 8 165.1, 162.3, 159.9,

144.4, 131.2, 131.1 (2C), 129.1 (2C), 88.5, 45.2, 12.6; MS (ESI'): m/z = 250 (M+-l,

100%; 35C1), 252 (M+-l, 30%; 37C1).

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ,CH N =

OH O

3.7.7. 4-(7>-Chloromethvl)benzovl-3-methvl-5-isoxazolone (T5t

This compound was synthesized according to a modified version of Sato’s

procedure (acyl transfer procedure ) . 69 Compound 12 (100 mg, 0.3984 mmol) was added

to the round bottom flask and purged with argon gas. The compound was dissolved in dry

benzene solvent (13 ml). The whole reaction set up was under argon blanket. While

stirring the solution, DMAP (230 mg, 1.88 mmol, 4.7 eq) was added to the reaction pot

and was stirred at 50-60°C for 16 hr. The reaction was monitored by TLC to confirm that

all the starting material was consumed. It was quenched with DI water (100 ml) and

aqueous layer was separated from benzene layer and was washed with benzene (3 x 30

ml). After acidifying with 4N HC1 to pH 1, the aqueous layer was extracted with

dichloromethane (3 x 50 ml). The combined organic layer was extracted with saturated

sodium bicarbonate solution (3 x 50 ml) and followed by dichloromethane extraction (3 x

100 ml). All the solvent was evaporated to obtain crude yellow solid, which was a

mixture of 4-(p-chloromethyl)benzoyl-3-methyl-5-isoxazolone (15) and 4-chloromethyl

benzoic acid (14). The crude yellow solid was washed with ether to get pure 4 -(p-

chloromethyl)benzoyl-3-methyl-5-isoxazolone (15) as yellow solid (20 mg, 20%). *H

NMR (CDCI3, 400 MHz) 5 7.63 (d, J = 8.44 Hz, aromatic 2H), 7.58 (d, J = 8.40 Hz,

aromatic 2H), 4.65 (s, 2H), 2.04 (s, 3H); MS (ESI'): m/z = 250 (M+-l, 100%; 35C1 ), 252

(NT-1 , 30%; 37C1).

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. O 0 3.7.8. 4-(p-Chloromethvltbenzovl-3-nhenvl-5-isoxazolone (IT). iV-(p-ChloromethvDbenzovl-3-phenvl-5-isoxazolone (181 ® & Q-(p-ChloromethyDbenzoYl-3-phenvl-5-isoxazolone (191

4-(p-Chloromethyl)benzoyl-3-phenyl-5-isoxazolone (17) was prepared according

to Imo’s procedure along with two other side products (18,19). 3-Phenyl-5-isoxazolone

(16, 1.0 g, 6.2 mmol) was dissolved in dry THF (40 ml) and purged with argon to make

sure there was no moisture in the reaction pot. Ca(OH )2 ( 1 .2 g, 16.14 mmol, 2 .6 eq) was

added to the stirring solution and stirring was continued for 5 minutes at room

temperature, then />-chloromethylbenzoyl chloride (3, 2.44 g, 12.92 mmol, 2.1 eq) was

added to the reaction mixture and reaction was stirred at room temperrature for half an

hour. Consumption of starting material was confirmed by TLC (2:1 hexane/ethyl acetate).

When all the starting material was consumed, the reaction was stopped and allowed to

cool to room temperature. When the reaction was quenched with 2N HC1,4-chloromethyl

benzoic acid (14, hydrolysis product of 4-chloromethylbezoyl chloride) was precipitated

out as fine glassy wool and was separated by filtration. The filtrate was extracted with

excess of dichloromethane (3 x 100 ml). The combined organic layer was washed with

DI water (200 ml), and dried with anhydrous MgSCV All the solvent was evaporated to

obtain pink solid, which is mixture of compounds (17, 18, and 19). Products were

separated by normal phase silica gel column chromatography ( 2 :1 hexane/ethyl acetate).

The desired 4-(p-chloromethyl)benzoyl-3-phenyl-5-isoxazolone (17) was the major

component (50%) and was obtained as a solid. !H NMR (CDCI 3, 400 MHz) 8 7.44 (d, J =

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 8.08 Hz, aromatic 2H), 7.28 (d, J = 6.60 Hz, aromatic 2H), 7.23-7.16 (m, aromatic 5H),

4.55 (s, 2H); MS (ESI'): m/z = 312 (M+-l, 100%; 3SC1 ), 314 (M+-l, 30%; 37C1); N-(p-

chloromethyl)benzoyl-3-phenyl-5-isoxazolone (18, 25%); *H NMR (CDCI 3, 400 MHz) 6

7.95 (d, J = 8.04 Hz, aromatic 2H), 7.56-7.46 (m, aromatic 7H), 5.76 (s, 1H), 4.63 (s,

2H); MS (ESI’): m/z = 312 (M+-1, 100%; 35C1 ), 314 (M+-l, 30%; 37C1).& 0-(p-

chloromethyl)benzoyl-3-phenyl-5-isoxazolone (19,15%) are minor products as solids.

O 3.7.9. iV-Chlorovalerovl-3-phenvl-5-isoxazolone (201 & 0-Chlorovalerovl-3-phenvl-5-isoxazolone (211 q

Phenylisoxazolone (16, 1.0 g, 6.2 mmol) was dissolved in dry THF (50 ml) and

purged with argon to make sure no moisture in the reaction pot. Ca(OH )2 (459 mg, 6 .2

mmol, 1 .0 eq) was added to the stirring solution and stirring was continued for 15 min. at

rt, then 5-chloroveleroyl chloride (2, 1.06 g, 0.876 ml, 6.82 mmol, 1.1 eq) was added

drop wisely. As soon as this reagent was added, reaction solution was slowly turned into

yellow color. Completion of the reaction was confirmed by TLC (2:1 hexane/ethyl

acetate). After 7 hrs of stirring at rt, when all the starting material was consumed, the

reaction was stopped. The excess of Ca(OH )2 was filtered off and all the THF solvent

was removed using rotary evaporator to obtain thick orange color semi solid compound

(mixture of A-acylated & O-acylated). Both compounds were separated using normal

phase silica gel column chromatography (2:1 hexane/ethyl acetate) to obtain pure N-

chloroveleroyl-3-methyl-5-isoxazolone (20, 85%) as solid. ]H NMR (CDCI 3,400 MHz) 5

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 7.55-7.43 (m, aromatic 5H), 5.58 (s, 1H), 3.54 (t, J = 5.48 Hz, 2H), 2.86 (t, J = 6.96 Hz,

2H), 1.89-1.81 (m, 4H); 13C NMR (CDC13, 100 MHz) 5 167.9, 166.1, 161.4, 131.7, 128.7

(2C), 128.4 (2C), 127.5, 96.4, 44.4, 34.4, 31.6, 21.0; MS (ESI'): m/z = 278 (M M , 100%;

35C1), 280 (M+-l, 30%; 37C1); Pure Ochloroveleroyl-3-phenyl-5-isoxazolone (21, 15%)

was obtained as yellow oil. 'H NMR (CDC13, 400 MHz) 8 7.77 (d, aromatic 2H), 7.48-

7.44 (m, aromatic 3H), 6.37 (s, 1H), 3.59 (t, J = 5.86 Hz, 2H), 2.70 (t, J = 7.14 Hz, 2H),

2.0-1.91 (m,4H).

3.7.10. 4-Chlorovalerovl-3-phenvl-5-isoxazolone (221

N-chloroveleroy 1-3-methyl-5-isoxazolone (20, 1.45 g, 5.2 mmol) was taken into a

round bottom flask and purged with argon thrice. Compound was dissolved in dry

benzene (25 ml) under the argon blanket, and then DMAP (635 mg, 5.2 mmol, 1.0 eq)

was added. The reaction mixture was stirred at 50°C for 30 minutes. Completion of the

reaction was confirmed by TLC (2:1 hexane/ethyl acetate). After the completion of the

reaction, stopped stirring and was allowed to cool to room temperature for a while. Then

the reaction was quenched with excess of water (50 ml), and 2N HC1 (50 ml) was added

to the flask. After the separation of benzene layer, the aqueous layer was washed with

benzene again (2 x 30 ml) and was extracted with dichloromethane (3 x 50 ml).

Combined organic layers were extracted with saturated sodium bicarbonate solution (3x

50 ml). The base solution was acidified to pH 1 with 2N HC1 solution. This acidic

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. solution was again extracted with dichloromethane (4 x 60 ml). Organic layer was

washed with water (2 x 50 ml) and all the dichloromethane solvent was evaporated using

rotary evaporator to obtain thick brown oily compound (22, 70%). 'H NMR (CDCI 3, 400

MHz) 5 11.18 (NH, 1H), 7.57-7.45 (m, aromatic 5H), 3.37 (t, J = 6.24 Hz, 2H), 2.34 (t, J

= 7.5 Hz, 2H), 1.73 (p, Ji = 6.96 Hz, J 2 = 8.01 Hz, 2H), 1.60 (p, Ji = 6.24 Hz, J 2 = 8.40

Hz, 2H); MS (ESI'): m/z = 278 (M+-l, 100%; 35C1), 280 (M+-l, 30%; 37C1).

OH O

3.7.11. General procedure for the synthesis of amnhinhilic acvlisoxazolones

Polyethylene glycol methyl ether was flushed with argon gas prior to dissolve in

dry THF. Small pieces of Na metal were added to reaction flask and stirred at room

temperature for about 12-14 hrs. The reaction mixture changed to light brown color from

colorless liquid. Excess undissolved Na metal was filtered off and the clear filtrate was

mixed with 4-haloacylisoxazolone (1.1 eq) in dry THF. The reaction mixture was

refluxed at 80°C for about 24 hr, when some solid (NaCl) was formed. The solid was

removed by filtration and the filtrate was concentrated by evaporating the excess THF to

obtain orange yellow brown thick waxy product (amphiphilic acylisoxazolone). The

product was characterized by LC/MS.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CHAPTER IV

LANTHANIDE METAL IONS SEPARATION BY HPLC METHOD USING AMPHIPHILIC ACYLPYRAZOLONES

4.1. Introduction

The acylpyrazolones were chosen as the chelating ligand structures for the

recognition of lanthanides by forming metal complexes due to their low pKa values (3.0-

4.0). Amphiphilic acylpyrazolone were synthesized from 4-haloacylpyrazolones as

described in Chapter 2. For the first time in the field of science, they have been employed

to investigate their lanthanide metal ion recognition efficacies through the HPLC

separation method. These ligands spontaneously self-assemble in the aqueous phase to

form spherical, dendritic and linear nanostructures. Their metal ion recognition could be

influenced by their self-assemblies. The significant details of these studies are discussed

in the results and discussion section of this chapter.

4.2. Instrumentation and experimental methods

4.2.1. HPLC system

The High Performance Liquid Chromatography (HPLC) instrument consisted of a

Perkin-Elmer quaternary series 200 LC pump, a Rheodyne model 7725 injection valve

equipped with a 20 pi sample loop and a Perkin-Elmer series 200 UV/Vis detector. Data

collection was performed with a Perkin-Elmer TOTAL CHROM Workstation. An Altex

A100 post column derivatization pump was used to pump the Arsinazo III (Figure 4.1)70’

71 indicator into the mobile phase.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.1. Chemical structure of arsinazo III indicator

4.2.2. pH meter

The pH measurements for the aqueous ligand solution were performed by an

ORION Ross glass electrode and an Accumet Basic AB15 model pH meter from Fisher

Scientific.

4.2.3. Transmission electron microscope (TEM)

A JEOL JEM-1230 Transmission Electron Microscope with an accelerated

voltage of 80 KV was used to observe nanoscale self-assembled ligand structures.

Samples were prepared on 400 mesh copper grid coated with carbon film. A drop of

sample solution (~ 10 pi) was carefully placed on to the grid surface and after about 2-5

minutes, the sample was air dried and excess solution was sucked back by the tip of filter

paper. Images were collected from the Western Michigan University and Washington

State University imaging centers.

4.2.4. HPLC column nacking

An Alltech HPLC Column Model 1666 Slurry Packer was used for the column

packing. This is a self-contained unit, designed to pack the column. 25 cm long columns

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. (4.6 mm inner diameter, 6.5 mm outer diameter) were purchased from Alltech Company,

were cut into 125 mm lengths, and the column ends were smoothed with rough salt paper.

The column and the end fittings were assembled with the reservoir and made sure there

was enough gas pressure (N 2) and solvent to pack. HPLC grade methanol was filtered

through 220 nm pore size Teflon filter papers before use for column packing.

Approximately 2.0 g of Cig silica gel (3 pm, 100 A) was dispersed in 20 ml of HPLC

grade methanol and kept in the ultrasonic bath for five minutes to make homogenous

slurry.

The slurry was carefully poured into the reservoir with the help of thin glass rod.

The reservoir was closed with a nitrogen gas inlet with 8000 psi pressure. The air supply

valve was turned to the “ON” position and the slurry packer pressure was maintained

around 6000 psi while the column was being packed. The prime valve was opened until

there was methanol flow to avoid air bubbles in the column, then the prime valve was

closed and the solvent flow needle valve was opened to the maximum. After pumping

500-600 ml of methanol through Cig silica gel column, solvent flow needle valve was

closed completely and air supply was turned to the “OFF” position to get the air pressure

zero psi. The reservoir was opened and the column was disconnected from the reservoir.

Silica gel on the top end (inlet of the column) of the column was smoothed with a spatula,

end fittings were attached, and tightening to ensure no leakage in the column while doing

experiment.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 4.2.5. Reagents

All amphiphilic acylpyrazolones and amphiphilic acylisoxazolone ligands were

synthesized. Sodium Perchlorate (ACS grade), Arsenazo III, and Chloroacetic Acid (ACS

grade) were purchased from Aldrich. Sodium Hydroxide (food grade) was purchased

from Mallinckrodt. All lanthanide metal ions were purchased from Aldrich in hydrated

chloride salt form. Cig silica gel was purchased from Alltech.

4.2.6. Procedure

Chloroacetic acid buffer solution of ionic strength 0.01 for pH 2-3 and formic acid

buffer solution of ionic strength 0.01 for pH 3-4 ,7 2 ,73 were prepared using HPLC grade

water. Milli-Q water was filtered through 220 nm pore size Teflon filter paper and was

used as HPLC grade water. Final ionic strength of the buffer solution was adjusted to 0.1

by adding 1.0M NaC 1 0 4 solution .74 Amphiphilic ligand was dissolved in a buffer solution

with desired pH and used as a mobile phase with 1 ml/min flow rate in HPLC system for

lanthanide metal ion separation studies. The mobile phase consisted of a suitable

concentration of ligand and a suitable pH. Arsenazo III (Figure 4.1) solution (pH: 3.45,

detection wavelength: 656 nm, flow rate: 0.5 ml/min) was used as a post column

derivatization for the detection of metal ions .7 0 ,71 Lanthanide metal ion solutions were

prepared from 0.01M (pH 3.5) stock solutions, and eventually diluted them to 1 x lO^M -

5 x 10'4M based on the metal ion peak height.

When the HPLC experiments were being performed, first, the Cis stationary

phase (reverse phase silica gel column) was equilibrated with the mobile phase for

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. approximately 2 to 8 hours depending on the ligand and its concentration. As the column

was getting equilibrated with ligand solution, 2 0 pi sample solution was manually

injected. When the experimental conditions were changed for the system such as pH and

concentration of the ligand (mobile phase), the column was washed with methanol to

remove the ligand and complexes, then re-equilibrated with the new mobile phase to

continue the next experiment. The Chromatographic separation experiments were

performed a minimum of 5 to 7 times at each condition for each ligand solution.

