Development of Synthetic Methodologies Towards Cyclic

DEVELOPMENT OF SYNTHETIC METHODOLOGIES TOWARDS CYCLIC

HYDROXAMIC ACID-BASED NATURAL PRODUCTS

RANJAN BANERJEE

A Dissertation Submitted to the Graduate Faculty of

WAKE FOREST UNIVERSITY GRADUATE SCHOOL OF ARTS AND SCIENCES

in Partial Fulfillment of the Requirements

for the Degree of

DOCTOR OF PHILOSOPHY

Chemistry

December 2010

Winston-Salem, North Carolina

Approved By:

S. Bruce King, Ph.D., Advisor

Pradeep K. Garg, Ph.D., Chairman

Rebecca W. Alexander, Ph.D.

Christa L. Colyer, Ph.D.

Paul B. Jones, Ph.D. ACKNOWLEDGEMENTS

I want to express my thanks and sincere gratitude to Dr. S. Bruce King, for all his

academic support and chemistry help, support, and advising throughout my Ph.D career. I

owe him a lot for helping me develop into an organic chemisty, guiding my projects and

his patience in helping me improve my scientific writing. I am really fortunate to have

shared his immense scientific knowledge and learn from his amazing mentoring ability. I

am also thankful to Dr. Paul Jones and Dr. Rebecca Alexander for being my committee

members.

I really want to thank Dr. Cynthia S. Day for crystallographic help and Dr. Marcus W.

Wright for NMR assistance. They have always been extremely helpful with any data

interpretation and experiment setup needed.

I thank my previous lab mates Dr. Sarah Knaggs, Dr. Weibin Chen, Dr. Mike Gorczyski,

Brad Poole and the present lab mates Mai Shoman, Raje Mukherjee, Dr. Richard Macri,

Dr. Mallinath Hadimani, Dr. Susan Mitroka, Craig Clodfelter, Julie Reisz and Jenna

DuMond for their assistance and friendship.

I appreciate the friendship and support I received from my Wake Forest friends Tanya

Pinder, Dr. Uli Bierbach, Lu Rao, Rajsekhar Guddneppanavar, Jayati Roychoudhuri,

Zhidong Ma, Samrat Dutta, Lindsey Davis, John Solano, R.P. Oates, Zhouli Zho and

Sandhya Bharti. I am really indebted to Subhasis De for his enormous help to start my

American life and really appreciate his friendship. I am also very thankful to my

roommates Saurav Sarma, Sebastial Berisha, Edison Munoz-Recuay, Angelo Malvestio,

Matt Koval, Joe Maye, Dhruv Gandhi, Anand Gondalerkar and Ben Rosenberg. My very special thanks and love to my beautiful girlfriend Erika Bechtold for bringing so much joy and happiness in life. Erika has always been an amazing support, my very best friend and the sweetest thing I have ever known. She has changed my life and I enjoy it every moment. I am also thankful to Erika’s roommates Charlie and Jeremy for their friendship and support.

My sincere love and gratitude goes to my father and mother for being the best parent in the world. Word cannot express my thankfulness for their endless love, countless sacrifices and tremendous encouragement throughout my life. I will be indebted to them forever and I thank God for blessing my life with their kind souls. I wish to dedicate this work to my parents and Erika.

TABLE OF CONTENTS

LIST OF FIGURES……………………………………………………………………

LIST OF TABLES……………………………………………………………………..

ABSTRACT……………………………………………………………………………

CHAPTER 1: INTRODUCTION………………………………………………………...1

1.1 Hydroxamic Acids………………………………………………………..1 1.2 Basic Structure…………………………………………………………....2 1.3 Biological Importance of Hydroxamic Acids…………………………….2 1.3.1 Enzyme Inhibitors…………………………………………………2 1.3.1.1 Matrix Metalloproteinase Inhibition……………………..2 1.3.1.2 Histone Deacetylase Inhibition…………………………..4 1.3.1.3 TNF-Alpha Converting Enzyme Inhibition……………...5 1.3.2 Therapeutic Uses of Hydroxamic Acids…………………………..6 1.3.2.1 Antimalarial Activity…………………………………….6 1.3.2.2 Antimelanogenic Agents………………………………....7 1.3.2.3 NO Donors……………………………………………….7

1.4 Biological Importance of Cyclic Hydroxamic Acids……………………..8

1.4.1. Iron Uptake by Siderophores……………………………………...8

1.4.2 Lipoxygenase Inhibitory Activity………………………………….9

1.4.3 Therapeutic Application of Cycllic Hydroxamic Acids………….10

1.4.3.1 Prostate Cancer Therapeutics……………………………10

1.4.3.2 Growth Inhibitors Tumor Cell Lines…………………....11

1.4.3.3 Analgesic Activity………………………………………12

1.5 Synthetic Approaches to Hydroxamic Acids…………………………….12

1.5.1 Simple Hydroxamic Acids………………………………………..12

1.5.2 Angeli-Rimini Reaction…………………………………………..13

1.5.3 Solid Phase Modification…………………………………………13

1.5.4 Oxidation of N-Boc protected Amides…………………………...14

1.5.5 N-Substituted Hydroxamic Acids………………………………...15

1.5.6 Solid Phase Synthetic Approach………………………………….16

1.5.7 Nitroso-ene Reactions…………………………………………….17

1.5.7.1 Intermolecular Nitroso-ene Reactions…………………..18

1.6 Synthesis of Cyclic Hydroxamic Acids…………………………………..20

1.6.1 Reductive Cyclization of Aliphatic Nitro Acids…………………..20

1.6.2 Synthesis of the Cobactin Core……………………………………20

1.6.3 Heterocycle Based Synthesis: Tungstate Catalyzed Oxidation of Tetrahydro Quinoline……………………………………………..21

1.6.4 Oxidation of Lactams to Cyclic Hydroxamic Acids……………...22

1.6.5 Phenyliodine(III) Bis Trifluoroacetate Mediated Ring-closure…...22

1.6.6 Photochemical Synthesis of Cyclic Hydroxamic Acids…………..23

1.6.7 Intramolecular Nitroso-ene Reaction……………………………...24

1.6.8 Ring Expansion of Cyclic Ketones ……………………………… 26

1.6.9 Piloty’s Acid-Based Rearrangements of Cyclic Ketones to Make Cobactin…………………………………………………………...27

1.7 Piloty’s Acid: A Nitroxyl Donor…………………………………………27

1.7.1 Nitroxyl (HNO) Chemistry……………………………………….28

1.8 Mycobactin – S…………………………………………………………..30

1.8.1 Biosynthesis of Mycobactins……………………………………………31

CHAPTER 2: SYNTHESIS OF CYCLIC HYDROXAMIC ACID THROUGH –NOH INSERTION OF KETONES……………………………………………34

2.1 Introduction………………………………………………………………35

2.2 Synthesis of N-Hydroxy Benzenesulfohydroxamic Acid (Piloty’s acid)...36

2.3 -NOH Insertion Reaction of Cyclic Ketones…………………………….36

2.3.1 Synthesis of N-hydroxy Piperidone………………………………36

2.3.2 -NOH Insertion Reaction in Cyclobutanones…………………….38

2.3.3 Solid phase modification………………………………………….42 2.4 Mechanism of –NOH insertion reaction………………………………….43

2.5 Scope of –NOH Insertion Reaction………………………………………49

2.5.1 Synthesis of O-Protected Cyclic Hydroxamic Acids…………….49

2.5.2 –NOH Insertion Reaction in α-Substituted Cyclopentanone…….50

2.5.3 -NOH Insertion in Cyclohexenone……………………………….51

2.5.4 Investigation of a Cobactin Synthesis Using the –NOH Insertion Reaction………………………………………………………...52

2.6 Experimental……………………………………………………………...54

CHAPTER 3: PROGRESS TOWARDS THE SYNTHESIS OF THE COBACTIN CORE AND MYCOBACTIC ACID UTILIZING NITROSO-ENE REACTION………………………………………………………….71

3.1 Intramolecular Nitroso-ene Reaction Approach to the Synthesis of Cobactin Core………………………………………………………….71

3.1.1 Retrosynthesis of Cobactin ……………………………………..71

3.1.2 Intramolecular Nitroso-ene Reactons: Results and Discussion…72

3.2 Structure Elucidation of Mycobactin …………………………………..76

iii

3.3 Investigation of Intermolecular Nitroso-ene Approach towards the Synthesis of Cobactin and Mycobactic Acid…………………………...76

3.3.1 Synthesis of the Precursor Alkene and Diels-Alder Cycloadduct (Acyl-nitroso Precursor) for a Nitroso-ene Reaction………………….78

3.3.2 Nitroso-ene Reaction…………………………………………...78

3.3.3 Progress Towards the Synthesis of Mycobactic Acid Utilizing a Nitroso-ene Reaction…………………………………………..83

3.4 Nitroso-ene Reactions of Acetyl-Nitroso Compounds…………………86

3.5 Nitroso-ene Reactions of Benzoyl Nitroso compounds………………...87

3.6 Experimental……………………………………………………………90

CHAPTER 4: SUMMARY …………………………………………………………...114

REFERENCES……………………………………………………………………….117

APPENDIX…………………………………………………………………………..136

SCHOLASTIC VITA………………………………………………………………...181

LIST OF FIGURES

Figure 1. X-ray Diffraction structure of N-hydroxy piperidone (85) …………. 38

Figure 2. X-ray Diffraction Structure of 89 …………………………………… 39

Figure 3. Figure 3. X-ray Diffraction Structure of 94…………………………. 42

Figure 4. X-ray Diffraction Structure of Compound 127 …………………….. 75

LIST OF TABLES

Table 1. Results of –NOH insertion with cyclopentanone towards N-hydroxy piperidone synthesis…………………………………………………. 37

Table 2. Results of –NOH Insertion into cyclobutanone ………………………40

Table 3. Gas Chromatography Results: Hydrolysis of acyloxy nitroso ………. 47

LIST OF ABBREVIATION

Boc tert-Butyloxy carbonyl

Cbz Carboxybenzyl

DCC N, N’-Dicyclohexylcabodiimide

DIBOA 2,4-dihydroxy-2H-1,4-benzooxazin-3(4H)-one

DMA Dimehtylanthracene

DMAP 4-Dimethylaminopyridine

DMD Dimethyldioxirane

DMF Dimehtylformamide

DMSO Dimehtylsulfoxide

DNA Deoxyribonucleic Acid

E Entgegen

EDC 1-Ethyl-3-(3-dimethylaminopropoyl)carbodiimide

FR900098 Fosmidomycin

GC Gas chromatography

HCl Hydrochloric acid

HDAC Histone deacetylase

HETE Hydroxyeicosatetraenoic acid

HNO Nitroxyl

HOAt 1-Hydroxy-7-azabenzotriazole

IC50 Half maximal inhibitory activity

MMP Matrix metalloproteinase

MS Mass Spectrometry

NMR Nuclear magnetic resonance

NO Nitric Oxide

vii

PIFA Phenylidoine (III) bis(trifluoroacetate)

Ppm Parts per million

SAHA Suberoylanilide hydroxamic acid

TACE Tumor necrosis factor alpha converting enzyme

TBDMSCl tert-Butyl dimethylsilyl chloride

TBDPSCl tert-Butyl chlorodiphenylsilane

TFA Trifluoroacteic acid

THF Tetrahydrofuran

TIMP Tissue inhibitor of metalloproteinases

TLC Thin layer chromatogrpahy

TMD Trifluoromethyl dioxiranes

TNF Tumor Necrosis Factor

TSA Tricostatin A

Z Zussamen

viii

Abstract

Hydroxamic acids are an important class of bioactive compounds with wide uses as anti-bacterial, or anti-inflammatory agents and a key component of many natural products, mainly siderophores (low-molecular-weight iron sequestering agents) in lower organisms. Hydroxamic acid based analogs may find potential therapeutic uses in the inhibition of siderophore biosynthesis. Our research targets to develop new synthetic

methodology towards making cyclic hydroxamic acids in a stereoselective and

regioselective fashion. Basic decomposition of Piloty’s acid transforms cyclic ketones

(mainly four and five membered) into ring-expanded cyclic hydroxamic acids in 20-69%

yield (Scheme 1). Mechanistic study reveals this reaction involves a C-nitroso intermediate 98 (Scheme 47) in the course of the rearrangement, which can also be generated by a separate hydrolysis reaction of acyloxy nitroso intermediate 103 (Scheme

51) leading to the ring-expansion product in 75-80% yield.

Nitroso-ene reactions regioselectively and stereoselectively functionalize olefins in the allylic position. We use various acyl nitroso species (general formula RCONO) as enophiles in an ene reaction with olefins to produce hydroxamic acids substituted at the nitrogen with an allylic group. This sequence allows the synthesis of many N-containing products with diverse structures. Amino acid-based hydroxamic acids are important components in mycobactin, hydroxamic acid-based siderophores produced by

Mycobacterium species. Mycobactin S inhibits growth against Mycobacterium tuberculosis and it consists of cobactin and mycobactic acid (Scheme 38), which contain

a hydroxamic acid residue derived from NЄ-hydroxylysine. The intermolecular nitroso-

ene reaction between a Nα-acyl-homoallyl-glycine ester (149) and a t-butyl nitroso

formate (136), followed by reduction and deprotection, yields Nα-acyl-NЄ-hydroxylysine

(152, Scheme 68). This sequence provides a key intermediate in route to cobactin, the seven-membered cyclic hydroxamic acid found in various mycobactins. A similar synthetic route using a long chain-derived acyl nitroso species supports a synthetic route to mycobactic acid. This research also involves exploring the scope of the nitroso-ene reaction of various acyl nitroso species with terminal alkene (149) to generate a series of

N-substituted hydroxamic acids.

CHAPTER 1

INTRODUCTION

1.1 Hydroxamic acids ( RCONR'OH )

Hydroxamic acids have been known since 1869 with the discovery of oxalohydroxamic acid by Lossen.1 Despite this early discovery, not much biological information about these important molecules was known for a long time. A tremendous amount of research attention has been given over the last couple of decades towards the synthesis and biomedical applications of hydroxamic acid containing organic molecules.2-5 Hydroxamic acids are powerful metal ion chelators,6 which possess a wide spectrum of biological activities, such as anti-bacterial, anti-fungal, anti- inflammatory, and anti-asthmatic properties,7-9 and are potent inhibitors of matrix metalloproteinases, a family of zinc-dependent enzymes associated with diseases like cancer, arthritis, nephritis and multiple sclerosis.10-13 Hydroxamic acids can find therapeutic potential as ribonucleoside reductase inhibitors blocking DNA biosynthesis and also act as histone deacetylase inhibitors, both common targets for cancer treatment.14 Hydroxamic acids play key roles in microbial iron consumption as siderophores, low molecular weight iron-sequestering agents produced by most microorganisms, especially under iron deficient conditions.15 Finally, hydroxamic acids have been shown to inhibit TNF (tumor necrosis factor)-alpha converting enzyme

(TACE), 16 to demonstrate anti-malarial activity and to act as donors of nitric oxide

(NO), an important biological signaling molecule.17

1.2 Basic Structure

Hydroxamic acids are N-hydroxy amides, derivatives of hydroxylamine and carboxylic acids. Cyclic hydroxamic acids (1) are generally N-hydroxy lactams. Two possible hydroxamic acid tautomers exist (Scheme 1), the keto isomer is predominant under acidic or neutral conditions, and the enol form is stable in alkaline conditions.

NMR studies have shown the presence of (E) and (Z) isomers extending the structural diversity. The structure of the hydroxamic acids allows these compounds to chelate various metal ions strongly.

Scheme 1. Structure of hydroxamic acids

1.3 Biological Importance of Hydroxamic Acids

1.3.1 Enzyme Inhibitors

1.3.1.1 Matrix Metalloproteinase Inhibition

Matrix metalloproteinases (MMPs) are a family of structurally related zinc containing enzymes that mediate the degradation of the extracellular matrix and tissue remodeling and are therefore targets for therapeutic inhibitors in inflammatory, malignant and degenerative diseases.18, 19 Under normal physiological conditions, the proteolytic activity of MMPs are controlled at three stages: transcription, activation of

2 zymogens (inactive enzyme precursors) and inhibition of the active forms by various tissue inhibitors (TIMPs). Under pathological conditions this equilibrium is shifted towards producing active MMPs leading to breakdown of connective tissues. MMPs are involved in morphogenesis, tissue remodeling (such as wound heaaling), apoptosis, cancer invasion, and arthritis, degradation of the blood - brain barrier, multiple schlerosis, dermatitis, congestive heart failure and Alzheimer’s disease. Originally,

MMPs were thought to be responsible for invasion and metastasis by matrix remodeling thereby allowing tumor cells to access blood and lymphatic vessels. The mechanism of action (Scheme 2) as proposed by Lovejoy et al, is mediated by coordination of the scissile amide carbonyl from the peptide to the active site zinc(III)

Scheme 2. Mechanism of proteolysis by MMPs 19

ion and successive attack of a water molecule on the carbonyl. Hydroxamic acids play a significant role in inhibition of MMPs by specifically binding the Zn atom in the enzyme active site as shown (3, Scheme 3).20 A representative hydroxamic acid based non-peptidyl MMP inhibitor is also shown (4, Scheme 3).

Scheme 3. Model inhibitors of MMPs

1.3.1.2 Histone Deacetylase Inhibition

Histone deacetylase and histone acetyl transferase are involved in chromatin structure modification and functional regulation of gene transcription.21 Reversible acetylation of the side chain amino groups of specific histone lysine residue plays important roles in chromatin remodeling. Evidence of aberrant histone deacetylase activity in leukemia has established the focus on developing both natural and synthetic histone decetylase inhibitors as potential therapeutic agents. Recent studies have shown that inhibition of histone deactylase has pronounced anti-cancer effects in several tumor cell lines by inhibiting cell growth and inducing apoptosis.22 One of the major families of histone deacetylases is the zinc-containing amido hydrolases, suggesting that a structural requirement for potent inhibitors is a specific Zn2+ binding functional group.

Indeed, one of the most effective naturally occurring histone deacetylase inhibitor is tricostatin A (TSA) (5, Scheme 4) that mimics the substrate and chelates zinc in the catalytic pocket as the main mechanism of inhibition.23 A major breakthrough in synthetic histone deacetylase inhibitors was the discovery of suberoylanilide hydroxamic acid (SAHA, 6, Scheme 4) which induces apoptosis in a variety of tumor cells and is currently in phase I clinical trials.24 Oxamflatin (7, Scheme 4), an antitumor

4 compound containing a hydroxamic acid, was found to be an inhibitor of mammalian histone deacetylase.25

Scheme 4. Natural and synthetic HDAC inhibitors

1.3.1.3 TNF-Alpha Converting Enzyme Inhibition (TACE)

Tumor necrosis factor-α (TNF-α) is a therapeutic target through inhibition of its formation for treatment of rheumatoid arthritis, meningitis and several other inflammatory diseases.26 Development of anti TNF-α agents has received tremendous attention for small molecule drug discovery.26 One approach is to establish small molecule inhibitors of the TNF-α converting enzyme (TACE) responsible for TNF processing. TACE is a member of the metalloproteinase family and its active site structure is very similar to MMPs. Hydroxamic acid based TACE inhibitors have been proven to be a class of potent anti-inflammatory agents and may lead to effective medicines in the future. Some of the representative examples are shown below.

Retrohydroxamates (N-substituted hydroxamic acids) are successfully incorporated in designing TACE inhibitors (8, Scheme 5) In early 1998, Pfizer reported compound 9

(Scheme 6) showing nanomolar activity for the inhibition of TNF-α release in a human blood assay.27 Macrocyclic hydroxamic acids (10, Scheme 5) with various aryl 5 substituents showed high selectivity for TACE inhibition over MMPs.28, 29 Anthranilic acid derivatives (11, Scheme 5) bearing an additional amine moiety also show promising results in a TACE inhibition study.28

Scheme 5. Different classes of TACE inhibitor templates

1.3.2. Therapeutic Uses of Hydroxamic Acids

1.3.2.1 Antimalarial activity

Hydroxamic acids also exert anti-malarial activity. The first hydroxamic acid based

Scheme 6. Hydroxamic Acid Based Antimalarial Agents

6 compound (12, Scheme 6) with profound activity was well studied by Hynes.30

Hydroxamic acids retard parasite growth by selective inhibition of DNA synthesis or protease activities. Phosphonohydroxamic acids, such as fosmidomycins (13, Scheme

6), are drug candidates for treatment of malaria, and are currently in phase II clinical trial.31, 32

1.3.2.2 Anti-melanogenic agents

The production of melanin in our body is mainly regulated by the enzyme tyrosinase

Tyrosinase is involved in catalyzing the oxidation of phenols and the production of melanin.33 Inhibitors of tyrosinase could be useful in the treatment of melanin hyperpigmentation. Metal ion chelators like kojic acid,34 flavonol,35 and N- nitrosohydoxylamines36 are all known tyrosinase inhibitors with their inhibitory activity coming from their binding Cu in the active site of tyrosinase. Kim et al exploited the metal chelation ability of hydroxamic acids and evaluated the potency of a group of anti-melanogenic agents (14 and 15) in a murine melanocyte cell line (Scheme 7).37

Scheme 7. Hydroxamic acids as anti-melanogenic agents

1.3.2.3 NO Donors

While most physiological roles of hydroxamic acids are due to their metal chelation ability, they were also found to be nitric oxide (NO) donors in recent studies.14

Hydroxamic acids readily transfers NO in presence of Ru(III) forming a stable

7 ruthenium-nitrosyl (II) complex (18, Scheme 8). These complexes result in vasorelaxation of rat aorta by NO mediated activation of guanylate cyclase.14

[Ru(HEDTA)Cl]- + RCONHOH [Ru(EDTA)(NO)Cl]2- + RCOOH

16 17 18 19 Scheme 8. Hydroxamic acids as NO Donor

1.4 Biological Importance of Cyclic Hydroxamic Acids

1.4.1 Iron Uptake by Siderophores

Iron plays crucial roles in almost every form of life on the earth. Although iron is one of the most abundant elements in Nature, its importance in physiological processes depends upon its assimilation. The most stable ionic form of iron is Fe (III), which is insoluble under physiological conditions. To circumvent this solubility problem many microbes, plants and even higher organisms synthesize low molecular weight, very specific iron-chelators called siderophores.38 Cyclic hydroxamic acids are the most common residues of the siderophores that scavenge iron in an iron-deficient environment. The competition for iron between host and bacteria is an important factor for determining bacterial infection. Structurally modified siderophores, may serve as antagonists of microbial growth by competitive iron-binding or inhibiting iron assimilation and show potential therapeutic value. This property was exploited to bring revolutionary changes in anti-bacterial and anti-fungal drug discovery.39 Naturally occurring siderophores such as mycobactine and pseudobactines (20, Scheme 9) are finding medicinal application as antibiotics.40 Biological studies have shown that mycobactin-S (21, Scheme 9) inhibits 99% growth of M.tuberculosis H37Rv at a

8 concentration of 12.5 µg/mL.41 Apart from their use as broad-spectrum antibiotics, siderophores could also be used for treatment of iron overload in the human body.

Desferrioxamine (22, Scheme 9) is still the drug of choice to remove excess iron

(binding specifically Fe (III)) for blood transfusions in thalassemic patients.42

NH O NH 2 H N N O H NH O O O N N O + Fe NH3 O O HO O HN H O O 5 O O HN O NH HO N O N H O O R'' H H HN HN HN 2 O N OH O 20 H NHR' HO 22 Desferrioxamines Fe(III) bound Pseudobactine

Scheme 9. Representative examples of siderophores

1.4.2 Lipoxygenase Inhibitory Activity

Lipoxygenases are enzymes involved in the oxidative metabolic pathway of unsaturated fatty acids. 5-, 8- and 12-Lipoxygenases are well characterized enzymes in the lipoxygenase enzyme family and a specific lipoxygenase oxidizes a specific position of the fatty acid to a produce a specific metabolite. Hydroxyeicosatetraenoic or

12-HETE (12(S)-5(Z),8(Z),10(E),14(Z)-), a metabolite produced by 12-lipoxygenase acid, exhibits a variety of biological activities. Cancer cells induce the release of this

9 arachidonate metabolite 12-HETE in high amounts. Cyclic hydroxamic acids are reported to be active inhibitors of 12-lipoxygenase and are useful for the treatment and prevention of cancer metastasis, inflammation, immune diseases and ischemic cardiovascular diseases.43 Some representative structures are shown below (23 and 24,

Scheme 10).

Scheme 10. Lipoxygenase inhibitors

1.4.3 Therapeutic Application of Cyclic Hydroxamic acids

1.4.3.1 Prostate Cancer Therapeutics

The development of new pharmacological agents to combat prostate cancer is a great challenge in biomedical research. The water soluble pollen extract Cernitin T-60, which contains the cyclic hydroxamic acid based structure DIBOA (2,4-dihydroxy-2H-

1,4-benzooxazin-3(4H)-one), shows striking growth inhibitory activity to a prostate cancer cell line.44 The chemical structure of DIBOA (25) and other analogous synthetic structures (26 and 27, Scheme 11) with potential inhibitory activity are shown in

Scheme 8. Natural and synthetic cyclic hydroxamic acid based structures are potential drug candidates for the clinical symptoms of prostatitis, prostatodynia and prostate cancer.

Scheme 11. Prostate Cancer Drug Candidates

1.4.3.2 Growth Inhibitors of Tumor Cell Lines

Amamistatin A and B (28 and 29, Scheme 12), cyclic hydroxamic acid containing natural products isolated from a strain of Nocardia, show antiproliferative activity

45 against three human tumor cell lines (IC50 0.24-0.56 µM). These linear lipopeptides contain a seven membered cyclic and a straight chain hydroxamic acid, similar in structure to formobactin and mycobactin. 46 47

Scheme 12. Tumor growth inhibitor Amamistatin A and B

1.4.3.3 Analgesic Activity

The elevated level of copper in blood relates to inflammation and arthritic diseases.48 Cyclic hydroxamic acids, such as 30 and 31 (Scheme 13) play an important role in chelating the copper (II) ion thereby alleviate inflammation and acts as analgesic agents. 49

Scheme 13. Cyclic hydroxamic acids for analgesic activity

1.5. Synthetic Approaches to Hydroxamic Acids

1.5.1 Simple Hydroxamic Acids

A common practice in the chemical synthesis of unsubstituted linear hydroxamic acids is to react an activated carboxylic acid with an O-protected hydroxylamine to produce a conveniently protected form of a hydroxamic acid (32, Scheme 14).

Scheme 14. General synthetic route to hydroxamic acids

1.5.2 Angeli-Rimini Reaction

In 1896, Angeli and Rimini discovered that benzenesulfohydroxamic acid (Piloty’s acid, named after O. Piloty) decomposes in basic condition in the presence of an aldehyde to yield the corresponding hydroxamic acid.50 This reaction is primarily known in the literature for detection of aldehydes but has never enjoyed preparative synthetic success because of low yields and purification problems. The mechanism of the reaction is also not well understood, but a proposed mechanism (Scheme 15) for this unique reaction describes nucleophilic attack of the N-anion of Piloty’s acid to an aldehyde followed by rearrangement to yield the desired product.51

O- O OH- PhSO2NHOH R CH NSO2Ph R H OH 33 34

R NHOH R CH N O O OH 35

Scheme 15. Angeli-Rimini reaction

1.5.3 Solid Phase Modification

While the Angeli-Rimini reaction has not found synthetic attention as a standard procedure to make hydroxamic acids, Andrea et al reported a solid state modification of this reaction to afford fair yields of hydroxamic acids.52 This methodology reported a solid-supported Piloty’s acid reagent (36, Scheme 16) that makes purification easier.

This new reagent can be prepared by shaking a pyridine solution of hydroxylamine hydrochloride with polystyrene sulfonyl chloride in dichloromethane at room

Scheme 16. Solid phase modification

temperature. Once the reaction between the poly-sulfohydroxamic acid polymer and aldehyde is done, the polymer can be filtered off and acidic work up yields the hydroxamic acid. This method is the only successful use of the Angeli-Rimini reaction to produce hydroxamic acids in organic synthesis.

1.5.4 Oxidation of N-Boc Protected Amides

Dimethyldioxirane (DMD) or methyl(trifluoromethyl)dioxiranes (TFD) are well known effective reagents for C-H or hetereoatom oxidation under mild conditions.53, 54

N-Boc protected peptides are reported to be oxidized by DMD or TFD (6 eq.) at 0ºC in

O H O O H O DMD or TFD in DCM N N O N O O N O H O 78% OH O 37

Scheme 17. Oxidation of amides to make hydroxamic acids

14 chlorinated solvents to generate N-hydroxy amides or hydroxamic acids (37, Scheme

17). These reactions are representative examples of amide oxidation to prepare hydroxamic acids.55

1.5.5 N-Substituted Hydroxamic Acids

Preparation of N-substituted hydroxamic acids follows a limited number of literature procedures.56, 57 One simple method is to prepare the O-protected analog (38,

Scheme 18) of the correctly N-substituted hydroxylamine followed by a substitution reaction with an acid halide (39, Scheme 18).

O O Base BnO X X N R' N H R' X OBn 38 39 40

Scheme 18. Preparation of N-substituted hydroxamic acid

Various synthetic pathways exist to make functionalized hydroxylamines. Using N- protected hydroxylamines as starting materials generally improves the yield. Scheme 19 shows different routes to functionalized hydroxylamines 38, which are difficult to prepare through the reaction of hydroxylamine and organic halides, especially if additional functional groups are present. Hydroxylamine 38 can be prepared from substituted aldehyde [42, X = CH2CO2Et ] by an imination reaction with NH2OBn and

58 further reduction of the intermediate 43 by NaBH3CN (Route A). Functionalized alkyl bromides [44, X= CHRP(O)(OEt)2] can be transformed into hydroxylamine 38 by

59 substitution reactions with a protected hydroxylamine (NH2OBn, Route B). Oxidation

15 of functionalized amines [45, X = R(NHR’)CO2Me) with dimethyldioxirane (Route

60 C) or imines [46, (X=(CH2)3COOH)] with peracid followed by a reaction with TFA

(Route D), also gives unprotected N-substituted hydroxylamines.61, 62

Scheme 19. Existing approaches to N-functionalized hydroxylamines 63

1.5.6 Solid Phase Synthetic Approach

Polymeric solid phase reagents are useful to synthesize substituted hydroxamic acids because of the easy separation of low-molecular weight products by filtration or selective precipitation. Usually O-linked polymer bound hydroxylamine (48, Scheme

20) is treated with ketones to form the corresponding oxime (49), which is subsequently reduced and acylated to give (51). Reductive cleavage of 51 would give the desired the desired N-substituted hydroxamic acid (52, Scheme 20). 64

Scheme 20. Solid State Synthesis of N-substituted Hydroxamic Acids

1.5.7 Nitroso-Ene Reactions

Over the last couple of decades, nitroso-ene reactions have been extensively used to synthesize various hydroxamic acids.65-69 This reaction typically involves a transient acyl nitroso species and an allylic component. Kirby et al reported a variety of C- nitroso carbonyl enophiles reacting in both intermolecular and intramolecular fashion giving a variety of hydroxamic acids.66, 67 Recent progress in the nitroso ene reactions has also been exploited to make hydroxamic acids such as FR900098 (55, Scheme 21), a N-substituted hydroxamic acid-containing antimalarial drug.34 Fokin and coworkers suggested this reaction typically involves a six membered transition state between an acyl nitroso component and the allylic part of an active alkene (Scheme 21).