4.3. HPLC separation results and discussion

Lanthanide metal ions separations were investigated in our research group by a

previous student using five types of water soluble surfactant (PEG 550, PEG750, PEG2000,

Brij35p and Triton X-100). All PEG surfactants are polydisperse in nature (average no. of

ethoxide units (n) is 12-45). The chemical structures of amphiphilic ligands and

surfactants that were used in the previous investigation are shown in the following figure

4.2 and 4.3.

'O B rij35P

"OTriton-X

Figure 4.2. Structures of a family of amphiphilic acylpyrazolone ligands

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. PEG5501 n - 12

PEG750, n = 17 n Me

PEG2000. n - 45

Brij35P, n = 13 HO n n-C-i2H25

Triton X - 100, n = 17 HO

Figure 4.3. Surfactants that are used for the preparation of amphiphilic ligands

In the previous studies, performed in our group, many kinds of amphiphilic

acylpyrazolone ligands were synthesized (not in high purity) with different kinds of R

groups (R, R', R" = electron withdrawing and donating group), and a variety of

surfactants that are polydisperse in nature (Figure 4.3). Having observed the previous

results of lanthanide metal ions separation with a variety of ligands, we came to a

conclusion that only two ligands such as valeroyl pyrazolone and benzoyl pyrazolone

with P E G 7 5 0 (Figure 4.4) provided good separation.

Figure 4.4. Chemical structure of HMPVP-750 and HBMPP-750

Control experiments were carried out in the previous studies with only 4-

haloacylpyrazolones (C2 and C3, Figure 4.5), and their amphiphilic ligands (HMPVP-

750 and HBMPP-750) for comparison of lanthanide metal ions separation. The

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. acylpyrazolone ligands solubilities in the aqueous phase are markedly different with their

amphiphilic derivatives being readily soluble whereas they are being insoluble. It is well

known in the field of biochemistry 75 that polyethylene glycols are widely used surfactants

to solubilize organic compounds in aqueous phase due to the activity of hydroxyl group.

Acylpyraozlones however will dissolve in the aqueous phase in the presence of excess of

PEG moiety. The HPLC separations of the lanthanide metal ions were compared and

confirmed that amphiphilic derivatives provided good separation while 4-

haloacylpyrazolones themselves did not.

I have continued this research project by synthesizing the above mentioned two

ligands (Figure 4.4) in high purity. Since these two ligands provided good separation with

polydisperse PEG 750, this research project was proceeded further by developing HPLC

separation methods for different PEG moieties such as monodisperse PEG 516, PEG913 . As

these monodisperse polymers have different physical properties they might influence the

self assembled structures and separation of metal ions. Other amphiphilic ligands such as

amphiphilic acylisoxazolone ligands, which have significance in the metal ion separation

as described in Chapter 3, were employed to investigate the separation of same lanthanide

metal ions that were separated by amphiphilic acylpyrazolone ligands. All results

pertaining to amphiphilic acylisoxazolones will be discussed in the next chapter.

The synthesis of all amphiphilic acylpyrazolone ligands (Figure 4.5) was

described in Chapter 2. These ligands are completely soluble in the aqueous phase and

self-assemble to form various interesting nanostructures as displayed in section 4.10.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. These ligands were dissolved in HPLC grade water and prepared as stock solution of 5 x

10'3M concentration. This stock solution was further diluted based on the experimental

conditions and used as the mobile phase in HPLC separation studies to understand the

influence of the mobile phase conditions such as ligand concentration, pH, ionic strength,

and structure of the ligand for the separation of the lanthanide metal ions.

PrNr OH O C3

N= OPEG750 a NNx?:^\^\^\^O PE G 75o Q r * ' \ ii OH O OH O HMPVP-750 HBMPP-750

c h 3 o p e g 516 N=( Y r P P ^ \ / ^ opEG516 C r . .. OH 0 OH O HMPVP-516 HBMPP-516

o p e g 913 -OPEG913 Cr"-1 I. OH O HMPVP-913 HBMPP-913

Figure 4.5. Parent acylpyrazolones (C2 and C3) and their amphiphilic derivatives (HMPVP-PEGX and HBMPP-PEGX)

After the performance of HPLC experiments, the resulting best HPLC separation

chromatograms of lanthanide metal ions for each ligand are shown in the following

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. figures. Ligand concentration, pH of the mobile phase, ligand structure and lanthanide

metal ions are showed on the chromatogram.

4.4. HPLC separation chromatograms by HMPVP-750 and results

Valeroylpyrazolone-PEG 75o (HMPVP-750) ligand provided very good lanthanide

metal ions separation (Figure 4.6). Experiments were carried with different ligand

concentrations with different pH. Figure 4.6 displayed some of best separation

chromatograms that are obtained at different ligand concentration (0.000015M,

0.000005M, 0.00001M, and 0.00002M) and at different pH (2.64, 2.93, 2.83, 2.68, and

2.79)

0.000015M, pH 2.64

CH N— 350 Ce, 3.33 OH 300 Pr, 4.67 13l Nd, 6.13 250 200 150 Sm , 18.63 Eu, 27.2 100 Ho, 50.07 50

0 0 20 40 60

Retention time (min)

Figure 4.6. HPLC separation chromatograms by HMPVP-750 ligand with different pH and concentrations

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00001 M, pH 2.83

270 i CH N= La, 2 .7 0 C e, 4 .2 3 Ph 220 - OH Pr, 6.67

Nd, 9.23 c 170 -

S 120 -

Sm, 3 2 .6 3

70 - Eu, 4 9 .3 3 Ho, 9 4 .10

0 20 40 60 80 100 120 140 Retention time (min)

0.00002M, pH 2.68 .CH

250 1, 2.80 (Ce,4.73 OH 200

Rr, 7.73 Md, 10.93 150

100

S m , 44.33 Eu, 72.00 50 Ho,170.0

0 0 20 40 60 100 120 140 160 180 Retention time (min)

Figure 4.6. - Continued

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.000005M, pH2.79 CH- N=< 250

200 -

Pr.

150 -

Ja 100 - Eu, 33.2 Tb, 48.4 Lu, 61.!

50 -

Retention time (Min)

Figure 4.6. - Continued

Amphiphilic valeroylpyrazolone-PEG 75o (HMPVP-750) provided excellent

baseline separation for the concentration range of 5 x 10"6M - 1.5 x 10'5M. This ligand

with high concentration (2 x 10"5M) also provided good separation but with low _i_o absorbance especially for heavy lanthanide metal ion (Ho ). The selectivity is good for a

I ■’> mixture of seven lanthanide metals (light, middle and heavy lanthanides). As Eu and

Ho+3 have very good separation with high selectivity, a mixture of eight lanthanide metal

ions (La+3, Ce+3, Pr+3, Nd+3, Sm+3, Eu+3, Tb+3, and Lu+3) was used and obtained good

separation for eight lanthanide metal ion mixture.

4.5. HPLC separation chromatograms by HMPVP-516 and results

Valeroylpyrazolone-PEG 5i6 (HMPVP-516) also provided excellent metal ion

separations like HMPVP-750 ligand, but the concentration range was very limited. The

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. separation behavior was same as HMPVP-750 with 5 x 10'6M, pH 2.75. For this

condition, it provided best baseline separation with good selectivity for light, middle and

heavy lanthanides (Figure 4.7).

325 -| La1Ce1Pr1.5Nd2.4Sm3.6Eu3.6Ho5.5-0.000005M-pH2.75 La, Z63 Ca, 4.20 N=< 275 PEG. OH

Pr, 6.60 225 - Nd, 9.13

175 -

125 - Sm , 30.77

Ho, 86.73

25 20 40 60 80 100 120 Retention time (min)

375 La1 Ce1 Pr1.5Nd2.4Sm3.6Eu3.6Tb5Lu6-0.000005M-pH2.75

L a, 2.60 N=* 325 -

C e, 4.00 OH

275 ^ Nd, 8.30 Pr,

o 225 -

125 S m , 27.53

Eu, 38.80 Tb, 57.57 75 ~

25 20 40 60 80 100 120 Retention time (min)

Figure 4.7. HPLC separation chromatograms by HMPVP-516 with different conditions

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00002M, pH 2.90 CH 270 N = - P h OH La, 3.00 220

170 - Ce, 5.67

120 - Pr, 9.93 Nd, 14.60

70 - Sm , 64.20 Eu, 90.60 Ho, 140

20 20 40 60 80 100 120 140 Retention time (min)

0.00001 M, pH 2.90 CH 300 -j N = Ph La, 2.60 250 - OH Ca, 3.97

3 200 - Pr, 6.90 Id, 8.00

100 - Sm, 26.07 Eu, 41.0 Ho, 74.2 50 -

10 20 30 40 50 60 70 80 90 Retention time (min)

Figure 4.7. - Continued

At higher concentration like 2 x 10'5M, HMPVP-516 showed good selectivity and

resolution for light lanthanide metal ions. In case of middle and heavy metal ions, there is

very poor selectivity with no resolution. This ligand provided very poor separation

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. efficiency with higher concentration, where as the low concentration ligand with longer

equilibration of column provided baseline separation for a mixture of eight lanthanide

metal ions. Ligand was accumulated (precipitated) on the top of the column when the

concentration of mobile phase was high, and made the peak flat and low absorbance.

4.6. HPLC separation chromatograms by HMPVP-913 and results

Valeroylpyrazolone-PEG 9 i3 (HMPVP-913) ligand also provided baseline

separation for the mixture of seven lanthanide metal ions, but with only at 5 x 10'6M and

1 x 10'5M. This ligand showed very good selectivity and efficient separation. When the

concentration of the ligand is high (1.5 x 10"5M), it was precipitated on the top of the

column that causes the high back pressure, result in the bad shape (flat and wider) of the

peak and poor selectivity for middle lanthanide metal ions (Figure 4.8).

0.00001 M, pH 2.80 N= 350 Ce, 3.63

a. 2.57. 300 Nd. 7.03 , 5.27 s 250 i 200

150 Sm, 21.93 Eu, 32.03 < 100 Ho, 57.53

Retention time (min)

Figure 4.8. HPLC separation chromatograms by HMPVP-913 with different conditions

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 350 n La1Ce1Pr1.SNd2.4Sm3.6Eu3.6Ho5.5-0.000005M-pH2.82

,CH3 N=< 300 - Co, 3.80

La, 2.57 OH

250 -

Pr, Nd, 7.I

8 200 -

150 -

Sm, 24.17 100 -

Ho, 50

40 50 60 70 80 Retention time (min)

0.00001, pH 2.90 CH N— 350 Ph

300 La, 2.87 OH

250 Ce, S.03

. - 200 Pr, 8.43 Nd, 12.17 5 150

< 100 Sm, 48.47 Eu, 68.70 H o , 1 3 4 .1 0

0 20 40 60 80 100 120 140 160 Retention time (min)

Figure 4.8. - Continued

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.000015M, pH 2.84

250 La, 3.13 CH: N= Ph 200 OH

Ce, 6.1

Pr, 11 Id, 16.43 £ 100 o nu> Sm, 72.03 Eu, 121.67 Ho, 125.13

80 100 120 140 160 180 200 Retention time (min)

Figure 4.8. - Continued

4.7. HPLC separation chromatograms by HBMPP-750 and results

Benzoylpyrazolone-PEG 75o (HBMPP-750) provided good separation for

lanthanide metal ions but separated up to five metal ions only. It (HBMPP-750) showed

very good selectivity and separation resolution with light, middle, and heavy metal ions.

More selectivity was observed with high concentration (1 x 10'5M), but poor selectivity

and poor baseline separation with low concentration (2.5 x 10‘6M) for light lanthanide

metal ions. It showed good selectivity and separation at higher pH 2.47, but poor

selectivity at lower pH 2.36 for light lanthanides (Figure 4.9).

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00001 M, pH 2.40 .CH; 270 La, 3.37 Ph OH 220

170

Ce, 6.20

5 120 Nd, 13.43

Sm, 50.37 Tb, 116.2

0 20 40 60 80100 120 140 Retention time (min)

0.000005M, pH 2.40 CH 320 i

La, 2.63

270 - OH

Ce, 3.73

= 220 Nd, 6.50

120 - Sm, 18.57

0 10 20 30 40 50 60 Retention time (min)

Figure 4.9. HPLC separation chromatograms by HBMPP-750 with different conditions

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.0000025M, pH 2.40

1400 i

Nd, 2.37 Ph 1200 - OH

1000 - Sm, 3.03

•| 800 - O La, 2. Tb, 3.87 « 600 - o <0 < 400 -

200 -

0 24 6 8 10 12 14 16 18 20 Retention time (min)

0.000008M, pH 2.40

CH 250 N =< ‘OP E G La, 3.43

200 -

D 10 150 - 1 Ce, 6.27

0 50 100 150 Retention time (min)

Figure 4.9. - Continued

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.000005NI, pH 2.47 CH;

300 i Ph La, 2.87 OH 250 -

200 - Ce, 4.50

Nd, 8.60

100 - Sm, 27.17

Tb, 58.40

Retention time (min)

0.000005M, pH 2.36 .CH; 370 i La, 2.53

320 - OH

270 - Ce, 3.33

£ 220 I, 5.40 170

120 - Sm , 14.77

Tb, 30.37

0 10 15 20 30 355 40 45 Retention time (min)

Figure 4.9. - Continued

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 4.8. HPLC separation chromatogram by HBMPP-516 and results

Benzoylpyrazolone-PEG 5i6 (HBMPP-516) surprisingly provided very poor

separation. It showed poor selectivity for light lanthanides (La+3, Ce+3, Nd+3) and did not

1-5 | separate well but showed good selectivity for middle and heavy metal ions (Sm , Ho ,

Figure 4.10).

0.000005M, pH 2.03 CH 80

Ce, Nd OH

La, 5.

Sm, 21.06

Ho, 47.70

Tim e(min)

Figure 4.10. HPLC separation chromatogram by HBMPP-516

4.9. HPLC separation chromatogram by HBMPP-913 and results

Benzoylpyrazolone-PEGc>i 3 (HBMPP-913) ligand separation behavior is similar to

HBMPP-516. It also showed poor selectivity for light lanthanide metal ions with poor

separation (Figure 4.11).

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00001 M, pH 2.15 CH N=- 70

OH 65 - Nd, 9.57

55 -

Sm , 29.37 Z0 50 - 1 45 - Ho, 89.07 .£ 40 -

35 -

100 Retention time(min)

Figure 4.11. HPLC separation chromatogram by HBMPP-913

All these experiments were carried out under identical conditions other than

concentration and pH for each ligand. Ligand solutions are with the concentration range

of 5 x 10'6M - 2 x 10"5M and pH 2.36 - 3.0 are compared. Having observed the best

HPLC separation chromatograms (above figures) with different amphiphilc ligand

solutions, the following conclusions were drawn:

1. Ligand selectivities and efficiencies for lanthanide metal ion separation are

very different.

2. Among the benzoylpyrazolone and valeroylpyrazolone ligands, the later

ligands provided better lanthanide metal ions separation.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 3. Valeroylpyrazolone ligands (HMPVP-750, HMPVP-516, and HMPVP-913)

provided excellent separation for a mixture of light, middle and heavy lanthanide metal

ions. The separation efficiency of this ligand with polydisperse PEG 750 (HMPVP-750) is

better than that of monodisperse PEG 516 and PEG913 moieties (HMPVP-516 and

HMPVP-913).