Scheme 21. Nitroso-ene Route to Antimalerial Agent FR900098

Both intra and intermolecular ene reactions offer a useful way to make hydroxamic acids. Acyl nitroso intermediates are very reactive in nature and are generated in situ by the oxidation of corresponding hydroxamic acid by oxidants like NaIO4, Bu4NIO4,

PhI(OAc)2, PhIO and stored as the Diels-Alder adducts of 9,10-dimethylanthracene or cyclopentadiene. 70 Heat exposure to these Diels-Alder adducts usually liberates the acyl nitroso component through a retro Diels-Alder reaction.

1.5.7.1 Intermolecular Nitroso-ene Reaction

The bimolecular ene reaction of an acyl nitroso species and a separate allylic component constitutes an efficient route to straight chain N-hydroxy amides by allylic

N-hydroxy amidation. The Kirby and Keck groups have done comprehensive research on these reactions.67, 71 C-Nitroso carbonyl compounds usually include acyl nitroso ketones,71 esters 67 or formamides 66 as enophiles (Scheme 22). The general procedure

Scheme 22. Intermolecular nitroso-ene route to N-subsituted hydroxamic acid of the bimolecular nitroso ene typically involves dissolving the acyl-nitroso precursor in the olefin as solvent and heating at reflux under an inert atmosphere until TLC monitoring indicates consumption of the precursor.71

The regiochemistry of intermolecular ene reaction is of special interest, since literature results demonstrate the possibility of multiple ene products from one substrate. Theoretical calculations reveal the nitroso-ene reaction could proceed through stepwise biradical intermediates instead of a typical concerted path.72, 73 Fokin and coworkers showed that allylic hydrogen abstraction in the case of mono substituted olefins renders a mixture of 1:1 (E : Z) hydroxamic acid products after the nitroso-ene between compound 53 and 54 (Scheme 23).63 However, with di, tri or tetrasubstituted olefins where several allylic hydrogens from different substituents are available, regiochemical issues become apparent. Tsutsumi and coworkers show that trisusbtituted olefins like 2-methyl-2 butene (55) yields the twix hydrogen (Scheme 23) abstraction product as the major regioisomer. 74

Scheme 23. Regiochemistry of intermolecular nitroso-ene reaction

1.6 Synthesis of Cyclic Hydroxamic Acids

1.6.1 Reductive Cyclization of Aliphatic Nitro Acids

The reductive cyclization of linear alkyl nitro compounds with a terminal acid group

(56, Scheme 24) is another method to synthesize five membered cyclic hydroxamic acids.75 This method is limited to the synthesis of five membered α-substituted cyclic hydroxamic acids from aliphatic nitro compounds.

O Zn,Ac O,AcOH R1 NO 2 2 N OH 2 60 -75 C R CO2H 46% R1 R2 56 57

Scheme 24. Nitro-reduction and cyclization route to a cyclic hydroxamic acid

1.6.2 Synthesis of the Cobactin Core

The Miller group reported the first total synthesis of mycobactin-S (21, Scheme 9), a biologically important bacterial siderophore that showed significant inhibitory activity in a human tuberculosis cell line.76 Mycobactin-S has a seven membered ring cyclic hydroxamic acid, the cobactin core, as a part of its structure. Miller’s reported method includes a six step synthesis from the Nα-Cbz-L-lysine (58, Scheme 25) as starting material. N-Cbz-lysine was converted to the tert-butyl ester (59) and then DMD oxidation provided nitrone (60, Scheme 25). Treatment of the nitrone with hydroxylamine hydrochloride followed by basic work-up, extraction and TFA hydrolysis gave hydroxylamine (62, Scheme 25). DCC-coupled ring closure in the presence of DMAP·HCl yielded 3-amino N-hydroxy azepanone (63, Scheme 25) in very low yield. O-Protection of the desired hydroxamic acid 63 with TBDMSCl or

Scheme 25. Synthesis of cobactin core76

TBDPSCl (for easy chromatographic purification) rendered the targeted cobactin hydroxamic acid residue 64 in moderate yield (50%, in last four steps). Miller’s

21 reported method for cobactin synthesis remains unclear as the key intermediate 62 was not characterized or isolated and the DCC cyclization to cobactin leads to poor yield.

1.6.3 Heterocycle Based Synthesis: Tungstate Catalyzed Oxidation of Tetrahydro quinoline

Catalytic oxidation of certain aromatic secondary amines (65, Scheme 26) can produce hydroxamic acids (66, Scheme 26).77 This method is limited to reduced quinoline systems.

Scheme 26. Catalytic oxidation of secondary amine yields hydroxamic acid

1.6.4 Oxidation of Lactams to Cyclic Hydroxamic Acids

Oxidation of cyclic amides produces cyclic hydroxamic acids.78 Sammes et al reported the synthesis of DIBOA (25) by the oxidation of the silylated lactam with a peroxo-molybdenum complex (Scheme 27) in the presence of dimethylformamide. This method produced the targeted hydroxamic acid in 33% yield and is mainly applicable to aromatic lactams.79

Scheme 27. Peroxo-molybdenum oxidation of lactams

1.6.5. Phenyliodine(III) Bis Trifluoroacetate Mediated Ring-closure

N-Acyl-N-alkyloxy nitrenium ions, which can be generated under mild conditions by treating N-methoxyamides with the hypervalent iodine compound, PIFA

[phenyliodine(III) bis trifluoroacetate], is another literature route to make cyclic hydroxamic acids. Work done by Wardrop et al demonstrates the PIFA mediated ring- closure method to build the bicylic five membered N-hydroxy lactam 67 (Scheme 28).80

Synthesis of a spirocyclic hydroxamic 68 (Scheme 28) was also reported by the same group of researchers using this method.80, 81 This method is limited to the use of disubstituted alkenes as substrates and trifluoroacetic acid as catalyst.

Scheme 28. PIFA mediated cyclization of nitrenium ion

1.6.6 Photochemical Synthesis of Cyclic Hydroxamic Acids

The photolysis of cyclic α-nitro ketones 69, which predominantly exist as the enol form in ethanol, generates cyclic hydroxamic acid 70 as one of the photo rearrangement products (11%). 82 According to this study with the steroidal α-nitro ketone (Scheme

29), the photoreaction proceeds through nitrogen insertion into the ring bearing the nitro group. Previous studies done by Reid et al suggested a nitro-nitrite rearrangement for the formation of the N-hydroxy imide.83, 84 Since this reaction suffers from a poor yield, it has not found larger synthetic attention.

Scheme 29. Photolysis of a α-nitroketone

1.6.7 Intramolecular Nitroso-ene Reaction

Intramolecular nitroso-ene reactions are a convenient way of making 5 and 6 membered cyclic hydroxamic acids (Scheme 30). These reactions find tremendous application in the total synthesis of Amaryllidaceae alkaloids and other cyclic hydroxamic acid- based natural products.85 Intramolecular nitroso-ene methodology is a two-step procedure to construct cyclic hydroxamic ring systems, the first step being the generation of the acyl-nitroso species and store it as a Diels-Alder cycloadduct, followed by a second step of thermal release of the acycl-nitroso species with the concomitant ene reaction.

Scheme 30. Intramolecular nitroso-ene reaction yields cyclic hydroxamic acids

However, synthetic application of this unique cyclization to build complex natural product requires some understanding of the regiochemical features of this reaction.

Keck et al reported the synthesis of five membered cyclic hydroxamic acid 71 (Scheme

30) and a bicyclic hydroxamic acid 72 being a key intermediate for (±) crinane synthesis.86 Two sets of allylic methylene groups (A and B) exist in compound 73, which gives two possible ene results (Scheme 30 ) corresponding to Type I and

Type II ene reactions according to the hypothesis of Oppolzer and Sneickus (Scheme

31).87

Scheme 31. Variants of the intramolecular nitroso-ene reaction

Type I generates the isolated fused spiro structure 74 (path A, Scheme 30) while the

Type II product or the fused bicycle 75 (path B, Scheme 30) has not been observed.

The nitroso-ene reaction of compound 76 generates the fused spirocyclic hydroxamic acid 77 in 85% yield and compound 78 in 15% yield (Scheme 30), suggesting the synthesis of seven membered ring hydroxamic acid by intramolecular nitroso-ene is achievable albeit in poor yield.

1.6.8 Ring Expansion of Cyclic Ketones

The Angeli-Rimini reaction generates hydroxamic acids from aldehydes and early

Italian literature reports cyclic ketones act in a similar fashion to make the corresponding ring-expanded cyclic hydroxamic acids.88, 89 An early patent indicates this method is useful for making 6 membered cyclic hydroxamic acids with or without substitution. The synthesis describes alkali metal hydroxide decomposition of Piloty’s acid in protic solvents to produce ring expanded hydroxamic acids (80, Scheme 32) after –NOH insertion in presence of saturated cyclic ketone .

Scheme 32. –NOH insertion into cyclic ketones under basic conditions

Interestingly the patent proposed a mechanism (Scheme 33) that features the anion of nitroxyl (HNO/NO-) attacking the ketone to yield a tetrahedral intermediate that rearranges to the product. However this mechanism differs from the proposed Angeli-

Rimini’s reaction as described before (Scheme 15). 90

Scheme 33. Proposed mechanism of ring-expansion

1.6.9 Piloty’s Acid-Based Rearrangements of Cyclic Ketones to Make Cobactin

Basic decomposition of Piloty’s acid in the presence of 2-bromo cyclohexanone yields the cobactin precursor 3-bromo-1-hydroxyazepanone in very low yield (1-2%)

(Scheme 34).91 These results show that while this pathway exists for seven membered ring formation, the yields are extremely low.

Scheme 34. –NOH insertion route to Cobactin

1.7 Piloty’s Acid: A nitroxyl donor

Benzenesulfohydroxamic acid or Piloty’s acid (33, Scheme 35) is known to decompose under basic conditions to produce nitroxyl (HNO) and a benzenesulfinic acid salt.92 It was first postulated by Angeli that the reaction might occur via elimination of HNO from the conjugate anion of Piloty’s acid in an analogy to

29 nitrohydroxamates or Angeli’s salt (Na2N2O3), another known source of HNO.

Scheme 35. Basic decomposition of Piloty’s acid : Generation of HNO

1.7.1 Nitroxyl (HNO) Chemistry

Nitroxyl (HNO), the one-electron reduced and protonated form of nitric oxide

(NO), (in HNO, N is in +1, formal oxidation state), has recently garnered increased attention based on its unique chemical and biological properties. Extensive theoretical and experimental studies indicate that singlet HNO, with a pKa = 11.4, exists as the dominant species of the HNO/NO- conjugate acid/base pair under physiological conditions (Scheme 36).93-95 Calculations predict NO- exists as a triplet species isoelectronic with oxygen. The spin change between 1HNO and 3NO- retards proton transfer from HNO and the high reduction potential eliminates the direct reduction of

NO as a source of in vivo HNO formation (Scheme 36).93, 94, 96

Scheme 36. Equilibrium predominance of 1HNO

Nitroxyl (HNO) represents the simplest possible nitroso compound (X-N=O, X = H) and demonstrates reactivity similar to C-nitroso compounds. The chemical reactions of

HNO have been reviewed and HNO generally reacts as an electrophile.96-99 Nitroxyl reacts rapidly with thiols as an electrophile to give N-hydroxysulfenamides that react with excess thiol to give disulfides and hydroxylamine or rearranges to sulfinamides

(Scheme 37). Calculations support the rapid reactions of HNO with thiols and other soft nucleophiles. HNO dimerizes to yield hyponitrous acid that dehydrates to nitrous oxide. Finally, nitroxyl reacts with oxidized metals (especially iron) to yield reduced nitrosyl complexes 93,96, 97

Scheme 37. Reactivity of nitroxyl with thiol

1.8 Mycobactin S

Mycobactins are a family of siderophores produced by mycobacteria including M. tuberculosis and M. smegmatis for promoting the iron uptake process and bacterial growth. All mycobactins have a nearly identical molecular nucleus, which includes two hydroxamic acid residues cobactin and mycobactic acid (83 and 84, Scheme 38) and a

2-hydroxyphenyl-oxazoline residue (Scheme 38). Detailed studies on mycobacterial growth factors and the structural elucidation of mycobactins led G. A. Snow to conclude that alternate or modified forms of the mycobactins could have anti- tuberculosis activity.47, 76 Mycobactin S acts as an antimycobacterial agent by disrupting the iron acquisition process in bacteria and hence finds tremendous therapeutic attention for mycobacterial infectious diseases. Mycobactin S also qualifies for better drug-delivery across the mycobacterial cell envelope. The Miller group verified the growth inhibitory activity of mycobactin S in a human tuberculosis cell line. Mycobactin S, isolated from M. smegmatis, consists of a seven membered cyclic hydroxamic acid residue cobactin (83) and a long chain hydroxamic acid residue mycobactic acid (84, Scheme 38).

Scheme 38. Structure of mycobactin S

This background work suggests that cyclic hydroxamic acids are biologically important molecules and potentially useful therapeutic agents but existing chemical methods to synthesize them are somewhat difficult and limited. Our research focuses on the development of synthetic methodology towards making cyclic hydroxamic acids and related natural products.

1.8.1 Biosynthesis of Mycobactins

Iron is essential for bacterial survival and is a seminal growth factor during infectious diseases in vertebrates.100 Under iron deficient condition, bacteria up-regulates the production of the enzymes that synthesize the siderophore mycobactin. The Walsh and coworkers have done a detailed study on identifying the mycobactin T biosynthetic

(mbt) gene clusters.101 The gene clusters, designated mbtA-J, contain isochorismate synthase, acetyl hydrolase, salicylate-AMP ligase, polyketide synthase, lysine-N- oxygenase and nonribosomal peptide synthatase. A major biosynthetic strategy in mycobactin synthesis is the use of non-ribosomal peptide synthatase assembly logic to assemble the hydroxmate and the oxazoline moieties. The non-ribosomal peptide synthetases are made during post translational modification of peptide carrier proteins and these enzymes mediate the generation of mycobactin peptide backbone. It is unknown whether lysine-N-oxygenase oxidizes the terminal amine of lysine residues befoe or after incorporation into mycobactin T The proposed linear biosynthesis is initiated with salicylic acid and terminated with intramolecular cyclization forming the seven membered ring hydroxamic acid.102

Research goals of this dissertation

1) Investigate the Piloty’s acid based –NOH insertion reaction methodology to

synthesize cyclic hydroxamic acid (mainly five and six membered) by –NOH

insertion and ring-expansion of a cyclic ketone as substrate

Scheme 39. Piloty’s acid based –NOH insertion of cyclic ketones

This –NOH insertion methodology research particularly aims to:

a) optimize the reaction conditions, yield and purification of the ring-expanded

cyclic hydroxamic acid as a –NOH insertion product

b) explore the regioselectivity and stereoselectivity of the insertion reaction using a

set of syn-bicyclic ketones as substrates

c) establish the mechanism of the insertion-ring expansion reaction, examining any

role of HNO

d) examine the scope of this unique ring-expansion reaction and study the

synthetic application to a cobactin core synthesis

2) Utilize nitroso-ene reactions towards the formal synthesis of cobactin and

mycobactic acid.

a) Investigate the scope of intramolecular nitroso-ene reactions towards cobactin

core synthesis. This research particularly aims to explore the alkene chain

lengths of different acyl nitroso intermediates

b) Investigate intermolecular nitroso-ene reactions towards the formal synthesis

of cobactin and mycobactic acid. This research mainly aims to:

I. explore the structure and chain lengths of different acyl nitroso species and

study their nitroso-ene reactions

II. investigate the nitroso-ene reactions of different acyl-nitroso enophiles to

synthesize the cobactin and mycobactic acid precursors from Nα-acyl-

homoallyl-glycine ester as the alkene substrate (scheme 40)

Scheme 40. Intermolecular nitroso-ene approach towards formal synthesis of cobactin and mycobactic acid

CHAPTER 2

SYNTHESIS OF CYCLIC HYDROXAMIC ACID THROUGH –NOH INSERTION OF

KETONES

Ranjan Banerjee and S. Bruce King*

Department of Chemistry,Salem Hall, Box 7486, Wake Forest University, Winston Salem,

North Carolina 27109

The work contained in this chapter was initially published in Organic Letters 2009, 11,

4580-4583. The manuscript, including figures and schemes, was drafted by Ranjan

Banerjee and edited by S. Bruce King. Since publication, changes in both format and content were made to adapt this work for the dissertation format. The research described herein was performed by Ranjan Banerjee. Crystal structure determination was performed by Dr. Cynthia S. Day. This work was supported by the American Chemical

Society Petroleum Research Fund (PRF 48660-AC1).

2.1 Introduction

Recent studies demonstrate the distinct biological character of nitroxyl (HNO)

compared to its redox partner nitric oxide (NO) as a signaling agent in the vascular system.103, 104 Nitroxyl exhibits different chemistry from NO by dimerizing and

dehydrating to nitrous oxide, by reacting with heme proteins through separate

mechanisms to NO and by rapidly condensing with thiols to yield disulfides and

sulfenamides.97, 98 This chemistry requires the use of HNO donors in fundamental

chemical and biochemical studies and focuses attention on these donors as potential

therapies for various conditions including congestive heart failure, cancer, alcoholism and

hemolytic disorders.103

Angeli’s salt (Na2N2O3) and Piloty’s acid (PhSO2NHOH, N-hydroxybenzene

sulfonamide, 33, Scheme 15) represent the two most widely used HNO sources for routine study. In addition to releasing HNO, Piloty’s acid reacts with aldehydes under basic conditions to give the corresponding hydroxamic acid and benzenesulfinic acid and this reaction forms the basis of the Angeli-Rimini test for the colorimetric identification of aldehydes (Scheme 15 and Scheme 41).52, 105 Early reports indicate that Piloty’s acid reacts with cyclopentanone to yield the corresponding cyclic hydroxamic acid (85,

Scheme 41) in low yield.88

Scheme 41. Angeli-Rimini reaction of aldehydes and ketones

Given our interest in the chemistry of HNO donors, we further examined this unusual reaction of Piloty’s acid with cyclic ketones and showed that under basic conditions N- hydroxybenzenesulfonamide reacts with small (four and five-membered) cyclic ketones to give the cyclic hydroxamic acid in moderate yields through a mechanism that includes a C-nitroso intermediate.

2.2 Synthesis of N-Hydroxy Benzenesulfohydroxamic Acid (Piloty’s acid)

Piloty’s acid is a commercially available compound, but we observed better results when freshly prepared and purified in the laboratory. It can be synthesized by dropwise addition of benzenesulfonyl chloride (86, Scheme 42) to a neutralized solution of hydroxylamine hydrochloride (87, Scheme 42) in methanol at 0°C with stirring at room temperature overnight.106 The crude Piloty’s acid (33) was purified by flash chromatography and stored in an airtight glass container in the freezer as it is vulnerable to air oxidation and decomposition at room temperature.

Scheme 42. Synthesis of Piloty’s Acid

2.3 -NOH Insertion Reaction of Cyclic Ketones

2.3.1 Synthesis of N-hydroxy Piperidone

Basic decomposition of Piloty’s acid was performed in the presence of cyclopentanone to yield the ring expanded N-hydroxy piperidone (85, Scheme 41). As

36 earlier reported,107 treatment of cyclopentanone with Piloty’s acid (0.9 equiv.) in ethanol with excess sodium hydroxide at room temperature yields 1-hydroxy piperdine-2-one

(85, Scheme 41, Entry 1, Table 1) in 18% yield. Table 1 shows that increasing the molar equivalents of Piloty’s acid increases the yield of 85 with two equivalents more than doubling and greater equivalents furthering increasing the isolated yield of 85 (Table 1,

Entries 5-8). The use of sodium methoxide as base or sodium hydride in a polar aprotic solvent (THF or DMF) results in product but does not improve the observed yield (Table

1, Entries 2-4).

Table 1. Results of –NOH insertion into cyclopentanone

Entry Equivalents of PA Base / Solvents Yield

1 0.9 2N NaOH / EtOH 18%

2 1.1 NaH / THF 11%

3 2 NaH / DMF 2%

4 2 NaOMe / MeOH 33%

5 2 2N NaOH / EtOH 40%

6 4 2N NaOH / EtOH 44%

7 6 2N NaOH / EtOH 59%

8 10 2N NaOH / EtOH 69%

Figure 1. X-ray Diffraction structure of N-hydroxy piperidone (85)

The structure of the N-hydroxy piperidone (85, Figure 1) was confirmed by NMR and

X-ray crystallography. Intermolecular H-bonding was observed between the carbonyl

oxygen and the H atom attached to the N-hydroxy oxygen of another molecule in the

same unit cell.

2.3.2 -NOH Insertion Reaction in Cyclobutanones

While treatment of cyclohexanone with Piloty’s acid in basic ethanol yields less than

5% of the corresponding cyclic hydroxamic acid (88, N-hydroxycaprolactam, Scheme

43), exposure of cyclobutanone to these conditions gives the five-membered ring

hydroxamic acid in 39% yield (89, Scheme 43). X-ray diffraction studies on crystalline

89 further confirms the structure of the ring-expanded hydroxamic acid (Figure 2).

Higher equivalents of Piloty’s acid yields a higher amount amount of 89 (Entry 1-2,

Table 2) in ethanol with excess sodium hydroxide as base. The use of sodium methoxide as base did not improve the yield (Entry 3, Table 2). Given the fact that Piloty’s acid decomposes to nitroxyl (HNO), the slow addition of Piloty’s acid (same equivalents) to

38 the reaction mixture does not improve yield. The volatility of cyclobutanone and the water solubility of 89 hinders isolation and purification, which account for the limited yield of 89, and these results suggest a relationship between ring size and yield.

Attempts to convert 2-hexanone to the corresponding hydroxamic acid under identical conditions gives less than 5% of N-hydroxy-N-propylacetamide (Scheme 43).

Scheme 43. –NOH insertion with various ketones

Figure 2. X-ray Diffraction Structure of 89

Table 2. Results of –NOH insertion into cyclobutanone

Entry Equivalents of PA Base / Solvents Yield

1 1.1 2N NaOH / EtOH 29%

2 2 2N NaOH / EtOH 39%

3 2 NaOMe / MeOH 32%

This success with cyclobutanone encouraged the application of this sequence to more substituted less-volatile cyclobutanones (Scheme 45). A standard two step sequence of a

[2 + 2] cycloaddition between an alkene and dichloroketene followed by Zn / AcOH

O O a b Cl 48% Cl 33% 90

O O a b Cl 28% Cl 34% 91

O O a b Cl Ph 46%Ph Cl 79% 92 O O a b Cl 5 19% 5 Cl 74% 5 93

a = i) Zn,ii) POCl3,iii) Cl3CCOCl, reflux /ether 0 b = i) Zn ii) CH3COOH, stirring at 60 C for 2hrs

Scheme 44. Synthesis of substituted cyclobutanones

reduction of the α,α-dichloroketone intermediate generates various substituted

cyclobutanone substrates (90 - 93 , Scheme 44).108, 109

Treatment of these cyclobutanones with Piloty’s acid (2 equiv.) in base forms the

corresponding cyclic hydroxamic acids in 30-59% yield ( 94 – 97, Scheme 45). The

decreased volatility of the substituted cyclobutanone substrates (compared to

cyclobutanone) and the decreased water solubility of the products likely improve the

isolated yields and facilitate purification. This sequence yields cyclic five-membered

ring hydroxamic acids derived from two unsymmetric bicyclic cyclobutanones (94 and

95) and from two symmetric substituted cyclobutanones (96 and 97). 1H and 13C NMR spectroscopy and high resolution mass spectrometry support the structures of 94-97 and

X-ray diffraction studies on crystals of 94 confirm the structure of this bicyclic hydroxamic acid (Figure 3). In addition to structural confirmation, the crystallography

Scheme 45. Reaction of Piloty’s acid with substituted cyclobutanones

studies also reveal two important features of this reaction 1) the –NOH inserts to the

more substituted side of the ketone demonstrating regioselectivity and 2) the –NOH

group adds stereoselectively with the overall ring juncture relative stereochemistry

remaining cis. NMR and chromatographic analysis of the crude reaction mixture do not

reveal the presence of any other hydroxamic acids. These results appear similar to the

Baeyer-Villiger oxidation of ketones with migration of the more electron-donating

branch with retention of configuration.110

Figure 3. X-ray Diffraction Structure of 94

2.3.3 Solid phase modification

Despite success in developing this method to synthesize cyclic hydroxamic acids, the

target molecules’ high polarity and difficulty in being chromatographed or recrystallized complicated purification. In an attempt to solve this problem, we explored the solid-phase

Angeli-Rimini reaction with polymer bound sulfohydroxamic acid as the reagent and cyclic ketones as substrate (Scheme 46). The reactions were done at room temperature, in dry THF using sodium methoxide as a base. Acidic work up was done with Dowex- 42

50WX2 ion exchange resin after filtration of the polymer bound reagent. The resin-

supported N-hydroxybenzene sulfonamide (36) was prepared as described before.52 Two equivalents of the modified reagent 36 reacted with cyclopentanone and cyclobutanone

(Scheme 46) to produce the corresponding hydroxamic acids in 9% and 35% yield respectively (Scheme 46). These results suggest solid phase modification does not improve the yield of the product over regular Piloty’s acid based ring-expansion reactions.

Scheme 46. Solid phase modification of –NOH insertion reaction

2.4 Mechanism of –NOH insertion reaction

Early work proposes two mechanistic possibilities for this unsual transformation. The first includes the direct addition of HNO or –NO to the ketone to form a C-nitroso

intermediate 98 that rearranges to the observed product (85, Scheme 47, path a). Further

experiments argue against this route and suggest instead that a nucleophilic addition of

the N-anion of Piloty’s acid to the cyclic ketone gives a tetrahedral intermediate 99 that

rearranges to the ring-expanded hydroxamic acid with the loss of benzenesulfinic acid

(Scheme 47, path b).111, 112 Such a mechanism finds direct precedence in the known reaction of aldehydes with Piloty’s acid to form hydroxamic acids (the Angeli-Rimini reaction, Scheme 41).112

Scheme 47. Proposed Mechanisms for Formation of 85

Experiments with O-benzyl p-toluene N-hydroxysulfonamide (100, Scheme 48), prepared by the treatment of p-tosyl chloride with O-benzylhydroxylamine, provide insight into the mechanism (Scheme 47). This compound cannot decompose to HNO

(Scheme 47, path a) but can form an N-anion that should react to give an O-alkyl cyclic hydroxamate (Scheme 47, path b). Reaction of 100 with cyclobutanone or cyclopentanone fails to yield any insertion product arguing against path a (Schemes 47 and 48) as the mechanism of product formation.

NaaOH SO2NHOBn No Reaction

Scheme 48. Reaction of O-protected Piloty’s Acid with Cyclopentanone

Treatment of 100 with NaH in THF followed by methyl iodide produces the N- methylated product (101, Scheme 49) verifying the nucleoophilic abiltiy of the anion of

100. X-ray diffraction studies on compound 101 cconfirms the structure (Scheme 49). This

SO2NHOCH2Ph SO2NMeOCH2Ph 1. 1.1 eq. NaH in THF 2. MeI, 74%

100 101

Scheme 49. N-methylation and X-ray diffraction structure of 101 result also eliminates the potential invovlement of O-nitrene intermediates that could form through the dissociation of the N-anion of Piloty’s acid and would give the 1-

(benzyloxy) piperidin-2-one product, which was not observed.

Further treatment of smaller O-protected Piloty’s acid analogs such as O-benzyl

methansulfonamide113 or O-methyl benzenesulfohydroxamic acid with cyclopentanone

under basic condition fails to yield any insertion product, indicating sterics do not play a

crucial role in tetrahedral intermediate (99, Scheme 47) formation during the insertion

reaction. In addition, incubation of cyclopentanone with Angeli’s salt in a

methanol/buffer mixture, conditions that clearly generate HNO as judged by gas

chromatographic identification of nitrous oxide, fail to produce 85 arguing against path a

(Scheme 50).114, 115 Previous pKa calculations also disfavor the direct reaction of –NO with cyclic ketones to form a C-nitroso intermediate and product (Scheme 47, path a).116

Scheme 50. Synthesis and reactions of other O-protected sulfonamides

These results led to the examination of acyloxy nitroso compounds as precursors to the C-nitroso intermediate (98, Scheme 51) and cyclic hydroxamic acids. Our previous work shows that hydrolysis of the cyclohexyl-derived acetoxy nitroso compound (102,

Scheme 51) forms cyclohexanone and HNO (as judged by nitrous oxide generation) through a C-nitroso intermediate and highlights these compounds as new HNO donors.117

GC data (Table 3) depicts a direct correlation between the ring size and HNO donating properties of 5-6 membered cyclic acyloxy-nitroso compounds. These GC data suggest

46 cyclopentyl derived acetoxy nitroso compounds are more prone to give ring-expanded hydroxamic acids upon hydrolysis, while cyclohexyl-derived acetoxy nitroso compounds are not because of low ring strain.

______

Table 3. Gas Chromatography Results: Hydrolysis of acyloxy nitroso

Entry Time Mols of N2O % yield

O N OAc 2 hrs 3.70E-08 6.81%

24hrs 3.55E-08 6.53%

2 hrs 2.89E-07 53.15%

24 hrs 2.71E-07 49.82%

Hydrolysis of the cyclopentyl-derived acetoxy nitroso compound (103), generated by the lead tetra-acetate oxidation of cyclopentanone oxime, yields cyclic hydroxamate (85,

Scheme 51) in 75% yield and only trace amounts of nitrous oxide (indicating little HNO formation, Scheme 51). These results provide evidence of a C-nitroso intermediate during the formation of 85 (Scheme 51) from cyclopentanone and Piloty’s acid in basic conditions and also show a ring size dependency on the reaction pathway. Scheme 47 depicts a mechanism where addition of the N-anion of Piloty’s acid generates a tetrahedral intermediate (99) that eliminates benzenesulfinic acid to give the C-nitroso

47 intermediate (98, path c, Scheme 47 and Scheme 51). This mechanism produces the orignially proposed C-nitroso intermediate (98) through this tetrahedral intermediate.

This C-nitroso intermediate either eliminates HNO to give the ketone (cyclohexyl) or rearranges to the cyclic hydroxamic acid (cyclobutyl, cyclopentyl).

Scheme 51. Modified Reaction Mechanism

Increasing the amount of Piloty’s acid would increase the amount of the reactive N- anion under equilibrium conditions and result in an increased yield of the observed product (Table 1). Previous results show slow addition of Piloty’s acid does not alter the yield of the product, suggesting the efficient generation of the N-anion of Piloty’s acid is critical to this method as the anion also decomposes to nitroxyl.92 The C-nitroso interemediate (98) shows structural similarity to the accepted peroxy intermediates in the

Baeyer-Villiger reaction and the observed regiochemistry and stereochemistry appears

consistent. Obviously, the ring strain of the C-nitroso intermediate plays a major

determinant in the product outcome. Unfavorable eclipsing interactions leading to ring

strain likely drive rearrangement of the smaller membered rings (cyclobutyl, cyclopentyl)

to larger rings that relieve these interactions.