4. Benzoylpyrazolone ligands (HBMPP-750, HBMPP-516 and HBMPP-913) also

separated the same mixture of metal ions at lower efficiency. Ligand concentration and

pH ranges are limited for these ligands.

5. In the case of valeroylpyrazolone ligand, we obtained good separation of

lanthanide metal ions with all PEG moieties, whereas benzoylpyrazolone ligand provided

better separation with polydisperse PEG than that of monodisperse PEG moieties.

6 . A possible explanation for these differences in the separation efficiency could

be from the differences in the ligand structure and the nature of the self-assemblies of

these ligands in the aqueous phase and on the stationary phase. A more detailed

explanation will be discussed in the following sections in this chapter.

4.10. Self-assembly behavior of amphiphilic acylpyrazolones in the aqueous phase

The nature of the self assemblies of all amphiphilic ligands in the aqueous phase

were examined by TEM images to gain an understanding of their behavior and its

influence on their metal ion selectivities. Ligand samples were prepared with 5 x 10’ M

stock solution. The unstained TEM images are displayed in the following figures.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.12. TEM im ages of valeroylpyrazolone-PEG 75o (HM PVP-750)

Figure 4.13. TEM im ages of valeroylpyrazolone-PEGs 16 (HM PVP-516)

Figure 4.14. TEM im ages of valeroylpyrazolone-PEG 9 i 3 (HMPVP-913)

Figure 4.15. TEM im ages of benzoylpyrazolone-PEG 75o (HBM PP-750)

7 8

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.16. TEM images of benzoylpyrazolone-PEGsie (HBMPP-516)

* . w. m» • ■ ■. * -

a' * * . c _ -v~.

► .‘..Furi

Figure 4.17. TEM images of benzoylpyrazolone-PEG 9 i3 (HBMPP-913)

The ligands HMPVP-750 and HMPVP-516 exclusively form nano-tubes, as well

as nano-rods as shown in figure 4.12 and 4.13. HMPVP-516 and HMPVP-913 self-

assemble as closely packed huge dendritic structures (4.13 and 4.14), which indicates that

these three ligand systems have the most ordered structures. Such structures like the

nano-tubes in the solution may be two dimensional analogs containing ordered layers on

the Cig silica surface. Mostly, Benzoylpyrazolone (HBMPP) ligand with all PEG

moieties forms vesicles, square and rectangular self-assemblies, trying to grow bigger

dendritic structures as shown figure 4.15, 4.16, and 4.17. The HMPVP-750 and HMPVP-

516 ligands might form a more closely packed monolayers on Cis silica surface as it

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. contains long alkyl side chain when compared to the benzoyl group of HBMPP-750. The

self-assembly of these ligands on Qg stationary phase is most likely different from each

other resulting in the observed differences in lanthanide metal ions separation.

All the above TEM images of amphiphilic ligands revealed mainly three types of

self-assemblies such as linear structures with long branches, dendritic structures with

similar to star shape, and spherical structures resembling vesicles. Formation of self-

assemblies could be due to the hydrogen bonding (intra and inter molecular), Van der

Waal interaction and electronic interactions.76'79 The formation of these nanostructures

could be rationalized by the formation of linear or branched dendrimers, micelle, and

vesicle self-assemblies as displayed in the following figure 4.18.

| - | |m 1111r i ij-| [ h * 111111 [ n 11111111 |_ | Q 11m 11 it H |_'111111111H L Me

H ,0 HI n C H 3 OH 0 HL Amphiphilic acylpyrazolone

+ + +

L = Ligand site C D

•~w w vR = P E G en d

Figure 4.18. Schematic representation of ligand self-assemblies in aqueous phase

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 1. Ligand site aggregations (type B and C) due to intermolecular hydrogen

bonding between the carbonyl group and the hydroxyl group, which is a strong

interaction and is thermodynamically favored, can provide micelle type of self-

assemblies.77 These micelles could have either the R group or ligand L on the surface

side.

2. The formation of linear fibrous, tubes, needles, rods like structures could be the

result of type A agglomerations, which include the intermolecular hydrogen bonding

between the ligand groups and Van der Waal interaction between the R groups. These are

dense structures where most likely the L to L and R to R assembly such as

LR RL LR....RL.80’81

3. Van der Waal interactions between the hydrophobic side chains R resulting in

tail to tail self-assembly and vesicle structures with the polar ligand group on the surface

of the vesicle. These would be derived from micelles with R group on the surface. This

could lead to vesicles (type D aggregation) with a nonpolar bilayer region.

4. Vesicles with a polar bilayer region (type E aggregation) can be formed due to

the intermolecular hydrogen bonding between the L groups on the surface of micelles and

free ligands to yield vesicles where the R group of the surfactants is on the surface.

5. Further self-assemblies of the L groups and R groups on the surface of the

vesicles could be leading to dendritic structures.76

These types of structures for amphiphilic ligands have been observed by others.

Spherical vesicle type structures have been identified by Johnsson for PEGs.82 Various

types of spherical and linear structures have been observed when alkylquinoline ligands,

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. macrocyclic amphiphilic ligands, and terpyridyl ligands derivatized with polyethylene

OA Q 1 0-1 glycols. ’ ’ ’ These self-assembled structures were found to be labile in the aqueous

environment.76 Our studies on acylpyrazolone and acylisoxazolone ligands with various

PEG moieties attached at the 4th position have shown the similar self-assemblies as it was

in the previous literature as discussed above. Self-assemblies at the nanoscale offer a new

approach to this challenging lanthanide metal ions separation, as properties at the

nanoscale are often dramatically different from molecular and bulk material properties.85,

Both polydisperse and monodisperse PEG moieties are employed in our research

project to synthesize amphiphilic acylpyrazolone and acylisoxazolone ligands. It is well

known that naturally occurring lipids such as Egg phosphotidyl choline (EggPC) is a

complex mixture of anionic and zwitterionic lipids with varying alkyl chain lengths and

forms self-assembly structures in the presence of metals like Mg+2. EggPC also forms

complex vesicle structures in the aqueous phase and forms linear structures as well in the

right conditions.87 As these amphiphilic acylpyrazolones and acylisoxazolones are

analogous to naturally occurring lipids (EggPC) and it is worth observing their analogous

behavior in the presence of MgC^.

The self-assembled structures of amphiphilic acylpyrazolones (HMPVP-750 and

HBMPP-750), which have the best selectivity and good separation for lanthanide metal

ions, have been investigated in the presence of MgCl 2 by TEM. A sample solution (1 x

10"3M) of the ligand was equilibrated with 1 x 10'3M MgCl 2 solution at pH 6.5 and 0.1

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ionic strength. This solution was mixed for overnight (12 hr), and took the TEM images

to understand the type of self-assemblies that are formed.

Figure 4.19. TEM images of HMPVP-750 with Mg+2 metal ion (12 hr solution)

TEM images showed for HMPVP-750 with Mg+2 metal ion solution after 12

hours all kinds of structures such as nano-tubes, needles, and vesicles (Figure 4.19). The

same solution even after one week showed same structures as shown in figure 4.20.

Figure 4.20. TEM images of HMPVP-750 with Mg+2 metal ion (one week solution)

The solution after 12 hr of HMPVP-516 with Mg+2 showed needles and vesicles (Figure 4.21), whereas the same solution after one week showed completely different

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. structures such as linear aggregation of vesicles and huge branched dendritic structures (Figure 4.22). These dense structures are formed probably due to the further agglomeration of the spherical and linear structures.

Figure 4.21. TEM images of HMPVP-516 with Mg+2 metal ion (12 hr solution)

Figure 4.22. TEM images of HMPVP-516 with Mg+2 metal ion (one week solution)

Aclypyrazolones with long chain monodisperse PEG moiety (HMPVP-913) in the

presence of Mg+2 metal (over night solution) ion also formed similar structures like

needles, vesicles, spikes, and broken glass tubes (Figure 4.23). One week old solution

showed dendritic structures and broken glass tubes (Figure 4.24) which the same

structures like HMPVP-516 ligand are.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.23. TEM images of HMPVP-913 with Mg+2 metal ion (12 hr solution)

Figure 4.24. TEM images of HMPVP-913 with Mg+2 metal ion (one week solution)

The self-assembly behavior of HBMPP ligands with Mg+2 metal ion was also

investigated to compare with other ligand solutions (HMPVP-PEGX). The HBMPP-750

ligand solution showed circular vesicle aggregation surrounded by needles, and broken

glass tubes for 12 hr solution (Figure 4.25), whereas after one week the same solution

showed crystal balls and broken glass tubes (Figure 4.26).

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.25. TEM images of HBMPP-750 with Mg+2 metal ion (12 hr solution)

Figure 4.26. TEM images of HBMPP-750 with Mg+2 metal ion (one week solution)

The HBMPP-516 solution showed same observation as that of HBMPP-750 in 12

hr solution (Figure 4.27), whereas one week old solution showed nice leaf-like dendritic

structures, circularly formed vesicle aggregations as multilayers, and multivesicle

vesicles (Figure 4.28).

Figure 4.27. TEM images of HBMPP-516 with Mg+2 metal ion (12 hr solution)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.28. TEM images of HBMPP-516 with Mg+2 metal ion (one week solution)

The HBMPP-913 ligand solution forms needles, vesicles, and vesicles in circular

layers as 12 hr solution (Figure 4.29), one week old solution formed nano-tubes, needle,

and glassy crystal colonies (Figure 4.30).

Figure 4.29. TEM images of HBMPP-913 with Mg+2 metal ion (12 hr solution)

Figure 4.30. TEM images of HBMPP-913 with Mg+2 metal ion (one week solution)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. It is also worth understanding the nature of self-assemblies formed in solution

when lanthanide metal ions are present. The middle series lanthanide metal ion, Sm+3,

was used to examine the structures by TEM. All images are shown for all the ligands in

the presence of Sm+3 metal ion solutions (12 hr solution and one week old solution).

Samarium ion (Sm+3) leads to the formation of nice nano-rods, very dense vesicles, and

needles as shown in figure (4.31 to 4.37).

Figure 4.31. TEM images of valeroylpyrazolone-PEG 75o (HMPVP-750) with Sm+3

Figure 4.32. TEM images of valeroylpyrazolone-PEGsi6 (HMPVP-516) with Sm+3

Figure 4.33. TEM images of valeroylpyrazolone-PEGc>i 3 (HMPVP-913) with Sm+3

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.34. TEM images of HBMPP-750 with Sm+3 metal ion

i ■3 Figure 4.35. TEM images of HBMPP-516 with Sm metal ion (one week solution)

Figure 4.36. TEM images of HBMPP-913 with Sm+3 metal ion

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 4.37. TEM images of HBMPP-913 with Sm+3 metal ion (one week solution)

4.11. Possible mechanism of lanthanide metal ion recognition and separation by amphiphilic ligands

In the HPLC experiments, mobile phase (ligand solution) passes through the Cig

silica gel column. Ligand can be adsorbed on the column that could lead to monolayer

formation while column was gradually getting equilibration. The self-assembly of ligands

can occur on Cig column, which can be a complex phenomenon similar to the self-

assembly in the aqueous phase (mobile phase). As ligand is adsorbed on the Cig, it can

also form complexes with metal ion like in mobile phase. The phenomenon of

complexation of metal ions is a pH dependent process.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. +

c D

CH;

•c h 3 h 3c 0 - ^ o ) ^ 0 ' lh

c h 3

CH;

Figure 4.38. Schematic representation of metal ion complexation and separation mechanism

A kind of ion-exchange process is involved at the time of metal complex

formation as schematically represented in figure 4.38. This metal ion complexation in the

mobile phase and on the Cis column seems to be a competitive process that eventually

leads to the separation of the metal ions. This metal complexation process always can be

expected to be influenced by the spontaneous self-assembly of the amphiphilic ligands in

the aqueous phase and on the Cis stationary phase.

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The results that are obtained from HPLC experiments for lanthanide metal ion

separation would substantially support the central hypothesis of the research, which is

that nanoscale self-assemblies in the aqueous phase and on the stationary phase would

provide excellent metal ion selectivities eventually separates lanthanide metal ions. In

general, HPLC experiments involve a multistage separation, which means differential

distribution of sample over multiple stages between two different phases. The distribution

ratio (D) or capacity factor of a metal ion, derived from retention time, of a metal ion

between a mobile phase and stationary phase can be influenced by stability constant of a

metal complex. It is very important to keep in mind that the stability constants of the

metal complexes and the kinetics of complex formation and dissociation could also

influence the separation of lanthanide metal ions by the self-assemblies of these

amphiphilic acylpyrazolones.

In order to understand the mechanism of lanthanide metal ion recognition by

amphiphilic acylpyrazolones, separation experiments through HPLC were conducted as a

function of ligand concentration (concentration dependent studies) at a constant pH, and

as a function of pH (pH dependent studies) at constant concentration (Table 4.1 and 4.2).

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.1. Amphiphilic valeroylpyrazolone concentrations vs. pH

Ligand Concentration (M) pH

HMPVP-750 5 x 10'6 2.83, 2.88, 2.93, and 2.98 1 x 10'5 2.68, 2.73, 2.78, and 2.83 1.5 x lO '5 2.60, 2.64, 2.68, and 2.73

2x 10'5, 1 x 10'5, and 1.5 x 10'5 2.68

HMPVP-516 5 x 10'6 2.85,2.90,2.95, and 3.00 2 x 10'5, 1 x 10'5, 1.5 x 10‘5, and 5 x 10’6 2.90

HMPVP-913 1 x 10’5 2.80, 2.85, 2.90, and 2.95 l x l O '5, 1.5 x lO '5, 1.25 x lO ’5, and 5 x 10'6 2.84

Table 4.2. Amphiphilic benzoylpyrazolone concentrations vs. pH

Ligand Concentration (M) pH

HBMPP-750 2.5 x 10'6, 5 x 10 6, 8 x 10'6, and 1 x 10'5 2.40 5 x 10'6 2.36, 2.40, 2.43, and 2.47

HBMPP-516 5 x 10'6 2.03 1 x 10'5 2.12 5 x 10'6 1.97

HBMPP-913 1 x 10'6 2.15 1 x 10'5 2.15 2 x 10'5 2.38 5 x 10'6 2.14

The results (retention time, distribution ratio, slope, and Kex) of these studies are

presented in the following section 4.12 as tables and figures. A detailed ligand

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. dependence and pH dependence data is compiled and presented to understand the

complexity of these dependences as indicated by the proposed mechanism in figure 4.38.

To understand the complexity of lanthanide metal ions separation and their

recognition by amphiphilc ligands, a simple equilibrium model (where the ligand does

not aggregate in the aqueous phase) was employed for the distribution of the metal ion

from the aqueous mobile phase to Cis stationary phase. This model 17'19,23> 26,28,40,43,88-93

has been used since long back for solvent extraction method for the separation of

lanthanide metal ions. Having considered the simple model, we can say that the

complexation of metal ion occurs in the aqueous phase and the distribution of complex

takes place from aqueous to stationary phase (Cis silica gel). The distribution of metal

complex can be expressed by equation 1 where the subscripts “aq” and “sp” represent

aqueous and stationary phases and the equilibrium constant Kex can be determined by

equation 2.

,« . K 0 v M (aq) 3HL(aq) - M L3(sp) + 3H +(aq) (Equation 1)

[M L3l

logKex = logD - 3pH - 3log[HL] (Equation 3)

logD = logKex + 3pH + 3log[HL] (Equation 4)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The [ML3]sp/[M+3]aq is the distribution ratio (D) for a given lanthanide metal ion.

The Kex can be rewritten in terms of D and written as a logarithmic expression as shown

in equation 3. This equation can also be rewritten as equation 4 for the dependence of

logD on pH and ligand concentration.

Complexation of metal ion was studied using the traditional “slope analysis”

method. The plots of logD vs. log[HL] at constant pH and logD vs. pH at constant ligand

concentration are closer to linear with slope values closer to 3, which tell us that the

formation of metal complex could likewise be a 1:3 (ML 3) ratio as mentioned in the

simple model.

4.12. Analysis of separation results through logD vs. log|~HLl and logD vs. pH plots of all amphiphilic ligands

The formation of ligand aggregations such as head to head and tail to tail

association have been envisaged by longer time equilibration of Qg stationary phase with

certain ligand concentrations at a desired pH. The separation of metal ions was performed

in nine to ten successive times of sample injection this means that the column gets ~14 to

16 hr equilibration by the time we inject the sample 10th time. As the column is getting

equilibrated for nine to ten successive times of sample injection, the retention times for

each metal ion will be increased. Thus the D (distribution ratio) values for each metal ion

will be increased with each successive run as shown in figure 4.39. The advantage of this

plot is that we can calculate any D value at any given minute for each lanthanide metal

ion by using the slope and intercept obtained from Real run time vs. Retention time plot

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. (Figure 4.40). The self-assembly nature of the ligand on the stationary phase as it gets

equilibration could increase the D value with increasing adsorption of ligand with

successive runs. At some point the column gets saturated.

The relationship between Retention time and Real run time, 0.00001 M, pH 2.78 HMPVP-750

Eu '■= 25

Sm a 15

Nd — Ce

100 200 300 400 500 600 700 Real run time (min)

Figure 4.39. Equilibration of the stationary phase (Ci8 column) with F1MPVP-750

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. La*3 C e+3

2.32 = 2.3 c 1 2.28 £ Z 2.26 ~ 2.24 c o 2.22 o oc c o_ 2.18 - £ 2.4 “ 2.16 - & 2.3 0 200 600400 800 0 200 400 600 800 Real run time (min) Real run time (min)

Pr43 Nd*3

4 5 3.5 4 3 2.5 3 2 1.5 2 1 1 0.5 0 0 0 200 400 600 800 0 200 400 600 800 Real run time (min) Real run time (min)

Sm«

12 2 0 -i .= 10 8 6 c 4 o 2 oc 0 'S 200 400 0 600 800 0 200 400 600 800 Real run time (min) Real run time (min)

30 25 20 15 10 5 0 0 200 400 600 800 Real run time (min)

Figure 4.40. Real run time vs. retention time plots of each lanthanide metal ion

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 4.12.1. LogD vs. loarHLl of HMPVP-750 at pH 2.68

The logD vs. log[HL] plots were drawn to analyze the dependence of distribution

ratio (D), which was calculated from experimental retention time (D = (tr-to)/to, where to =

2.17, from Na 2Cr2C>7 retention time known as dead time), as a function of ligand

concentration at a pH 2.68. Ligand concentrations are 1 x 10~5M, 1.5 x 10'5M, and 2 x 10‘

5M. All the calculated D values and logD values are shown in table format for each

lanthanide metal ion in the following tables. Selectivity of the ligand for the adjacent

lanthanide metal ions are tabulated at different concentrations (Table 4.5).

Table 4.3. D values for each lanthanide metal ion at pH 2.68 as a function of ligand concentration

D values [HL] log[HL] La Ce Pr Nd Sm Eu Ho 1 x 10'5 -5.00 0.10 0.31 0.65 1.03 4.00 5.87 10.14

1.5 x lO '5 -4.82 0.28 0.67 1.43 2.26 9.10 13.65 25.25

2 x 10'5 -4.59 0.59 1.36 2.94 4.62 21.10 33.76 69.58

Table 4.4. LogD values for each lanthanide metal ion at pH 2.68 as a function of ligand concentration

logD values [HL] log[HL] La Ce Pr Nd Sm Eu Ho 1 x 10'5 -5.00 -1.63 -0.51 -0.18 0.01 0.60 0.77 1.01

1.5 x 10'5 -4.82 -1.10 -0.18 0.16 0.35 0.96 1.14 1.40

2 x 10'5 -4.59 -0.73 -0.13 0.47 0.66 1.32 1.53 1.84

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.5. Selectivity (a) for adjacent lanthanide metal ions

a values [HL] Ce/La Pr/Ce Nd/Pr Sm/Nd Eu/Sm Ho/Eu 1 x 10's 2.95 2.14 1.57 3.89 1.47 1.73

1.5 x 10'5 2.39 2.16 1.58 4.03 1.50 1.85

2 x 10‘5 2.32 2.16 1.91 3.75 1.60 2.06

Sm*3

1.2 •

0.8 a f 0.6 y = 2.3725x + 12.447 R2 ■ 1 0.4

0.2

-5.05 -5 -4.95 -4.9 -4.85 -4.8 -4.75 -4.7 -4 65 Ioq [HL]

H o *3

y "2.7433x+14.697

0.6 - 0.4 • 0.2 • i------1------1------,------,------,------,------1------6 - •5.06 -5 -495 -49 -465 -48 -4.75 -4.7 -465

Figure 4.41. LogD vs. log[HL] plots of each lanthanide metal ion

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The values are calculated from experimentally obtained slopes from logD vs.

log[HL] plots and are shown in table 4.6. The difference in the Kex values for adjacent

lanthanide metal ions indicates that this ligand has good selectivity and separation.

Table 4.6. LogKex values for each metal ion as a function of ligand concentration

losK« values [HL] La Ce Pr Nd Sm Eu Ho 1 x 10'5 0.41 1.72 2.33 2.67 3.99 4.90 6.34

1.5 x lO '5 0.39 1.69 2.29 2.63 3.18 4.52 6.00

2.0x1 O'5 0.39 1.58 2.33 2.67 4.00 4.91 6.35

4.12.2. LogD vs. pH of HMPVP-750 at 1 x 10'5M

The dependence of distribution ratio (D) was analyzed as a function of pH (2.68,

2.73, 2.78, and 2.83) at constant ligand concentration. The D values and logD values are

calculated from experimental values of retention time and tabulated in the following

tables (Table 4.7 and 4.8). Selectivity values are tabulated for adjacent lanthanide metal

ions in table 4.9.

Table 4.7. D values for each lanthanide metal ion at different pH

D values pH La Ce Pr Nd Sm Eu Ho 2.68 0.06 0.23 0.50 0.79 3.03 3.68 6.01

2.73 0.07 0.27 0.59 0.93 3.62 4.26 6.85

2.78 0.13 0.41 0.88 1.32 5.38 6.39 10.32

2.83 0.24 0.66 1.43 2.25 8.93 10.62 17.41

100

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.8. LogD values for each lanthanide metal ion at different pH

loeD values pH La Ce Pr Nd Sm Eu Ho 2.68 -1.22 -0.64 -0.30 -0.10 0.48 0.57 0.78

2.73 -1.13 -0.57 -0.23 -0.03 0.56 0.63 0.84

2.78 -0.89 -0.39 -0.05 0.12 0.73 0.81 1.01

2.83 -0.62 -0.18 0.16 0.35 0.95 1.03 1.24

Table 4.9. Selectivity (a) for adjacent lanthanide metal ions

a values pH Ce/La Pr/Ce Nd/Pr Sm/Nd Eu/Sm Ho/Eu 2.68 3.75 2.21 1.58 3.83 1.22 1.63

2.73 3.70 2.19 1.56 3.89 1.18 1.61

2.78 3.17 2.16 1.50 4.06 1.19 1.62

2.83 2.79 2.16 1.57 3.97 1.19 1.64

The logD values are plotted against pH (Figure 4.39) to obtain the slopes for each

lanthanide metal ion at approximate time intervals. The experimentally obtained slopes,

displayed in table 4.10, were used to calculate the KeX values and compared among the

lanthanide metal ions (Table 4.11), which indicated good separation with good

selectivity.

101

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. La*3

216 2.68 2.7 2.72 2.74 2.76 2.78 2.8 2.82 2 84 -0.2 -0.4 O -0.6 a .2 -0.8 -1

* y*4.0516x-t2.t29 RJ = 1 y = 3.1508x-9.1245 R2 = 1 pH

Pr*3 N d*3

0.2

o o s 0a>

y = 3 .0 7 3 4 x - 8.5734 R2 = 1

y = 3.1155x- 7.8266

y=3.1617x-8.0303R2= 1

y = 3.1293x-7.654

Figure 4.42. LogD vs. pH plots of each lanthanide metal ion

102

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.10. Slopes for each lanthanide metal ion

La Ce Pr Nd Sm Eu Ho Ligand depend 2.97 2.14 2.15 2.16 2.37 2.49 2.74

pH depend 4.05 3.15 3.07 3.03 3.16 3.12 3.13

Table 4.11. LogKex values for each lanthanide metal ion at different pH

Ios Kpv values pH La Ce Pr Nd Sm Eu Ho 2.68 0.39 1.63 2.02 2.20 3.41 4.01 4.98

2.73 0.37 1.56 1.95 2.13 3.34 3.94 4.91

2.78 0.31 1.57 1.81 2.13 3.34 3.94 4.89

2.83 0.38 1.62 2.02 2.20 3.41 4.02 4.98

Having obtained good selectivity and separation for 1 x 10'5M concentration of

HMPVP-750, other concentrations such as 5 x 10'6M (low concentration) and 2 x 10'5M

(high concentration) were also tried to understand the separation mechanism. Somehow

the lower and higher concentration provided poor separation with low absorption values

and good selectivities. The slopes are closer to three as described in simple model. As

described in equation 4, we can explain the separation factor of lanthanide metal ions by

Kex values. The separation factor for a given two pair of metal ions is expressed by the

differences in their logKex values.5,33,42 If the difference of logKeX value of two adjacent

metal ions is big, then the separation is good. All KeX values are tabulated for all

103

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. amphiphilic ligands as a function of concentration and pH, This data indicates the

separation of metal ions.

4.12.3. LogD vs. loarHLl for HMPVP-516 at pH 2.90

Distribution ratio (D values) and their logD values are calculated from

experimentally obtained retention time and are tabulated (Table 4.12 and 4.13) for each

lanthanide metal ion at different ligand concentrations.

Table 4.12. D values as a function of ligand concentration at pH 2.90

D values [HL] La Ce Pr Nd Sm Eu Ho 5 x 10'6 0.02 0.04 0.10 0.17 0.55 0.70 3.61

1 x 10'5 0.22 0.32 0.66 1.01 3.67 5.43 24.15

1.5 x 10'5 0.51 0.80 1.62 2.52 9.95 14.78 46.34

2 x 10'5 0.84 1.37 3.01 4.78 — — —

Table 4.13. LogD values as a function of ligand concentration at pH 2.90

losD values [HL] log[HL] La Ce Pr Nd Sm 1 ii Ho 5 x 10"6 -5.30 -1.67 -1.44 -0.99 -0.77 -0.26 -0.16 0.56

1 x 10'5 -5.00 -0.65 -0.49 -0.18 0.00 0.57 0.74 1.38

1.5 x 10'5 -4.82 -0.29 -0.10 0.21 0.40 1.00 1.17 1.67

2x 10'5 -4.70 -0.08 0.14 0.48 0.68 --

104

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. The logD vs. log[HL] plots were drawn to calculate the slopes and values for

each lanthanide metal ion.(Figure 4.43)

La+3

-4,8. -5.2 -5.3 -5.4

0.4

P r+ 3 Nd

-1.5

>«T38x + 12.041 -0.5 I - -0-4 R2 = 0.9986 -4.7 ^5.1 -5.2 -5.3 -5.4 "4 :6 -0 .2 -4 .7 -4 .8 -4.1 -5.2 -5.3 -5.4

0.2 0.4

0.8

-5.1 -5 .2 -5 .4

0.2 - 2.6415x + 13.752 0 .4 0.6

1.2 lo g [H L ] lo g [ H L ]

Ho+3

-5.4 -5.3 -52 -5.1 •5 -4..9 log[HL]

Figure 4.43. LogD vs. log[HL] plots of each lanthanide metal ion

105

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.14. Selectivity (a) for adjacent metal ions at different ligand concentration

a values [HL] Ce/La Pr/Ce Nd/Pr Sm/Nd Hu Sm Ho Hu 5 x 10'b 1.70 2.84 1.64 3.27 1.26 5.18

1 x 10'5 1.45 2.06 1.52 3.64 1.48 4.45

1.5 x 10'5 1.57 2.04 1.55 3.95 1.48 3.14

2 x 10‘5 1.64 2.20 1.59 -

Table 4.15. LogKex values for each metal ion as a function of ligand concentration

loaKpv values [HL] La Ce Pr Nd Sm Eu Ho

5 X 10‘6 3.08 5.12 6.04 6.63 7.16 7.97 8.43

1 x 10'5 3.29 5.14 5.95 6.58 7.09 8.00 8.38

1.5 x 10'5 3.18 5.05 5.84 6.51 7.03 7.96 8.16

2 x 10'5 3.06 4.94 5.72 6.41 --

4.12.4. LogD vs. p H of HMPVP-516 at 5 x 10'6M

Experimentally obtained D values were used to draw the logD vs. pH plots at 5 x

10'6M concentration. Plots for each lanthanide metal ion are shown in figure 4.44. The

slopes obtained from this plots are close to three and comply with the slope of simple

model (Equation 1 and Equation 3). All slopes (Table 4.19) and Kex values (Table 4.20)

as a function of pH and ligand concecentration are displayed as average values of

different real runtimes.