In summary, these results show that the N-anion of Piloty’s acid reacts with four and

five-membered ring cyclic ketones to form tetrahedral intermediates that rearrange to C-

nitroso species and ultimately a cyclic hydroxamic acid (depending on ring size) in

moderate yields. These experiments suggest a regio and stereoselective rearrangement

similar to the Baeyer-Villiger reaction. This chemistry shows the complexity of the

reactions of N-hydroxysulfonamides and may prove useful for the synthesis of various

cyclic hydroxamic acid containing compounds, such as siderophores,9, 102, 118, 119 natural iron-chelating compounds that show promise as new antibiotics. As such, these studies also compliment other methods for the direct conversion of ketones to the corresponding

N-OH amides.120-123 This work aids in the structural development of new HNO (both

Piloty’s acid and acyloxy nitroso-based) donors as the reactivity of the N-anion must be

considered for Piloty’s acid-based donors and the structures, particularly ring size, must

be taken into account for new acyloxy nitroso compounds.

2.5 Scope of –NOH Insertion Reaction

2.5.1 Synthesis of O-Protected Cyclic Hydroxamic Acids

Initial ring-expansion of cyclopentanone generates N-hydroxy piperidone (85), which

was used for further transformations including O-acylation and O-alkylation reactions

(Scheme 52) to prepare O-protected cyclic hydroxamic acids. Treatment of N-hydroxy

piperidone (85) with acetic anhydride in the presence of DMAP produces O-acetoxy piperidone (104, Scheme 52) in 63% yield. Treatment of N-hydroxy piperidone with

methyl iodide in the presence of sodium hydride produces O-methyl piperidone (105,

Scheme 52) in 58% yield. Cyclic Weinreb amides 124, 125 such as compound 105 (Scheme

52) are useful synthetic intermediates for ring-opening transformations by Grignard

reagents or other nucleophiles.

Scheme 52. O-protection of N-hydroxy piperidone

2.5.2 –NOH Insertion Reaction in α-Substituted Cyclopentanone

Preliminary results show that an excess of Piloty’s acid greatly increases yield (Table

1), which indicates inefficiencies in generating the reactive species. Apart from the use of

excess amounts of Piloty’s acid another limitation is that these –NOH insertions show the

best results with unsubstituted cyclic ketones as substrates. Treatment of Piloty’s acid

with 2-methyl cyclopentanone under identical –NOH insertion condition yields only trace

amounts of ring expanded cyclic hydroxamic acid 106 (Scheme 53). However this

problem may be overcome through the hydrolysis of the acyloxy nitroso compounds of

the corresponding cyclic ketone. Previous results (Scheme 51) demonstrate that the

hydrolysis of the cyclopentyl-derived acetoxy-nitroso compound 103 produces the ring-

expanded hydroxamic acid 85. Treatment of hydroxylamine hydrochloride with 2-

methyl cyclopentanone yields 2-methyl cyclopentanone oxime (Scheme 53) and

Pb(OAc)4 oxidation gives the corresponding acetoxy nitroso compound (107, Scheme

53). Hydrolysis of 107 gives 6-methyl-N-hydroxy-piperidone (106, Scheme 53) in 77%

yield.

Scheme 53. –NOH Insertion in 2-Methyl Cyclopentanone

2.5.3 -NOH Insertion in Cyclohexenone

Treatment of cyclohexenone (108, Scheme 54) with Piloty’s acid under the identical –

NOH insertion conditions installs an oxime group at the 3-position of the ketone and

generates 3-hydroxyimino cyclohexanone (109, Scheme 54). This reaction provides support for the –NOH insertion mechanism (Scheme 47) and validates the generation of a

C-nitroso intermediate after Michael addition of the N-anion of Piloty’s acid (Scheme

54). This reaction is a unique example of introducing an oxime group at the 3-position of

cyclohexenone and opens a new avenue of oxo-amination reactions of cyclic ketones.

Scheme 54. Piloty’s acid based –NOH insertion reaction of cyclohexenones

2.5.4 Investigation of a Cobactin Synthesis Using the –NOH Insertion Reaction

The final objective of this project was to apply the –NOH insertion methodology to

the preparation of a small biologically relevant natural product, cobactin (83, Scheme

38). We proposed a short synthesis of cobactin from an α-amino cyclohexanone using

this Piloty’s acid based –NOH insertion methodology (Scheme 55). This synthesis would

consist of three steps, aminohydroxylation126, 127 of cyclohexene to give a cyclic amino

alcohol (110, Scheme 55) followed by oxidation to yield an α-amino cyclohexanone

(111) and finally the –NOH insertion reaction with the precursor ketone to yield the

cobactin core 112 (Scheme 55).

Scheme 55. Prosposed route to cobactin core using Piloty’s acid based –NOH insertion

method

Previous results describe that the –NOH insertion works best with four and five

memebered ring ketones. To explore this pathway in a five membered substrate a new route was designed, where Sharpless aminohydroxylation of cyclopentene provided the

N-Boc protected α-amino alcohol (113, Scheme 56) in 76% yield and Dess-Martin

oxidation gave the precursor ketone (114, Scheme 56) in 40% yield. Treatment of 114

with Piloty’s acid under identical –NOH insertion condition afforded only trace amounts

of the ring-expanded hydroxamic acid (115, Scheme 56). These results show the Piloty’s acid based –NOH insertion method may not be an efficient route for the ring-expansion of an α-subsituted cyclic ketones, suggesting the cobactin synthesis will not be an efficient synthetic application of this method.

Scheme 56. Exploring Piloty’s acid rearrnagement reaction with α-amino-substituted

cyclopentanones

2.6 Experimental

General. Melting points (mp) were measured on a Mel-Temp capillary melting point

apparatus and are uncorrected. Analytical TLC was performed on silica gel plates with

QF-254 indicator. Visualization was accomplished with UV light, FeCl3, phosphomolybdic acid, and/or dinitrophenylhydrazine stain. Extraction and chromatography solvents were technical grade. All reactions were performed under an

1 13 inert atmosphere of dry argon. H NMR and C NMR were recorded in CDCl3 and deuterated DMSO on a Bruker Avance 300 MHz and 500 MHz NMR Spectrometer.

Chemical shifts are given in ppm (δ); multiplicities are indicated by s (singlet), d

(doublet), t (triplet), q (quartet), m (multiplet) and b (broadened). Low resolution mass spectra were obtained using an Agilent GC/MS system consisting of a 5973 mass selective detector interfaced to a 6850 gas chromatograph or from HT Laboratories (San

Diego, CA) and data are reported in m/z. Elemental analysis were performed by Atlantic

Microlab, Inc. (Atlanta, GA).

N-Hydroxybenzene sulfonamide (Piloty’s Acid, 33).128 A solution of potassium

carbonate (27.7 g, 0.2 mol) in water (30 mL) was added dropwise to a solution of

hydroxylamine hydrochloride (13.6 g, 0.2 mol) in water: methanol (3:2, 50 mL) at 0°C

with vigorous magnetic stirring for 1 h. Ice cold methanol (200 mL) was added in a

single portion to this mixture followed by dropwise addition of benzenesulfonyl chloride

(35.3 g, 0.2 mol) over 1 h. After 18 h of stirring the mixture was filtered and the methanol

was removed under vacuum. The water residue was acidified to neutral pH, extracted

with ethyl acetate (300 mL), concentrated under vacuum and purified by flash

chromatography on SiO2 to give 33 as a white solid (9.25 g, 27%); Rf = 0.176 (3:1 pet.

1 ether : EtOAc); mp 108°C. H NMR (DMSO-d6) 9.62-9.68 (m, 2H), 7.85-7.92 (m, 2H),

13 7.61-7.78 (m, 3H); C NMR (DMSO-d6) 137.12, 133.01, 128.84, 127.98.

1-Hydroxypiperidine-2-one (85).107, 129 A solution of degassed sodium hydroxide (2N,

80 mmol, 40 mL) was added to a solution of cyclopentanone (1 mL, 11.3 mmol) in EtOH

(20 mL) at 0°C. To this mixture, a solution of Piloty’s acid (3.91 g, 22.6 mmol) in EtOH

(20 mL) was added dropwise over 30 min and the reaction mixture was stirred at room

temperature for 18 h. The ethanol was removed under vacuum and the water layer was

extracted with diethyl ether to remove unreacted ketone. The aqueous layer was acidified

to pH 5.5 with 2N HCl and extracted with CHCl3 (6 x 50 mL). The CHCl3 layer was

dried with MgSO4 and concentrated to give a semisolid crude residue that was purified by

flash chromatography to give 85 as a reddish white solid (0.52 g, 40%); Rf = 0.37 (95:5

CHCl3 : MeOH). Pure white crystals of N-hydroxy piperidone were obtained by

1 subliming the product (45°C, 0.05mm); mp 48-50°C. H NMR (CDCl3) 9.39 (1H, b),

3.63 (t, J = 5.93 Hz, 2H), 2.45 (t, J = 6.47 Hz, 2H), 1.95 (m, 2H), 1.82 (m, 2H); 13C

+ (CDCl3) NMR 165.39, 49.96, 31.54, 23.58, 21.13; GC MS m/z 115 (M ). Anal. Calcd.

for (C5H9NO2. 0.5H2O) : C 48.36%, H 8.12%, N 11.29%. Found: C 48.36%, H 8.11%, N

11.19%.

O OH N

1-Hydroxyazepan-2-one (88).119 A solution of degassed sodium hydroxide (2N, 80

mmol, 40 mL) was added to a solution of cyclohexanone (1.11 g, 11.3 mmol) in EtOH

(20 mL) at 0°C. To this mixture, a solution of Piloty’s acid (3.91 g, 22.6 mmol) in EtOH

(20 mL) was added dropwise over 30 min and the reaction mixture was stirred at room

temperature for 18 h. The ethanol was removed under vacuum and the water layer was

extracted with diethyl ether to remove the unreacted ketone. The aqueous layer was

acidified to pH 5.5 with 2N HCl and extracted with CHCl3 (5 x 50 mL). The CHCl3 layer was dried with MgSO4 and concentrated to give a solid crude residue that was purified by flash chromatography to give the hydroxamic acid 88 as a reddish white solid (60 mg,

1 4%) ; Rf = 0.41 (95:5 CHCl3 : MeOH). H NMR (CDCl3) 8.13 (b, 1H), 3.72 (b, 2H),

13 2.54-2.52 (m, 2H), 1.77-1.65 (m, 6H); C NMR (CDCl3) 166.93, 50.51, 34.1, 29.69,

26.65, 22.94; GC MS m/z 129 (M+).

1-Hydroxypyrrolidine-2-one (89). A solution of degassed sodium hydroxide (2N, 80

mmol, 40 mL) was added to a solution of cyclobutanone (1 g, 14.27 mmol) in EtOH (20

mL) at 0°C. To this mixture, a solution of Piloty’s acid (4.94 g, 28.53 mmol) in EtOH (20

mL) was added drop wise over 30 min. The reaction mixture was stirred at 0°C for 4 h

and at room temperature for another 18 h. The ethanol was removed under vacuum and

the water layer was extracted with diethyl ether to remove the unreacted cyclobutanone.

The aqueous layer was acidified to pH 5.5 with 2N HCl and extracted with CHCl3 (6 x 50

mL). The CHCl3 layer was dried with MgSO4 and was concentrated to a solid crude

residue that was purified by flash chromatography to give 89 as a pale white solid

product (0.565 g, 39%) ; Rf = 0.4 (95:5 CHCl3 : MeOH). Pure white crystals were

1 collected after sublimation (40°C, 0.05mm); mp 77-79°C. H NMR (CDCl3) 10.41 (b,

1H), 3.67 (t, J = 7.19 Hz, 2H), 2.41 (t, J = 7.57 Hz, 2H), 2.061 (q, J = 7.26 Hz, 2H); 13C

+ NMR (CDCl3) 170.92, 49.16, 28.55, 15.56; GC MS m/z 101(M ). Anal. Calcd. for

C4H7NO2 : C 47.50%, H 6.98%, N 13.86%, Found : C 47.72%, H 7.08%, N 13.75%.

2-Hydroxyhexhydrocyclopenta[c]pyrrole-1(2H)-one (94). A solution of degassed

sodium hydroxide (2N, 80 mmol, 40 mL) was added to a solution of

bicylco[3.2.0]heptane-6-one (0.68 g, 6.18 mmol) in EtOH (20 mL) at 0°C. To this

mixture, a solution of Piloty’s acid (2.14 g, 12.36 mmol) in EtOH (20 mL) was added dropwise over 30 min. The reaction was stirred at 0°C over 4 h and over 18 h at room temperature. The ethanol was removed under vacuum and the water layer was extracted with diethyl ether to remove unreacted ketone. The aqueous layer was acidified to pH 5.5 with 2N HCl and extracted with CHCl3 (6 x 50 mL).The CHCl3 layer was dried and

concentrated to give a solid crude residue that was purified by flash chromatography to

give 94 as a pale white solid product (0.35 g, 41%); Rf = 0.45 (95:5 CHCl3: MeOH,). The

product was further purified by sublimation (50°C, 0.05mm) to give white crystalline

1 solid; mp 76°C. H NMR (CDCl3) 10.39 (b, 1H), 4.21 (b, 1H), 2.29-2.1 (m, 2H), 2.78-

13 2.63 (m, 1H), 2.076-1.97 (m, 1H), 1.8-1.48 (m, 5H); C NMR (CDCl3) 169.52, 65.59,

+ 35.89, 35.03, 32.67, 30.61, 23.72; GC MS m/z 141 (M ). Anal. Calcd. for C7H11NO2 : C

59.54% , H 7.86%, N 9.93%, Found : 59.57%, 7.99%, 9.90%.

OH N O

2-Hydroxyoctahydro-1H-isoindol-1-one (95). A solution of degassed sodium hydroxide

(2N, 80 mmol, 40 mL) was added to a solution of octahydro-1H-inden-1-one (0.69 g,

5.56 mmol) in EtOH (20 mL) at 0°C. To this reaction mixture, a solution of Piloty’s acid

(1.926 g, 11.1 mmol) in EtOH (20 mL) was added dropwise over 30 min and the reaction

mixture was stirred at 0°C for 4 h and for 18 h at room temperature. The ethanol was

removed under vacuum and the water layer washed with diethyl ether. The aqueous layer

was acidified to pH 5.5 with 2N HCl and extracted with CHCl3 (6 x 50 mL). After drying

with MgSO4, the CHCl3 layer was concentrated to give a solid crude residue that was

purified by flash chromatography to give 95 (0.41 g, yield 48%); Rf = 0.46 (95:5 CHCl3:

MeOH). Sublimation (55°C, 0.05mm) afforded a white crystalline solid; m.p 78°C. 1H

NMR (CDCl3) 9.56 (b, 1H), 3.74 (q, J = 4.95 Hz, 2H), 2.45-2.25 (m, 2H), 2.12-1.92 (m,

13 2H), 1.76-1.64 (m, 2H), 1.59-1.25 (m, 5H); C NMR (CDCl3) 172.53, 59.1, 35.79, 29.8,

+ 28.39, 25.83, 23.24, 20.69; GC MS 155 (M ). Anal. Calcd. for C8H13NO2 : C 61.95%, H

8.45%, N 9.03%, Found : C 61.60%, H 8.62%, N 8.93%.

1-Hydroxy-4-phenylpyrrolidin-2-one (96). A solution of degassed sodium hydroxide

(2N, 80 mmol, 40 mL) was added to a solution of 3-phenyl cyclobutanone (0.67 g, 4.6

mmol) in EtOH (20 mL) at 0°C. To this mixture a solution of Piloty’s acid (1.59 g, 9.2

mmol) in EtOH (20 mL) was added dropwise over 30 min, stirred at 0°C for 4 h and at

room temperature overnight. The ethanol was removed under vacuum and the water layer

washed with diethyl ether to remove the unreacted ketone. The aqueous layer was

acidified to pH 5.5 with 2N HCl and extracted with CHCl3 (5 x 50 mL), was dried and

concentrated to give a solid crude residue that was purified by flash chromatography to

give 96 as a reddish product (0.3 g, 37%); Rf = 0.65 (95:5 CHCl3: MeOH). Sublimation

1 (60°C, 0.05 mm); mp 84°C. H NMR (CDCl3) 9.94 (b, 1H), 7.38-7.23 (m, 5H), 3.67-3.56

(pentet, J = 8.21 Hz, 1H), 4.04 (t, J = 8.78 Hz, 1H), 3.76-3.71 (dd, J = 22.29, 7.18 Hz,

1H), 2.82-2.92 (dd, J =17.04, 9.26 Hz, 1H), 2.52-2.60 (dd, J = 17.04, 7.73 Hz, 1H) ; 13C

NMR (CDCl3) 169.63, 141.64, 129.05, 127.42, 126.76, 55.63, 36.67, 34.68. GC MS m/z

+ 177 (M ). Anal. Calcd. for (C10H11NO2, 0.1H2O): C 67.08%, H 6.31%, N 7.82%, Found :

C 67.17%, H 6.31%, N 7.62%.

4-Hexyl-1-hydroxypyrrolidine-2-one (97). A solution of degassed sodium hydroxide

(2N, 80 mmol, 40 mL) was added to a solution of 3-hexyl cyclobutanone (0.892 g, 5.79

mmol) in EtOH (20 mL) at 0°C. A solution of Piloty’s acid (1.926 g, 11.1 mmol) in

EtOH (20 mL) was added dropwise over 30 min and the reaction mixture was stirred at

0°C over 4 h and then over 18 h at room temperature. The ethanol was removed under

vacuum and the water layer washed with diethyl ether. The aqueous layer was acidified

to pH 5.5 by 2N HCl and extracted with CHCl3 (5 x 50 mL), dried with MgSO4 and concentrated to give a liquid residue that was purified by flash chromatography to afford

1 97 as a liquid (0.64 g, yield 59%); Rf = 0.65 (95:5 CHCl3: MeOH). H NMR (CDCl3)

10.49 (b,1H), 3.74 (t, J = 8.79 Hz, 1H), 3.37-3.23 (dd, J = 9.1, 6.75 Hz, 1H), 2.58-2.48

(dd, J = 16.49, 8.89 Hz, 1H), 2.43-2.26 (septet, J = 7.24 Hz, 1H), 2.12-2.01 (dd, J

=16.486, 7.085 Hz, 1H),1.58-1.35 (m, 2H), 1.34-1.15 (m, 8H), 0.92-0.84 (m, J = 6.52 Hz,

13 3H); C NMR (CDCl3) 170.17, 54.61, 35.08, 34.92, 31.71,29.17, 29.14,27.09, 22.59;

+ ESI MS m/z 187.2 (M+2H ). Anal. Calcd. for (C10H19NO2, 0.1 mol H2O): C 64.19%, H

10.35%, N 7.49%, Found : C 64.32%, H 10.36%, N 7.19%.

N-(Benzyloxy)-4-methylbenzenesulfonamide (100). A solution of potassium carbonate

(1.38 g, 10 mmol) in water (15 mL) was added dropwise to a solution of O-benzyl hydroxylamine hydrochloride (1.59 g, 10 mmol) in water: methanol (3:2, 25 mL) at 0°C.

After stirring vigorously for 1 h, ice cold MeOH (100 mL) was added in a single portion followed by dropwise addition of benzenesulfonyl chloride (1.90 g, 10 mmol). After 24 h at room temperature, the methanol was removed; the water layer acidified to neutral pH and extracted with ethyl acetate. The organic layer was dried over MgSO4, filtered and concentrated to give a crude residue that was purified by flash chromatography to give

1 100 as a white solid (1.39 g, 51%); Rf = 0.46 (85:15 pet. ether : EtOAc). mp 72°C, H

NMR 7.8 (d, J = 8.25 Hz, HA and HA’ of AA’BB’ spin-system), 7.34-7.31 (m, 7H), 6.89

(b, 1H), 4.97 (s, 1H), 2.43 (s, 1H); 13C NMR 145.53, 135.92, 134.33, 130.38, 129.96,

129.29, 129.20, 129.15, 80.02, 22.30. ESI MS m/z 276 (M-H-) Anal. Calcd. for

C14H15NSO3 : C 60.63%, H 5.46%, N 5.05%; Found: C 60.66%, H 5.35%, N 4.87%.

N-(Benzyloxy)-N,4-dimethylbenzenesulfonamide (101). A solution of MeI (0.068 g,

0.48 mmol) in THF (10 mL) was added dropwise into a mixture of NaH (0.014 g, 0.58 mmol) and 100 (0.134 g, 0.48 mmol) in dry THF (40 mL) that had stirred for 30 min..

The reaction mixture was stirred for 18 h and quenched with water (10 mL). The THF was removed under vacuum and the aqueous layer acidified to pH 7 and extracted with

EtOAc (2 x 30 mL). The organic layer was dried, filtered and concentrated to give a solid residue that was purified by flash chromatography to give 101 as a white solid (0.109 g,

78%); Rf = 0.4 (90:10 pet. ether: EtOAc). The product was crystallized from pet. ether /

1 EtOAc (95:5). mp 57°C, H NMR (CDCl3) 8.29 (d, J = 8.29 Hz, 2H), 7.4-7.32 (m, 7H),

13 5.02 (s, 2H), 2.66 (s, 3H), 2.43 (s, 3H); C NMR (CDCl3) 144.79, 135.6, 129.78,

129.54, 129.49, 129.34, 128.58, 128.49, 78.62, 40.14, 21.69.

1-Nitroso-cyclohexyl-acetate (102).117 A solution of cyclohexanone oxime (2.73 g,

24.12 mmol) in CH2Cl2 (50 mL) was added drop wise with stirring to a solution of lead

tetraacetate (10.69 g, 24.12 mmol) in DCM (100 mL) at 0°C. A blue color gradually

appeared with the addition of the oxime. After 1 h at 0°C, the reaction mixture was

warmed to room temperature and stirred for another 2 h, water (30 mL) was added, and

the organic layer was washed with (2 x 30 mL) water and saturated sodium bicarbonate

solution (2 x 30 mL). The organic layer was dried over MgSO4, the solvent evaporated

and the residue purified by flash chromatography to give 102 as a bright blue liquid (2.09

1 13 g, 51%); Rf = 0.68 (20:1 pet.ether : EtOAc). H NMR (CDCl3) 2.19-1.29 (m, 13H); C

NMR (CDCl3) 21.68 (CH2), 21.99 (2CH2), 25.09 (2CH2), 29.72 (CH3), 124.09 (O-C-N),

169.28 (C=O) Anal. Calcd. for C8H13NO3 : C 56.13%, H 7.65%, N 8.18%; Found: C

56.07%, H 7.84%, N 8.05%.

1-Nitroso-cyclopentyl-acetate (103). A solution of cyclopentanone oxime (0.495 g, 5

mmol) in CH2Cl2 (10 mL) was added dropwise with stirring to a solution of lead

tetraacetate (2.216 g, 5 mmol) in DCM (20 mL) at 0°C. A blue color gradually appeared

with the addition of the oxime. After 1 h at 0°C, the reaction mixture was warmed to

room temperature. After 2 h at room temperature, water (10 mL) was added, and the

organic layer was washed with water (2 x 10 mL) and saturated sodium bicarbonate

solution (2 x 10 mL).The organic layer was dried over MgSO4, the solvent evaporated

and the residue purified by flash chromatography to give 103 as a bright blue liquid

1 (0.360 g, 49%); Rf = 0.62 (20:1 pet. ether: EtOAc). H NMR (CDCl3) 2.28-1.81 (m,

13 11H); C NMR (CDCl3) 21.38 (2CH2), 25.73 (2CH2), 34.21 (CH3), 131.19 (O-C-N),

169.67 (C=O) Anal. Calcd. for C7H11NO3 : C 53.49%, H 7.05%, N 8.91%; Found: C

53.89%, H 7.29%, N 8.69%.

2-Oxopiperidin-1-yl acetate (104). A solution of acetic anhydride (0.709 g, 5 mmol) in

CH2Cl2 (10 mL) was added dropwise into a mixture of DMAP (0.027 g, 0.22 mmol) and

N-hydroxy piperidone (0.51 g, 4.43 mmol) in dry CH2Cl2 (150 mL) over 30 min. The

reaction mixture was stirred for another 12 h. This solution was thoroughly washed with

water, brine (30 mL). The organic layer was dried over MgSO4, the solvent was removed

under vacuum and the residue was purified by flash chromatography to give 104 as a

1 white solid (0.39 g, 63%); Rf = 0.81 (95:5 CHCl3:MeOH). H NMR (CDCl3) 3.565 (t, J =

5.99 Hz, 2H), 2.47 (t, J = 6.56 Hz, 2H), 2.15 (s, 3H), 2.02-1.885 (m, 2H), 1.86-1.75 (m,

13 2H); C NMR (CDCl3) 166.44, 164.83, 51.72, 33.45, 24.42, 21.62, 19.07.

1-Methoxypiperidin-2-one (105). A solution of MeI (0.709 g, 5 mmol) in THF (10 mL)

was added dropwise into a mixture of NaH (0.12 g, 4.98 mmol) and N-hydroxy

piperidone (0.521 g, 4.53 mmol) in dry THF (40 mL) that had stirred for 30 min. The

reaction mixture was stirred for 12 h and quenched with water (10 mL). The THF was

removed under vacuum and the aqueous layer acidified to pH 7 and extracted with CHCl3

(3 x 40 mL). The organic layer was dried, filtered and concentrated to give a solid residue that was purified by flash chromatography to give 105 as a reddish liquid (0.339 g,

1 58%); Rf = 0.32 (EtOAc). H NMR (CDCl3) 3.79 (s, 3H), 3.61 (t, J = 6.01 Hz, 2H), 2.47

13 (t, J = 6.46 Hz, 2H), 2.01-1.93 (m, 2H), 1.87-1.74 (m, 2H); C NMR (CDCl3) 167.48,

62.77, 50.75, 35.33, 26.21, 23.42.

2-Methyl-1-nitrosocyclopentyl acetate (107). A solution of 2-methyl cyclopentanone- oxime (0.97 g, 8.58 mmole) in CH2Cl2 (10 mL) was added dropwise with stirring to a

solution of lead tetraacetate (3.805 g, 8.58 mmole) in CH2Cl2 (20 mL) at 0°C. A blue

color gradually appeared with the addition of the oxime. After 1 h at 0°C, the reaction

mixture was warmed to room temperature. After 2 h at room temperature, water (10 mL)

was added, and the organic layer washed with water (2 x 10 mL) and saturated sodium

bicarbonate solution (2 x 10 mL).The organic layer was dried over MgSO4, the solvent evaporated under vacuum and the residue purified by flash chromatography to give 107

1 as a bright blue liquid (0.968 g, 66%); Rf = 0.44 (20:1 pet. ether: EtOAc). H NMR

13 (CDCl3) 2.92-2.71 (m, 1H), 2.47-1.77 (m, 11H); C NMR (CDCl3) 168.91, 130.68,

46.21, 32.67, 23.53, 20.9, 12.12.

1-Hydroxy-6-methylpiperidin-2-one (106). A solution of 2-methyl-1-nitrosocyclopentyl

acetate (0.524 g, 4.07 mmol) in MeOH (10 mL) was added to an aqueous solution of 2M

NaOH (10 mL). The blue color gradually disappeared and after 18 h stirring at room

temperature, the MeOH was removed under vacuum. The aqueous layer was acidified to pH 6 and extracted with CHCl3 (3 x 50 mL). The organic layer was washed with water,

brine and dried over MgSO4. The solvent was removed under vacuum and the residue was purified by flash chromatography to give 106 as a reddish brown solid (0.335 g,

1 78%); Rf = 0.62 (20:1 pet. ether: EtOAc). H NMR (CDCl3) 8.45 (b, 1H), 3.89-3.77 (m,

1H), 2.6-2.29 (m, 2H), 2.12-1.92 (m, 1H), 1.89-1.77 (m, 1H), 1.75-1.55 (m, 2H), 1.35 (d,

13 J = 6.34 Hz, 3H); C NMR (CDCl3) 164.82, 55.16, 31.05, 30.82, 19.18, 18.206.

3-(Hydroxyimino)cyclohexanone (109). A solution of degassed sodium hydroxide (2N,

40 mmol, 20 mL) was added to a solution of cyclohexenone (0.238 g, 2.5 mmol) in EtOH

(20 mL) at 0°C. To this mixture, a solution of Piloty’s acid (0.866 g, 5 mmol) in EtOH

(20 mL) was added dropwise over 30 min and the reaction mixture was stirred at room

temperature for 18 h. The ethanol was removed under vacuum and the water layer was

extracted with diethyl ether to remove unreacted ketone. The aqueous layer was acidified

to pH 6 with 2N HCl and extracted with CHCl3 (3 x 50 mL). The CHCl3 layer was dried

with MgSO4 and concentrated to give a semisolid crude residue that showed a brown spot in FeCl3 stain by TLC. The residue was purified by flash chromatography to give a

yellow liquid (0.063 g, 20%) as an inseparable mixture of syn and anti oxime product

1 (109); Rf = 0.45 (EtOAc). H NMR (CDCl3) 8.39 (b, 1H), 3.48 (s, 1H), 3.22 (s, 1H), 2.73

(t, J = 6.57 Hz, 1H), 2.565-2.47 (m, 2H), 2.41-2.37 (m, 1H), 1.99-1.85 (m, 2H); 13C NMR

(CDCl3) 206.96, 206.02, 155.58, 155.13, 45.95, 40.98, 40.84, 30.05, 24.00, 21.37, 19.34.

tert-Butyl (1-hydroxy-2-oxopiperidin-3yl)carbamate (115). A solution of degassed

sodium hydroxide (2N, 80 mmol, 40 mL) was added to a solution of tert-butyl (2-

oxocyclopentyl)carbamate (113, Scheme 56) (0.6 g, 3.015 mmol) in EtOH (20 mL) at

0°C. To this mixture a solution of Piloty’s acid (1.044 g, 6.03 mmol) in EtOH (20 mL)

was added dropwise over 30 min and the reaction mixture was stirred at room

temperature for 18 h. The ethanol was removed under vacuum and the water layer was

extracted with diethyl ether to remove the unreacted ketone. The aqueous layer was

acidified to pH 5.5 with 2N HCl and extracted with CHCl3 (5 x 50 mL). The CHCl3 layer was dried with MgSO4 and concentrated to give a solid crude residue that was purified by flash chromatography to give 115 as a reddish solid (0.06 g, 4%) ; Rf = 0.24 (95:5 CHCl3

1 13 : MeOH). H NMR (CDCl3) 5.46 (b,1H), 5.24 (b, 1H), 2.475 (t, J = 5.61 Hz, 2H), C-

NMR (CDCl3) 165.85, 155.55, 79.7, 68.14, 34.46, 32.24, 30.52, 19.01, LCMS m/z 229

(M+).

N-hydroxy-N-propylacetamide. A solution of degassed sodium hydroxide (2N, 80

mmol, 40 mL) was added to a solution of 2-pentanone (1.13 g, 11.3 mmol) in EtOH (20

mL) at 0°C. To this reaction mixture, a solution of Piloty’s acid (3.91 g, 22.6 mmol) in

EtOH (20 mL) was added dropwise over 30 min and this mixture was stirred at 0°C for 4

h and for 18 h at room temperature. The ethanol was removed under vacuum and the

water layer washed with diethyl ether. The aqueous layer was acidified to pH 5.5 with 2N

HCl and extracted with CHCl3 (6 x 50 mL). After drying with MgSO4, the CHCl3 layer was concentrated to a solid crude residue and purified by flash chromatography to give the N-hydroxy-N-propylacetamide as a thick red liquid (.04 g, 3%); Rf = 0.44 ( EtOAc);

1 H NMR (CDCl3) 3.60 (t, J = 6.69 Hz, 2H), 2.1 (s, 3H), 1.76-1.62 (m, 2H), 1.45-1.28 (m,

2H), 0.95 (t, J = 7.23 Hz, 3H).

CHAPTER 3

PROGRESS TOWARDS THE SYNTHESIS OF THE COBACTIN CORE AND

MYCOBACTIC ACID UTILIZING NITROSO-ENE REACTION

Ring-expansion reactions of cyclic ketones through –NOH insertion fails to generate the seven-membered ring cobactin core (Scheme 55 and Scheme 56). As an alternative approach, this research focuses on investigating the nitroso-ene reaction for making a synthetic cobactin analog. The basics of the nitroso ene reaction have already been discussed in the context of making linear and cyclic hydroxamic acid analogs in the introduction.34 Our research goal is to utilize this reaction to synthesize cobactin and

mycobactic acid analogs. The specific aims of this project include investigating both the

intramolecular and intermolecular nitroso-ene reaction towards the formal synthesis of

cobactin and mycobactic acid.

3.1 Intramolecular Nitroso-ene Reaction Approach to the Synthesis of Cobactin Core

3.1.1 Retrosynthesis of Cobactin

A synthesis of the cobactin core (83, Scheme 57), a seven membered ring hydroxamic

acid residue of the natural product mycobactin S, would be approached using nitroso-ene

cyclization methodology. We envisioned that the cobactin core A could arise by

reduction of the unsaturated analog B, which could be synthesized in one step using an

intramolecular nitroso-ene cyclization of the linear hydroxamic acid C (Scheme 57).

Scheme 57. Retrosynthesis of cobactin using intramolecular nitroso-ene reaction

3.1.2 Intramolecular Nitroso-ene Reactons : Results and Discussion

The intramolecular nitroso-ene cyclization is one of the most useful tools to synthesize cyclic hydroxamic acid frameworks. While nitroso-ene reactions are immensely useful to make five or six membered ring hydroxamic acids, only a few reports exist for the seven- membered ring synthesis.65, 71, 86

To explore the proposed retrosynthetic route (Scheme 58), the initial group of experiments were directed to generate the proper chain length acyl nitroso species 116

(Scheme 58) and evaluate the nitroso-ene cyclization to construct a cobactin core.

Scheme 58. Proposed synthesis of cobactin core

To investigate this pathway with the α-unsubstituted acylnitroso species 116 (Scheme

58), the synthetic scheme began with the esterification of commercially available 5-

hexenoic acid (118, Scheme 59) to the corresponding methyl ester (119) by treatment of

methyl iodide and potassium carbonate in acetone. Treatment of 119 with excess hydroxylamine hydrochloride in base gave the corresponding hydroxamic acid (120,

Scheme 59).130 Oxidation of 120 with tetrabutylammonium periodate generates the

acylnitroso intermediate (116) which was trapped by 9, 10-dimethylanthracene (9,10-

O O O 1. NH OH HCl 1. K2CO3 (3eq.) 2 OH OMe NHOH 3 2. MeI (5eq.), 3 2. K2CO3,MeOH 3 118 Acetone 119 55% 120 35% 1. Bu4NIO4 2. 9,10-DMA 52% O O OH N N Reflux in benzene or toluene 3 + O or sealed tube heating 117 121 X

Scheme 59. Attempted synthesis of cobactin core

DMA) to give a Diels-Alder bicycloadduct (121). Thermolysis of the cycloadduct 121

did not produce the seven membered hydroxamic acid 117 (Scheme 59).

O N O H

Scheme 60. Nitroso-ene mechanism: Improper alignment

The failure of this cyclization may be due to the unfavorable deformation needed to

attain the intramolecular nitroso ene geometry required for the desired product (Scheme

60). The alkyl chain may not be long enough to allow proper alignment of the nitroso

species and the bridging alkene group in 116 (Scheme 58) for a productive nitroso-ene reaction.

In order to circumvent this problem, we examined an alternative pathway where the acyl nitroso species is derived from a 6-octenoic hydroxamic acid (124, Scheme 61). This

substrate has added flexibility with an extra methylene group and two carbon homologous alkyl chain (123, Scheme 61). Ozonolysis of cyclohexene using the procedure reported by Li et. al, yields 122 (Scheme 61) in 52% yield.131 Compound 122

was converted to the homologous two carbon (Z) alkene 123 by Wittig olefination.132

Treatment of 123 with

Scheme 61. Intramolecular nitroso-ene reaction of 126 generates eight membered ring

hydroxamic acids

74 excess hydroxylamine hydrochloride in the presence of excess K2CO3 afforded the hydroxamic acid (124) in 77% yield. Tetrabutyylammonium periodate oxidation of 124, followed by the Diels-Alder reaction with 9,10-DMA gave the cycloadduct (125, 56%,

Scheme 61). Thermolysis of the cycloadduct (125) in refluxing toluene yielded an unusual eight membered ring hydroxamic acid presumably as a nitroso-ene product 127

(Path a, Scheme 61) in 40% yield, while the formation of the vinyl substituted cobactin core 128 (Path b, Scheme 61) was not observedd. X-ray diffraction studies on crystals of

127 (Figure 4) confirms the structure, which is also supported by NMR and mass- spectroscopic data. While this intramolecular nitroso-ene reacction scheme doesn’t support the synthesis of a cobactin core, it provides a unique route to make eight membered ring hydroxamic acid.

Figure 4. X-ray diffraction structure of compound 127

3.2 Structure Elucidation of Mycobactin

The first structure elucidation of the hydroxamic acid-based siderophore mycobactin

was achieved by G. A. Snow in 1954.133 Amino acid-based hydroxamic acids are

important components in mycobactin. Mycobactin S exhibits growth inhibitory activity

against Mycobacterium tuberculosis and it consists of cobactin and mycobactic acid

(Scheme 62), both of which contain a hydroxamic acid residue derived from

NЄ-hydroxylysine. Mycobactin S is the closest structural relative of mycobactin T (found

in tuberculosis causing bacteria), differing in variable long chain substituents R1 (Scheme

62).76

Scheme 62. Structure elucidation of mycobactin S

3.3 Investigation of Intermolecular Nitroso-ene Approach towards the Synthesis of

Cobactin and Mycobactic Acid

Since the intramolecular nitroso-ene reactions failed to give an efficient formal synthesis of the cobactin core, we proposed an intermolecular nitroso-ene methodology 76

as an alternative. Scheme 63 shows this route to cobactin and mycobactic acid using the

intermolecular nitroso-ene reaction. The ene reaction between a Nα-protected homoallyl-

glycine ester 129 and a tert-butyl C-nitroso formate ester would generate the hydroxamic

acid (130, Scheme 63). Hydrogenation of 130 would yield the saturated hydroxamic acid

derivative (131) and deprotection by HCl would give the Nα-protected NЄ-hydroxylysine

(132, Scheme 63). A DCC-DMAP coupled cyclization should generate the seven membered ring hydroxamic acid (133, Scheme 63) as reported.76 This sequence provides

a short route to a key intermediate of cobactin synthesis. A similar synthetic route using

a long chain-derived acyl nitroso species supports the synthesis of a precursor of

mycobactic acid (84, Scheme 63).

Scheme 63. Intermolecular nitroso-ene reaction route to cobactin and mycobactic acid

3.3.1 Synthesis of the Precursor Alkene and Diels-Alder Cycloadduct ( Acyl-nitroso

Precursor) for a Nitroso-ene Reaction

To investigate the proposed synthetic route to cobactin (Scheme 63), we started with

the synthesis of a properly substituted alkene (135, Scheme 64) and a precursor of an acyl

nitroso compound (136, Scheme 64). Alkylation of the Schiff base protected glycine

ethyl ester with 4-bromo-1-butene (Scheme 64) gave the precursor alkene (ethyl 2-

((diphenylmethylene)amino)hex-5-enoate) in 89% yield.134 Tetrabutylammonium periodate oxidation of tert-butyl hydroxycarbamate followed by a Diels-Alder reaction with 9,10-DMA yielded the cycloadduct (136) as a precursor acyl nitroso species (71%,

Scheme 64).

Scheme 64. Synthesis of the precursor alkene and acyl nitroso component

3.3.2 Nitroso-ene Reaction

Intermolecular nitroso-ene reactions were performed by heating a solution of ethyl 2-

((diphenylmethylene)amino)hex-5-enoate and the precursor acylnitroso compound (136)

in toluene at reflux in a sealed tube for 4 h. This nitroso-ene reaction successfully generates the hydroxamic acid (137) in 45% yield (Scheme 65). The best results were obtained using an excess (4 eq.) amount of alkene to trap the in situ generated tert-butyl

C-nitrosoformate ester species (Scheme 65). Nitroso-ene reactions of terminal alkenes are reported to produce a mixture of E and Z regioisomers and these reactions appears to give inseparable mixtures of E/Z stereoisomers.63 The nitroso-ene product 137 (Scheme 65)

was isolated as a mixture of E/Z stereoisomers after chromatographic purification. As the

next synthetic step requires reduction, chromatographic separation or determination of the stereochemistry of the alkene (137) was not performed.

Scheme 65. Intermolecular nitroso-ene reaction

Hydrogenation of 137 in presence of Pd/C (10 wt. %) catalyst in MeOH reduced the olefin as well as the imino protecting group, giving a mixture of 138 and 139 (Scheme

66). Deprotection of the imino group of 137 with strong acid in MeOH yielded 140 which failed to generate the hydrogenated product (141, Scheme 66), as the unprotected hydroxylamine group may poison the hydrogenation catalyst. 79

Scheme 66. Results of hydrogenation on the nitroso-ene product

These results led to the design of an alkene with a different N-protecting group. N-

Acetyl homoallyl glycine ester (145, Scheme 67) was synthesized from diethyl acetamidomalonate (142) by alkylation with 1-bromo butene followed by β-keto- decarboxylation and esterification of the free acid 144 (Scheme 67).135 The nitroso-ene reaction of 145 with the cycloadduct (136, Scheme 67) afforded 146 in 46% yield as a mixture of E/Z stereoisomers after chromatographic separation. Hydrogenation of 146

with Pd/C catalyst yielded the saturated hydroxamic acid (147, 89%, Scheme 66).

However, acid hydrolysis failed to generate the free acid needed for cyclization from 147

and only produced the hydroxylamine (148).

Scheme 67. Synthesis and nitroso-ene reactions of N-acetyl homo allyl glycine ethyl ester

The precursor alkene (144) was modified to an acid cleavable N-acetyl tert-butyl

homoallyl glycine ester (149, Scheme 68) using a published method.70 The nitroso-ene

reaction of 149 with the cycloadduct (136) under identical conditions afforded the

hydroxamic acid (150, Scheme 68) as a mixture of E and Z stereoisomers in 46% yield.

Hydrogenation of 150 under 2.5 atm pressure with Pd/C (10 wt.%) catalyst led to the

saturated hydroxamic acid (151, Scheme 68, 89%). Treatment of 151 with TFA in

CH2Cl2 (1:1) at room temperature for 3 h yields the deprotected precursor hydroxylamine

(152, Scheme 68), which failed to generate a seven membered cobactin core 153 after

either a DCC-DMAP (dicyclohexylcarbodiimide - dimethyl aminopyridine) or EDC–

HOAt (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide – 1-hydroxy-7-aza benzotriazole) coupled cyclization, both methods reported by Miller.60, 136 Nevertheless, these results show that the intermolecular nitroso-ene reactions provides a short route to generate the key intermediate 152, which could be potentially useful to make the cobactin core.

Scheme 68. Synthetic approach to the cobactin core using an intermolecular nitroso ene reaction

The nitroso-ene product (150, Scheme 68), with its high polarity and iron chelating ability, was difficult to purify by chromatography. Moreover mass-spectroscopic data

(see experimental) reveals 152 also binds iron strongly. To make a less polar non- chelating tert-Butyl dimethyl silyl (TBDMS) protected analog of 152, 150 was treated with TBDMSCl in presence of imidazole as base to give the O-protected hydroxamic acid (154, Scheme 69). Hydrogenation of 154 under 1.5 atm. pressure with Pd/C (10 wt.

%) yielded the saturated O-protected hydroxamic acid (155, Scheme 69), which failed to produce the TBDMS protected hydroxylamine (156) under mild orthophosphoric acid deprotection condition.

Scheme 69. Attempted synthesis of TBDMS protected hydroxylamine

3.3.3 Progress Towards the Synthesis of Mycobactic Acid Utilizing a Nitroso-ene

reaction

Mycobactic acid consists of a long chain derived [R1 = -(CH2)12CH3] N-substituted

hydroxamic acid along with a 2-hydroxyphenyl oxazoline residue. Both residues are

essential for bacterial siderophore activity and the long alkyl chain is important for lipid

solubility of mycobactins during iron transport through the cell membrane.102

Intermolecular nitroso-ene reactions between the precursor alkene (149, Scheme 70) and a long chain derived acyl nitroso species provides a key intermediate 161 towards the synthesis of mycobactic acid 84 (Scheme 63). The synthesis of the long chain N-

Scheme 70. Intermolecular nitroso-ene route to formal synthesis of mycobactic acid

substituted hydroxamic acid residue started with the acyl nitroso precursor (159, Scheme

70). Ethyl myristate (157, Scheme 70) was converted to N-hydroxy tetradecanamide

(158, Scheme 70) by treatment with a large excess of hydroxylamine hydrochloride and potassium hydroxide in EtOH at room temperature. Tetrabutylammonium periodate

84 oxidation of 158 and Diels-Alder reaction with 9,10-DMA gave the cycloadduct (159,

Scheme 70). Thermolysis and concomitant nitroso-ene reaction of 159 with 149 (Scheme

70) under identical conditions for 6 h provided the hydroxamic acid (160) in 23% yield.

After chromatographic purification, 160 appeared to be a mixture of E and Z stereoisomers. Hydrogenation of 160 gave the saturated long chain hydroxamic acid

(161), which is a precursor of mycobactic acid (84, Scheme 70).

Since the long chain derived acyl-nitroso-ene product 160 is difficult to chromatograph and this complicated purification led to a poor yield, we tried protecting the hydroxamic acid as a TBDMS ester. tert-Butyl dimethylsilyl (TBDMS) protection of the hydroxamic acid group of cobactin has been reported by Miller.76 Using this method, treatment of 160 with TBDMSCl resulted in a stable O-protected nitroso-ene product

(162, Scheme 71) in 30% yield with improved chromatographic purification still as a mixture of E/Z stereoisomers.

Scheme 71. TBDMS protection of hydroxamic acid group in long chain derived nitroso-ene product

3.3.4 The Nitroso-ene Chemistry of Long Chain-Derived Acyl Nitroso Species

The next set of experiments verifies the reactivity of the long chain-derived acyl nitroso species. Treatment of the long chain derived acyl-nitroso cycloadduct (159, Scheme 72) with cyclohexene as solvent under identical nitroso-ene reaction conditions afforded 163

(Scheme 72) in 33% yield, further proving the ability of the long chain- derived acyl nitroso species to undergo nitroso-ene reactions, however the yield remained below 40%.

Scheme 72. Nitroso-ene reaction of a long chain-derived acyl nitroso species with

cyclohexene

3.4 Nitroso-ene Reactions of Acetyl-Nitroso Compounds

A small acyl nitroso compound was screened as an enophile in the nitroso-ene reaction with the N-acetyl tert-butyl homoallyl glycine ester (149, Scheme 73).

Tetrabutylammonium periodate oxidation of acetohydroxamic acid generates the acetyl nitroso intermediate that reacts with 9,10-DMA to give a Diels-Alder cycloadduct (164,

Scheme 73). The nitroso-ene reaction between 164 and alkene (149) under identical nitroso-ene conditions afforded the hydroxamic acid after chromatographic purification

(165, 29%, Scheme 73), as a mixture of E and Z stereoisomers. The nitroso-ene product

(165) being highly polar, complicated purification and resulted in low isolation. TBDMS protection of the hydroxamic acid (165) gave the O-protected product (166, 33%, Scheme

73) with ease in purification, still as a mixture of E/Z stereoisomers.

Scheme 73. Nitroso-ene reactions of acyl-nitroso species with the precursor alkene 149

3.5 Nitroso-ene reactions of Benzoyl nitroso compounds

Benzoyl nitroso compounds were screened to examine the scope of the nitroso-ene reactions. Oxidation of benzohydroxamic acid with tetrabutylammonium periodate gave the benzoyl nitroso intermediate and the in situ Diels-Alder reaction with 9,10-DMA yields the cycloadduct (167, 53%, Scheme 74). The nitroso-ene reaction between 167 and the alkene (149) generates the hydroxamic acid (168, 20%, Scheme 74) as a mixture of

E/Z stereoisomers. These results demonstrate poor nitroso-ene ability of the benzoyl nitroso species.

Scheme 74. The nitroso-ene reaction of benzoyl nitroso species

Given the research goal of a short synthetic route to cobactin core and mycobactic acid using nitroso-ene reactions, we wanted to examine the general ability of the acyl nitroso compounds to undergo ene reactions. The last two experiments reveal the scope of the nitroso-ene reactions of a short acetyl-nitroso and benzoyl nitroso intermediates, with benzoyl nitroso compounds being the least efficient. In each case the hydroxamic acids (165 and 168) are difficult to purify and complicated chromatographic separation results in low yield. TBDMS protection of the hydroxamic acid in 165 facilitates purification, while it has not been attempted for 168. Previous results towards cobactin and mycobactic acid synthesis suggest the tert-butyl C-nitroso formate esters show the best nitroso-ene reactivity with 149, while the long chain derived acyl nitroso intermediates are not very efficient enophiles. This failure to afford an efficient nitroso- ene reaction could be attributed to the fact that the folding of the long chain alkyl group may hinder the acyl nitroso fragment accessing the alkene, thus affecting its nitroso-ene ability. In general acyl nitroso species are transient, exceptionally reactive and

electrophiles.137 All nitroso-ene reactions were performed in non-nucleophilic solvents like toluene and benzene under inert atmosphere. This study shows the nitroso-ene reactivity profile of a series of acyl nitroso intermediates with terminal alkenes generating a series of N-substituted hydroxamic acids, which could further be useful for synthesizing various substituted hydroxylamine derivatives.

3.6 Experimental

General. Analytical TLC was performed on silica gel plates with QF-254 indicator.

Visualization was accomplished with UV light, FeCl3, bromocresol green, potassium permanganate, phosphomolybdic acid, and/or dinitrophenylhydrazine stain. Extraction and chromatography solvents were technical grade. All reactions were performed under

1 13 an inert atmosphere of dry argon. H NMR and C NMR were recorded in CDCl3 and deuterated DMSO on a Bruker Avance 300 MHz and 500 MHz NMR Spectrometer.

Chemical shifts are given in ppm (δ); multiplicities are indicated by s (singlet), d

(doublet), t (triplet), q (quartet), m (multiplet) and b (broadened). Low resolution mass spectra were obtained using an Agilent LCMS system consisting of an 1100 LC/MSD detector or from the UNC Mass Spectrometry Laboratories (UNC-Chapel Hill, NC) using a BioToF-NT instrument and data are reported in m/z.

1-(9,10-dimethyl-9,10-dihydro-9,10-(epoxyimino)anthracen-11-yl)hex-5-en-1-one

(121). A solution of 120 (0.133 g, 1.03 mmol) in CHCl3 (9 mL) was added dropwise to a solution of 9, 10-dimethylanthracene (0.53 g, 2.57 mmol) and Bu4NIO4 (0.49 g, 1.13 mmol) in CHCl3 (13 mL). The solution was stirred for 90 min and poured into aq.

Na2S2O3 solution (20 mL) and extracted with CH2Cl2 (2 x 20 mL). The extracts were dried over MgSO4 and purified by flash chromatography (15% EtOAc : 85% pet.ether,

1 Rf = 0.74) to give as 121 a white solid ( 0.18 g, 52%); H-NMR (CDCl3, 300 MHz)

7.49-7.46 (m, 2H), 7.38-7.36 (m, 2H), 7.30-7.26 (m, 4H), 5.75-5.54 (m, 1H), 4.91-4.79

(m, 2H), 2.71 (s, 3H), 2.23 (s, 3H), 2.16 (t, J = 7.42 Hz, 2H), 1.81 (q, J = 7.16 Hz,

13 2H), 1.55 (s, 2H), 1.39 (p, J = 7.35 Hz, 2H); C-NMR (CDCl3, 300 MHz) 178.52,

140.54, 140.42, 138.07, 138.00, 126.9, 126.59, 120.09, 120.01, 113.94, 113.89, 79.60,

64.15, 36.74, 34.12, 24.15, 17.66, 15.92.

Methyl-6-oxo-hexanoate (122).138 Ozone was bubbled through a mixture of cyclohexene (2.46 g, 3.04 mL, 30 mmol) and anhydrous sodium carbonate (0.82 g, 7.74

91 mmol) in CH2Cl2 (90 mL) and methanol (18 mL) was added at -78°C until a faint blue color appeared.139, 140 Argon was bubbled through the mixture until the blue color discharged. The cooling bath was removed and the mixture was allowed to warm-up to room temperature. After filtration, benzene (30 mL) was added and the mixture was concentrated to a volume of ~20 mL. The resulting viscous liquid was diluted with

CH2Cl2 (80 mL) and cooled to 0°C. Et3N (4.5 g, 6.2 mL, 44.48mmol) and Ac2O (8.53 g

7.59 mL, 83.57 mmol) were sequentially added dropwise and the mixture was stirred at

0°C for 30 min and then at room temperature overnight. The organic phase was washed with 0.1M aq. HCl (2 x 60 mL), 10% aq. NaOH (2 x 60 mL), water (60 ml) and dried over MgSO4. After filtration, solvent was removed under vacuum and the product was isolated as a crude oil and purified by flash chromatography (30% EtOAc : 70% pet. Ether, Rf = 0.65) to give a colorless oil (2.25 g, 52%) with spectroscopic and analytical details were identical to those reported.138

(Z)-Methyl oct-6-enoate (123). A NaHMDS solution in THF (1.1 g, 6 mmol) was added to a suspension of ethyl triphenyl phosphonium bromide (1.86 g, 5 mmol) in distilled THF (50 mL) under nitrogen and was stirred at room temperature for 30 min.

After 30 min, the solution color changed from red to orange, and the solution was cooled to -80°C and methyl-6-oxohexanoate (0.72 g, 5 mmol) was added. After 16 h at room temperature, the solution was heated to reflux for 3 h, the THF was evaporated

92 under vacuum and water (20 mL) added to the residue. The mixture was extracted with

Et2O (3 x 50mL) and dried over magnesium sulfate to give a crude mixture that was purified by flash chromatography (25% Et2O:75% pet. ether, Rf = 0.95) to give 123 as a

1 colorless volatile liquid (0.45 g, 58%); H-NMR (CDCl3, 300 MHz) 5.54-5.34 (m, 2H),

3.7 (s, 3H), 2.35 (t, J = 7.5 Hz, 2H), 2.08 (q, J = 7.35 Hz, 2H), 1.74-1.6 (m, 5H), 1.47-

13 1.37 (m, 2H); C-NMR (CDCl3, 300 MHz) 173.36, 129.78, 123.96, 52.12, 34.88,

29.95, 27.4, 25.56, 14.49.

(Z)-N-Hydroxyoct-6-enamide (124). A solution of potassium hydroxide (12.12 g.,

216 mmol) in MeOH (70 mL) was added dropwise to a solution of hydroxylamine hydrochloride (7.5 g, 108 mmol) in MeOH (70 mL) at 0°C. This solution was stirred for 20 min and a solution of methyl-5-heptenoate (1.69g, 10.8 mmol) in MeOH (35 mL) was added dropwise and stirred for 2 h. Water (45 mL) was added and the pH of the solution adjusted to 6 with conc. HCl. This acidified solution was extracted with

CHCl3 (3 x 50 mL), dried over MgSO4 and the solvent was removed under vacuum. The crude mixture was purified by flash chromatography (1:1 EtOAc : pet. ether, Rf = 0.48)

1 to give 124 as a light brown thick liquid (1.3 g, 77%); H-NMR (CDCl3, 500MHz) 9.03

(b, 1H), 5.49-5.28 (m, 2H), 2.16 (t, J = 7.36 Hz, 2H), 2.04 (q, J = 7.21 Hz, 2H), 1.68-

13 1.58 (m, 5H), 1.42-1.31 (m, 2H); C-NMR (CDCl3, 300 MHz) 172.44, 130.99, 125.49,

35.19, 31.27, 28.77, 27.39, 15.28. Homonuclear decoupling NMR experiment revealed the Z configuration of the alkene (124).

(Z)-1-(9,10-Dimethyl-dihydro-9,10-(epoxyimino)anthracen-11-yl)oct-6-en-1-one

(125). A solution of (Z)-N-hydroxy-6-octenamide (124, 0.338 g, 2.15 mmol) in CHCl3

(9 mL) was added dropwise to a solution of 9, 10-dimethylanthracene (1.11 g, 5.38 mmol) and Bu4NIO4 (1.02 g, 2.36 mmol) in CHCl3 (13 mL). The solution was stirred for 90 min and poured into an aq. Na2S2O3 solution (20 mL) and extracted with CH2Cl2

(2 x 20 mL). The extracts were dried over MgSO4, solvent was removed under vacuum and purified by flash chromatography (15% EtOAc : 85% pet.ether, Rf = 0.78) to give

1 125 as a white solid ( 0.438 g, 56%); H-NMR (CDCl3, 300 MHz) 7.51-7.48 (m, 2H),

7.41-7.37 (m, 2H), 7.32-7.27 (m, 4H), 2.74 (s, 3H), 2.27 (s, 3H), 2.19 (t, J = 7.17 Hz,

2H), 1.96-1.89 (dd, J = 7.32, 7.07 Hz, 2H), 1.56 (d, J = 6 Hz, 3H), 1,38-1.28 (m, 2H),

13 1.2-1.1 (m, 2H); C-NMR (CDCl3, 300 MHz) 180.07, 142.17, 142.06, 131.50, 128.56,

128.25, 124.93, 122.58, 121.75, 81.25, 65.78, 38.50, 31.20, 28.93, 25.91, 19.32, 17.58,

15.28.

(Z)-1-Hydroxy-8-methyl-1,4,5,8-tetrahydroazocin-2(3H)-one (127). A solution of

125 (0.198 g, 0.54 mmol) in toluene (250 mL) was refluxed for 20 min until TLC

95 revealed a high Rf UV active spot corresponding to 9,10-DMA and a broad polar spot that turned purple in FeCl3 stain. After solvent evaporation under vacuum, the remaining crude yellow solid was purified by flash chromatography (a gradient of 35%

EtOAc : 65% pet. ether to 95% CHCl3: 5% MeOH, Rf = 0.22) to give a reddish brown thick liquid that was further purified by sublimation at 45°C under reduced pressure

1 (0.05 mm) to give 127 as a white crystalline solid (0.026 g, 30%); H-NMR (CDCl3,

300 MHz) 7.87 (b, 1H), 5.86-5.69 (m, 2H), 4.82-4.73 (m, 1H), 2.68-2.6 (m, 2H), 2.45-

13 2.25 (m, 2H), 1.94-1.84 (m, 1H), 1.5 (d, J = 6.75 Hz, 3H), C-NMR (CDCl3, 300

MHz) 172.35, 134.66, 130.70, 53.39, 34.54, 28.38, 23.66, 18.72. HRMS (ESI) m/z

156.1025 (M+H)+ Expected 155.0946

tert-Butyl-9,10-dimethyl-9,10-dihydro-9,10-(epoxyimino)anthracene-11-carboxy latE (136).70 A solution of tert-butyl N-hydroxycarbamate (0.5 g, 3.75 mmol) in DMF

(5 mL) was added to a solution of 9,10-dimethylanthracene (0.516 g, 2.5 mmol) and tetrabutylammonium periodate (1.625 g, 3.75 mmol) in CHCl3 (13 mL) at 5°C. This solution was allowed to warm to room temperature, stirred for 24 h, poured into ethyl acetate (100 mL) and washed with saturated sodium thiosulphate, brine and water. The organic extract was dried over MgSO4 and filtered to give a crude yellow solid that was purified by flash chromatography (5% EtOAc : 95% pet.ether, Rf = 0.38) to give an off

96 white solid 136 (0.6 g, 71%) with spectroscopic and analytical details identical to those reported.70

Ethyl-6-((tert-butoxycarbonyl)(hydroxyl)amino)-2-((diphenylmethylene)amino) hex-4-enoate (137). A solution of ethyl 2-((diphenylmethylene)amino)hex-5-enoate

(135, 2.65 g, 8.26 mmol) and 136 (0.696 g, 2.07 mmol) in toluene (5 mL) was heated in a sealed tube to reflux for 4 h. The reaction mixture was cooled in an ice bath to crystallize 9,10-DMA, which was filtered. The dark brown filtrate showed a crude mixture of the excess alkene and a hydroxamic acid by TLC. The crude mixture was purified by flash chromatography (25% EtOAc : 75% pet. ether, Rf = 0.26) to give 137

1 as a thick red liquid (0.417 g, 45%); H-NMR (CDCl3, 300 MHz) 7.63-7.02 (m, 10H),

5.64-5.49 (m, 2H), 4.22-4.09 (m, 4H), 3.98-3.96 (m, 1H), 2.71-2.57 (m, 2H), 1.47-1.44

13 (m, 10H), 1.28-1.23 (t, J = 7.14, 3H); C-NMR (CDCl3, 500 MHz) 172.12, 171.16,

157.18, 139.83, 136.70, 130.91, 130.74, 129.27, 129.16, 129.03, 128.86, 128.41,

128.17, 128.12, 127.23, 82.20, 65.60, 61.33, 52.64, 37.01, 28.64, 28.59, 28.53, 14.55.

MS (ESI) m/z 453 (M)+.

Ethyl-6-((tert-butoxycarbonyl)(hydroxy)amino)-2-((diphenylmethylene)amino) hexanoate (139). A solution of 137 (0.1 g, 0.22 mmol) in MeOH (20 mL) was slowly added to 10 wt. % Pd/C (0.009 g) in a hydrogenation flask. This reaction mixture was subjected to hydrogenation using a Parr-hydrogenator under 2.5 atm. pressure for 24 h.

The reaction mixture was filtered through celite and the filtrate concentrated to give a crude mixture of hydroxamic acid products (TLC revealed two violet spots in FeCl3).

The crude product was purified by flash chromatography (25% EtOAc : 75% pet. ether,

1 Rf = 0.42) to give 139 as a thick red liquid (0.02 g, 20%); H-NMR (CDCl3, 500 MHz)

7.92-7.16 (m, 10H), 4.80 (s, 1H), 4.24-4.18 (q, J = 7.22Hz, 2H), 3.54-3.43 (m, 1H),

13 3.24-3.17 (m, 1H), 1.77-1.16 (m, 18H); C-NMR (CDCl3 , 500 MHz) 172.12, 171.17,

157.17, 139.84, 136.70, 130.90, 130.75, 129.16, 128.86, 128.41, 128.17, 128.12,

127.23, 82.21, 65.6, 61.33, 52.64, 37.01, 28.597, 14.54; MS (ESI) m/z 457 (M+2)+

Ethyl 2-amino-6-(hydroxyamino)hex-4-enoate (140). A solution of HCl (1.25M) in

MeOH (16 mL) was added to 139 (0.441 g, 1.33 mmol) and stirred at room temperature for 12 h. The solution was concentrated under vacuum, dissolved in water (5 mL), and extracted with diethyl ether (20 mL). The water layer was lyophilized to give 140 as a dark brown liquid (0.076 g, 30%). 1H-NMR (MeOD, 300 MHz) 6.09-5.95 (m, 1H),

5.92-5.78 (m, 1H), 4.33 (q, J = 7.14, 2H), 4.2 (t, J = 6.3, 1H), 3.91-3.82 (m, 2H), 2.91-

13 2.67 (m, 2H), 1.33 (t, J = 7.12, 3H); C-NMR (CDCl3 , 300 MHz) 168.11, 133.77,

123.87, 63.68, 53.52, 53.21, 34.51, 14.83. MS (ESI) m/z 189 (M)+.