106

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.16. D values for each lanthanide metal ion at different pH

D values pH La Ce Pr Nd Sm Eu Ho 2.90 0.03 0.16 0.33 0.56 1.05 1.41 2.77

2.95 0.04 0.18 0.39 0.60 1.17 1.60 3.06

3.00 0.07 0.29 0.64 0.98 1.98 2.80 5.03

Table 4.17. LogD values for each lanthanide metal ion at different pH

loaD values pH La Ce Pr Nd Sm Eu Ho 2.90 -1.49 -0.80 -0.48 -0.25 0.02 0.15 0.44

2.95 -1.35 -0.75 -0.41 -0.22 0.07 0.20 0.49

3.00 -1.17 -0.53 -0.19 -0.01 0.30 0.45 0.70

Table 4.18. Selectivity (a) for adjacent lanthanide metal ions at different pH

a values pH Ce/La Pr/Ce Nd/Pr Sm/Nd Eu/Sm Ho/Eu 2.90 4.87 2.09 1.69 1.88 1.35 1.97

2.95 3.98 2.19 1.54 1.94 1.37 1.91

3.00 4.33 2.18 1.53 2.02 1.41 1.79

107

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. -0 .8 5

-0 .8

-0 .7 5 -1.4 y = 3.2448x- 10.907 y = 2.7355X - 8.7639 R2 = 1 -0 .7 R2 = 1

-0 .6 5 -1.2 - - 0.6 -

- 0 .5 5 - 2.9 2.92 2.94 2.96 2.98 3.02 2 9 2.92 2.94 2.96 2 .9 8 3 .0 2 -0 .5

P r 3

-0.6 - 0 .3

-0.5 - 0 .2 5 y = 2.464X - 7.4299 -0.4 - 0 .2 R2 = 1

» -0.3 9 - 0 .1 5

y = 2.9014x-893te^ -0.2 - 0.1 -

-0.1 • - 0 .0 5 - 2. 08 2.9 2.92 2.94 2.96 2.98 3 3.02 2.38 2.9 2.92 2.94 2.96 2.98 3.02

Sm+3 Eu+3

0 .35 0 .5 0 .4 5 0 .4 0 .2 5 0 .3 5 0.2 0 .3 • 0 .2 5 » 0.15 0.2 0 .1 5 0.1 0.05 0 .0 5 0 2.9 2.92 2.94 2.96 2.98 3.02 2.92 2.94 2.96 2.98 3 3 .0 2

PH PH

0 .8 -| y = 2.5912x- 7.1008

0 .3 -

0.2 - 0.1 - 0 -I-— i ------,------,------,------■ 2.88 2.9 2.92 2.94 2.96 2.98 3 3.02

p H

Figure 4.44. LogD vs. pH plots of each lanthanide metal ion

108

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.19. Slopes for each metal ion as a function of concentration and pH

La Ce Pr Nd Sm Eu Ho Ligand depend 2.67 2.64 2.44 2.41 2.64 2.51 2.37

pH depend 3.24 2.74 2.90 2.46 2.78 2.99 2.59

Table 4.20. LogKex values for each lanthanide metal ion as a function of pH

logKpv values PH La Ce Pr Nd Sm Eu Ho 2.90 2.90 3.91 5.21 6.07 7.58 8.10 6.85

2.95 2.71 3.96 5.18 6.01 7.51 8.04 6.79

3.00 2.82 3.93 5.21 6.10 7.62 8.16 7.31

4.12.5. LogD vs. logfHLl of HMPVP-913 at pH 2.84

The D values and their log values were calculated from experimentally obtained

retention time for each lanthanide metal ion and are tabulated in table 4.21 and 4.22.

Ligand selectivity for each lanthanide metal ion is displayed in table 4.23. The logD

values are plotted (Figure 4.45) as a function of ligand concentration at constant pH 2.84

and obtained slopes are used to calculate the Kex values (Table 4.24) for each lanthanide

metal ion.

109

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.21. D values for each lanthanide metal ion at different ligand concentration

D values [HL] log[HL] La Ce Pr Nd Sm Eu Ho 5 x 10‘b -5.30 0.06 0.22 0.39 0.59 2.31 3.16 5.58

1 x 10'5 -5.00 0.25 1.01 2.17 3.43 13.93 20.51 36.72

1.25 x 1 O'5 -4.90 0.40 1.64 3.70 5.47 22.09 35.58 61.16

1.5 x 10‘5 -4.82 0.63 2.49 5.55 8.36 41.67 66.25 85.17

Table 4.22. LogD values for each lanthanide metal ion at different ligand concentration

logD values [HL] log[HL] La Ce Pr Nd Sm Eu Ho 5 x 10"6 -5.30 -1.22 -0.67 -0.41 -0.23 0.36 0.50 0.75

1 x 10'5 -5.00 -0.60 0.00 0.34 0.54 1.14 1.31 1.57

1.25 x 10'5 -4.90 -0.39 0.21 0.57 0.74 1.34 1.55 1.79

1.5 x 10'5 -4.82 -0.20 0.40 0.74 0.92 1.62 1.82 1.93

Table 4.23. Selectivity (a) for adjacent lanthanide metal ions at different concentration

a values [HL] Ce/La Pr/Ce Nd/Pr Sm/Nd Eu/Sm Ho/Eu 5 x 10'b 3.55 1.82 1.49 3.93 1.37 1.77

1 x 10'5 4.03 2.16 1.58 4.07 1.47 1.79

1.25 x 1 O'5 4.05 2.26 1.48 4.04 1.61 1.72

1.5 x 10'5 3.96 2.23 1.51 4.98 1.59 1.29

110

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ■1.4 r - 0 .8 - -0.6 y = 2.1063x + 9.9425 - -0.4 = 2.2192x + 11.098 -0.8 _/;8-0.2 -4.9 -5.2 -5.3 -5.4 -0.6 -0.4 - 0.2 -4.9 -5.1 -5.2 -5.3 -5.4 L 0.6 log[HL]

r -0.4 -0.4 -5.1 -5.2 -5.4 -4.9

0.2 0.2 y = 2.4264x + 12.64 y =2.4217x + 12.436 0.4

0.6 0.8

log[HL]

Sm' 1.8 y = 2.5803x + 14.03; y = 2.7296x + 14.96; 1.4 1.2

oat 0.8 e 0.6 0.4 0.5 0.2

-5.4 -5.3 -5.2 -5.1 -4.9 -4.8 -5.4 -5.3 -5.2 -5.1 -4.9 -4.8 log[HL] log[HL]

Ho+3

2.5

y = 2.5259X + MA5*

0.5

-5.4 -5.3 -5.2 -5.1 -4.9 -4.8 log[HL]

Figure 4.45. LogD vs. log[HL] plots of each lanthanide metal ion

111

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.24. LogKex values as a function of ligand concentration

logKPY values [HL] La Ce Pr Nd Sm Eu Ho 5 x 10'6 3.94 4.47 5.69 5.78 7.07 7.89 7.00

1 x 10'5 3.92 4.47 5.71 5.81 7.07 7.88 7.06

1.25 x 1 O'5 3.92 4.47 5.70 5.78 7.02 7.85 7.03

1.5 x 10'5 3.95 4.47 5.69 5.77 7.10 7.91 6.98

4.12.6 LogD vs. p H of HMPVP-913 at 1 x 10'5M

Distribution ratio (D) values and logD values were calculated as in the case of the previous ligands and are tabulated (Table 4.25 and 4.26). The logD values are plotted (Figure 4.46) as a function pH (2.80, 2.85, 2.90. and 2.95) at constant concentration. The experimentally obtained slopes were used to calculate the KeX values (Table 4.29) and compared them among the lanthanide metal ions.

Table 4.25. D values for each lanthanide metal ion at different pH

D values pH La Ce Pr Nd Sm Eu Ho 2.80 0.04 0.26 0.59 0.92 3.50 5.26 9.89

2.85 0.06 0.33 0.72 1.13 4.42 6.67 12.45

2.90 0.12 0.55 1.22 1.85 7.05 10.91 20.89

2.95 0.14 0.62 1.35 2.11 8.11 12.46 25.08

112

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.26. LogD values for each lanthanide metal ion at different pH

loeD values pH La Ce Pr Nd Sm Eu Ho 2.80 -1.35 -0.59 -0.23 -0.03 0.54 0.72 1.00

2.85 -1.23 -0.49 -0.14 0.05 0.65 0.82 1.10

2.90 -0.92 -0.26 0.09 0.27 0.85 1.04 1.32

2.95 -0.86 -0.21 0.13 0.32 0.91 1.10 1.40

y=3.553x- 11.307 R2 = 1

- 0.22 PH

Pr+3 Nd+3

y y = 2.6213x - 7.5751

Eu+3 1 1.2 0.9 1.1 0.8 1 y y = 2.5937x -6.72 q 0.9 y = 2.6752x-6.7718 $ 0.7 £ 0.8 R2"1 0.6 0.7 0.6 0.6 0.4 2.75 2.8 2.85 2.9 2.95 3 2.75 2.8 2.85 2.9 2.95 3 pH

Ho+3

1.6. 1.5 1.4 1.3 a s 12 1.1 1 0.9 0.82.75 2.85 2.9 2.95 3 PH

Figure 4.46. LogD vs. pH plots of each lanthanide metal ion

113

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.27. Selectivity (a) for adjacent lanthanide metal ions at different pH

a values pH Ce/La Pr/Ce Nd/Pr Sm/Nd Eu/Sm Ho/Eu 2.80 5.84 2.27 1.57 3.79 1.50 1.88

2.85 5.48 2.22 1.57 3.90 1.51 1.87

2.90 4.64 2.21 1.51 3.81 1.55 1.92

2.95 4.52 2.17 1.56 3.85 1.54 2.01

Table 4.28. Slopes for each lanthanide metal ion

La Ce Pr Nd Sm Eu Ho Ligand depend 2.12 2.22 2.42 2.45 2.58 2.73 2.53

pH depend 3.55 2.74 2.62 2.57 2.57 2.68 2.87

Table 4.29. LogKex values for each lanthanide metal ion as a function of pH

logKpy values pH La Ce Pr Nd Sm Eu Ho 2.80 3.25 4.39 5.74 5.72 6.76 6.56 7.00

2.85 3.24 4.40 5.78 5.76 6.80 7.87 6.70

2.90 3.30 4.47 5.83 5.81 6.86 7.93 6.78

2.95 3.22 4.37 5.72 5.70 6.73 7.80 6.65

114

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 4.12.7. LogD vs. logrHLl of HBMPP-750 at pH 2.40

The distribution ratio (D) and its log values were calculated from the

experimentally obtained retention time. All D values and logD values are displayed in the

following tables (Table 4.30 and 4.31). Selectivity values of HBMPP-750 for each

lanthanide metal ion were calculated from D values and displayed in table 4.32. Slopes

were obtained (Table 4.37) by plotting all logD values as a function of ligand

concentration (Figure 4.47) at constant pH 2.40

Table 4.30. D values for each lanthanide metal ion at different ligand concentration

D values [HL] log[HL] La Ce Nd Sm Tb 2.5 x 10'6 -5.60 0.05 0.15 0.27 —

5 x 10'6 -5.30 0.05 0.26 0.83 2.73 5.79

8 x 10‘6 -5.10 0.26 0.89 2.40 8.91 19.10

1 x 10'5 -5.00 0.28 0.98 2.65 10.30 22.12

Table 4.31. LogD values for each lanthanide metal ion at different ligand concentration

logD values [HL] log[HL] La Ce Nd Sm Tb 2.5 x 10’b -5.60 -1.34 -0.82 -0.57 —

5 x 10'6 -5.30 -1.30 -0.58 -0.08 0.44 0.76

8 x 10'6 -5.10 -0.58 -0.05 0.38 0.95 1.28

1 x 10'5 -5.00 -0.55 -0.01 0.42 1.01 1.34

115

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. -1.4 r -0 .7

- 1.2 • - 0.6 • -0 .5 a ■ -0 .4 -O.i y=2.6584x + 12.837 y = 2.0078x+, ? - -0 .3 -0 .( - - 0.2 :9-0.1 -5 .2 -5 .3 -5 .4 .0,2 -5.05 -5.1 -5.15 -5.2 -5.25 -5.3 -5.35 0.1 lo g [H L ] log[H L ]

N d '

-1.5

)46x + 15.621

5 4 5 . 5. 5 751-5.2 -5.4 -5 .5 -5 .6 -5.7-5.1

0.5

Tb+3

-0 .5 -5 .1 -5.2 -5.3 -5.4 -5.5 -5.7

0 .5 y=3.2592x + 17.816 R2 = 1

lo g [H L ]

Figure 4.47. LogD vs. log[HL] plots of each lanthanide metal ion

Table 4.32. Selectivity (a) for adjacent metal ions at different ligand concentration

a values [HL] Ce/La Nd/Ce Sm/Nd Tb/Sm 3.30 1.79 2.5 x 10‘6 ——

5 x 10’6 5.23 3.17 3.28 2.12

8 x 10’6 3.37 2.71 3.71 2.14

1 x 10‘5 3.45 2.70 3.89 2.15

116

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Kex values were calculated from experimentally obtained slopes using equation 3.

All Kex values were tabulated in their log values (Table 4.33).

Table 4.33. LogKex values for each lanthanide metal ion as a function of concentration

losKnY values [HL] La Ce Nd Sm Tb

2.5 x 10'6 —— 8.89 10.38 11.11

5 x 10"6 8.03 8.86 9.20 10.69 11.38

8x 10'6 7.94 8.77 9.09 10.57 11.26

1 x 10"5 7.69 8.51 8.83 10.33 11.00

4.12.8. LogD vs. p H of HBMPP-750 at 5 x 10'6M i All D values and logD values were calculated at different pH from experimentally

obtained retention time for each lanthanide metal ion and displayed in table 4.34 and

4.35. The logD values were plotted (Figure 4.48) as a function of pH to obtain the slopes

for each lanthanide metal ion. All slopes are tabulated in table (Table 4.37).

Table 4.34. D values for each lanthanide metal ion at different pH

D values pH La Ce Nd Sm Tb 2.36 0.07 0.27 0.73 2.77 6.03

2.40 0.08 0.34 1.03 3.49 7.45

2.43 0.11 0.42 1.18 4.31 9.40

2.47 0.16 0.57 1.55 5.85 12.94

117

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.35. LogD values for each lanthanide metal ion at different pH

loeD values pH La Ce Nd Sm Tb 2.36 -1.13 -0.57 -0.14 0.44 0.78

2.40 - 1 .1 0 -0.47 0 .0 1 0.54 0.87

2.43 -0.96 -0.37 0.07 0.63 0.97

2.47 -0.81 -0.25 0.19 0.77 1 .1 1

L a +3 C e * 3

- 1 .4 - - 0 .7 -

- 1 .2 - - 0 .6 -

- 0 .5 -

a - ° - 8 y = 3.1062x- 8.4999^ a R2 = 1 lo g D

■2 -0 .6 o o> o = 2.9436X - 7 .5 2 4 ^ -0 .4 y -0 .2 R2 = 1 © c cvf1 34 2.36 2.38 2.4 2.42 2.44 2.46 2 .4 8 -°v 34 2.36 2.38 2.4 2.42 2.44 2.46 2 .4 8

lo g [H L ] lo g [ H L ]

0 .2 5 -> 0.2 0 .1 5 0 .7 0.1 0.6 i.9 9 6 5 y=2.9573x-6.5451 O 0 .0 5 R2 = 1 R2 = 1 2 0 .4 0 .3 2 .4 2 .4 5 2 .5 -o.o£- 0.2 - 0.1 ■ 0.1 - 0 .1 5 • 2 .3 4 - 0.2 • 2.36 2.38 2.4 2.42 2.44 2 .4 6 2 .4 8 lo g [ H L ]

TbH

y = 3.0382x-6.4027 R * = 1

2 . 3 5 2.4 2 .4 5 2.5

Figure 4.48. LogD vs. pH plots of each lanthanide metal ion

118

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 4.36. Selectivity (a) for adjacent metal ions at different pH

a values pH Ce/La Nd/Ce Sm/Nd Tb/Sm 2.36 3.68 2.70 3.80 2.17

2.40 4.22 3.07 3.39 2.14

2.43 3.82 2.80 3.65 2.18

2.47 3.61 2.74 3.77 2 .2 1

Table 4.37. Slopes for each lanthanide metal ion

La Ce Nd Sm Tb

Ligand depend 2 .6 6 2 .0 1 3.01 3.13 3.26

pH depend 3.11 2.94 2.94 2.96 3.04

Table 4.38. LogKex values for each lanthanide metal ion at different pH

loaKev values

PH La Ce Nd Sm Tb 2.36 8.05 8.87 9.17 10.69 11.40

2.40 8.03 8 .8 6 9.20 10.69 11.38

2.43 8 .0 0 8.85 9.17 10.67 11.37

2.47 8.06 8.87 9.18 10.70 11.40

119

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 4.13. Discussion of separation mechanism

The Kex values for the light, middle, and heavy lanthanide metal ions taken

together with the slopes for pH and ligand dependence indicate a complex mechanism for

separation than they are predicted by the simple equilibrium model. According to the

observed selectivity & Kex values, we can say that almost all of the amphiphilic ligands

that we have synthesized provided good separation. The slopes drawn from logD vs. pH

and logD vs. [HL] are almost closer to 3, which indicate that the ratio of the metal to

ligand could be 1:3. Our experimental values for slope are always a nonintegral and

approximately 3, which was correlated to the value of simple extraction model. The

reason for slopes not be a simple integer could be the extent of metal-ligand complex

formation and its distribution between stationary phase and aqueous phase over a multi

stage. When the mobile phase (ligand solution) passing through the Cis column, ligand

could be adsorbed on the surface of stationary phase (Cjg silica gel) and results in either

monolayer or multiple layers of the ligand were formed. This kind of self-assembly can

occur through tail to tail or head to head association as showed in figure 4.18.