Ethyl 2-acetamido-6-((tert-butoxycarbonyl)(hydroxy)amino)hex-4-enoate (146). A solution of ethyl 2-acetamidohex-5-enoate (145, 1.64 g, 8.24 mmol) and 136 (0.634 g,

2.059 mmol) in toluene (5 mL) was heated to reflux for 4 h in a sealed tube. The reaction mixture was cooled in an ice bath to crystallize the 9, 10-DMA, which was filtered. The reddish brown filtrate showed a crude mixture of the excess alkene and a

99 hydroxamic acid by TLC. The crude mixture was purified by flash chromatography

(1:1 EtOAc : pet. ether, Rf = 0.19) to yield 146 as a thick brown liquid (0.3 g, 45%);

1 H-NMR (CDCl3, 300 MHz) 6.18 (d, J = 7.85 Hz, 1H), 5.7-5.49 (m, 2H), 4.76-4.58 (m,

1H), 4.21 (q, J = 7.14 Hz, 1H), 4.11-3.9 (m, 2H) 2.69-2.55 (1H, m), 2.47-2.34 (m, 1H),

13 2.01 (s, 3H), 1.49 (s, 9H), 1.286 (t, J = 7.13 Hz, 3H); C-NMR (CDCl3, 500 MHz)

172.13, 170.64, 157.10, 129.17, 128.57, 82.14, 62.05, 52.35, 52.22, 36.02, 28.66, 23.53,

14.52; HRMS (ESI) m/z (M+Na)+ 353.1689 Expected 330.3767.

Ethyl 2-acetamido-6-((tert-butoxycarbonyl)(hydroxy)amino)hexanoate (147). A solution of 146 (0.175 g, 0.53 mmol) in MeOH (34 mL) was slowly added to 10 wt. %

Pd/C (0.02 g) in a hydrogenation flask. This reaction mixture was subjected to hydrogenation using a Parr-hydrogenator under 2.5 atm. pressure for 24 h. The reaction mixture was filtered through celite and the isolated extract gave pure product (147)

1 after solvent evaporation (0.172 g, 98%); H-NMR (CDCl3, 300 MHz) 6.22 (d, J = 8.08

Hz, 1H), 4.69-4.53 (m, 1H), 4.175 (q, J = 7.15 Hz, 2H), 3.61-3.35 (m, 2H), 2.02 (s,

13 3H), 1.94-1.32 (m, 15H), 1.27 (t, J = 7.13, 3H); C-NMR (CDCl3, 500 MHz) 172.85,

170.53, 157, 81.50, 61.66, 51.76, 49.71, 32.47, 28.47, 26.14, 23.24, 22.12, 14.28.

100

tert-Butyl-2-acetamidohex-5-enoate (149). A solution of 2-acetamidohex-5-enoic acid

(144, 0.495 g, 2.89 mmol)135 in dry toluene (20 mL) was added to N, N-dimethyl formamide di-tert butyl acetal (2.78 mL, 11.608 mmol) at 80°C. The solution was stirred at 80°C for 20 min and allowed to cool to room temperature.127 The reaction mixture was concentrated under reduced pressure and purified by flash chromatography (1:1 EtOAc : pet.ether, Rf = 0.46) to give 149 as a colorless oil (0.48 g,

1 73%) that solidified in the freezer. H-NMR (CDCl3, 300 MHz) 6.02 (d, J = 7.42 Hz,

1H), 5.86-5.73 (m, 1H), 5.12-4.95 (m, 2H), 4.56-4.49 (m, 1H), 2.15-1.87 (m, 5H), 1.81-

13 1.67 (m, 2H), 1.48 (s, 9H); C-NMR (CDCl3, 300 MHz) 170.75, 168.64, 136.64,

115.10, 82.25, 52.76, 32.67, 32.6, 30.08, 28.74, 24.05. MS (ESI) m/z 227 (M)+.

tert-Butyl-2-acetamido-6-((tert-butoxycarbonyl)(hydroxyl)amino)hex-4-enoate

(150). A solution of tert-butyl-2-acetamidohex-5-enoate (149) (2.42 g, 10.68 mmol) and 136 (0.9 g, 2.67 mmol) in toluene (5 mL) in a sealed tube was heated to reflux for 4

101 h. The reaction mixture was cooled in an ice bath to crystallize the 9,10-DMA, which was filtered. The dark brown filtrate showed a crude mixture of the excess alkene and a hydroxamic acid by TLC. The crude mixture was purified by flash chromatography

1 (1:1 EtOAc : pet. ether, Rf = 0.5) to yield 150 as a thick brown liquid ( 0.4 g, 41%); H-

NMR (CDCl3, 300 MHz) 6.18 (d, J = 7.63 Hz, 1H), 5.73-5.51 (m, 2H), 4.65-4.48 (m,

1H), 4.17-3.91 (m, 2H), 2.68-2.53 (m, 1H), 2.42-2.3 (m, 1H), 2.01 (s, 3H), 1.48 (s, 9H),

13 1.46 (s, 9H) ; C-NMR (CDCl3, 300 MHz) 170.15, 169.51, 156.2, 128.38, 127.83,

82.73, 81.86, 52.87, 52.74, 36.5, 29.21, 28.89, 24.15; MS (ESI) m/z 391 (M+Na)+.

tert-Butyl 2-acetamido-6-(tertbutoxycarbonyl)(hydroxyamino)hexanoate (151). A solution of 150 (0.458 g, 1.28 mmol) in MeOH (40 mL) was slowly added to 10 wt. %

Pd/C (0.051 g) in a hydrogenation flask. This reaction mixture was subjected to hydrogenation using a Parr-hydrogenator under 2.5 atm. pressure for 24 h. The reaction mixture was filtered through celite and the filtrate gave 151 after solvent evaporation

1 (0.455 g, 99%); H-NMR (CDCl3, 300 MHz) 6.12 (d, J = 8.11 Hz, 1H), 4.58-4.49 (m,

1H), 3.6-3.36 (m, 2H), 2.02 (s, 3H), 1.88-1.69 (m, 2H), 1.64-1.56 (m, 1H), 1.47-1.33

13 (m, 21H); C-NMR (CDCl3, 300 MHz) 170.9, 169.29, 159.99, 82.35, 81.41, 52.42,

50.03, 33.29, 29.1, 28,74, 26.71, 24.03, 22.71; HRMS (ESI) m/z 383.2158 (M+Na)+

Expected 360.2260.

102

tert-Butyl-2-acetamido-6-(hydroxyamino)hexanoic acid (152). A solution of

TFA/CH2Cl2 (5.5mL / 5.5mL) was added to 151 (0.441g, 1.33 mmol) and stirred at room temperature for 1.5 h. The solution was concentrated under vacuum, dissolved in water (5mL), and extracted with diethyl ether (20mL). The water layer was lyophilized to give 152 as a foamy brown solid (0.25 g, 92%); 1H-NMR (MeOD, 300 MHz) 4.5-4.4

(m, 1H), 3.29-3.23 (t, J = 7.67 Hz, 2H), 2.07-1.25 (m, 6H); 13C-NMR (MeOD,

300MHz) 173.64, 172.06, 53.32, 51.84, 32.24, 24.37, 24.15, 22.68, HRMS (ESI) m/z

206.1561 (M+2)+ Expected (M)+ 204.1110, MS (ESI) m/z 204 (M)+, 260 (M+Fe)+.

tert-Butyl-2-acetamido-6-((tert-butoxycarbonyl)((tert-butyldimethylsilyl)oxy) amino) hex-4-enoate (154). A solution of 150 (0.325 g, 0.91 mmol) in DMF (5 mL) was added to a mixture of TBDMSCl (0.34 g, 2.26 mmol) and imidazole (0.306 g, 4.5 mmol).76 After stirring the reaction mixture overnight at 35°C, EtOAc (40 mL) was added and the organic layer was washed with water and brine, dried over MgSO4 and

103 filtered to give a brown crude product that was purified by flash chromatography (25%

EtOAc :75% pet. ether, 0.3) to yield 154 as a colorless liquid (0.17g, 40%); 1H-NMR

(CDCl3, 500 MHz) 5.87 (d, J = 7.82 Hz, 1H), 5.56-5.47 (m, 1H), 5.41-5.31 (m, 1H),

4.43-4.41 (m, 1H), 3.88-3.69 (m, 2H), 2.40-2.35 (m, 2H), 1.89 (3H, s), 1.36-1.34 (m,

13 18H), 0.83 (s, 9H), 0.04 (s, 6H); C-NMR (CDCl3, 300 MHz) 170.866, 169.76, 158.45,

128.95, 128.42, 82.42, 81.71, 55.1, 52.39, 35.38, 28.58, 28.39, 26.18, 23.61, 18.04, -

4.74; MS (ESI) m/z 497 (M+Na)+.

tert-Butyl-2-acetamido-6-((tert-butoxycarbonyl)((tert-butyldimethylsilyl)oxy) amino) hexanoate (155). A solution of 154 (0.14 g, 0.296 mmol) in MeOH (20 mL) was slowly added to 10 wt. % Pd/C (0.016 g) in a hydrogenation flask. This reaction mixture was subjected to hydrogenation using a Parr-hydrogenator under 2.5 atm. pressure for 24 h. The reaction mixture was filtered through celite and the filtrate

1 concentrated to give 155 (0.135 g, 96%) after solvent evaporation ; H-NMR (CDCl3,

300 MHz) 6.01 (d, J = 7.74 Hz, 1H), 4.50-4.44 (m, 1H), 3.42-3.37(m, 2H), 2.0 (s, 3H),

1.89-1.74 (m, 1H), 1.75-1.57 (m, 3H), 1.47 (s, 9H), 1.45 (s, 9H), 1.35-1.24 (m, 2H),

13 0.94 (s, 8H), 0.14 (s, 5H), 0.002 (s, 1H); C-NMR (CDCl3, 300 MHz) 170.73, 166.65,

104

157.04, 82.09, 81.17, 53.04, 53.04, 52.45, 33.06, 28.98, 28.73, 26.57, 24.02, 23.16,

18.64, -3.95; HRMS (ESI) m/z 497.3023 (M)+ expected 474.3125.

N-Hydroxytetradecanamide (158). A solution of KOH (4.49 g, 80 mmol) in EtOH

(50 mL) was added dropwise to a solution of hydroxylamine hydrochloride (2.64 g, 80 mmol) in EtOH (50 mL) at 0°C. This reaction mixture was stirred for 30min and ethyl myristate (2.048 g, 8 mmol) in ethanol (25 mL) was added dropwise. After 4 h the

EtOH was removed under vacuum and water (30 mL) was added into it. The aqueous solution was neutralized to pH 6 with 2N HCl and extracted with EtOAc : CHCl3

(150:150 mL). The organic layer was concentrtated under vacuum and purified by flash chromatography (1:1 EtOAc : hexane, Rf = 0.44) to give 158 as a white flaky solid

(0.68 g, 35%); 1H-NMR (MeOD, 300 MHz) 2.08 (t, J = 7.57 Hz, 2H), 1.68-1.53 (m,

2H), 1.29 (b, 20H), 0.93-0.88 (m, 3H); 13C-NMR (MeOD, 300 MHz) 171.50, 33.99,

33.27, 30.99, 30.85, 30.70, 30.63, 30.51, 30.41, 27.06, 24.04, 14.86.

105

1-(9,10-Dimethyl-9,10-dihydro-9,10-(epoxyimino)anthracen-11-yl)tetradecan-1- one (159). A solution of 158 (0.329 g, 1.357 mmol) in DMF (5 mL) was added dropwise to a solution of 9, 10-dimethylanthracene (0.7 g, 3.39 mmol) and Bu4NIO4

(0.644 g, 1.487 mmol) in CHCl3 (13 mL). The solution was stirred for 90 min and then poured into a Na2S2O3 solution (20 mL) and extracted with EtOAc : CHCl3 (100:100 mL). The extracts were dried over MgSO4, concentrated under vacuum to a yellow solid residue and purified by flash chromatography (silica gel, 5% EtOAc : 95%

1 pet.ether, Rf = 0.69) to give 159 as a white solid (0.35 g, 58%); H-NMR (CDCl3, 300

MHz) 7.47-7.41 (m, 2H), 7.37–7.32 (m, 2H), 7.26-7.20 (m, 4H), 2.7 (s, 3H), 2.21 (s,

13 3H), 2.13 (t, J = 7.36 , 2H), 1.24 (b, 22H), 0.89-0.85 (m, 3H); C-NMR (CDCl3, 300

MHz) 179.89, 141.35, 141.23, 127.56, 127.24, 121.51, 120.67, 79.65, 63.99, 36.50,

32.07, 29.82, 29.80, 29.75, 29.57, 29.55, 29.50, 29.15, 24.02, 22.84, 16.99, 15.23,

14.29; MS (ESI) m/z 446 (M)+.

106

tert-Butyl 2-acetamido-6-(N-hydroxyteradecanamido)hex-4-enoate (160). A solution of tert-butyl 2-acetamidohex-5-enoate (149, 0.198 g, 0.88 mmol) and 159

(0.98 g, 0.22 mmol) in toluene (5 mL) in a sealed tube was heated to reflux for 4 h. The reaction mixture was cooled in an ice bath to crystallize the 9,10-DMA, which was filtered. The brown filtrate showed a crude mixture of the excess alkene and a hydroxamic acid by TLC. The crude mixture was purified by flash chromatography

(eluted with a gradient of 1:1 EtOAc : pet.ether to 95% CHCl3 : 5% MeOH, Rf = 0.35

1 in EtOAc) to yield 160 as a brown liquid product (0.022 g, 23%); H-NMR (CDCl3,

500 MHz) 6.01-6.55 (m, 1H), 5.95-5.23 (m, 2H), 4.9-4.45 (m, 1H), 4.32-3.57 (m, 2H),

2.69-1.96 (m, 6H), 1.71-1.57 (m, 2H), 1.57-1.41 (m, 9H), 1.39-1.13 (m, 21H), 0.9 (t, J

13 = 6.8 Hz, 3H); C-NMR (CDCl3, 500 MHz) 175.98, 173.13, 171.05, 169.73, 130.25,

127.92, 127.02, 114.01, 82.57, 68.79, 52.378, 41.95, 41.18, 36.93, 36.093, 35.61, 32.06,

29.81, 29.79, 29.74, 29.61, 29.47, 29.38, 29.28, 28.23, 28.16, 26.68, 22.83, 14.26; MS

(ESI) m/z 467.5 (M)+.

107

tert-butyl 2-acetamido-6-(N-hydroxytetradecanamido)hexanoate (161). A solution of 160 (0.015 g, 0.032 mmol) in MeOH (20 mL) was slowly added to 10 wt. % Pd/C

(0.003 g) in a hydrogenation flask. This reaction mixture was subjected to hydrogenation using a Parr-hydrogenator under 2.5 atm. pressure for 24 h. The reaction mixture was filtered through celite and the filtrate gave pure 161 after solvent

1 evaporation (0.016 g, 99%, Rf = 0.43 in EtOAc); H-NMR (CDCl3, 300 MHz) 6.5-5.92

(m, 1H), 4.68-4.23 (m, 1H), 3.79-3.41 (m, 2H), 2.70-1.92 (m, 6H), 1.91-1.01 (m, 34H),

13 0.96-0.73 (m, 5H); C-NMR (CDCl3, 300 MHz) 173.38, 171.75, 169.93, 82.14, 72.15,

52.3, 42.85, 31.89, 29,62, 29.45, 29.32, 29.22, 27.98, 22.66, 14.08.

tert-Butyl 2-acetamido-6-(N-((tert-butyldimethylsilyl)oxy)tetradecanamido) hex-4- enoate (162). A solution of 160 (0.065 g, 0.128 mmol) in DMF (5 mL) was added to a mixture of TBDMSCl (0.048 g, 0.32 mmol) and imidazole (0.044 g, 0.64 mmol).76

108

After stirring the reaction mixture overnight at 35°C, EtOAc (40 mL) was added and the organic layer was washed with water and brine, dried over MgSO4, filtered and concentrated under vacuum to give a brown crude product that was purified by flash chromatography (silica gel, 50% EtOAc : 50% pet. ether, 0.23) to give 162 as a

1 yellowish liquid (0.22 g, 30%); H-NMR (CDCl3, 500 MHz) 6.023 (d, J = 7.9 Hz, 1H),

5.83-5.4 (m, 2H), 4.69-4.43 (m, 1H), 4.36-3.84 (m, 3H), 2.99-3.75 (m, 2H), 2.73-2.4

(m, 2H), 2.39-2.2 (m, 1H), 2.01 (s, 3H), 1.85-1.74 (m, 1H), 1.7-1.54 (m, 3H), 1.52-1.08

13 (m, 29H), 1.025-0.73 (8H), 0.213 (s, 3H), 0.0.15 (3H); C-NMR (CDCl3, 500 MHz)

175.23, 174.78, 174. 21, 171.58, 171.04, 170.91, 169.75, 130.50, 128.48, 128.05,

128.035, 83.19, 82.37, 71.66, 71.30, 70.89, 55.71, 53.82, 53.09, 52.13, 38.51, 38.19,

37.33, 35.24, 34.28, 33.21, 32.78, 32.485, 32.06, 29.83, 29.79, 29.66, 29.58, 29.56,

29.5, 29.46, 28.50, 28.16, 28.12, 25.89, 25.79, 23.36, 22.84, 22.6, 19.98, 14.8, 14.28,

13.96, 0.14, -3.43, -4.49; MS (ESI) m/z 583 (M)+.

N-(cyclohex-2-en-1-yl)-N-hydroxytetradecanamide (163). A solution of cyclohexene

(5 mL) and 159 (0.2 g, 0.45 mmol) was heated in a sealed tube to reflux for 4 h. The reaction mixture was cooled in an ice bath to crystallize the 9,10-DMA, which was filtered. The filtrate showed a crude mixture of the excess alkene and a hydroxamic acid by TLC (purple spot in FeCl3). The crude mixture was purified by flash chromatography (25% EtOAc: 75% pet.ether, Rf = 0.0.63 ) to yield 163 as a reddish

1 white solid (0.048 g, 33%); H-NMR (CDCl3, 300 MHz) 6.00 (d, J = 8.53 Hz, 1H),

109

5.58 (d, J = 8.52 Hz, 1H), 4.45 (b, 1H), 2.55-2.33 (m, 1H), 2.23-1.8 (m, 5H), 1.77-1.61

13 (m, 3H), 1.43-1.17 (m, 20H), 1.00-0.833 (m, 4H); C-NMR (CDCl3, 300 MHz)

172.57, 169.92, 167.47, 141.03, 132.31, 131.19, 127.96, 126.02, 125.18, 71.73, 70.06,

55.04, 53.37, 46.37, 44.55, 34.66, 31.91, 29.64, 29.49, 29.35, 26.85, 25.56, 24.22,

22.67, 21.05, 14.1 MS (ESI) m/z 324 (M+1)+.

1-(9,10-Dimethyl-9,10-dihydro-9,10-(epoxyimino)anthracen-11-yl)ethanone (164)

A solution of acetohydroxamic acid (0.437 g, 5.82 mmol) in DMF (5 mL) was added dropwise to a solution of 9, 10-dimethylanthracene (0.8 g, 3.88 mmol) and Bu4NIO4

(0.644 g, 5.82 mmol) in CHCl3 (30 mL). The solution was stirred for 90 min and then poured into a Na2S2O3 solution (20 mL) and extracted with EtOAc:CHCl3 (200:200 mL). The extracts were dried over MgSO4, concentrated under vacuum to a solid residue and purified by flash chromatography (25% EtOAc :75% pet.ether, Rf = 0.5) to give 164 as a white solid (0.852 g, 52%) with spectroscopic and analytical details were identical to those reported.70

110

tert-Butyl 2-acetamido-6-(N-hydroxyacetamido)hex-4-enoate (165). A solution of tert-butyl 2-acetamidohex-5-enoate (0.88 g, 3.88 mmol) and 164 (0.255 g, 0.97 mmol) in toluene (5 mL) was heated in a sealed tube to reflux for 4 h. The reaction mixture was cooled down in an ice bath to crystallize the 9,10-DMA and it was filtered. The filtrate showed a crude mixture of the excess alkene and a hydroxamic acid by TLC

(purple spot in FeCl3 stain due to the presence of hydroxamic acid). The crude mixture was purified by flash chromatography (a gradient of 1:1 EtOAc: pet.ether to 5% MeOH

1 in CHCl3, Rf = 0.20 in EtOAc) to yield 165 as a thick brown oil (0.084 g, 29%); H-

NMR (500 MHz, CDCl3) 6.11 (d, J =7.69 Hz, 1H), 5.69-5.50 (m, 2H), 4.89-4.37 (m,

2H), 4.23-4.04 (m, 1H), 2.77-2.43 (m, 1H), 2.36-1.91(m, 7H), 1.48 (s, 9H) ; 13C-NMR

(CDCl3, 300 MHz) 169.96, 169.90, 168.89, 128.89, 128.39, 127.30,125.53, 82.57,

52.61, 35.88, 29.96, 24.13, 21.35, 19.47; MS (ESI) m/z 323 (M+Na)+.

tert-Butyl-2-acetamido-6-(N-((tert-butyldimethylsilyl)oxy)acetamido)hex-4-enoate

(166) A solution of 165 (0.075 g, 0.25 mmol) in DMF (5 mL) was added to a solid

111 mixture of TBDMSCl (0.112 g, 0.75 mmol) and imidazole (0.102 g, 1.5 mmol). After stirring the reaction mixture overnight at 35°C, EtOAc (40 mL) was added to dilute it.

The organic layer was washed with water and brine, dried over MgSO4 , filtered and concentrated under vacuum to give a brown crude product that was purified by flash chromatography (95% CHCl3 : 5% pet. ether, Rf = 0.53) to yield 166 as a yellow liquid

1 (0.035 g, 33%); H-NMR (CDCl3, 300 MHz) 6.04 (d, J = 6.56 Hz, 1H), 5.63-5.44 (m,

2H), 4.59-4.46 (m, 1H), 4.2-3.95 (m, 2H), 2.65-2.44 (m, 2H), 2.08-2.01 (m, 6H), 1.47

13 (s, 9H), 0.97 (s, 9H), 0.22 (m, 5H), 0.013 (s, 1H); C-NMR (CDCl3, 300 MHz)

170.31, 169.90, 168.8, 127.76, 127.69, 82.42, 60.86, 52.61, 35.88, 30.55, 28.91, 26.64,

24.12, 22.32, 22.00, 18.83, 15.25, -3.29, -3.38, MS (ESI) m/z 414 (M)+.

(9,10-Dimethyl-9,10-dihydro-9,10-(epoxyimino)anthracen-11-yl)(phenyl) methanone (167) A solution of benzohydroxamic acid (0.559 g, 4.08 mmol) in DMF

(5 mL) was added dropwise to a solution of 9, 10-dimethylanthracene (0.56 g, 2.72 mmol) and Bu4NIO4 (1.77 g, 4.08 mmol) in CHCl3 (20 mL). The solution was stirred for 90 min and then poured into a Na2S2O3 solution (20 mL) and extracted with EtOAc

(200 mL). The extracts were dried over MgSO4, filtered, concentrated under vacuum to give a solid residue that was purified by flash chromatography (5% EtOAc : 95% pet.ether, Rf = 0.5) to give 167 as a white solid ( 0.528 g, 38%) with spectroscopic and analytical details were identical to those reported.70

112

tert-Butyl 2-acetamido-6-(N-hydroxybenzamido)hex-4-enoate (168). A solution of tert-butyl 2-acetamidohex-5-enoate (149, 0.687 g, 3.026 mmol) and 167 (0.258 g, 0.75 mmol) in toluene (5 mL) was heated in a sealed tube to reflux for 4 h. The reaction mixture was cooled in an ice bath to crystallize 9,10-DMA, which was filtered. The filtrate showed a crude mixture of the excess alkene and a hydroxamic acid by TLC

(purple spot in FeCl3 due to presence of hydroxamic acid). The crude mixture was concentrated under vacuum and purified by flash chromatography (gradient of 1:1

EtOAc: pet.ether to 5% MeOH in CHCl3, Rf = 0.34 in EtOAc) to yield 168 as a thick

1 brown liquid (0.049 g, 20%); H-NMR (CDCl3, 500 MHz) 8.20-7.8 (m, 1H), 7.73-7.33

(m, 4H), 6.47-5.98 (b, 1H), 5.95-5.25 (m, 2H), 4.73-3.97 (m, 2H), 3.87-3.36 (m, 1H),

1.98 (s, 3H), 1.55-1.069 (m, 10H), 0.97-0.76 (m, 1H) MS (ESI) m/z 361.5 (M)+.

113

CHAPTER 4

SUMMARY

To summarize, this dissertation shows a detailed study of developing methods towards

the synthesis of cyclic hydroxamic acids and its application to cobactin synthesis. The

first stage of the research project featured the development of synthetic methodology

towards making cyclic hydroxamic acids from cyclic ketones by –NOH insertion and the mechanistic details of this method. Basic decomposition of Piloty’s acid in presence of cyclic ketones yields cyclic hydroxamic acids (mainly five and six membered) in 20-69%

yield with stoichiometric increase in Piloty’s acid amounts. Piloty’s acid-based

rearrangement reaction with two unsymmetric bicyclic ketones (90 and 91, Scheme 44)

reveals the regioselective nature of the reaction as the –NOH insertion always happens

from the most substituted side of the substrate ketones. The reaction is stereoselective in

nature as the syn configuration of the substrates (90, Scheme 44) retained after the –NOH

insertion. Mechanistic studies show that the –NOH insertion reaction involves a C-nitroso

intermediate (98). The C-nitroso intermediate can be generated by the hydrolysis of

acyloxy nitroso compound (103, Scheme 51) forming the N-hydroxypiperidone in 75%

yield. This discovery is seminal to perform an efficient synthesis of cyclic hydroxamic

acids (five and six membered) and could serve as a potential alternative of Piltoy’s acid

based -NOH insertion reaction. Piloty’s acid-based –NOH insertion did not work with α-

substituted ketones and failed to generate seven membered rings like cobactin. However,

hydrolysis of acyloxy nitroso intermediate generated from α-methyl cyclopentanones

114

(107, Scheme 53) makes the ring-expanded product in 78% yield. Overall, –NOH insertion fails to generate cobactin, still it is widely applicable to synthesize weinreb amides, selective oxoaminated products and could be vastly useful to make various amino acids and diverse group of hydroxylamines.

The second part of the developmental research featured the general application of intramolecular and intermolecular nitroso-ene reactions to make various hydroxamic acids based structures and their potential uses in the synthesis of siderophore mycobactin

S. Intramolecular nitroso-ene reaction of acyl-nitroso species derived from 6-hexenoic hydroxamic acid (120, Scheme 59) failed to generate any cyclized product, while the two carbon homologous acyl nitroso species (124, Scheme 61) derived from (Z)-N-

Hydroxyoct-6-enamide (126, Scheme 61) gave an unprecedented eight membered ring hydroxamic acid (127, Scheme 61). The next developmental stage of this project was exploring inter-molecular nitroso-ene reactions between model alkene 149 and acyl nitroso precursor 136 towards generating the key intermediate N€-hydroxy lysine (152,

Scheme 68). Unfortunately, the key intermediate 152 failed to produce cobactin core after DCC-DMAP coupled cyclization. The long chain hydroxamic residue of mycobactin, mycobactic acid synthesis was approached using this nitroso-ene technology with the model alkene 149 and a long chain-derived acyl nitroso precursor (159, Scheme

70). This method successfully generated the key intermediate 161 which can be useful to

make mycobactic acid. Nitroso-ene reactions of acyl nitroso and benzoyl nitroso species

with the model alkene 149 to produce the corresponding linear unsaturated hydroxamic

acids in 29% and 20% yields.

115

Overall, this research establishes a new method to make cyclic hydroxamic acids using –

NOH insertion technique into cyclic ketones, its potential synthetic applications and utilization of nitroso-ene reactions to build hydroxamic acid components of siderophore

Mycobactin S.

116

References

1. Lossen, H., Liebigs Annalen 1869, 150, 314-322.

2. Hogg, J. H.; Ollmann, I. R.; Haeggstrom, J. Z.; Wetterholm, A.; Samuelsson, B.;

Wong, C. H., Amino Hydroxamic Acids as Potent Inhibitors of Leukotriene a(4)

Hydrolase. Bioorganic & Medicinal Chemistry 1995, 3, (10), 1405-1415.

3. Spasojevic, I.; Armstrong, S. K.; Brickman, T. J.; Crumbliss, A. L.,

Electrochemical behavior of the Fe(III) complexes of the cyclic hydroxamate siderophores alcaligin and desferrioxamine E. Inorganic Chemistry 1999, 38, (3), 449-

454.

4. Codd, R., Traversing the coordination chemistry and chemical biology of hydroxamic acids. Coordination Chemistry Reviews 2008, 252, (12-14), 1387-1408.

5. Saban, N.; Bujak, M., Hydroxyurea and hydroxamic acid derivatives as antitumor drugs. Cancer Chemotherapy and Pharmacology 2009, 64, (2), 213-221.

6. Kurzak, B.; Kozlowski, H.; Farkas, E., Hydroxamic and Aminohydroxamic Acids

and Their Complexes with Metal-Ions. Coordination Chemistry Reviews 1992, 114, (2),

169-200.

7. Weber, G., Biochemical Strategy of Cancer-Cells and the Design of

Chemotherapy - Gha-Clowes-Memorial-Lecture. Cancer Research 1983, 43, (8), 3466-

3492.

8. Miller, M. J., Hydroxamate Approach to the Synthesis of Beta-Lactam

Antibiotics. Accounts of Chemical Research 1986, 19, (2), 49-56.

9. Miller, M. J., Syntheses and Therapeutic Potential of Hydroxamic Acid Based

Siderophores and Analogs. Chemical Reviews 1989, 89, (7), 1563-1579.

117

10. Whittaker, M.; Floyd, C. D.; Brown, P.; Gearing, A. J. H., Design and therapeutic

application of matrix metalloproteinase inhibitors. Chemical Reviews 1999, 99, (9), 2735-

2776.

11. Cheng, M. Y.; De, B.; Pikul, S.; Almstead, N. G.; Natchus, M. G.; Anastasio, M.

V.; McPhail, S. J.; Snider, C. E.; Taiwo, Y. O.; Chen, L. Y.; Dunaway, C. M.; Gu, F.;

Dowty, M. E.; Mieling, G. E.; Janusz, M. J.; Wang-Weigand, S., Design and synthesis of

piperazine-based matrix metalloproteinase inhibitors. Journal of Medicinal Chemistry

2000, 43, (3), 369-380.

12. Park, J. D.; Kim, D. H., Reversed hydroxamate-bearing thermolysin inhibitors

mimic a high-energy intermediate along the enzyme-catalyzed proteolytic reaction

pathway. Bioorganic & Medicinal Chemistry Letters 2003, 13, (19), 3161-3166.

13. Molteni, V.; He, X. H.; Nabakka, J.; Yang, K. Y.; Kreusch, A.; Gordon, P.;

Bursulaya, B.; Warner, I.; Shin, T.; Biorac, T.; Ryder, N. S.; Goldberg, R.; Doughty, J.;

He, Y., Identification of novel potent bicyclic peptide deformylase inhibitors. Bioorganic

& Medicinal Chemistry Letters 2004, 14, (6), 1477-1481.

14. Marmion, C. J.; Griffith, D.; Nolan, K. B., Hydroxamic acids - An intriguing family of enzyme inhibitors and biomedical ligands. European Journal of Inorganic

Chemistry 2004, (15), 3003-3016.

15. Johnstone, R. W., Histone-deacetylase inhibitors: Novel drugs for the treatment of cancer. Nature Reviews Drug Discovery 2002, 1, (4), 287-299.

16. Muri, E. M. F.; Nieto, M. J.; Sindelar, R. D.; Williamson, J. S., Hydroxamic acids as pharmacological agents. Current Medicinal Chemistry 2002, 9, (17), 1631-1653.

118

17. Marmion, C. J.; Murphy, T.; Docherty, J. R.; Nolan, K. B., Hydroxamic acids are nitric oxide donors. Facile formation of ruthenium(II)-nitrosyls and NO-mediated activation of guanylate cyclase by hydroxamic acids. Chemical Communications 2000,

(13), 1153-1154.

18. Wilhelm, S. M.; Collier, I. E.; Kronberger, A.; Eisen, A. Z.; Marmer, B. L.; Grant,

G. A.; Bauer, E. A.; Goldberg, G. I., Human-Skin Fibroblast Stromelysin - Structure,

Glycosylation, Substrate-Specificity, and Differential Expression in Normal and

Tumorigenic Cells. Proceedings of the National Academy of Sciences of the United States of America 1987, 84, (19), 6725-6729.

19. Senior, R. M.; Griffin, G. L.; Fliszar, C. J.; Shapiro, S. D.; Goldberg, G. I.;

Welgus, H. G., Human 92-Kilodalton and 72-Kilodalton Type-Iv Collagenases Are

Elastases. Journal of Biological Chemistry 1991, 266, (12), 7870-7875.

20. Lovejoy, B.; Hassell, A. M.; Luther, M. A.; Weigl, D.; Jordan, S. R., Crystal-

Structures of Recombinant 19-Kda Human Fibroblast Collagenase Complexed to Itself.

Biochemistry 1994, 33, (27), 8207-8217.

21. Kim, H. M.; Lee, K.; Park, B. W.; Ryu, D. K.; Kim, K.; Lee, C. W.; Park, S. K.;

Han, J. W.; Lee, H. Y.; Lee, H. Y.; Han, G. H., Synthesis, enzymatic inhibition, and cancer cell growth inhibition of novel delta-lactam-based histone deacetylase (HDAC) inhibitors. Bioorganic & Medicinal Chemistry Letters 2006, 16, (15), 4068-4070.

22. Conti, C.; Leo, E.; Eichler, G. S.; Sordet, O.; Martin, M. M.; Fan, A.; Aladjem,

M. I.; Pommier, Y., Inhibition of Histone Deacetylase in Cancer Cells Slows Down

Replication Forks, Activates Dormant Origins, and Induces DNA Damage. Cancer

Research 2010, 70, (11), 4470-4480.

119

23. Finnin, M. S.; Donigian, J. R.; Cohen, A.; Richon, V. M.; Rifkind, R. A.; Marks,

P. A.; Breslow, R.; Pavletich, N. P., Structures of a histone deacetylase homologue bound to the TSA and SAHA inhibitors. Nature 1999, 401, (6749), 188-193.

24. Richon, V. M.; Emiliani, S.; Verdin, E.; Webb, Y.; Breslow, R.; Rifkind, R. A.;

Marks, P. A., A class of hybrid polar inducers of transformed cell differentiation inhibits histone deacetylases. Proceedings of the National Academy of Sciences of the United

States of America 1998, 95, (6), 3003-3007.

25. Kim, Y. B.; Lee, K. H.; Sugita, K.; Yoshida, M.; Horinouchi, S., Oxamflatin is a novel antitumor compound that inhibits mammalian histone deacetylase. Oncogene 1999,

18, (15), 2461-2470.

26. Echchannaoui, H.; Leib, S. L.; Neumann, U.; Landmann, R. M. A., Adjuvant

TACE inhibitor treatment improves the outcome of TLR2(-/-) mice with experimental pneumococcal meningitis. Bmc Infectious Diseases 2007, 7, -.

27. Kleinman, E. F.; Campbell, E.; Giordano, L. A.; Cohan, V. L.; Jenkinson, T. H.;

Cheng, J. B.; Shirley, J. T.; Pettipher, E. R.; Salter, E. D.; Hibbs, T. A.; DiCapua, F. M.;

Bordner, J., Striking effect of hydroxamic acid substitution on the phosphodiesterase type

4 (PDE4) and TNF alpha inhibitory activity of two series of rolipram analogues:

Implications for a new active site model of PDE4. Journal of Medicinal Chemistry 1998,

41, (3), 266-270.

28. Xue, C. B.; Voss, M. E.; Nelson, D. J.; Duan, J. J. W.; Cherney, R. J.; Jacobson, I.

C.; He, X. H.; Roderick, J.; Chen, L. H.; Corbett, R. L.; Wang, L.; Meyer, D. T.;

Kennedy, K.; DeGrado, W. F.; Hardman, K. D.; Teleha, C. A.; Jaffee, B. D.; Liu, R. Q.;

Copeland, R. A.; Covington, M. B.; Christ, D. D.; Trzaskos, J. M.; Newton, R. C.;

120

Magolda, R. L.; Wexler, R. R.; Decicco, C. P., Design, synthesis, and structure-activity

relationships of macrocyclic hydroxamic acids that inhibit tumor necrosis factor alpha

release in vitro and in vivo. Journal of Medicinal Chemistry 2001, 44, (16), 2636-2660.

29. Xue, C. B.; He, X. H.; Corbett, R. L.; Roderick, J.; Wasserman, Z. R.; Liu, R. Q.;

Jaffee, B. D.; Covington, M. B.; Qian, M. X.; Trzaskos, J. M.; Newton, R. C.; Magolda,

R. L.; Wexler, R. R.; Decicco, C. P., Discovery of macrocyclic hydroxamic acids

containing biphenylmethyl derivatives at P1 ', a series of selective TNF-alpha converting

enzyme inhibitors with potent cellular activity in the inhibition of TNF-alpha release.

Journal of Medicinal Chemistry 2001, 44, (21), 3351-3354.

30. Hynes, J. B., Hydroxylamine derivatives as potential antimalarial agents. 1.

Hydroxamic acids. Journal of Medicinal Chemistry 1970, 13, (6), 1235-7.

31. Kurz, T.; Geffken, D.; Wackendorff, C., Carboxylic acid analogues of

fosmidomycin. Zeitschrift Fur Naturforschung Section B-a Journal of Chemical Sciences

2003, 58, (5), 457-461.

32. Schluter, K.; Walter, R. D.; Bergmann, B.; Kurz, T., Arylmethyl substituted

derivatives of Fosmidomycin: Synthesis and antimalarial activity. European Journal of

Medicinal Chemistry 2006, 41, (12), 1385-1397.

33. Penalver, M. J.; Hiner, A. N. P.; Rodriguez-Lopez, J. N.; Garcia-Canovas, F.;

Tudela, J., Mechanistic implications of variable stoichiometries of oxygen consumption during tyrosinase catalyzed oxidation of monophenols and o-diphenols. Biochimica Et

Biophysica Acta-Protein Structure and Molecular Enzymology 2002, 1597, (1), 140-148.

34. Soni, M. G.; White, S. M.; Flamm, W. G.; Burdock, G. A., Safety evaluation of dietary aluminum. Regulatory Toxicology and Pharmacology 2001, 33, (1), 66-79.

121

35. Kubo, I.; Nihei, K.; Shimizu, K., Oxidation products of quercetin catalyzed by

mushroom tyrosinase. Bioorganic & Medicinal Chemistry 2004, 12, (20), 5343-5347.

36. Shiino, M.; Watanabe, Y.; Umezawa, K., Synthesis of N-substituted N-

nitrosohydroxylamines as inhibitors of mushroom tyrosinase. Bioorganic & Medicinal

Chemistry 2001, 9, (5), 1233-1240.

37. Rho, H. S.; Baek, H. S.; Ahn, S. M.; Yoo, J. W.; Kim, D. H.; Kim, H. G.,

Hydroxamic Acid Derivatives as Anti-melanogenic Agents: The Importance of a Basic

Skeleton and Hydroxamic Acid Moiety. Bulletin of the Korean Chemical Society 2009,

30, (2), 475-478.

38. Ito, S.; Nardi, G.; Palumbo, A.; Prota, G., Isolation and Characterization of

Adenochrome, a Unique Iron(Iii)-Binding Peptide from Octopus-Vulgaris. Journal of the

Chemical Society-Perkin Transactions 1 1979, (11), 2617-2623.

39. Bravo, H. R.; Lazo, W., Antialgal and antifungal activity of natural hydroxamic

acids and related compounds. Journal of Agricultural and Food Chemistry 1996, 44, (6),

1569-1571.

40. Owotoki, W.; Geffken, D.; Kurz, T., Synthesis of 1-hydroxypyrrolidin-2,5-dione

derivatives of the phosphonic-hydroxamic acid antibiotic SF-2312. Australian Journal of

Chemistry 2006, 59, (4), 283-288.

41. Xu, Y. P.; Miller, M. J., Total syntheses of mycobactin analogues as potent antimycobacterial agents using a minimal protecting group strategy. Journal of Organic

Chemistry 1998, 63, (13), 4314-4322.

42. Dobbin, P. S.; Hider, R. C., Iron Chelation-Therapy. Chemistry in Britain 1990,

26, (6), 565-568.

122

43. Johnson, K. V. H. C. R., Cyclic Hydroxamic Acid. United States Patent 1994.

44. Roberts, K. P.; Iyer, R. A.; Prasad, G.; Liu, L. T.; Lind, R. E.; Hanna, P. E.,

Cyclic hydroxamic acid inhibitors of prostate cancer cell growth: Selectivity and structure activity relationships. Prostate 1998, 34, (2), 92-99.

45. Fennell, K. A.; Mollmann, U.; Miller, M. J., Syntheses and biological activity of amamistatin B and analogs. Journal of Organic Chemistry 2008, 73, (3), 1018-1024.

46. Murakami, Y.; Kato, S.; Nakajima, M.; Matsuoka, M.; Kawai, H.; ShinYa, K.;

Seto, H., Formobactin, a novel free radical scavenging and neuronal cell protecting substance from Nocardia sp. Journal of Antibiotics 1996, 49, (9), 839-845.

47. Snow, G. A., Mycobactins: iron-chelating growth factors from mycobacteria.

Bacteriological Reviews 197034, 99-125.

48. Tanaka, K.; Matsuo, K.; Nakanishi, A.; Hatano, T.; Izeki, H.; Ishida, Y.; Mori,

W., Syntheses and Anti-Inflammatory and Analgesic Activities of Hydroxamic Acids and

Acid Hydrazides. Chemical & Pharmaceutical Bulletin 1983, 31, (8), 2810-2819.

49. Tanaka, K.; Matsuo, K.; Nakanishi, A.; Kataoka, Y.; Takase, K.; Otsuki, S.,

Syntheses of Cyclic Hydroxamic Acid-Derivatives, and Their Chelating Abilities and

Biological-Activities. Chemical & Pharmaceutical Bulletin 1988, 36, (7), 2323-2330.

50. Angeli, A., Gazzetta Chimica Italiana 1896, 26, 17-25.

51. Hassner, A.; Wiederkher, R.; Kascheres, A. J., Reaction of Aldehydes with N-

Hydroxybenzenesulfonamide. Journal of Organic Chemistry 1970, 35, (6), 1962-1964.

52. Porcheddu, A.; Giacomelli, G., Angeli-Rimini's reaction on solid support: A new approach to hydroxamic acids. Journal of Organic Chemistry 2006, 71, (18), 7057-7059.

123

53. Murray, R. W.; Jeyaraman, R., Dioxiranes - Synthesis and Reactions of

Methyldioxiranes. Journal of Organic Chemistry 1985, 50, (16), 2847-2853.

54. Cassidei, L.; Fiorentino, M.; Mello, R.; Sciacovelli, O.; Curci, R., O-17 and C-13

Identification of the Dimethyldioxirane Intermediate Arising in the Reaction of

Potassium Carbonate with Acetone. Journal of Organic Chemistry 1987, 52, (4), 699-

700.

55. Rella, M. R.; Williard, P. G., Oxidation of peptides by methyl(trifluoromethyl)dioxirane: The protecting group matters. Journal of Organic

Chemistry 2007, 72, (2), 525-531.

56. Liu, Y.; Jacobs, H. K.; Gopalan, A. S., Reactions of N-Benzyloxycarbamate

Derivatives with Stabilized Carbon Nucleophiles: A New Synthetic Approach to

Polyhydroxamic Acids and Other Hydroxamate-Containing Mixed Ligand Systems.

Journal of Organic Chemistry 2009, 74, (2), 782-788.

57. AlKhatib, H. S.; Taha, M. O.; Aiedeh, K. M.; Bustanji, Y.; Sweileh, B., Synthesis and in vitro behavior of iron-crosslinked N-methyl and N-benzyl hydroxamated derivatives of alginic acid as controlled release carriers. European Polymer Journal 2006,

42, (10), 2464-2474.

58. Kurz, T.; Schluter, K.; Kaula, U.; Bergmann, B.; Walter, R. D.; Geffken, D.,

Synthesis and antimalarial activity of chain substituted pivaloyloxymethyl ester analogues of Fosmidomycin and FR900098. Bioorganic & Medicinal Chemistry 2006,

14, (15), 5121-5135.

124

59. Ramurthy, S.; Miller, M. J., Framework-reactive siderophore analogs as potential cell-selective drugs. Design and syntheses of trimelamol-based iron chelators. Journal of

Organic Chemistry 1996, 61, (12), 4120-4124.

60. Hu, J. D.; Miller, M. J., A New Method for the Synthesis of N-Epsilon-Acetyl-N-

Epsilon-Hydroxy-L-Lysine, the Iron-Binding Constituent of Several Important

Siderophores. Journal of Organic Chemistry 1994, 59, (17), 4858-4861.

61. Lin, Y. M.; Miller, M. J., Practical synthesis of hydroxamate-derived siderophore components by an indirect oxidation method and syntheses of a DIG-siderophore conjugate and a biotin-siderophore conjugate. Journal of Organic Chemistry 1999, 64,

(20), 7451-7458.

62. Wu, T. Y. H.; Hassig, C.; Wu, Y. Q.; Ding, S.; Schultz, P. G., Design, synthesis,

and activity of HDAC inhibitors with a N-formyl hydroxylamine head group. Bioorganic

& Medicinal Chemistry Letters 2004, 14, (2), 449-453.

63. Fokin, A. A.; Yurchenko, A. G.; Rodionov, V. N.; Gunchenko, P. A.; Yurchenko,

R. I.; Reichenberg, A.; Wiesner, J.; Hintz, M.; Jomaa, H.; Schreiner, P. R., Synthesis of

the antimalarial drug FR900098 utilizing the nitroso-ene reaction. Organic Letters 2007,

9, (21), 4379-4382.

64. Robinson, D. E.; Holladay, M. W., Convenient preparation of O-linked polymer-

bound N-substituted hydroxylamines, intermediates for synthesis of N-substituted

hydroxamic acids. Organic Letters 2000, 2, (18), 2777-2779.

65. Keck, G. E.; Webb, R., Carbon-Nitrogen Bond Formation Via Acyl-Nitroso

Compounds - Intra-Molecular Ene Processes. Tetrahedron Letters 1979, (14), 1185-1186.

125

66. Christie, C. C.; Kirby, G. W.; Mcguigan, H.; Mackinnon, J. W. M., C-

Nitrosoformamides, a New Class of Transient Dienophiles Formed by Oxidation of N-

Hydroxyureas. Journal of the Chemical Society-Perkin Transactions 1 1985, (11), 2469-

2473.

67. Kirby, G. W.; Mcguigan, H.; Mclean, D., Intramolecular Ene Reactions of

Transient, Allylic, and Homoallylic C-Nitrosoformate Esters. Journal of the Chemical

Society-Perkin Transactions 1 1985, (9), 1961-1966.

68. Corrie, J. E. T.; Kirby, G. W.; Mackinnon, J. W. M., Reactions of Transient-C-

Nitrosocarbonyl Compounds with Dienes, Monoolefins, and Nucleophiles. Journal of the

Chemical Society-Perkin Transactions 1 1985, (4), 883-886.

69. Kirby, G. W.; Mcguigan, H.; Mackinnon, J. W. M.; Mclean, D.; Sharma, R. P.,

Formation and Reactions of C-Nitrosoformate Esters, a New Class of Transient

Dienophiles. Journal of the Chemical Society-Perkin Transactions 1 1985, (7), 1437-

1442.

70. Jenkins, N. E.; Ware, R. W.; Atkinson, R. N.; King, S. B., Generation of acyl nitroso compounds by the oxidation of N-acyl hydroxylamines with the Dess-Martin periodinane. Synthetic Communications 2000, 30, (5), 947-953.

71. Keck, G. E.; Webb, R. R.; Yates, J. B., A Versatile Method for Carbon-Nitrogen

Bond Formation Via Ene Reactions of Acylnitroso Compounds. Tetrahedron 1981, 37,

(23), 4007-4016.

72. Adam, W.; Bottke, N.; Engels, B.; Krebs, O., An experimental and computational

study on the reactivity and regioselectivity for the nitrosoarene ene reaction: Comparison

126

with triazolinedione and singlet oxygen. Journal of the American Chemical Society 2001,

123, (23), 5542-5548.

73. Leach, A. G.; Houk, K. N., The mechanism and regioselectivity of the ene reactions of nitroso compounds: a theoretical study of reactivity, regioselectivity, and kinetic isotope effects establishes a stepwise path involving polarized diradical intermediates. Organic & Biomolecular Chemistry 2003, 1, (8), 1389-1403.

74. Fakhruddin, A.; Iwasa, S.; Nishiyama, H.; Tsutsumi, K., Ene reactions of acyl nitroso intermediates with alkenes and their halocyclization. Tetrahedron Letters 2004,

45, (51), 9323-9326.

75. Thomas, A.; Rajappa, S., Synthesis of Cyclic Hydroxamic Acids from Aliphatic

Nitro-Compounds. Tetrahedron 1995, 51, (38), 10571-10580.

76. Hu, J. D.; Miller, M. J., Total synthesis of a mycobactin S, a siderophore and growth promoter of Mycobacterium smegmatis, and determination of its growth inhibitory activity against Mycobacterium tuberculosis. Journal of the American

Chemical Society 1997, 119, (15), 3462-3468.

77. Murahashi, S. I.; Oda, T.; Sugahara, T.; Masui, Y., Tungstate-Catalyzed

Oxidation of Tetrahydroquinolines with Hydrogen-Peroxide - a Novel Method for the

Synthesis of Cyclic Hydroxamic Acids. Journal of Organic Chemistry 1990, 55, (6),

1744-1749.

78. Atkinson, J.; Arnason, J.; Campos, F.; Niemeyer, H. M.; Bravo, H. R., Synthesis and Reactivity of Cyclic Hydroxamic Acids - Resistance Factors in the Gramineae. Acs

Symposium Series 1992, 504, 349-360.

127

79. Matlin, S. A.; Sammes, P. G.; Upton, R. M., Oxidation of Trimethylsilylated

Amides to Hydroxamic Acids. Journal of the Chemical Society-Perkin Transactions 1

1979, (10), 2481-2487.

80. Wardrop, D. J.; Bowen, E. G.; Forslund, R. E.; Sussman, A. D.; Weerasekera, S.

L., Intramolecular Oxamidation of Unsaturated O-Alkyl Hydroxamates: A Remarkably

Versatile Entry to Hydroxy Lactams. Journal of the American Chemical Society 2010,

132, (4), 1188-1189.

81. Wardrop, D. J.; Burge, M. S., Nitrenium ion azaspirocyclization-spirodienone

cleavage: A new synthetic strategy for the stereocontrolled preparation of highly

substituted lactams and N-hydroxy lactams. Journal of Organic Chemistry 2005, 70,

(25), 10271-10284.

82. Suginome, H.; Kurokawa, Y., Photoinduced Molecular-Transformations .104.

Pathways of the Photorearrangements of 5-Membered Cyclic Steroidal Alpha-Nitro

Ketones to N-Hydroxy Cyclic Imides, Cyclic Hydroxamic Acid, and Cyclic Imide.

Journal of Organic Chemistry 1989, 54, (25), 5945-5953.

83. Cridland, J. S.; Moles, P. J.; Reid, S. T.; Taylor, K. T., Photochemistry of Cyclic and Acyclic Nitroalkenes. Tetrahedron Letters 1976, (49), 4497-4500.

84. Moles, P. J.; Reid, S. T., Thermal-Decomposition of Alpha-Nitro Ketones.

Tetrahedron Letters 1976, (26), 2283-2286.

85. Keck, G. E.; Webb, R. R., Alkaloid Synthesis Via Intramolecular Ene Reaction .2.

Application to Dl-Mesembrine and Dl-Dihydromaritidine. Journal of Organic Chemistry

1982, 47, (7), 1302-1309.

128

86. Keck, G. E.; Webb, R. R., Alkaloid Synthesis Via Intramolecular Ene Reactions

.1. Application to (+/-)-Crinane. Journal of the American Chemical Society 1981, 103,

(11), 3173-3177.

87. Oppolzer, W.; Snieckus, V., Intra-Molecular Ene Reactions in Organic-Synthesis.

Angewandte Chemie-International Edition in English 1978, 17, (7), 476-486.

88. Panizzi, L. D., G.; Tardella, P.A.; d'Abbiero, L., Ric. Sci. 1961, 1, 312-318.

89. Saturated N-hydroxylactams. Societa Farmaceutici Italia 1964, (GB 955945).

90. Hassner, A. W., R.; Kascheres, A. J., Reaction of aldehydes with N-

hydroxybenzenesulfonamide. Acetal formation catalyzed by nucleophiles. Journal of

Organic Chemistry 1970.

91. D. ST.C.Black, R. F. C. B., A. M. Wade, SYNTHETIC STUDIES RELATED TO

MYCOBACTINS. Australian Journal of Chemistry 1972, 25, 2429-44.

92. Bonner, F. T.; Ko, Y. H., Kinetic, Isotopic, and N-15 Nmr-Study of N-

Hydroxybenzenesulfonamide Decomposition - an Hno Source Reaction. Inorganic

Chemistry 1992, 31, (12), 2514-2519.

93. Shafirovich, V.; Lymar, S. V., Spin-forbidden deprotonation of aqueous nitroxyl

(HNO). Journal of the American Chemical Society 2003, 125, (21), 6547-6552.

94. Shafirovich, V.; Lymar, S. V., Nitroxyl and its anion in aqueous solutions: Spin states, protic equilibria, and reactivities toward oxygen and nitric oxide. Proceedings of the National Academy of Sciences of the United States of America 2002, 99, (11), 7340-

7345.

129

95. Bartberger, M. D.; Fukuto, J. M.; Houk, K. N., On the acidity and reactivity of

HNO in aqueous solution and biological systems. Proceedings of the National Academy of Sciences of the United States of America 2001, 98, (5), 2194-2198.

96. Fukuto, J. M.; Dutton, A. S.; Houk, K. N., The chemistry and biology of nitroxyl

(HNO): A chemically unique species with novel and important biological activity.

Chembiochem 2005, 6, (4), 612-619.

97. Fukuto, J. M.; Bartberger, M. D.; Dutton, A. S.; Paolocci, N.; Wink, D. A.; Houk,

K. N., The physiological chemistry and biological activity of nitroxyl (HNO): The neglected, misunderstood, and enigmatic nitrogen oxide. Chemical Research in

Toxicology 2005, 18, (5), 790-801.

98. Miranda, K. M., The chemistry of nitroxyl (HNO) and implications in biology.

Coordination Chemistry Reviews 2005, 249, (3-4), 433-455.

99. Fukuto, J. M.; Switzer, C. H.; Miranda, K. M.; Wink, D. A., Nitroxyl (HNO):

Chemistry, biochemistry, and pharmacology. Annual Review of Pharmacology and

Toxicology 2005, 45, 335-355.

100. Wuest, W. M.; Sattely, E. S.; Walsh, C. T., Three Siderophores from One

Bacterial Enzymatic Assembly Line. Journal of the American Chemical Society 2009,

131, (14), 5056-+.

101. Quadri, L. E. N.; Sello, J.; Keating, T. A.; Weinreb, P. H.; Walsh, C. T.,

Identification of a Mycobacterium tuberculosis gene cluster encoding the biosynthetic

enzymes for assembly of the virulence-conferring siderophore mycobactin. Chemistry &

Biology 1998, 5, (11), 631-645.

130

102. Vergne, A. F.; Walz, A. J.; Miller, M. J., Iron chelators from mycobacteria (1954-

1999) and potential therapeutic applications. Natural Product Reports 2000, 17, (1), 99-

116.

103. Switzer, C. H.; Flores-Santana, W.; Mancardi, D.; Donzelli, S.; Basudhar, D.;

Ridnour, L. A.; Miranda, K. M.; Fukuto, J. M.; Paolocci, N.; Wink, D. A., The emergence of nitroxyl (HNO) as a pharmacological agent. Biochimica Et Biophysica

Acta-Bioenergetics 2009, 1787, (7), 835-840.

104. Irvine, J. C.; Ritchie, R. H.; Favaloro, J. L.; Andrews, K. L.; Widdop, R. E.;

Kemp-Harper, B. K., Nitroxyl (HNO): the Cinderella of the nitric oxide story. Trends in

Pharmacological Sciences 2008, 29, (12), 601-608.

105. Rimini, E., Atti R. Accad. dei Lincei Roma 1901, 10, 355-362.

106. Scholz, J. N., Engel, P.S.,Glidewell,C.Whitemire, K.H., Tetrahedron 1989, 45,

7695-7708.

107. Panizzi, L. D. M., G.; Tardella, P. A.; d'Abbiero, L. , Action of nitroxyl on ketonic

compounds. I. Cyclic ketones. . Ricerca Sci. 1961, 1, (IIA), 312-318.

108. Krepski, L. R.; Hassner, A., Cycloadditions .24. Improved Procedure for Addition of Dichloroketene to Unreactive Olefins. Journal of Organic Chemistry 1978, 43, (14),

2879-2882.

109. Eckehard Volker Dehmlow*, J. K., Monika Buchholz, and Dirk Hannemann,

Preparation of Cyclobutyl Group Carrying Cyclobutanones and Related Synthetic

Building Blocks

J. Prakt. Chem. 2000, 342, 409-413.

131

110. Renz, M.; Meunier, B., 100 years of Baeyer-Villiger oxidations. European

Journal of Organic Chemistry 1999, (4), 737-750.

111. DiMaio, G. T., P. A. , Gazz. Chim. Ital 1966, 96, 526-31.

112. Smith, P. A. S., Hein, G.E., The Alleged Role of Nitroxyl in Certain Reactions of

Aldehydes and Alkyl Halides. Jorunal of American Chemical Society 1960, 82, 5731-

5738.

113. Charles Z Ding, J. T. H., Katerina Leftheris, Rajeev S. Bhide, Preparation of non-

imidazole benzodiazepine inhibitors of farnesyl protein transferase. United States Patent

2002, US 6458783 B1.

114. King, S. B.; Nagasawa, H. T., Chemical approaches toward generation of

nitroxyl. Nitric Oxide, Pt C 1999, 301, 211-220.

115. King, S. B. a. N., H. T., Chemical approaches towards the generation of nitroxyl.

Methods in Enzymology 1998, 301, (part C), 211-220.

116. Bartberger, M. D.; Liu, W.; Ford, E.; Miranda, K. M.; Switzer, C.; Fukuto, J. M.;

Farmer, P. J.; Wink, D. A.; Houk, K. N., The reduction potential of nitric oxide (NO) and its importance to NO biochemistry. Proceedings of the National Academy of Sciences of the United States of America 2002, 99, (17), 10958-10963.

117. Sha, X.; Isbell, T. S.; Patel, R. P.; Day, C. S.; King, S. B., Hydrolysis of acyloxy nitroso compounds yields nitroxyl (HNO). Journal of the American Chemical Society

2006, 128, (30), 9687-9692.

118. Neset, S. M.; Benneche, T.; Undheim, K., Synthesis of Cyclic Hydroxamic Acids

by Oxidation of Secondary Amines with Dimethyldioxirane. Acta Chemica Scandinavica

1993, 47, 1141-1143.

132

119. Black, D. S. C.; Brown, R. F. C.; Wade, A. M., Synthetic Study Related to

Mycobactins. Australian Journal of Chemistry 1972, 25, 2429-44.

120. Schnur, R. C.; Howard, H. R., 1,2,3,4-Tetrasubstituted Isoquinoline Acetic-Acids.

Tetrahedron Letters 1981, 22, (30), 2843-2846.

121. Vedejs, E.; Sano, H., Synthesis of N-Methoxy and N-H Aziridines from Alkenes.

Tetrahedron Letters 1992, 33, (23), 3261-3264.

122. Krow, G. R.; Szczepanski, S. W.; Kim, J. Y.; Liu, N.; Sheikh, A.; Xiao, Y. S.;

Yuan, J., Regioselective functionalization. 7. Unexpected preferences for bridgehead migration in Schmidt rearrangement syntheses of novel 2,6-diazabicyclo[3.2.x]alkan-3- ones (x=1-3). Journal of Organic Chemistry 1999, 64, (4), 1254-1258.

123. Hilmey, D. G.; Paquette, L. A., Promoter-dependent course of the Beckmann rearrangement of stereoisomeric spiro[4.4]nonane-1,6-dione monoximes. Organic Letters

2005, 7, (10), 2067-2069.

124. Nahm, S.; Weinreb, S. M., N-Methoxy-N-Methylamides as Effective Acylating

Agents. Tetrahedron Letters 1981, 22, (39), 3815-3818.

125. De Luca, L.; Giacomelli, G.; Taddei, M., An easy and convenient synthesis of

Weinreb amides and hydroxamates. Journal of Organic Chemistry 2001, 66, (7), 2534-

2537.

126. Andersson, M. A.; Epple, R.; Fokin, V. V.; Sharpless, K. B., A new approach to osmium-catalyzed asymmetric dihydroxylation and aminohydroxylation of olefins.

Angewandte Chemie-International Edition 2002, 41, (3), 472-475.

133

127. Atkinson, R. N.; Moore, L.; Tobin, J.; King, S. B., Asymmetric synthesis of

conformationally restricted L-arginine analogues as active site probes of nitric oxide

synthase. Journal of Organic Chemistry 1999, 64, (10), 3467-3475.

128. Scholz, J. N.; Engel, P. S.; Glidewell, C.; Whitmire, K. H., Reaction of

Hydroxylamine with Benzenesulfonyl Chloride - X-Ray Crystal-Structure of Pilotys Acid

and Other Benzenesulfonylhydroxylamines. Tetrahedron 1989, 45, (24), 7695-7708.