120

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CHAPTERV

LANTHANIDE METAL IONS SEPARATION BY HPLC METHOD USING AMPHIPHILIC ACYLISOXAZOLONES

5.1. Introduction

Since we have succeeded in the separation of lanthanide metal ions using

amphiphilic acylpyrazolone ligands, this work was continued with other compounds

analogous to pyrazolone structure and properties such as amphiphilic acylisoxazolones.

As these acylisoxazolones have lower pKa values (1.0-2.0) than those of

acylpyrazolones, they have been proclaimed as chelating ligands in the solvent extraction

method for the separation of lanthanide metal ions, ’ ’ ’ and other metals. ’ ’ ’

Amphiphilic acylisoxazolone ligands have been synthesized as described in Chapter 3

and employed them to investigate their lanthanide metal ion recognition efficacies

through the HPLC separation method. We have envisaged that these ligands might also

spontaneously self-assemble in the aqueous phase to form variety of nanostructures like

amphiphilic acylpyrazolone ligands. Their ability to bind to lanthanide metal ion and how

their self-assemblies affect the metal ion separation were investigated. The significant

details of these studies are discussed in the results and discussion section of this chapter.

5.2. Instrumentation and experimental methods

HPLC system, column packing and all other experimental conditions such as

preparation of mobile phase, preparation of metal ion solution, and TEM image analysis

were carried out same as that of amphiphilic acylpyrazolone ligand system.

121

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 5.3. Results and discussion Amphiphilic acylisoxazolone ligands were synthesized using monodisperse PEG

moieties (PEG 516, PEG913 ) and polydisperse PEG 750. The chemical structures of these

amphiphilic ligands are shown in figure 5.1.

,CH OH OH N= N = N = 0 ^ OPEG7 5 0 0 ^ / OPEG516

OH 0 OH 0 HMVIS-750 HMVIS-516 ° H ° HMVIS-913

JP h Ph Ph N=r N = 0 ^ OPEG7 5 0 OPEG516 OPEG913

OH 0 OH 0 OH 0 HPVIS-750 HPVIS-516 HPVIS-913

OPEG7 5 0 M_ OPEG516 N OPEG913

OH 0 HBPIS-750 OH 0 HBPIS-516 OH 0 HBPIS-913

750 N OPEG913

OH 0 OH 0 OH 0 HBMIS-750 HBMIS-516 HBMIS-913

Figure 5.1. Chemical structures of synthesized amphiphilic acylisoxazolone ligands

After the performance of HPLC experiments, the resulting best HPLC separation

chromatograms of lanthanide metal ions for each ligand are shown in the following

section. Ligand concentration, pH of the mobile phase, ligand structure and lanthanide

metal ions are showed on the chromatogram. Separation chromatograms indicated that

amphiphilic acylisoxazolone ligands did not separate the mixture of seven lanthanide

122

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. metal ions, instead they separated a mixture of five lanthanide metal ions with good

separation resolution and selectivity.

5.4. HPLC separation chromatograms bv HBPIS-750. HBPIS-516. HBPIS-913, HPVIS- 750. and HPVIS-516 ligands

All amphiphilic benzoylisoxazolones (HBPIS-750, HBPIS-516, HBPIS-913) did

not provide any peak on the chromatogram for La+3 metal ion. The regular metal ion peak

could not be seen on the chromatograms (Figure 5.2, 5.3, and 5.4). Since these ligands

have two electron withdrawing groups (phenyl rings) on the isoxazolone ring, the ligands

might have very low pKa values (<1.0), and would result in strong complexation with

high association constant. Another possibility would be the metal complex could have

been strongly adsorbed on the column, which might be causing very slow distribution

between stationary phase and mobile phase. The peak for metal complex that comes out

from the column might not be able to see on the chromatogram as the peak is too wide

and flat. Other ligands such as the amphiphilic valeroylisoxazolones (HPVIS-750, and

HPVIS-516) also did not provide any separation (Figure 5.5 and 5.6) for a mixture of five

lanthanide metal ions (La+3, Ce+3, Nd+3, Sm+3, Ho+3).

La+3, 0.000005M, pH 2.13

30 -I------,------,------,______,______, o 5 10 15 20 25 Retention time (min)

Figure 5.2. HPLC separation chromatogram by HBPIS-750 (5 x 10'6M)

123

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. La+3, 0.000001 M, pH 2.13 Ph

45 - OH 0

8 o E o0) c <0 f 30 - M -Q < 25

0 10 20 3040 50 60 Retention time (min)

Figure 5.3. HPLC separation chromatogram by HBPIS-516 (1 x 10'6M)

La+3, 0.000001 M,pH2.08

N— 100 - |

90 - OH

z> < 80 - 08 1 70 - oa> 60 -

o <0 50 - •O < 40

■5 5 15 25 3545 55 Retention time (min)

Figure 5.4. HPLC separation chromatogram by HBPIS-916 (1 x 10'6M)

124

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00002M, pH 2.14 N = 2500 OPEG7 5 0 OH 0

2000 -

« 1500 - £ o

o A<0 < 500 -

0 2 3 4 56 7 Retention time (min)

Figure 5.5. HPLC separation chromatogram by HPVIS-750 (2 x 10'5M)

0.000005M, pH 2.14

900 n

800 * OH

700 -

600 - o 500 -

400 -

300 -

200 -

100 -

Retention time (min)

Figure 5.6. HPLC separation chromatogram by HPVIS-516 (5 x 10'6M)

125

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 5.5. HPLC separation chromatograms by HMVIS-750 and results

Methylvaleroylisoxazolone-PEGvso (HMVIS-750) ligand provided good

separation, but not as good as amphiphilic pyrazolone ligands. In the initial trials, a

mixture of three lanthanide metal ions (La+3, Sm+3, Ho+3) has been used as a sample

solution. This ligand provided very good selectivity for light, middle, and heavy

lanthanide metal ions (Figure 5.7). In the following trials a mixture of five lanthanide

I q I ^ I i t o metal ions (La , Ce , Nd , Sm , and Ho ) was used, and obtained good separation

with good selectivity and separation resolution (Figure 5.8).

0.00002M, pH 2.18 CH

OH 0 120 La, 7.80 Sm, 20.20

S 100 Ho, 30.97

Retention time (min)

Figure 5.7. HPLC separation chromatogram by HMVIS-750 (2 x 10'5M)

126

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00004, pH 2.16 .CH N— 120

110 OH O 100 Nd, 31.63 S m , 47.03 90 La, 16.63 Ce, 24.8: Ho, 71.10 80

30 20 40 60 80 100 Retention time (min) Figure 5.8. HPLC separation chromatogram by HMVIS-750 (4 xlO°M)

Table 5.1. D values at different ligand concentration

D values [HL] La Ce Nd Sm Ho 2 x 10"5 2.59 - - 8.31 13.27

4 x 10'5 6.66 10.44 13.58 20.67 31.76

Table 5.2. Selectivity (a) for adjacent lanthanide metal ions at different ligand concentration

a values [HL] Sm/La Ho/Sm

2 x 10'5 2.59 - - 8.31

a values Ce/La Nd/Ce Sm/Nd Ho/Sm 4 x 10"5 6.66 10.44 13.58 20.67

127

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. D values (Table 5.1) and Selectivity values (Table 5.2) indicate that the HMVIS-

750 ligand showed good selectivity and provided good separation for a mixture of five

lanthanide metal ions.

5.6. HPLC separation chromatograms by HMVIS-516 and results

Methylvaleroylisoxazolone-PEG 5i6 (MVIS-516) ligand also provided good

separation. High concentration (5 xlO'5M) ligand provided good separation with good

selectivity (Figure 5.9) for a mixture of five lanthanide metal ions. Since it has separated

five metal ions with good separation resolution, one more middle lanthanide (Eu+3) was

added to the mixture of five metal ion samples. The separation chromatogram (Figure

5.10) indicated that this ligand has poor selectivity for middle lanthanides.

0.00005M, pH 2.12 CH: N = -

130 OH O 120 Nd, 23.83 Smj 3 5 1 0 La, 12.43 110 Ce, 18. Ho, 52.40 100

100 Retention time (min)

Figure 5.9. HPLC separation chromatogram by HMVIS-516 (5 x 10'5M)

128

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00005M, pH 2.12 ,CH N= 150 OH 0 130 - Sm, 37.27 Nd, 25.76 Eu, 43.20 La,* 13.40 i 110 - Ce, 20.0 SI Ho, 57.17

70 -

50 -

Retention time (min)

Figure 5.10. HPLC separation chromatogram for six metal ions by HMVIS-516

Separation of a mixture of five lanthanide metal ions was tried with other ligand

concentrations (1 x 10'5M and 2 x 10'5M). These ligand concentrations provided not good

baseline separation and selectivity (Figure 5.11 and 5.12).

CH N=<

La. 15.76 70 - OH O Ce, 32.37 Sm. 60.23 — 65-

Ho. 81.70

■e 50-

< 45 -

40 -

100 120 R etention time (min)

Figure 5.11. HPLC separation chromatogram by HMVIS-516 (1 x 10"5M)

129

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00002M , pH 2.19 ,CH N= 150 i Nd. 12.70 Sm, 17.97 OH 0 130 - Ce, 1 0 .1 Ho, 26.43 La, 7.40

Retention time (min)

Figure 5.12. HPLC separation chromatogram by HMVIS-516 (2 x 10'5M)

5.7. HPLC separation chromatograms bv HMVIS-913 and results

Methylvaleroylisoxazolone-PEG 9 i3 (HMVIS-913) ligand showed good selectivity

with poor baseline separation, when compared with HMIVS-750 and HMVIS-516.

Separation conditions and chromatograms were shown in the following figure 5.6. A

mixture of five lanthanide metal ions (La+3, Ce+3, Nd+3, Sm+3, and Ho+3) could only be

separated by this ligand (HMVIS-913).

130

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0.00001 M, pH 2.19 ,CH N = 100 n OH O 90 - Nd, 31.33 80 - Sm , 49.80 La, 15.33 Ce, 23 j Ho, 77.43 70 -

60 -

50 -

40 -

100 120 Retention time (min)

0.00002M, pH 2.19 CH N = r 95 -i

Nd, 25.70 OH O 85 - La, 12.70 Ce, 19. Sm ,41.20

75 - Ho, 76.50 65 -

45 -

100 Retention time (min)

Figure 5.13. Separation chromatograms for five metal ions by HMVIS-913

131

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ,CH: 0.000005M, pH 2.20

110 N5?’ 18-27 S m . 27.23 OH 0 100 Ho, 42.47 La. 9.87

Retention time (min)

Figure 5.13. - Continued

5.8. LogD vs. loefHLl of HMVIS-516 at pH 2.19

Distribution ratio (D) values and their log values were calculated from

experimentally obtained retention time for each lanthanide metal ion. All D values and

logD values are tabulated in the following table (Table 5.3 and 5.4). Selectivity for each

lanthanide metal ion was calculated from D values and tabulated in table 5.5. All logD

values were plotted against ligand concentration at constant pH 2.19 to obtain the slopes.

These experimentally obtained slopes were used to calculate the values. All KeX

values are tabulated in their log values in table 5.6.

132

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 5.3. D values at different ligand concentrations at pH 2.19

D values [HL] log[HL] La Ce Nd Sm Ho 2x 10'5 -4.70 0.55 0.79 1.05 1.60 2.50

5 x 10'5 -4.30 2.69 3.90 4.70 6.61 8.21

Table 5.4. LogD values as a function of ligand concentration at pH 2.19

loaD values [HL] log[HL] La Ce Nd Sm Ho 2 x lO'* -4.70 -0.26 -0.10 0.02 0.20 0.40

5 x 10'5 -4.30 0.43 0.59 0.67 0.82 0.91

Table 5.5. Selectivity (a) for adjacent lanthanide metal ions at different ligand concentration

a values [HL] Ce/La Nd/Ce Sm/Nd Ho/Sm 2 x 10'5 1.458 1.32 1.53 1.57

5 x 10’5 1.457 1.21 1.41 1.24

133

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. .0.4

-4.8 4.3 -42

log[HL] log[HL]

4.8 -4.7 -4.6 -4.5 -4.4 -4.3 -4.1 -4 -3.9

log[HLI log[HL]

y=1.3527

lo g [H L ]

Figure 5.14. LogD vs. log[HL] plots of each lanthanide metal ion

Table 5.6. LogKex values for each metal ion as a function of ligand concentration

loalCvvalues [HL] La Ce Nd Sm Ho 2 x lO’5 3.635725 3.786711 3.891543 3.295629 2.546

5 x 10’5 3.635733 3.786708 3.891498 3.295558 2.542

134

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 5.9. LogD vs. pH of HMVIS-516 at 5 x 10'5M

All D values were calculated from experimentally obtained retention time for

each lanthanide metal ion and were tabulated in the following tables (Table 5.7 and 5.8).

Selectivity of this ligand (HMVIS-516) for each lanthanide metal ion was calculated and

displayed in table 5.9. All logD values were plotted against pH to obtain slopes (Table

5.10). All Kex values were calculated from experimentally obtained slopes and tabulated

in table 5.11.

Table 5.7. D values for each lanthanide metal ion as a function of pH

D values pH La Ce Nd Sm Ho 2.12 3.310 5.240 6.832 10.144 15.793

2.19 4.113 6.441 8.193 12.879 19.391

Table 5.8. LogD values for each lanthanide metal ion as a function of p]

loeD values pH La Ce Nd Sm Ho 2.12 0.52 0.72 0.83 1.01 1.20

2.19 0.61 0.81 0.91 1.11 1.29

Table 5.9. Selectivity (a) for adjacent lanthanide metal ions at different pH

a values pH Ce/La Nd/Ce Sm/Nd Ho/Sm 2.12 1.58 1.30 1.49 1.56

2.19 1.57 1.27 1.57 1.51

135

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Table 5.10. Slopes for each lanthanide metal ion

La Ce Nd Sm Ho pH depend 1.35 1.28 1.13 1.48 1.27

Ligand depend 1.74 1.74 1.64 1.55 1.30

Table 5.11. LogKex values for each lanthanide metal ion at different pH

logKpv values PH La Ce Nd Sm Ho 2.12 3.78816 3.96255 4.08269 3.5169 2.81163

2.19 3.78813 3.96252 4.08268 3.5168 2.81164

In the case of amphiphilic acylisoxazolones also the Kex values for the light,

middle, and heavy lanthanide metal ions taken together with the slopes for pH and ligand

dependence indicate a more complex mechanism for separation than they are predicted

by the simple equilibrium model.