129. Neset, S. M.; Benneche, T.; Undheim, K., Synthesis of Cyclic Hydroxamic Acids

by Oxidation of Secondary-Amines with Dimethyldioxirane. Acta Chemica Scandinavica

1993, 47, (11), 1141-1143.

130. Sparks, S. M.; Chow, C. P.; Zhu, L.; Shea, K. J., Type 2 intramolecular N-

acylnitroso Diels-Alder reaction: Scope and application to the synthesis of medium ring

lactams. Journal of Organic Chemistry 2004, 69, (9), 3025-3035.

131. Li, G. Y.; Che, C. M., Highly selective intra- and intermolecular coupling reactions of diazo compounds to form cis-alkenes using a ruthenium porphyrin catalyst.

Organic Letters 2004, 6, (10), 1621-1623.

132. Appel, M.; Blaurock, S.; Berger, S., A Wittig reaction with 2-furyl substituents at the phosphorus atom: Improved (Z) selectivity and isolation of a stable oxaphosphetane intermediate. European Journal of Organic Chemistry 2002, (7), 1143-1148.

133. Snow, G. A., J. Chem. Soc 1954, 2588.

134. Andrei, D.; Wnuk, S. F., S-adenosylhomocysteine analogues with the carbon-5 '

and sulfur atoms replaced by a vinyl unit. Organic Letters 2006, 8, (22), 5093-5096.

135. Lin, Y. A.; Chalker, J. M.; Floyd, N.; Bernardes, G. J. L.; Davis, B. G., Allyl

sulfides are privileged substrates in aqueous cross-metathesis: Application to site-

134

selective protein modification. Journal of the American Chemical Society 2008, 130,

(30), 9642-9643.

136. Walz, A. J.; Miller, M. J., Synthesis and biological activity of hydroxamic acid-

derived vasopeptidase inhibitor analogues. Organic Letters 2002, 4, (12), 2047-2050.

137. Kirby, G. W., Electrophilic C-Nitroso-Compounds. Chemical Society Reviews

1977, 6, (1), 1-24.

138. Bitar, A. Y.; Frontier, A. J., Formal Synthesis of (+/-)-Roseophilin. Organic

Letters 2009, 11, (1), 49-52.

139. Schreiber, S. L.; Claus, R. E.; Reagan, J., Ozonolytic Cleavage of Cycloalkenes to

Terminally Differentiated Products. Tetrahedron Letters 1982, 23, (38), 3867-3870.

140. Gorczynski, M. J.; Huang, J. M.; King, S. B., Regio- and stereospecific syntheses

and nitric oxide donor properties of (E)-9- and (E)-10-nitrooctadec-9-enoic acids.

Organic Letters 2006, 8, (11), 2305-2308.

135

APPENDIX A

CRYSTAL STRUCTURE ANALYSIS REPORT AND TABLES FOR COMPOUND

C5H9NO2 (85)

Wake Forest X-Ray Facility Reference Code a52m

Performed by Dr. Cynthia Day Wake Forest University

136

EXPERIMENTAL

Colorless rectangular-parallelepiped-shaped crystals of C5H9NO2 are, at 193(2)

15 K, orthorhombic, space group Pbca – D 2h (No. 61) (1) with a = 15.900(3) Å, b = 3 7.187(1) Å, c = 19.995(4) Å, V = 2284.9(8) Å and Z = 16 molecules {dcalcd = 1.339

3 -1 g/cm ; a(MoK ) = 0.103 mm }. A full hemisphere of diffracted intensities (1868 20-second frames with a  scan width of 0.30) was measured for a single-domain specimen using graphite-monochromated MoK radiation (= 0.71073 Å) on a Bruker SMART APEX CCD Single Crystal Diffraction System (2). X-rays were provided by a fine-focus sealed x-ray tube operated at 50kV and 30mA. Lattice constants were determined with the Bruker SAINT software package using peak centers for 5101 reflections. A total of 21446 integrated reflection intensities having 2((MoK )< 58.70 were produced using the Bruker program SAINT(3); 3116 of these were unique and gave Rint = 0.058 with a coverage which was 99.6% complete. The relative transmission factors ranged from 0.966 to 0.990. The Bruker software package SHELXTL was used to solve the structure using “direct methods” techniques. All 2 stages of weighted full-matrix least-squares refinement were conducted using Fo data with the SHELXTL Version 6.12 software package(4). The final structural model incorporated anisotropic thermal parameters for all nonhydrogen atoms and isotropic thermal parameters for all hydrogen atoms. Hydroxyl hydrogen atoms were located from a difference Fourier map and refined as independent isotropic atoms. The remaining hydrogen atoms were included into the structural model as idealized atoms (assuming sp3-hybridization of the carbon atoms and C-H bond lengths of 0.99 Å). The isotropic thermal parameters for H2O and H4O refined to final values of 0.069(5) and 0.066(5)Å2, respectively. The isotropic thermal parameters of the remaining hydrogen atoms were fixed at values 1.2 times the equivalent isotropic thermal parameter of the carbon atom to which they are covalently bonded. One of the two independent molecules shows disorder involving two of the ring carbon atoms with two orientations in the crystal; the major orientation is adopted 71

137

% of the time and the minor orientation is adopted 29 % of the time. The major (71%)

orientation is specified by carbon atoms C8 and C9 and hydrogen atoms H7A, H7B, H8A,

H8B, H9A, H9B, H10A and H10B; the minor (29%) orientation is specified by carbon

atoms C8’ and C9’ and hydrogen atoms H7C, H7D, H8’A, H8’B, H9’A, H9’B, H10C and H10D, respectively. A total of 172 parameters were refined using no restraints, 3116 data and weights

of w = 1/ [2(F2) + (0.0816 P)2], where P = [Fo2 + 2Fc2] / 3. Final agreement factors

at convergence are: R1(unweighted, based on F) = 0.048 for 2332 independent

“observed” reflections having 2(MoK )< 58.70 and I>2(I); R1(unweighted, 2 based on F) = 0.062 and wR2(weighted, based on F ) = 0.130 for all 3116 independent reflections having 2(MoK )< 58.70. The largest shift/s.u. was 0.000 in the final refinement cycle. The final difference map had maxima and minima of 0.30 and -0.16 e-/Å3, respectively.

Acknowledgment The authors thank the National Science Foundation (grant CHE-0234489) for funds to purchase the x-ray instrument and computers.

References

(1) International Tables for Crystallography, Vol A, 4th ed., Kluwer: Boston (1996). (2) Data Collection: SMART Software Version 5.628 (2002). Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373 USA. (3) Data Reduction: SAINT Software Version 6.36a (2002). Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373 USA. (4) G. M. Sheldrick (2000). SHELXTL Version 6.12 Reference Manual. Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373 USA.

138

Table 1. Crystal data and structure refinement for C5H9NO2

Identification code a52m2m Empirical formula C5 H9 N O2 Formula weight 115.13 Temperature 193(2) K Wavelength 0.71073 Å Crystal system Orthorhombic 15 Space group Pbca – D 2h (No. 61) Unit cell dimensions a = 15.900(3) Å b = 7.187(1) Å c = 19.995(4) Å Volume 2284.9(8) Å3 Z 16 Density (calculated) 1.339 g/cm3 Absorption coefficient 0.103 mm-1 F(000) 992 Crystal size 0.34 x 0.13 x 0.10 mm3 Theta range for data collection 3.82 to 29.35° Index ranges -20≤h≤21, -9≤k≤9, -27≤l≤27 Reflections collected 21446 Independent reflections 3116 [R(int) = 0.0576] Completeness to theta = 29.35° 99.6 % Absorption correction None Max. and min. transmission 0.9897 and 0.9657 Refinement method Full-matrix least-squares on F2 Data / parameters 3116 / 172 Goodness-of-fit on F2 0.980 Final R indices [I>2sigma(I)] R1 = 0.0478, wR2 = 0.1214 R indices (all data) R1 = 0.0620, wR2 = 0.1295 Largest diff. peak and hole 0.304 and -0.165 e-/Å3

------R1 =  ||Fo| - |Fc|| /  |Fo| wR2 = {  [w(Fo2 - Fc2)2] /  [w(Fo2)2] }1/2

139

Table 2. Atomic coordinates (x 104) and equivalent isotropic displacement parameters

(Å2x 103) for C5H9NO2. U(eq) is defined as one third of the trace of the orthogonalized Uij tensor. ______x y z U(eq) ______O(1) 771(1) -1432(1) 1876(1) 51(1) O(2) 1634(1) 441(1) 958(1) 42(1) N(1) 1448(1) 1205(1) 1583(1) 32(1) C(1) 1045(1) 131(2) 2020(1) 33(1) C(2) 1867(1) 2964(2) 1714(1) 45(1) C(3) 1534(1) 3864(2) 2342(1) 58(1) C(4) 1473(1) 2487(2) 2905(1) 50(1) C(5) 899(1) 926(2) 2704(1) 44(1) O(3) 656(1) 1986(1) 69(1) 38(1) O(4) -278(1) 3562(1) -876(1) 45(1) N(2) -609(1) 3080(1) -254(1) 36(1) C(6) -95(1) 2329(2) 193(1) 31(1) C(7) -1490(1) 3599(2) -171(1) 47(1) C(8) -1727(1) 3574(3) 593(1) 50(1) C(9) -1378(1) 1853(4) 925(1) 45(1) C(10) -458(1) 1901(2) 872(1) 38(1) C(8') -1920(3) 2496(7) 316(2) 40(1) C(9') -1411(3) 2866(10) 928(3) 41(1) ______

140

Table 3. Bond lengths [Å] and angles [°] for C5H9NO2 ______O(1)-C(1) 1.239(1) O(3)-C(6) 1.245(1)

O(2)-N(1) 1.397(1) O(4)-N(2) 1.394(1)

O(2)-H(2O) 0.88(2) O(4)-H(4O) 0.87(2)

N(1)-C(1) 1.329(2) N(2)-C(6) 1.325(2) N(1)-C(2) 1.453(2) N(2)-C(7) 1.459(2)

C(1)-C(5) 1.501(2) C(6)-C(10) 1.506(2) C(2)-C(3) 1.508(2) C(7)-C(8) 1.572(3) C(3)-C(4) 1.501(2) C(8)-C(9) 1.510(3) C(4)-C(5) 1.502(2) C(9)-C(10) 1.467(3)

C(7)-C(8') 1.430(5) C(10)-C(9') 1.670(6) C(8')-C(9') 1.492(7)

N(1)-O(2)-H(2O)106.4(12) N(2)-O(4)-H(4O) 105.7(12)

C(1)-N(1)-O(2) 117.46(10) C(6)-N(2)-O(4) 118.00(10) C(1)-N(1)-C(2) 127.41(10) C(6)-N(2)-C(7) 128.25(11) O(2)-N(1)-C(2) 113.98(9) O(4)-N(2)-C(7) 113.62(10)

O(1)-C(1)-N(1) 122.96(11) O(3)-C(6)-N(2) 122.47(11) O(1)-C(1)-C(5) 120.10(11) O(3)-C(6)-C(10) 120.49(11) N(1)-C(1)-C(5) 116.90(10) N(2)-C(6)-C(10) 117.04(11) N(1)-C(2)-C(3) 111.24(11) N(2)-C(7)-C(8) 109.75(12) C(4)-C(3)-C(2) 111.34(13) C(9)-C(8)-C(7) 110.38(18) C(3)-C(4)-C(5) 109.32(12) C(10)-C(9)-C(8) 108.4(2) C(1)-C(5)-C(4) 115.72(11) C(9)-C(10)-C(6) 116.85(13)

C(8')-C(7)-N(2) 113.3(2) C(8')-C(9')-C(10) 111.3(4) C(7)-C(8')-C(9') 101.5(4) C(6)-C(10)-C(9') 108.9(2) ______141

Table 4. Anisotropic displacement parameters (Å2x 103) for C5H9NO2. The anisotropic displacement factor exponent takes the form: -22[ h2 a*2U11 + ... + 2 h k a* b* U12 ] ______U11 U22 U33 U23 U13 U12 ______O(1) 70(1) 38(1) 46(1) -6(1) 5(1) -18(1) O(2) 40(1) 55(1) 30(1) 0(1) 3(1) 10(1) N(1) 31(1) 36(1) 29(1) 1(1) 1(1) -1(1) C(1) 33(1) 31(1) 35(1) 1(1) 0(1) -2(1) C(2) 47(1) 38(1) 49(1) 8(1) 1(1) -11(1) C(3) 70(1) 36(1) 69(1) -9(1) 3(1) -11(1) C(4) 54(1) 53(1) 43(1) -14(1) 0(1) -6(1) C(5) 55(1) 42(1) 36(1) -4(1) 9(1) -8(1) O(3) 36(1) 42(1) 35(1) -1(1) 2(1) 1(1) O(4) 63(1) 41(1) 31(1) 5(1) -4(1) -11(1) N(2) 42(1) 33(1) 33(1) 2(1) -2(1) -1(1) C(6) 37(1) 24(1) 31(1) -5(1) -2(1) -3(1) C(7) 42(1) 41(1) 59(1) 3(1) -10(1) 8(1) C(8) 43(1) 54(1) 53(1) 4(1) 1(1) 19(1) C(9) 37(1) 49(1) 49(1) 10(1) 5(1) 7(1) C(10) 42(1) 39(1) 31(1) -1(1) 2(1) 1(1) C(8') 33(2) 44(3) 45(3) -8(2) -3(2) -2(2) C(9') 36(3) 43(3) 44(3) 0(3) 7(2) 9(3) ______

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Table 5. Hydrogen coordinates (x 104) and isotropic displacement parameters (Å2x

10 3) for C5H9NO2. ______x y z U(eq) ______

H(2O) 1288(12) 970(30) 672(9) 69(5) H(2A) 1780 3812 1331 54 H(2B) 2479 2747 1762 54 H(3A) 1911 4895 2475 70 H(3B) 971 4394 2252 70 H(4A) 1251 3109 3310 60 H(4B) 2038 1990 3010 60 H(5A) 957 -90 3035 53 H(5B) 311 1379 2727 53 H(4O) -458(11) 2720(30) -1152(8) 66(5) H(7A) -1852 2714 -418 57 H(7B) -1585 4859 -356 57 H(7C) -1521 4923 -36 57 H(7D) -1780 3471 -606 57 H(8A) -1497 4697 814 60 H(8B) -2347 3596 642 60 H(9A) -1548 1820 1401 54 H(9B) -1600 724 703 54 H(10A) -238 677 1017 45 H(10B) -245 2843 1191 45 H(10C) -87 2405 1226 45 H(10D) -501 537 934 45 H(8'A) -1912 1160 196 48 H(8'B) -2510 2908 373 48 H(9'A) -1707 2359 1324 49 H(9'B) -1351 4226 991 49 ______

143

Table 6. Torsion angles [°] for C5H9NO2 ______O(2)-N(1)-C(1)-O(1) -6.27(17) C(2)-N(1)-C(1)-O(1) -173.16(12) O(2)-N(1)-C(1)-C(5) 176.17(10) C(2)-N(1)-C(1)-C(5) 9.27(18) C(1)-N(1)-C(2)-C(3) -22.01(18) O(2)-N(1)-C(2)-C(3) 170.71(11) N(1)-C(2)-C(3)-C(4) 46.37(17) C(2)-C(3)-C(4)-C(5) -59.18(17) O(1)-C(1)-C(5)-C(4) 160.80(13) N(1)-C(1)-C(5)-C(4) -21.57(18) C(3)-C(4)-C(5)-C(1) 46.40(18) O(4)-N(2)-C(6)-O(3) -2.47(16) C(7)-N(2)-C(6)-O(3) -178.17(12) O(4)-N(2)-C(6)-C(10) 177.28(9) C(7)-N(2)-C(6)-C(10) 1.58(18) C(6)-N(2)-C(7)-C(8') -28.4(3) O(4)-N(2)-C(7)-C(8') 155.8(2) C(6)-N(2)-C(7)-C(8) 13.0(2) O(4)-N(2)-C(7)-C(8) -162.85(13) N(2)-C(7)-C(8)-C(9) -44.4(2) C(7)-C(8)-C(9)-C(10) 61.8(3) C(8)-C(9)-C(10)-C(6) -48.1(3) O(3)-C(6)-C(10)-C(9) -163.59(17) N(2)-C(6)-C(10)-C(9) 16.7(2) O(3)-C(6)-C(10)-C(9') 169.6(3) N(2)-C(6)-C(10)-C(9') -10.1(3) N(2)-C(7)-C(8')-C(9') 59.2(4) C(7)-C(8')-C(9')-C(10) -69.3(5) C(6)-C(10)-C(9')-C(8') 46.0(5) ______

144

Table 7. Hydrogen bonds for C5H9NO2 [Å and °] ______D-H...A d(D-H) d(H...A) d(D...A) <(DHA) ______O(2)-H(2O)...O(3) 0.88(2) 1.73(2) 2.6101(13) 176.3(18) O(4)-H(4O)...O(1)#1 0.868(18) 1.789(18) 2.6383(14) 165.4(18) ______Symmetry transformations used to generate equivalent atoms: #1 -x,-y,-z

145

APPENDIX B

CRYSTAL STRUCTURE ANALYSIS REPORT AND TABLE FOR COMPOUND

C4H7NO2

Wake Forest X-Ray Facility Reference Code: a66n-2008

Performed by Dr. Cynthia Day Wake Forest University

146

EXPERIMENTAL

Colorless single crystals of C4H7NO2 are, at 193(2) K, monoclinic, space group

5 P21/n (an alternate setting of P21/c – C 2h (No. 14)) with a = 10.471(1) Å, b = 7.0758(9)

-3 Å, c = 12.929(2) Å, β = 95.461(2)° , V = 953.6(2) Å3, and Z = 8 {dcalcd = 1.409gcm ;

-1 μa(MoK ) = 0.113 mm }. A full hemisphere of diffracted intensities (1968 30-second frames with an  scan width of 0.30) was measured for a single-domain specimen using graphite-monochromated MoK radiation (= 0.71073 Å) on a Bruker SMART APEX CCD Single Crystal Diffraction System. X-rays were provided by a fine-focus sealed x-ray tube operated at 50kV and 30mA. Lattice constants were determined with the Bruker SAINT software package using peak centers for 4024 reflections having 7.82˚ ≤ 2θ ≤ 60.01˚. A total of 9994 integrated reflection intensities having 2((MoK )≤ 60.10 were produced using the

Bruker program SAINT; 2756 of these were unique and gave Rint = 0.033 with a coverage which was 98.7% complete. The data were corrected empirically (SADABS) for variable absorption effects using equivalent reflections; the transmission factors ranged from 0.8179 to 0.8925. The Bruker software package SHELXTL was used to solve the structure using “direct methods” techniques. All stages of weighted full-matrix least-squares 2 refinement were conducted using Fo data with the SHELXTL Version 6.12 software package. The resulting structural parameters have been refined to convergence {R1 (unweighted, based on F) = 0.0465 for 2330 independent reflections having 2Θ(MoK ) < o 2 2 60.10 and F >2σ(F )} {R1 (unweighted, based on F) = 0.0530 and wR2 (weighted, based on F2) = 0.1261 for all 2756 reflections} using counter-weighted full-matrix least-squares techniques and a structural model which incorporated anisotropic thermal parameters for all nonhydrogen atoms. Hydrogen atoms were located from a difference Fourier map and refined as independent isotropic atoms whose parameters were allowed to vary in least- squares refinement cycles. A total of 183 parameters were refined using no restraints and 2756 data. The largest shift/s.u. was 0.000 in the final refinement cycle. The final

147

3 difference map had maxima and minima of 0.405 and -0.199 e-/Å , respectively. Acknowledgment

The authors thank the National Science Foundation (grant CHE-0234489) for funds to purchase the x-ray instrument and computers. References

(5) International Tables for Crystallography, Vol A, 4th ed., Kluwer Academic Publishers: Boston (1996). (6) Data Collection: SMART (Version 5.628) (2002). Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373 USA. (7) Data Reduction: SAINT (Version 6.36A) (2003). Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373, USA. (8) G. M. Sheldrick (2006). SADABS-2006/3. BRUKER AXS Inc., 5465 East Cheryl Parkway, Madison, WI 53711-5373 USA (9) G. M. Sheldrick (2001). SHELXTL (Version 6.12). Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373 USA.

148

Table 1. Crystal data and structure refinement for C4H7NO2

Identification code a66n2m Empirical formula C4 H7 N O2 Formula weight 101.11 Temperature 193(2) K Wavelength 0.71073 Å Crystal system Monoclinic 5 Space group P21/n (an alternate setting of P21/c – C 2h (No. 14)) Unit cell dimensions a = 10.4711(13) Å b = 7.0758(9) Å, β = 95.461(2)° c = 12.9288(16) Å Volume 953.6(2) Å3 Z 8 Density (calculated) 1.409 g/cm3 Absorption coefficient 0.113 mm-1 F(000) 432 Crystal size 0.26 x 0.21 x 0.06 mm3 Theta range for data collection 3.91 to 30.05° Index ranges -14≤h≤14, -9≤k≤9, -18≤l≤17 Reflections collected 9994 Independent reflections 2756 [R(int) = 0.0328] Completeness to theta = 30.05° 98.7 % Absorption correction Multi-scan (SADABS) Max. and min. transmission 0.9932 and 0.9712 Refinement method Full-matrix least-squares on F2 Data / parameters 2756 / 183 Goodness-of-fit on F2 1.032

Final R indices [2330 I>2σ(I)] R1 = 0.0465, wR2 = 0.1199

R indices (all 2756 data) R1 = 0.0530, wR2 = 0.1261 Largest diff. peak and hole 0.405 and -0.199 e- /Å3 ------

R1 =  ||Fo| - |Fc|| /  |Fo| 2 2 2 2 2 1/2 wR2 = {  [w(Fo - Fc ) ] /  [w(Fo ) ] }

149

Table 2. Atomic coordinates a,b ( x 104) and equivalent isotropic displacement

parameters (Å2x 103) for C4H7NO2

______x y z U(eq) c ______O(1A) 5748(1) 2949(1) 5550(1) 34(1) O(2A) 3323(1) 1280(1) 5476(1) 37(1) N(1A) 3885(1) 2008(1) 4637(1) 31(1) C(1A) 5070(1) 2692(1) 4719(1) 28(1) C(2A) 3312(1) 1639(2) 3588(1) 36(1) C(3A) 4136(1) 2903(2) 2956(1) 35(1) C(4A) 5412(1) 3080(2) 3632(1) 32(1) O(1B) 1813(1) 3593(1) 6404(1) 40(1) O(2B) 2043(1) 5393(1) 4507(1) 36(1) N(1B) 1552(1) 6200(1) 5360(1) 31(1) C(1B) 1414(1) 5212(2) 6218(1) 30(1) C(2B) 912(1) 8021(2) 5273(1) 34(1) C(3B) 689(1) 8398(2) 6407(1) 38(1) C(4B) 661(1) 6436(2) 6897(1) 37(1) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit. b Atoms are labeled in agreement with Figures 1 and 2. c U(eq) is defined as one third of the trace of the orthogonalized Uij tensor.

150

a,b Table 3. Bond lengths [Å] and angles [°] for C4H7NO2

______O(1A)-C(1A) 1.2433(12) O(1B)-C(1B) 1.2356(14) O(2A)-N(1A) 1.3820(11) O(2B)-N(1B) 1.3830(11) O(2A)-H(2A) 0.873(19) O(2B)-H(2B) 0.887(18) N(1A)-C(1A) 1.3267(14) N(1B)-C(1B) 1.3309(13) N(1A)-C(2A) 1.4527(14) N(1B)-C(2B) 1.4519(14) C(1A)-C(4A) 1.5081(14) C(1B)-C(4B) 1.5070(15) C(2A)-C(3A) 1.5315(16) C(2B)-C(3B) 1.5306(16) C(2A)-H(2A1) 0.981(15) C(2B)-H(2B1) 0.957(16) C(2A)-H(2A2) 0.984(16) C(2B)-H(2B2) 0.992(15) C(3A)-C(4A) 1.5310(16) C(3B)-C(4B) 1.5270(17) C(3A)-H(3A1) 0.979(16) C(3B)-H(3B1) 0.974(16) C(3A)-H(3A2) 0.995(16) C(3B)-H(3B2) 1.022(18) C(4A)-H(4A1) 0.980(16) C(4B)-H(4B1) 0.989(17) C(4A)-H(4A2) 0.965(14) C(4B)-H(4B2) 0.89(2)

N(1A)-O(2A)-H(2A) 103.7(12) N(1B)-O(2B)-H(2B) 102.6(11)

C(1A)-N(1A)-O(2A) 122.53(9) C(1B)-N(1B)-O(2B) 121.76(9) C(1A)-N(1A)-C(2A) 116.12(9) C(1B)-N(1B)-C(2B) 116.40(9) O(2A)-N(1A)-C(2A) 119.80(9) O(2B)-N(1B)-C(2B) 120.54(8)

O(1A)-C(1A)-N(1A) 125.16(9) O(1B)-C(1B)-N(1B) 125.97(10) O(1A)-C(1A)-C(4A) 127.68(9) O(1B)-C(1B)-C(4B) 127.27(10) N(1A)-C(1A)-C(4A) 107.17(8) N(1B)-C(1B)-C(4B) 106.75(9) N(1A)-C(2A)-C(3A) 100.93(9) N(1B)-C(2B)-C(3B) 101.05(9) N(1A)-C(2A)-H(2A1)109.4(9) N(1B)-C(2B)-H(2B1) 108.3(9) C(3A)-C(2A)-H(2A1)114.9(9) C(3B)-C(2B)-H(2B1) 113.1(9) N(1A)-C(2A)-H(2A2)111.1(9) N(1B)-C(2B)-H(2B2) 107.9(9) C(3A)-C(2A)-H(2A2)111.2(8) C(3B)-C(2B)-H(2B2) 114.2(8) H(2A1)-C(2A)-H(2A2) H(2B1)-C(2B)-H(2B2) 111.4(13) C(4A)-C(3A)-C(2A)104.39(9) C(4B)-C(3B)-C(2B) 104.44(9) C(4A)-C(3A)-H(3A1)110.6(9) C(4B)-C(3B)-H(3B1) 114.1(10) C(2A)-C(3A)-H(3A1)114.1(9) C(2B)-C(3B)-H(3B1) 111.3(10)

151

C(4A)-C(3A)-H(3A2) 109.4(9) C(2A)-C(3A)-H(3A2) 108.0(8) H(3A1)-C(3A)-H(3A2) 110.2(12) C(1A)-C(4A)-C(3A) 104.17(9) C(1A)-C(4A)-H(4A1) 108.8(9) C(3A)-C(4A)-H(4A1) 112.1(9) C(1A)-C(4A)-H(4A2) 109.9(7) C(3A)-C(4A)-H(4A2) 113.7(8) H(4A1)-C(4A)-H(4A2) 108.0(11) C(4B)-C(3B)-H(3B2) 108.5(10) C(2B)-C(3B)-H(3B2) 109.1(9) H(3B1)-C(3B)-H(3B2) 109.3(13) C(1B)-C(4B)-C(3B) 104.45(9) C(1B)-C(4B)-H(4B1) 107.3(10) C(3B)-C(4B)-H(4B1) 113.7(10) C(1B)-C(4B)-H(4B2) 112.4(12) C(3B)-C(4B)-H(4B2) 114.5(12) H(4B1)-C(4B)-H(4B2) 104.5(15) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit. b Atoms are labeled in agreement with Figures 1 and 2.

152

a,b,c Table 4. Anisotropic displacement parameters (Å2x 103) for C4H7NO2

______U11 U22 U33 U23 U13 U12 ______O(1A) 35(1) 36(1) 29(1) -2(1) 0(1) 0(1) O(2A) 43(1) 31(1) 38(1) 5(1) 13(1) -1(1) N(1A) 34(1) 30(1) 29(1) 0(1) 5(1) -2(1) C(1A) 31(1) 22(1) 29(1) -2(1) 3(1) 4(1) C(2A) 36(1) 36(1) 34(1) -4(1) -2(1) -4(1) C(3A) 41(1) 37(1) 27(1) -1(1) 0(1) 1(1) C(4A) 34(1) 33(1) 28(1) -1(1) 5(1) 1(1) O(1B) 45(1) 35(1) 40(1) 7(1) 7(1) 6(1) O(2B) 38(1) 40(1) 30(1) -6(1) 5(1) -1(1) N(1B) 32(1) 32(1) 29(1) 0(1) 4(1) 3(1) C(1B) 27(1) 33(1) 30(1) 1(1) -1(1) -3(1) C(2B) 36(1) 28(1) 36(1) 1(1) -1(1) -1(1) C(3B) 41(1) 33(1) 41(1) -5(1) 7(1) 1(1) C(4B) 41(1) 39(1) 32(1) 0(1) 6(1) 2(1) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit. b 2 2 2 2 The form of the anisotropic thermal parameter is: exp[-22 (U11h a* + U22k b* + 2 2 U33l c* + 2U12hka*b* + 2U13hla*c* + 2U23klb*c*)]. c Atoms are labeled in agreement with Figures 1 and 2.

153

Table 5. Hydrogen coordinates ( x 104) and isotropic displacement parameters (Å2x 10 3) a,b for C4H7NO2

______x y z U(eq) ______H(2A) 2833(17) 2200(30) 5656(15) 55(5) H(2A1) 2398(14) 1970(20) 3538(12) 39(4) H(2A2) 3400(14) 300(20) 3403(11) 43(4) H(3A1) 4273(15) 2400(20) 2270(13) 47(4) H(3A2) 3718(14) 4170(20) 2884(12) 42(4) H(4A1) 6038(15) 2140(20) 3451(12) 44(4) H(4A2) 5804(12) 4310(20) 3603(10) 30(3) H(2B) 2796(17) 5970(30) 4500(13) 58(5) H(2B1) 133(15) 7880(20) 4830(12) 40(4) H(2B2) 1502(14) 8930(20) 4980(11) 37(3) H(3B1) -89(15) 9130(20) 6458(12) 49(4) H(3B2) 1457(16) 9120(30) 6758(13) 52(4) H(4B1) -210(16) 5890(20) 6884(13) 52(4) H(4B2) 967(18) 6390(30) 7561(16) 61(5) ______a All hydrogen atoms were located in a difference Fourier map and included in the

structural model as independent isotropic atoms whose parameters were allowed to

vary in least-squares refinement cycles.

b Hydrogen atoms are labeled with the same numerical and literal subscript(s) as their

respective oxygen or carbon atoms.