5.10. Self-assembly behavior of the amohiphilic acvlisoxazolone ligands

The self assembly behavior of all amphiphilic acylisoxazolone ligands in the

aqueous phase were examined by TEM images to gain an understanding of their

influence on their lanthanide metal ion selectivities. Ligand samples were prepared in

similar manner to that of amphiphilic acylpyrazolone ligands. TEM images are displayed

in the following figures (Figure 5.15 to 5.23).

136

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 5.15. TEM images of benzoylphenylisoxazolone-PEG7 5 o (HBPIS-750)

Figure 5.16. TEM images of benzoylphenylisoxazolone-PEGsie (HBPIS-516)

Figure 5.17. TEM images of benzoylphenylisoxazolone-PEGgo (HBPIS-913)

137

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 5.18. TEM images of phenylvaleroylisoxazolone-PEGyso (HPVIS-750)

Figure 5.19. TEM images of phenylvaleroylisoxazolone-PEG 5i6 (HPVIS-516)

Figure 5.20. TEM images of phenylvaleroylisoxazolone-PEG 9 i3 (HPVIS-913)

138

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Figure 5.21. TEM images of methylvaleroylisoxazolone-PEG 75 o (MVIS-750)

Figure 5.22. TEM images of methylvaleroylisoxazolone-PEG 5i6 (MVIS-516)

Figure 5.23. TEM images of methylvaleroylisoxazolone-PEGgn (MVIS-913)

139

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. In case of amphiphilic acylisoxazolone ligands, the similar self-assemblies have

been observed when compared with amphiphilic acylpyrazolone ligands. Amphiphilic

acylisoxazolones formed different nanostructures among themselves, which can be

attributed to the different substituents on the isoxazolone ring. The HBPIS-750 ligand

solution showed linear self-assemblies like broken nano-tubes, needles and vesicles

(Figure 5.15) which are similar to amphiphilic valeroylpyrazolone ligands, whereas the

HBPIS-516, HBPIS-913 ligands showed vesicles with linear aggregation and dendritic

structures (Figure 5.16 and 5.17). Amphiphilic phenyl valeroylisoxazolones (HPVIS-750,

HPVIS-516, and HPVIS-913) showed vesicles with linear aggregation and hying to grow

dendritic structures (Figure 5.18, 5.19, and 5.20)

Amphiphilic methylvaleroylisoxazolone ligands (HMVIS-750, HMVIS-516, and

HMVIS-913) formed linear structures such as tubes, nano-rods dendritic structures, small

thick needles, and linear fibers (Figure 5.21, 5.22, and 5.23). Though these ligands

provided better separation for a mixture of lanthanide metal ions when compared with

amphiphilic HBPIS and HPVIS ligands, the calculated slopes from experimental D

values are not closer to three. As these ligands have lower pKa values than that of

pyrazolones, they should have higher pKa values than that of pyrazolones. That means,

the separation mechanism of these ligands could be much more complex than

pyrazolones.

140

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CHAPTER VI

SUMMARY AND FUTURE DIRECTIONS

6.1. Summary

A family of new 4-haloacylpyrazolones (C2, 02, C3, 03, C4, 04, and 05) and

4-haloacylisoxazolone (9, 10, 12, 13, 15, 17, 18, 19, 20, 21, and 22) compounds have

been synthesized and fully characterized. The 4-acylated products were synthesized

efficiently from TV-acylated compounds using an effective acyl-transfer inducing reagent

such as DMAP with high yield. Single crystal X-ray structural analysis of

acylpyrazolones and acylisoxazolones was performed to understand their molecular

structures in terms of bond angles, bond lengths and the position of the OH group in the

ligands. These crystal structures indicated that the pyrazolones exist in the enol form with

the OH group on the pyrazolone ring, whereas isoxazolone is in the yS-diketone form

(existence of keto and enol forms is a function of the substituents present on the ring and

nature of the solvent in which they dissolve). Different kinds of substituents such as

electron withdrawing groups (benzoyl, phenyl), and electron donating groups (methyl,

valeroyl) are introduced at 3rd and 4th positions of the pyrazolone and isoxazolone rings

along with the different types of PEG moieties (monodisperse and polydisperse) to

prepare variety of amphiphilic ligands.

Novel amphiphilic acylpyrazolones (Figure 4.5) and amphiphilic

acylisoxazolones (Figure 5.1) have been synthesized and characterized. To substantiate

the central hypothesis of the research, namely, that organized nanoscale self-assemblies

141

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. can provide excellent trivalent lanthanide metal ion selectivities, these amphiphilic

acylpyrazolones and amphiphilic acylisoxazolone ligands have been employed for the

separation of a mixture of light, middle, and heavy lanthanide metal ions. The self-

assembly nature of these ligands in the aqueous phase has been characterized by TEM.

These TEM images indicated that the amphiphilic ligands spontaneously form spherical,

dendritic, and linear (fibers, tubes, and rods) nanoscale self-assembled structures in the

aqueous phase, which can influence their metal ion separation efficacies. After analyzing

the HPLC separation results for lanthanide metal ions, we can say that structure of the

amphiphilic ligand and nature of the PEG moiety also play a key role, along with the self-

assembly property of the amphiphilic ligands.

Several types of amphiphilic acylpyrazolones and amphiphilic acylisoxazolones

were used for the separation of lanthanide metal ions. Some of them provided best

separations in terms of efficacies, selectivities, and resolution. Amphiphilic

valeroylpyrazolone ligands (HMPVP-750, HMPVP-516, and HMPVP-913) efficiently

separated a mixture of seven light, middle, and heavy lanthanide metal ions (La+3, Ce+3,

Pr+3, Nd+3, Sm+3, Eu+3, and Ho+3) with baseline separation. HMPVP-750 and HMPVP-

516 ligands even provided a best baseline separation for a mixture of eight lanthanide

metal ions (La+3, Ce+3, Pr+3, Nd+3, Sm+3, Eu+3, Tb+3, and Lu+3), Whereas amphiphilic

benzoylpyrazolone (HBMPP-750) provided a best separation of a mixture of five

I I q ^ * 3 I 1 _i_o ______lanthanide metal ions (La , Ce , Nd , Sm , and Tb ) with baseline separation. This

ligand with monodisperse PEG moieties (HBMPP-516 and HBMPP-913) could also

separate a mixture of five lanthanide metal ions with poor selectivity and poor resolution.

142

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Amphilic benzoylphenylisoxazolone ligands (HBPIS-750, HBPIS-516, and

HBPIS-913) provided no metal ion peak on the separation chromatogram. Amphiphilic

phenylvaleroylisoxazolone ligands (HPVIS-750, HPVIS-516) ligands provided no

separation for the mixture of five lanthanide metal ions in various conditions.

Amphiphilic methylvaleroylisoxazolone ligands (HMVIS-750, HMVIS-516, and

HMVIS-913) provided better separation for a mixture of five lanthanide metal ions when

compared with HBPIS and HPVIS ligands.

According to the HPLC separation chromatograms and results, we can say that

the separation mechanism (metal ion recognition) is complex, which could be mainly

influenced by the structure of amphiphilic ligand, nature of self-assemblies of these

ligands in the aqueous mobile phase as well as on the stationary phase. The distribution

ratio of a metal ion exhibited different values (gradually increases as column gets

equilibrated with ligand) with the equilibration of column. This indicates us that each

ligand has its own behavior in the mobile phase and on the stationary phase with different

types of self-aggregation that eventually leads to form nanostructures and helps in

separation of trivalent lanthanide metal ions. This self-assembly behavior was confirmed

by TEM image analysis. The self-assembly behavior of these ligands in the presence of 4-9 4-3 metal ions (Mg and Sm ) revealed that the formation of nanostructures are much larger

in the presence of metal ions. Aging these solutions in the presence of metal ions leads to

form more extensive nanostructures.

143

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Most of the times, spherical structures like vesicles are formed by amphiphilic

benzoylpyrazolone ligands (HBMPP-PEGX) Amphiphilic benzoylisoxazolone ligands

self-assembled as vesicles and linear structures like needles and nano-rods. All the time,

linear structures are formed by amphiphilic valeroylpyrazolone and valeryoylisoxazolone

ligands. Dendritic structures are observed by HMPVP-516 and HMVIS-516 ligands.

6.2. Future directions

So far, this research project has described the self-assembly behavior and

lanthanide metal ion recognition property of novel amphiphilic acylpyrazolones and

amphiphilic acylisoxazolones. It identified that the amphiphilic acylpyrazolones have

excellent recognition capabilities for trivalent lanthanide metal ions, and is extended to

investigate the metal ion recognition property of amphiphilic acylisoxazolones, and

studied some of their lanthanide metal ion recognition properties. Still, many

fundamental concepts need to be understood to employ these novel systems as efficient

separation tools.

The self-assembly behavior of the ligands on the Cig silica surface needs to be

characterized to understand their behavior on stationary phase. The high resolution TEM

images should be taken to understand their self-assembly nature very well. More studies

on the amphiphilic acylisoxazolones should be carried out to understand their separation

mechanism as they provided poor separation with poor selectivity. Separation mechanism

of HMVIS-516 was studied as a function of ligand concentration and pH. Other types of

amphiphilic acylisoxazolone ligands have been synthesized (HMVIS-750, HMVIS-913,

144

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. HBMIS-750, HBMIS-516, HBMIS-913, HPVIS-750, HPVIS-516, HPVIS-913). Further

studies have to be done for their separation mechanism for a better understanding of

lanthanide metal ion recognition. Different types of substituents can also be tried in the

acyl side chain to change the acidity of the ligand, which influences the separation of

metal ions. Studies on the TEM imaging of amphiphilic isoxazolone ligands (HMVIS-

750 and HMVIS-516) and their metal ion complexes need to be performed to discern

their nanoscale structures. The self assembly behavior of amphiphilic benzoylisoxazolone

ligands (HBPIS-750, HBPIS-516, HBPIS-913) on stationary phase as well as in the

mobile phase has to be studied by TEM image analysis. Our separation results, slopes and

KeX values as a function of pH and ligand concentration, indicated that these ligands have

much more complex mechanism when compared with amphiphilic acylpyrazolones.

145

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78. Sek, S.; Misicka, A.; Bilewicz, R. "Effect of interchain hydrogen bonding on electron

transfer through alkanethiol monolayers containing amide bonds". J. Phys. Chem. B

2000,104(22), 5399-5402.

156

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 79. Lim, R.; Li, J.; Li, S. F. Y.; Valiyaveettil, S. "The formation of two-dimensional

supramolecular chiral lamellae by diamide molecules at the solution/graphite interface: A

scanning tunneling microscopy study" Langmuir 2000,16(17), 7023-7030.

80. Schmatloch, S.; Berg, A. M. J.; Alexeev, A. S.; Hofmeier, H.; Schubert, U. S.

"Soluble high-molecular-mass poly(ethylene oxide)s via self-organization"

Macromolecules 2003, 36(26), 9943-9949.

81. Ringler, P.; Schluz, G. "Self-assembly of proteins into designed networks" Science

2003, 302(5642), 106-109.

82. Johnsson, M.; Hansson, P.; Edwards, K. "Spherical micelles and other self-assembled

structures in dilute aqueous mixtures of poly(ethylene glycol) lipids" J. Phys. Chem. B

2001,105(35), 8420-8430.

83. Lin, Z.; He, M.; Scriven, L. E.; Davis, H. T. "Vesicle formation in electrolyte

solutions of anew cationic siloxane surfactant" J. Phys. Chem. 1993, 97(14), 3571-3578.

84. Marques, E.; Khan, A.; Miguel, M. G.; Lindman, B. "Self-assembly in mixtures of a

cationic and an anionic surfactant: the sodium dodecyl sulfate-

didodecyldimethylammonium bromide-water system" J. Phys. Chem. 1993, 97(18),

4729-4736.

85. Sarzanini, C. "Liquid chromatography: a tool for the analysis of metal species" J.

Chromatogr. A 1999, 850, 213-228.

157

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 86. Yoshita, Z.; Kimura, T.; Meguro, Y., "Recent progress in actinide separation

chemistry". 2nd ed.; World scientific, Singapore. 1997.

87. Akashi, K.; Miyata, H.; Itoh, H.; Kinosita, K. "Preparation of giant liposomes in

physiological conditions and their characterization under an optical microscope".

Biophysical Journal 1996, 71(6), 3242-3250.

88. Buonomenna, M. G.; Molinari, R.; Drioli, E. "Selective mass transfer of iron (III) in

supported liquid membrane using highly acidic extractants, 3-phenyl-4-acyl-5-

isoxazolone" Desalination 2002,148(1-3), 257-262.

89. Atanassova, M.; Dukov, I. L. "Solvent extraction and separation of lanthnoids with

mixtures of chelating extractant and l-(2-pyridylazo)-2-naphthol" Separ. Puri. Technol.

2006, 49, 101-105.

90. Umetani, S.; Matsui, M. "Solvent extraction of alkaline-earth metals with 4-acyl-5-

pyrazolones and polydentate phosphine oxides" Anal. Chem. 1992, 64, 2288-2292.

91. Umetani, S.; Matsui, M.; Kihara, S. "The synergestic extraction of alkaline earth

metals with 4-benzoyl-3-methyl-l-phenyl-5-pyrazolone and bis(diphenylphosphinyl)

methane" Chem. Let. 1986, 1545-1548.

92. Jordanov, V. M.; Atanssova, M.; Dukov, I. L. "Solvent extraction of lanthanides with

l-phenyl-3-methyl-4-benzoyl-5-pyrazolone" Separ. Sci. Technol. 2002, 37(14), 3349-

3356.

158

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 93. Xiong, Y.; Wang, X.; Li, D. "Synergestic extraction and separation of Heavy

lanthanide my mixtures of bis(2,4,4-trimethylpentyl)phosphinic acid and 2-ethylhexyl

phosphinic acid mono-2-ethylhexyl ether" Separ. Sci. Technol 2005, 40, 2325-2336.

159

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. APPENDICES

160

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Appendix A

'H NMR, 13C NMR, IR, and LC/MS spectral data of acylpyrazolones

161

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. PH N =

J\___

Ph' OH 0 C2 (13C NMR in CDCI3)

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ,CH 9 0 n N = Ph OH O 80 - (IR)

70 -

50 -

4 0 -

3 0 -

20 4000 3500 3000 2500 2000 1500 1000 500 wave number (cm-1)

LCMS (ESIfve) spectrum of compound C2, (M+ ion + mwt of Na = 293 + 23) fciten. (x 1 .0 0 0.000)

823 899.9E7 1043 2 5 0 5 0 0 7 5 0 1 2 5 0 rrVz

LCMS (ESI've) spectrum of compound C2, (M+ ion = 293) nten.(x1.000.000)

2.0

1 .5

1.0

2M4 6Q5 0.0^ 111 174 2271 gloi 4 0 9 AQA 775 8 4 8 1 0 4 3 1 1 3 7 1.22.7 1348 5 0 0 7 5 0 1000

163

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH N =

0 Cl 0 02 (1H NMR in CDCI3)

____

/ " j

lil - - iK JI'M ' "■ 'M A \ aassjf iiiiiii i8H------5535555 3333 X : parts per NCilUon t 1H

.CH

0 Cl 0 02 (13C NMR in CDCI3)

mnA JL mnmiA mm 1 9 & 0 isos r,170^ IMA ISOjO j 140^ I38j«^ ,1300 110* 100* , A ir r r Rtf*m II 1 i X ; parti p tr M U jpa: 13C

164

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 80 ,CH N=< Ph 70

(IR) 60

000 0 3 5 0 0 3 0 0 0 2 5 0 0 2 0 0 0 1500 1000 5 0040 wave number (cm-1)

P h" OH 0 C3 (1H NMR in CDCI3)

' ' 7 ,: mm iillll I

X t parts p«r MilUon : 1H

165

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 60 Ph OH o

50 (IR)

0 3900 3400 2900 2400 1900 1400 900 400

166

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LCMS (ESI+ve) spectrum of compound C3, (M+ ion + mwt of Na = 326 + 23) friten.Cx1.000.00C0______

349 5.0

4 . 0

3.0

2.0 301 5

3 2 7 '7 1.0 381 32S 544 641 723 13.84 0.0 185 203 22_ 8,58 1° 56 1200 250 500 750 1000 1250 rrVz LCMS (ESI ) spectrum of compound C3, (M ion = 326) friten.Cx1.000.0001

3.0 325

2.5

2.0

1.5

1.0

0.5 53 0.0^ 157 217 3Q1[354 411 477 ____ 5^7 6?3 73^ 843___ 9 4 3 1.Q21 1 1 . 6 7 l 2 3 p 1374 250 500 750 1000 1250 mCz

CH N = r

03 (1H NMR in CDCI3)

167

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH

0 03 (13C NMR in CDCI3)

•0 m m * man0 n m W m m 12M 11M MM , «m _ m o r tai m A SB5S CR*

CH 90 N=< Ph

80 -

70 -

60 -

50 -

40 -

30 -

20 4 0 0 0 3500 3000 2500 2000 1000 5001500 w a v e n u m b er (cm -1)

168

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LCMS (ESI+ve) spectrum of compound 03, (M+ ion + mwt of Na = 326 + 23) Inten, ( x 1 0 0 ,0 0 0 ) 4.0

3,5 197 3,0 ; 2,5

2,0

175 261 3)1 1.0 371 0.5 0.01 11,16 14,47 100 200 300 400 500 600 700 800 900 1000 1100 1200 1300 1400 m /z LCMS (ESI've) spectrum of compound 03, (Mf ion = 326)

1.75 257 325 1.50

1,25 361 1.00

0.75 3 >7 344 0.50 635 169 0.25 731 1491 0.001 11,15 m 100 200 300 400 500 600 700 800 900 1000 1100 1200 1300 1400 nVz

N—

04 (1H NMR in CDCI3)

169

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ,CH3 N=- Ph 2

0 04 (13C NMR in CDCI3)

ptrXUMloa * X3C

.CH

A ATs

170

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. ,c h 3 N = (

OH 0 C4 (13C NMR in CDCI3)

90 n

N=< I 85 Ph

80 (IR)

2 70 -

I- 65 -

60 -

55 -

50 4000 3500 3000 2500 2000 1500 1000 500 wave number (cm-1)

171

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LCMS (ESI+ve) spectrum of compound C4, (M+ ion = 336) [nten.(x100.0Q01

6 9 5 W 2 5 5 10,33 2 1 9 T 2 9 1 1 5 7 449 517 557 1719 771 821 9 2 8 ... TiL 1197 1283 , 13L91

CH OH 0 C5 (1H NMR in CDCI3)

S8* I I ! ! ! I ll

172

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 1 CH 1 i Ph CH 1 i- OH 0 C5 (13C NMR in CDCI3)

Ii|M I|irM NiilX

CH. N = i Ph- 50 'CH; OH O

(IR) 40

0 -f- 4000 3500 3000 2500 2000 1500 1000 500 wave num ber (cm-1)

173

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LCMS (ESI+ve) spectrum of compound C5, (M^ ion + mwt of Na = 258 + 23) tilen.(x1,000,000) 10.0 281 7.5

5.0

25S 5,3 407 175 :?9 4M4S7 5?9 . fi.517Q? 813 043 1p15 1137 1423 , 1510 1888 250 500 750 1000 1250 1500 1750 nVz

LCMS (ESLve) spectrum of compound C5, (M+ ion = 258) Inten.(x1,000,000)

24Q 318. 461 ;5?7 10.98 . 1284 1377 1487 1,580 1Q52 . 1308

( H NMR in CDCI3)

L i * -- M i illiiilliliili X i parts par Million 1 1H

174

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH /N—

0 CH 0 05 (13C NMR in CDCI3)

MfcwMMMw* Um MAMM I N I U M 1TM U U l u H J , 7M 5M 4M i m A i3**! r a ™ s i £ S III I Si ii X i p— —r MM— 113C

,CH N=i 80 P h CH 70 OR)

0 4 0 0 0 3 5 0 0 3 0 0 0 2 5 0 0 2000 15 0 0 1000 5 0 0 wave number (cm-1)

175

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LCMS (ESI+ve) spectrum of compound 05, (Mf ion + mwt of Na = 258 + 23) hten.(x1,000,000)

..259

ffi,...... 5896t5. 737795 874 9791039 1325 1459 1S79 1848 250 500 750 1000 1250 1500 1750 LCMS (ESI’ve) spectrum of compound 05, (M4 ion = 258) hlen.(x100,000)_____ 257 5.0 '173

2.5

OC4 427 T . lOPO1985 0.0 II...X W ,,589 671.. .832 940 i 10411)08118? 1284 13^3 1fi7Q 1724 1877 \ 250 500 750 1000 1250 1500 1750 mfz

06 H NMR in CDCI3)

176

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. .CH N =

(13C NMR in CDCI3 )

rtPWUfcimlfl UM IMu* . N4 r tai me A i n LCMS (ESI^6) spectrum of compound 06 , (M+ ion -+■ mwt of Na = 292 + 23)

1.75

1.50

1.25 433 1.00

0.75 293 0.50 26T

0.25 0.001 9^5 1Q45 . 11^3 1208 1 3 5 3 1 471 100 200 300400 500 600 700 800 900 1000 1100 1200 1300 1 4 0 0 nVz LCMS (ESI've) spectrum of compound 06, (M+ ion = 292)

291 1.50

1.25 257

1.00

0.75 361

0.50

0.25 0.001 100 200 300400 500 600 700 800 900 1000 1100 1200 1300 1400 nVz

177

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. .CH

Ph 0 OH

(1H NMR in CDCI 3 )

N="

0 OH

C6 (13C NMR in CDCI3)

X i pwrti per M H aa: 13C

178

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LCMS (ESI've) spectrum of compound C6, (M+ ion = 292) Inten.fx100.000t______

897 1015 Z x 1,,%6[7958?1.8^ SL ,108? 11,79 4311 1467 100 200 300 400 500 600 700 800 900 1000 1100 1200 1300 1400 m/z

179

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Appendix B

*H NMR, 13C NMR, IR, and LC/MS spectral data of acylisoxazolones

180

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH

JCHj, N=<

0 8

(13C NMR in CDCI3 )

AnAiMi mjtlU. 1904 1804 1704 1004 1304 M04 19M 1204 1104 so.# m o 3M 304 104 0 A EPSill X : p « ti perMUU ob : 13C

181

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Wavenumber cm -1

LC/MS (ESI+ve) spectrum of compound 8 (MT1- ion + mwt of Na = 9 9 + 23) Inten.(x1,000,000)

234

154 100

1482 93g ; 10431099 1167 1ff1 1424N 250 500 750 1000 1250 irVz

LC/MS (ESI've) spectrum of compound 8 (M+ ion = 99) hten.(xlOO.OOO) 2 0

19i

? 1495 sJ 133 . 1!! 2J7 352 486 577 671 742 921 969 r 11^9 1955 1381 144K 250 500 750 1000 1250 mIz

182

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH

( H NMR in CDCI3 )

Cl .CH,

0 9

(13C NMR in CDCI3)

ft Mfta ■* A

2*0.0 190.0 ISM 17M, , I«M ISM 14M ISM 12M 11M 10M 9M «M9M

IIIPPJS X : part* pw MMtw: 13C

183

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (ESLve) spectrum of compound 10 ((M4" ion = 217) Inten(x1,000.000) 2.

216; 1.

0. 83, ,1$2l.. 250 500 750 1000 m/z

(1H NMR in CDCI3)

184

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH HN a

a 10 (13C NMR in MeOD)

is^xC r llilililil 1

LC/MS (ESI+ve) spectrum of compound 10 ((M+ ion + wmt of Na = 217 + 23)

. Jnten.(x 1 ,0 0 0 ,0 0 0 )

0.2-'65, i 128 , 182 ,279 j , ¥1 , 1421, (473 5p5,530, 582 161,7 , 676 : 729757 100 200 300 400 500 600 700 m/z

LC/MS (ESI've) spectrum of compound 10 ((M+ ion = 217) Q jnterfxl ,000,000)

5.0

, g s f , m , 100 200 300 400 500 600 700 m/z

185

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH

CH 3 N =

a 11 (13C NMR in CDCI3)

186

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (ESI+ve) spectrum of cyclised compound 11 ((M4 ion = 181)

4.0T 182

3.0

2.0

1.0 1493 412a 500 750 1000 1250 mte LC/MS (ESI've) spectrum of cyclised compound 11 ((M+ ion = 181 + 18) titen.(x1,000,000)______

198

250 500 750 1000 1250

(1H NMR in CDCI3)

liiii

187

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0 Cl -CM, .CH,

(13C NMR in CDCI3)

if"

LC/MS (ESI4^6) spectrum o f compound 15, (M+ ion + mwt of Na = 251 +23)

100 2.5

!0 513 674

100 150 200 250 300 350 400450 500 550 600 650 m/z

LC/MS (ESI've) spectrum of compound 15, (M+ ion = 251) hten.(x1,000,000) 250

0.5

403421 446 631 658 100 150 200 300 350400 450 500 550 600

188

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. CH HN

0 13 (13C NMR in CDCI3)

sis

189

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. 0 OH 15 (1H NMR in CDCI3)

jLJ JU...... A i nr S ill

X - . f i i M f r MUnilH LC/MS (ESI+ve) spectrum of compound 15, (M+ ion + mwt of Na = 251 +23) hten.(x1.000,000) 1.25

1.00 292 0.75 274 0.50 185 306 351 100 523 0.25 2/6 338 363 173 91 116 ;l217 250 l.ip l 331,1 [I,!,,.. 4)4436460 J lf,, . . I , . , ,613 ...... 67.6, . 728 763 787. 822 0.00+ 100 200 300 400 500 600 700 800 m/z

LC/MS (ESLve) spectrum of compound 15, (M+ ion = 251)

3.0

250

100 200 300 500 600 700400 800 m/z

190

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. (1H NMR in CDCI3 )

LC/MS (ESFve) spectrum of compound 18, (M+ ion = 313) Inten.(x1,000,000)

647

J450. 250 500 750 1000

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. OH

(1H NMR in MeOD)

AM m m

LC/MS (ESI've) spectrum of compound 17, (M ion = 313) hten.(x1.000.000)

3Q8 397 , 480 538 812 ?64 101168

192

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Ph

0 20 (13C NMR in CDCk)

DM UM

193

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (ESLve) spectrum of compound 20, (M+ ion = 279) hten.fxl ,000,000V 2.0;' ~ : 278

2 JO 0.5

243 294 ?31 363 , . 4?1 ^07 542 5B3 629 674 ; . 796 956984 200 300 400 500 600 700 800 900 1000 nVz

(1H NMR in CDCI3)

IU u

m m

LC/MS (ESI've) spectrum of compound 22, (M+ ion = 279) hten.(x100,000)______278 5.0

579 874 55 242 581 8^8 /86 1?0 214|27l2 390 492 [ 622 7Q8 756 1.1 1076 11,56 133Q 14,54 250 500 750 1000 1250 nVz

194

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (ESI've) spectrum of compound 22, (M+ ion + Na metal = 279 + 23) Inteitfxl ,000,000)

o .g U ^ 5345?3627B67 600 700 800 900 m/z

195

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. Appendix C

LC/MS spectral data of PEGs, amphiphilic acylpyrazolones, and amphiphilic acylisoxazolones

196

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (PDA chromatogram) of polydiesperse PEG-750 x 10,000,000)

2.25 TIC

2.00

1.75

1.50

1.25

1.00

0.75

0.50

0.25

0.0 5.0 10.0 15.0 20.0 25.0 30.0 35.040.0 45.0 50.0

LC/MS (ESI+ve) spectrum of polydiesperse PEG-750 Inten.(x1,000.000) 2.00 583 1.75 627

1.50 539 1.25 495 759 1.00

0.75 451 803 0.50 407 62 2 191 ] 9?79?9 ■2 1 2 . 108611.42 1ffi2 1276 250 500 750 1000 1250

LC/MS (PDA chromatogram) of monodiesperse PEG-516 (1 lmer)

197

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (ESI+ve) spectrum of monodiesperse PEG-516 Inten. (x 10.000.000)

1finS 17tin 250 500 750 1000 1250 1500 1750 mtz

LC/MS (PDA chromatogram) of monodiesperse PEG-913 (20mer)

x 10,000,000) WCi

0.7

0.6

0.5

0.2

0.0 5.0 10.0 15.0 20.0 25.0 30.0 35.0 40.0 45.0 50.0 55.0

LC/MS (ESI+ve)j+ve spectrum of monodiesperse PEG-913 Inten.(x1,000,000) 2.25

2.00

1.75

1.50

1.25

1.00

0.75

0.50

0.25 61 0.00- / 129 __ 203_ 323 413 ?7 543___622 671 _22°___878 I s 996 1157 1717 1348

198

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (ESI+ve) spectrum of HMPVP-913 (M+ ion + mwt of Na = 1169 + 23) Inten.(x100,000)

0.5 0

0. 25 1139 10,32 10.85 1010 126 2 13,51 14,02 ooJ 1193 1240 1296 1394 1417

LC/MS (ESLve) spectrum of HBPIS-516 (M+ ion = 793) hten.(x1,000.000)

2 . 339

2 .

1 .

0. 490 7|2 1489 0.olfisaL 163 220; 294 if 4154?9 609 701 748 950 . 1QS6 1203 1788 1391 250 500 750 1000 1250 m/z

LC/MS (ESrve) spectrum of HBPIS-913 (M+ ion = 1189)

hten.(x100,000) 7.5 339

353 5.0

2.5 490 459 1188 1487 76 193 213 2g4 33' 387. 5?7 609 6f3 746 796 866 937 1013 1248 ,1385 0 0 250 500 750 1000 1250 m/z

199

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission. LC/MS (ESLve) spectrum of HMVIS-516 (M+ ion = 697) Inten(x100,000)

3. 696

2.0: 180

1 .(L ,361 498 7 91. 892 11059 11,96 0. ■ill Mil I. I, mT. — . - - i — ,< U— i ‘ 14^1160^1703 1897 250 500 750 1000 1250 1500 1750 m/z

LC/MS (ESLve) spectrum of HMVIS-913 (M+ ion = 1094) Inten/xl 0,000)

i 7. ' ______m o ' j _ :

5. 180

. 3' 2 1 6 ‘ 409484 H i 785 1004 0. -4 3 ? . r 1 1 1 — -f ...... - 1 —1 [■ \ I1 " i" I ■1 1 1,1 250 500 750 1000 1250 1500 1750 m/z

LC/MS (ESI've) spectrum of HP VIS-516 (M+ ion - 759) Inten.(x100,000) 7. 242 758 5.

2 .

0. 1805 1927 250 500 750 1000 1250 1500 1750 m/z

LC/MS (ESI've) spectrum of HP VIS-913 (M+ ion = 1156) Inten.(x10,000) 5.

2 .

0. 250 500 750 1000 1250 1500 1750 m/z

200

Reproduced with permission of the copyright owner. Further reproduction prohibited without permission.