154

Table 6. Torsion angles [°] for C4H7NO2

______O(2A)-N(1A)-C(1A)-O(1A) -8.16(15) C(2A)-N(1A)-C(1A)-O(1A) -173.88(10) O(2A)-N(1A)-C(1A)-C(4A) 171.81(8) C(2A)-N(1A)-C(1A)-C(4A) 6.09(12) C(1A)-N(1A)-C(2A)-C(3A) -20.91(12) O(2A)-N(1A)-C(2A)-C(3A) 172.96(9) N(1A)-C(2A)-C(3A)-C(4A) 25.99(11) O(1A)-C(1A)-C(4A)-C(3A) -168.30(10) N(1A)-C(1A)-C(4A)-C(3A) 11.74(11) C(2A)-C(3A)-C(4A)-C(1A) -23.67(11) O(2B)-N(1B)-C(1B)-O(1B) 8.15(16) C(2B)-N(1B)-C(1B)-O(1B) 175.15(10) O(2B)-N(1B)-C(1B)-C(4B) -170.82(9) C(2B)-N(1B)-C(1B)-C(4B) -3.82(12) C(1B)-N(1B)-C(2B)-C(3B) 18.93(12) O(2B)-N(1B)-C(2B)-C(3B) -173.91(9) N(1B)-C(2B)-C(3B)-C(4B) -25.20(11) O(1B)-C(1B)-C(4B)-C(3B) 167.76(11) N(1B)-C(1B)-C(4B)-C(3B) -13.29(12) C(2B)-C(3B)-C(4B)-C(1B) 24.10(12) ______

155

Table 7. Inter-molecular Hydrogen bonds for C4H7NO2 [Å and °]

______D-H...A d(D-H) d(H...A) d(D...A) <(DHA) ______O(2A)-H(2A)...O(1B) 0.873(19) 1.802(19) 2.6419(12) 160.8(18) O(2B)-H(2B)...O(1A)#10.887(18) 1.714(18) 2.6007(12) 178.4(17) ______Symmetry transformations used to generate equivalent atoms: #1 -x+1,-y+1,-z+1

156

APPENDIX C

CRYSTAL STRUCTURE ANALYSIS REPORT AND TABLE FOR COMPOUND

C7H11NO2

Wake Forest X-Ray Facility Reference Code: a11m

Performed by Dr. Cynthia Day Wake Forest University

157

EXPERIMENTAL

Table 1. Crystal data and structure refinement for C7H11NO2

Identification code a11m Empirical formula C7 H11 N O2 Formula weight 141.17 Temperature 193(2) K Wavelength 0.71073 Å Crystal system Monoclinic 5 Space group P2(1)/c - C 2h (No. 14) Unit cell dimensions a = 13.040(3) Å b = 10.086(3) Å, β = 92.336(3)° c = 10.802(3) Å Volume 1419.5(6) Å3 Z 8 Density (calculated) 1.321 g/cm3 Absorption coefficient 0.097 mm-1 F(000) 608 Crystal size 0.18 x 0.12 x 0.08 mm3 Theta range for data collection 4.03 to 25.00° Index ranges -15≤h≤15, 0≤k≤11, 0≤l≤12 Reflections collected 3820 Independent reflections 3822 [R(int) = 0.0000] Completeness to theta = 25.00° 99.6 % Refinement method Full-matrix least-squares on F2 Data / parameters 3822 / 190 Goodness-of-fit on F2 1.194

Final R indices [I>2sigma(I)] R1 = 0.0684, wR2 = 0.1301

R indices (all data) R1 = 0.0845, wR2 = 0.1365 Largest diff. peak and hole 0.213 and -0.172 e-/Å3 ------

R1 =  ||Fo| - |Fc|| /  |Fo| 2 2 2 2 2 1/2 wR2 = {  [w(Fo - Fc ) ] /  [w(Fo ) ] }

158

a,b Table 3. Bond lengths [Å] and angles [°] for C7H11NO2

______O(1A)-N(1A) 1.387(2) O(1B)-N(1B) 1.383(2)

O(1A)-H(1A) 0.94(2) O(1B)-H(1B) 0.90(3)

O(2A)-C(1A) 1.237(2) O(2B)-C(1B) 1.230(3)

N(1A)-C(1A) 1.327(3) N(1B)-C(1B) 1.321(3)

N(1A)-C(2A) 1.454(3) N(1B)-C(2B) 1.456(3)

C(1A)-C(4A) 1.495(3) C(1B)-C(4B) 1.496(3)

C(2A)-C(5A) 1.519(3) C(2B)-C(5B) 1.520(3) C(2A)-C(3A) 1.549(3) C(2B)-C(3B) 1.553(3) C(3A)-C(7A) 1.524(3) C(3B)-C(7B) 1.520(3) C(3A)-C(4A) 1.528(3) C(3B)-C(4B) 1.529(3) C(5A)-C(6A) 1.518(3) C(5B)-C(6B) 1.518(4) C(6A)-C(7A) 1.519(3) C(6B)-C(7B) 1.506(4)

N(1A)-O(1A)-H(1A) N(1B)-O(1B)-H(1B) 104.9(18)

C(1A)-N(1A)-O(1A) C(1B)-N(1B)-O(1B) 121.56(17) C(1A)-N(1A)-C(2A) C(1B)-N(1B)-C(2B) 117.72(18) O(1A)-N(1A)-C(2A) O(1B)-N(1B)-C(2B) 120.34(18)

O(2A)-C(1A)-N(1A) O(2B)-C(1B)-N(1B) 126.4(2) O(2A)-C(1A)-C(4A) O(2B)-C(1B)-C(4B) 126.2(2) N(1A)-C(1A)-C(4A) N(1B)-C(1B)-C(4B) 107.37(19

159

N(1A)-C(2A)-C(5A) 111.66(18) N(1A)-C(2A)-C(3A) 102.36(17) C(5A)-C(2A)-C(3A) 106.47(18) C(7A)-C(3A)-C(4A) 114.47(19) N(1B)-C(2B)-C(5B) 112.04(19) N(1B)-C(2B)-C(3B) 102.35(18) C(5B)-C(2B)-C(3B) 105.7(2) C(7B)-C(3B)-C(4B) 113.6(2) C(7A)-C(3A)-C(2A) 104.70(18) C(4A)-C(3A)-C(2A) 105.73(17) C(1A)-C(4A)-C(3A) 106.69(18) C(6A)-C(5A)-C(2A) 103.55(19) C(5A)-C(6A)-C(7A) 102.39(19) C(6A)-C(7A)-C(3A) 104.17(19) C(7B)-C(3B)-C(2B) 104.7(2) C(4B)-C(3B)-C(2B) 105.41(17) C(1B)-C(4B)-C(3B) 106.87(19) C(6B)-C(5B)-C(2B) 104.6(2) C(7B)-C(6B)-C(5B) 102.0(2) C(6B)-C(7B)-C(3B) 104.6(2) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit. b Atoms are labeled in agreement with Figures 1 and 2.

160

Table 7. Inter-molecular Hydrogen bonds for C7H11NO2 [Å and °]

______D-H...A d(D-H) d(H...A) d(D...A) <(DHA) ______O(1A)-H(1A)...O(2B)#10.94(2) 1.71(3) 2.648(2) 178(2) O(1B)-H(1B)...O(2A) 0.90(3) 1.69(3) 2.584(2) 173(3) ______Symmetry transformations used to generate equivalent atoms: #1 -x+1,y+1/2,-z+1/2

161

APPENDIX D

CRYSTAL STRUCTURE ANALYSIS REPORT AND TABLE FOR COMPOUND

C15H17SO3N

Wake Forest X-Ray Facility Reference Code: a92ktab

Performed by Dr. Cynthia Day Wake Forest University

162

Table 1. Crystal data and structure refinement for C15H17SO3N Identification code a92k Empirical formula C15 H17 N O3 S Formula weight 291.36 Temperature 193(2) K Wavelength 0.71073 Å Crystal system Monoclinic 5 Space group P21/c - C 2h (No. 14) Unit cell dimensions a = 16.8053(17) Å b = 7.8504(8) Å, β = 105.649(2)° c = 11.6413(12) Å Volume 1478.9(3) Å3 Z 4 Density (calculated) 1.309 g/cm3 Absorption coefficient 0.225 mm-1 F(000) 616 Crystal size 0.29 x 0.26 x 0.19 mm3 Theta range for data collection 4.16 to 30.04° Index ranges -23≤h≤23, -10≤k≤11, -16≤l≤16 Reflections collected 14854 Independent reflections 4292 [R(int) = 0.0321] Completeness to theta = 30.04° 99.1 % Absorption correction Multi-scan (SADABS) Max. and min. transmission 0.9585 and 0.9376 Refinement method Full-matrix least-squares on F2 Data / parameters 4292 / 183 Goodness-of-fit on F2 1.139 Final R indices [I>2sigma(I)] R1 = 0.0579, wR2 = 0.1444 R indices (all data) R1 = 0.0635, wR2 = 0.1485 Largest diff. peak and hole 0.552 and -0.246 e-/Å-3

163

Table 2. Atomic coordinates (x 104) and equivalent isotropic displacement parameters

(Å2x 103) for C15H17SO3N. U(eq) is defined as one third of the trace of the orthogonalized Uij tensor. ______x y z U(eq) ______S(1) 2242(1) 3423(1) 3160(1) 30(1) O(1) 2640(1) 1791(2) 3364(1) 45(1) O(2) 2129(1) 4251(2) 2035(1) 41(1) O(3) 2420(1) 6311(1) 4073(1) 34(1) N(1) 2843(1) 4712(2) 4179(1) 31(1) C(1) 1281(1) 3303(2) 3482(1) 28(1) C(2) 636(1) 4344(2) 2874(1) 34(1) C(3) -115(1) 4255(2) 3145(2) 41(1) C(4) -231(1) 3173(2) 4030(2) 39(1) C(5) 429(1) 2178(2) 4644(2) 39(1) C(6) 1182(1) 2214(2) 4371(1) 34(1) C(7) -1054(1) 3078(3) 4315(2) 58(1) C(8) 3033(1) 4141(3) 5425(2) 44(1) C(9) 2932(1) 7609(2) 3738(2) 37(1) C(10) 3662(1) 8065(2) 4755(1) 31(1) C(11) 4443(1) 7464(3) 4803(2) 46(1) C(12) 5106(1) 7893(4) 5763(2) 64(1) C(13) 4987(2) 8920(4) 6652(2) 67(1) C(14) 4209(2) 9511(3) 6615(2) 55(1) C(15) 3545(1) 9085(2) 5666(2) 38(1) ______

164

Table 3. Bond lengths [Å] and angles [°] for C15H17SO3N ______S(1)-O(2) 1.4277(13) S(1)-N(1) 1.6748(14) S(1)-O(1) 1.4341(13) S(1)-C(1) 1.7565(16)

O(3)-N(1) 1.4309(17) O(3)-C(9) 1.453(2) N(1)-C(8) 1.468(2)

C(1)-C(6) 1.387(2) C(10)-C(11) 1.382(2) C(1)-C(2) 1.389(2) C(10)-C(15) 1.384(2) C(2)-C(3) 1.383(3) C(11)-C(12) 1.389(3) C(3)-C(4) 1.389(3) C(12)-C(13) 1.369(4) C(4)-C(5) 1.387(3) C(13)-C(14) 1.376(4) C(5)-C(6) 1.385(2) C(14)-C(15) 1.383(3)

C(4)-C(7) 1.509(3) C(9)-C(10) 1.500(2)

O(2)-S(1)-O(1) 119.97(9) O(2)-S(1)-C(1) 108.50(8) O(2)-S(1)-N(1) 106.30(7) O(1)-S(1)-C(1) 109.29(8) O(1)-S(1)-N(1) 104.94(8) N(1)-S(1)-C(1) 107.08(7)

N(1)-O(3)-C(9) 108.77(12) O(3)-N(1)-C(8) 108.95(13) O(3)-N(1)-S(1) 106.19(9) O(3)-C(9)-C(10) 111.90(14) C(8)-N(1)-S(1) 116.39(11)

C(6)-C(1)-C(2) 120.56(15) C(11)-C(10)-C(9) 121.05(17) C(6)-C(1)-S(1) 119.67(12) C(15)-C(10)-C(9) 119.25(16) C(2)-C(1)-S(1) 119.73(12) C(10)-C(11)-C(12) 119.7(2) C(3)-C(2)-C(1) 119.31(16) C(13)-C(12)-C(11) 120.2(2) C(2)-C(3)-C(4) 121.25(17) C(12)-C(13)-C(14) 120.3(2) C(5)-C(4)-C(3) 118.32(16) C(13)-C(14)-C(15) 119.9(2) C(5)-C(4)-C(7) 120.78(19) C(14)-C(15)-C(10) 120.12(19) C(3)-C(4)-C(7) 120.91(19) C(6)-C(5)-C(4) 121.54(16) C(5)-C(6)-C(1) 118.99(16) C(11)-C(10)-C(15) 119.69(17) 165

Table 4. Anisotropic displacement parameters (Å2x 103) for C15H17SO3N. The anisotropic displacement factor exponent takes the form: -2π2[ h2 a*2U11 + ... + 2 h k a* b* U12 ] ______U11 U22 U33 U23 U13 U12 ______S(1) 35(1) 24(1) 31(1) -2(1) 9(1) -2(1) O(1) 47(1) 27(1) 63(1) -5(1) 16(1) 4(1) O(2) 51(1) 44(1) 30(1) -2(1) 14(1) -9(1) O(3) 32(1) 24(1) 44(1) -3(1) 7(1) 0(1) N(1) 31(1) 24(1) 35(1) -1(1) 4(1) 0(1) C(1) 32(1) 24(1) 25(1) -1(1) 4(1) -4(1) C(2) 41(1) 30(1) 30(1) 3(1) 4(1) 2(1) C(3) 37(1) 39(1) 43(1) -5(1) 2(1) 4(1) C(4) 37(1) 38(1) 41(1) -16(1) 11(1) -10(1) C(5) 46(1) 38(1) 34(1) -1(1) 11(1) -13(1) C(6) 39(1) 30(1) 31(1) 5(1) 3(1) -5(1) C(7) 46(1) 65(1) 70(2) -29(1) 27(1) -15(1) C(8) 47(1) 42(1) 34(1) 5(1) -4(1) -4(1) C(9) 45(1) 26(1) 36(1) 2(1) 3(1) -5(1) C(10) 36(1) 25(1) 33(1) 1(1) 9(1) -5(1) C(11) 42(1) 52(1) 49(1) 5(1) 19(1) 3(1) C(12) 33(1) 87(2) 69(2) 26(1) 8(1) -2(1) C(13) 60(1) 79(2) 46(1) 19(1) -13(1) -33(1) C(14) 80(2) 47(1) 32(1) -3(1) 5(1) -18(1) C(15) 48(1) 31(1) 36(1) -1(1) 12(1) -2(1) ______

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Table 5. Hydrogen coordinates (x 104) and isotropic displacement parameters (Å2x

10 3) for C15H17SO3N ______x y z U(eq) ______

H(2) 710 5108 2279 41 H(3) -560 4948 2717 49 H(5) 363 1454 5266 47 H(6) 1624 1504 4787 41 H(7A) -966 2790 5159 87 H(7B) -1396 2199 3820 87 H(7C) -1334 4182 4151 87 H(8A) 3262 2985 5487 66 H(8B) 2527 4142 5689 66 H(8C) 3439 4913 5928 66 H(9A) 2595 8642 3470 44 H(9B) 3129 7188 3062 44 H(11) 4527 6761 4181 55 H(12) 5642 7473 5802 77 H(13) 5444 9226 7299 80 H(14) 4129 10211 7240 66 H(15) 3008 9493 5639 46 ______

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Table 6. Torsion angles [°] for C15H17SO3N ______C(9)-O(3)-N(1)-C(8) 115.63(14) C(9)-O(3)-N(1)-S(1) -118.29(11) O(2)-S(1)-N(1)-O(3) 57.37(12) O(1)-S(1)-N(1)-O(3) -174.55(10) C(1)-S(1)-N(1)-O(3) -58.46(11) O(2)-S(1)-N(1)-C(8) 178.79(13) O(1)-S(1)-N(1)-C(8) -53.12(15) C(1)-S(1)-N(1)-C(8) 62.96(14) O(2)-S(1)-C(1)-C(6) 164.73(13) O(1)-S(1)-C(1)-C(6) 32.25(15) N(1)-S(1)-C(1)-C(6) -80.92(14) O(2)-S(1)-C(1)-C(2) -17.58(15) O(1)-S(1)-C(1)-C(2) -150.06(13) N(1)-S(1)-C(1)-C(2) 96.78(13) C(6)-C(1)-C(2)-C(3) -1.5(2) S(1)-C(1)-C(2)-C(3) -179.18(13) C(1)-C(2)-C(3)-C(4) 1.4(3) C(2)-C(3)-C(4)-C(5) 0.2(3) C(2)-C(3)-C(4)-C(7) -179.65(17) C(3)-C(4)-C(5)-C(6) -1.6(3) C(7)-C(4)-C(5)-C(6) 178.16(17) C(4)-C(5)-C(6)-C(1) 1.5(3) C(2)-C(1)-C(6)-C(5) 0.1(2) S(1)-C(1)-C(6)-C(5) 177.74(12) N(1)-O(3)-C(9)-C(10) -72.79(17) O(3)-C(9)-C(10)-C(11) 103.30(19) O(3)-C(9)-C(10)-C(15) -75.81(19) C(15)-C(10)-C(11)-C(12) -0.2(3) C(9)-C(10)-C(11)-C(12) -179.28(18) C(10)-C(11)-C(12)-C(13) -0.6(3) C(11)-C(12)-C(13)-C(14) 1.2(4) C(12)-C(13)-C(14)-C(15) -0.8(3) C(13)-C(14)-C(15)-C(10) 0.0(3) C(11)-C(10)-C(15)-C(14) 0.5(3) 168

C(9)-C(10)-C(15)-C(14) 179.60(17) ______

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APPENDIX E

CRYSTAL STRUCTURE ANALYSIS REPORT AND TABLE FOR COMPOUND

C8H13NO2

Wake Forest X-Ray Facility Reference Code: a71q-2010

Performed by Dr. Cynthia Day Wake Forest University

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EXPERIMENTAL

5 Crystals of C8H13NO2 are, at 173(2) K, monoclinic, space group P21/c – C 2h (No. 14) with a = 4.8089(12) Å, b = 21.598(5) Å, c = 8.156(2) Å, β = 106.278(3)°, V =

-3 -1 813.1(3)Å3, and Z = 4 {dcalcd = 1.268gcm ; μa(MoK ) = 0.091 mm }. A full hemisphere of diffracted intensities (1968 30-second frames with an  scan width of 0.30) was measured for a single-domain specimen using graphite-monochromated

MoK radiation (= 0.71073 Å) on a Bruker SMART APEX CCD Single Crystal Diffraction System. X-rays were provided by a fine-focus sealed x-ray tube operated at 50kV and 30mA. Lattice constants were determined with the Bruker APEX2 software package using peak centers for 1087 reflections having 7.69˚ ≤ 2θ ≤ 42.80˚. A total of 7741 integrated reflection intensities having 2((MoK )≤ 55.74 were produced using the

Bruker program SAINT; 1938 of these were unique and gave Rint = 0.051 with a coverage which was 99.8% complete. The data were corrected empirically for variable scaling and absorption effects using the SADABS program; the estimated minimum and maximum transmission values reported were 0.6422 and 0.7460. The Bruker software package SHELXTL was used to solve the structure using “direct methods” techniques. All stages of weighted full-matrix least-squares 2 refinement were conducted using Fo data with the SHELXTL software package. The resulting structural parameters have been refined to convergence {R1 (unweighted, based on F) = 0.0488 for 1354 independent reflections having 2Θ(MoK ) < 55.74o and 2 2 2 F >2σ(F )} {R1 (unweighted, based on F) = 0.0787 and wR2 (weighted, based on F ) = 0.1152 for all 1938 reflections} using counter-weighted full-matrix least-squares techniques and a structural model which incorporated anisotropic thermal parameters for all nonhydrogen atoms. All hydrogen atoms were located from a difference Fourier map and included in the structural model as individual isotropic atoms whose parameters were allowed to vary in least-squares refinement cycles. A total of 152 parameters were refined using no restraints and 1938 data. The largest shift/s.u. was 0.000 in the final refinement cycle. The final difference map had maxima and minima of 0.228 and

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3 -0.169 e-/Å , respectively.

Acknowledgment

The authors thank the National Science Foundation (grant CHE-0234489) for funds to purchase the x-ray instrument and computers.

References

International Tables for Crystallography, Vol A, 4th ed., Kluwer Academic Publishers: Boston (1996). Data Collection: SMART (Version 5.628) (2002). Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373 USA. (10) Data Reduction: SAINT (Version 7.66A) (2009). Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373, USA. (11) G. M. Sheldrick (2008). SADABS (Version 2008/1). Program for Empirical Absorption Correction of Area Detector Data. University of Göttingen, Germany. (12) G. M. Sheldrick (2008). SHELXTL. Bruker-AXS, 5465 E. Cheryl Parkway, Madison, WI 53711-5373 USA. .

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Table 1. Crystal data and structure refinement for C8H13NO2

Identification code a71q Empirical formula C8 H13 N O2 Formula weight 155.19 Temperature 173(2) K Wavelength 0.71073 Å Crystal system Monoclinic 5 Space group P21/c – C 2h (No. 14) Unit cell dimensions a = 4.8089(12) Å b = 21.598(5) Å, β = 106.278(3)° c = 8.156(2) Å Volume 813.1(3) Å3 Z 4 Density (calculated) 1.268 g/cm3 Absorption coefficient 0.091 mm-1 F(000) 336 Crystal size 0.17 x 0.10 x 0.03 mm3 Theta range for data collection 3.85 to 27.87° Index ranges -6≤h≤6, -28≤k≤28, -10≤l≤10 Reflections collected 7741 Independent reflections 1938 [R(int) = 0.0506] Completeness to theta = 27.87° 99.8 % Absorption correction Multi-scan (SADABS) Max. and min. transmission 0.7460 and 0.6422 Refinement method Full-matrix least-squares on F2 Data / parameters 1938 / 152 Goodness-of-fit on F2 1.018

Final R indices [1354 I>2σ(I) data] R1 = 0.0488, wR2 = 0.1030

R indices (all data) R1 = 0.0787, wR2 = 0.1152 Largest diff. peak and hole 0.228 and -0.169e-/Å3 ------

R1 =  ||Fo| - |Fc|| /  |Fo| 2 2 2 2 2 1/2 wR2 = {  [w(Fo - Fc ) ] /  [w(Fo ) ] }

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Table 2. Atomic coordinates a,b ( x 104) and equivalent isotropic displacement

parameters (Å2x 103) for C8H13NO2

______x y z U(eq) c ______O(1) 5896(2) 3237(1) 5685(2) 32(1) O(2) 5023(2) 2519(1) 3050(2) 36(1) N(1) 3277(3) 3271(1) 4387(2) 27(1) C(1) 3143(3) 2911(1) 3008(2) 27(1) C(2) 1960(3) 3892(1) 4263(2) 27(1) C(3) 3590(4) 4328(1) 3432(2) 31(1) C(4) 2975(4) 4419(1) 1762(2) 37(1) C(5) 565(4) 4123(1) 426(2) 40(1) C(6) 1001(4) 3435(1) 105(2) 35(1) C(7) 566(4) 2983(1) 1468(2) 31(1) C(8) 1842(4) 4110(1) 6010(2) 36(1) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit. b Atoms are labeled in agreement with Figures 1 and 2. c U(eq) is defined as one third of the trace of the orthogonalized Uij tensor.

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a,b Table 3. Bond lengths [Å] and angles [°] for C8H13NO2

______O(1)-N(1) 1.4018(16) O(1)-H(1) 0.91(2)

O(2)-C(1) 1.2324(19) C(3)-C(4) 1.325(2)

N(1)-C(1) 1.355(2) N(1)-C(2) 1.474(2)

C(1)-C(7) 1.505(2) C(2)-C(8) 1.516(2) C(2)-C(3) 1.503(2) C(5)-C(6) 1.533(3) C(4)-C(5) 1.494(3) C(6)-C(7) 1.537(2)

C(2)-H(2) 0.937(16) C(6)-H(6B) 0.963(18) C(3)-H(3) 0.975(18) C(7)-H(7A) 0.969(18) C(4)-H(4) 0.97(2) C(7)-H(7B) 0.973(17) C(5)-H(5A) 0.992(19) C(8)-H(8A) 0.95(2) C(5)-H(5B) 0.982(19) C(8)-H(8B) 0.989(18) C(6)-H(6A) 0.973(18) C(8)-H(8C) 0.982(18)

N(1)-O(1)-H(1)103.8(14)

C(1)-N(1)-O(1)114.87(13) C(4)-C(3)-C(2) 124.19(17) C(1)-N(1)-C(2)123.13(13) C(4)-C(3)-H(3) 121.0(11) O(1)-N(1)-C(2)112.18(11) C(2)-C(3)-H(3) 114.7(11) O(2)-C(1)-N(1)120.56(14) C(3)-C(4)-C(5) 126.18(18) O(2)-C(1)-C(7)121.33(15) C(3)-C(4)-H(4) 118.1(12) N(1)-C(1)-C(7)118.04(15) C(5)-C(4)-H(4) 115.7(12)

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N(1)-C(2)-C(3) 109.40(13) N(1)-C(2)-C(8) 110.29(14) C(3)-C(2)-C(8) 112.98(15) N(1)-C(2)-H(2) 105.1(10) C(3)-C(2)-H(2) 111.1(10) C(8)-C(2)-H(2) 107.6(10) C(4)-C(5)-C(6) 115.11(15) C(4)-C(5)-H(5A) 109.9(10) C(6)-C(5)-H(5A) 109.7(10) C(4)-C(5)-H(5B) 110.1(11) C(6)-C(5)-H(5B) 106.4(11) H(5A)-C(5)-H(5B)105.1(14) C(5)-C(6)-C(7) 116.07(15) C(5)-C(6)-H(6A) 109.8(10) C(7)-C(6)-H(6A) 109.9(10) C(5)-C(6)-H(6B) 108.2(10) C(7)-C(6)-H(6B) 105.0(10) H(6A)-C(6)-H(6B)107.5(14) C(1)-C(7)-C(6) 114.95(14) C(1)-C(7)-H(7A) 111.8(10) C(6)-C(7)-H(7A) 108.2(10) C(1)-C(7)-H(7B) 106.1(10) C(6)-C(7)-H(7B) 107.0(10) H(7A)-C(7)-H(7B)108.5(14) C(2)-C(8)-H(8A) 110.5(11) C(2)-C(8)-H(8B) 109.7(11) H(8A)-C(8)-H(8B)111.1(16) C(2)-C(8)-H(8C) 110.4(10) H(8A)-C(8)-H(8C)106.6(15) H(8B)-C(8)-H(8C)108.5(15) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit. b Atoms are labeled in agreement with Figures 1 and 2.

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a,b,c Table 4. Anisotropic displacement parameters (Å2x 103) for C8H13NO2

______U11 U22 U33 U23 U13 U12 ______O(1) 29(1) 34(1) 26(1) 3(1) -1(1) -3(1) O(2) 38(1) 37(1) 32(1) -2(1) 7(1) 6(1) N(1) 26(1) 28(1) 23(1) 0(1) 0(1) -1(1) C(1) 29(1) 27(1) 24(1) 3(1) 9(1) -6(1) C(2) 25(1) 30(1) 26(1) 0(1) 7(1) 0(1) C(3) 36(1) 25(1) 34(1) -2(1) 13(1) 2(1) C(4) 46(1) 29(1) 39(1) 5(1) 20(1) 6(1) C(5) 48(1) 44(1) 27(1) 10(1) 11(1) 14(1) C(6) 36(1) 44(1) 22(1) -2(1) 4(1) 3(1) C(7) 30(1) 34(1) 28(1) -5(1) 6(1) -5(1) C(8) 39(1) 38(1) 31(1) -4(1) 11(1) -3(1) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit. b 2 2 2 2 The form of the anisotropic thermal parameter is: exp[-22 (U11h a* + U22k b* + 2 2 U33l c* + 2U12hka*b* + 2U13hla*c* + 2U23klb*c*)]. c Atoms are labeled in agreement with Figures 1 and 2.

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Table 5. Hydrogen coordinates ( x 104) and isotropic displacement parameters (Å2x a,b 10 3) for C8H13NO2 ______x y z U(eq) ______H(1) 5510(50) 2968(10) 6450(30) 59(7) H(2) 50(40) 3839(7) 3590(20) 25(4) H(3) 5240(40) 4531(8) 4220(20) 38(5) H(4) 4210(40) 4695(9) 1350(20) 48(6) H(5A) 210(40) 4356(8) -660(20) 38(5) H(5B) -1250(40) 4152(8) 750(20) 39(5) H(6A) 2900(40) 3372(7) -70(20) 29(4) H(6B) -420(40) 3315(7) -930(20) 34(5) H(7A) -1150(40) 3108(8) 1780(20) 32(5) H(7B) 220(30) 2576(8) 940(20) 32(5) H(8A) 940(40) 3809(9) 6530(20) 44(5) H(8B) 810(40) 4511(9) 5900(20) 37(5) H(8C) 3810(40) 4168(8) 6770(20) 36(5) ______a All hydrogen atoms were located from a difference Fourier map and included in the

structural model as individual isotropic atoms whose parameters were allowed to

vary in least-squares refinement cycles. b Hydrogen atoms are labeled with the same numerical subscript(s) as their respective

oxygen or carbon atoms with an additional literal subscript (a, b or c) to distinguish

between hydrogens bonded to the same carbon atom.

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a Table 6. Torsion angles [°] for C8H13NO2 ______

O(1)-N(1)-C(1)-O(2) 10.7(2) C(2)-N(1)-C(1)-O(2) 153.80(14) O(1)-N(1)-C(1)-C(7) -172.23(13) C(2)-N(1)-C(1)-C(7) -29.2(2) C(1)-N(1)-C(2)-C(3) -68.76(19) O(1)-N(1)-C(2)-C(3) 75.17(16) C(1)-N(1)-C(2)-C(8) 166.38(14) O(1)-N(1)-C(2)-C(8) -49.69(17) N(1)-C(2)-C(3)-C(4) 88.99(19) C(8)-C(2)-C(3)-C(4) -147.73(17) C(2)-C(3)-C(4)-C(5) 1.4(3) C(3)-C(4)-C(5)-C(6) -72.4(2) C(4)-C(5)-C(6)-C(7) 75.3(2) O(2)-C(1)-C(7)-C(6) -89.25(19) N(1)-C(1)-C(7)-C(6) 93.74(19) C(5)-C(6)-C(7)-C(1) -82.4(2) ______a The numbers in parentheses are the estimated standard deviations in the last significant digit.

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Table 7. Hydrogen bonds for C8H13NO2 [Å and °]

______d(D-H) d(H...A) d(D...A) <(DHA) ______O(1)-H(1)...O(2)#1 0.91(2) 1.74(2) 2.6485(18) 175(2) ______Symmetry transformations used to generate equivalent atoms: #1 x,-y+1/2,z+1/2

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SCHOLASTIC VITA

RANJAN BANERJEE

BORN December, 1980 Howrah, West Bengal, India

EDUCATION Ph.D, Organic Chemistry, Wake Forest University Winston Salem, NC, December 2010

MS, Inorganic Chemistry, Indian Institute of Technology-Kharagpur, June 2005

B.Sc., University of Calcutta, Kolkata, July 2003

SCHOLASTIC AND PROFESSIONAL EXPERIENCE

Research Assistant 2006-2010

Teaching Assistant 2005-2006

AWARDS AND HONORS

Royal Society of Science Summer Research Fellowship, 2004 IISc Bangalore, India

“First Class” recognition from the University of Calcutta 2003 for outstanding undergraduate academic performance

PROFESSIONAL SOCIETIES:

American Chemical Society 2007-present

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CONFERENCE PRESENTATIONS

Ranjan Banerjee and S. Bruce King “Utilizing the Nitroso-ene Reaction for the Synthesis of Hydroxamic Acid-Based Natural Products” Abstract Submitted for 240th ACS National Meeting, Boston, MA, August 22-26, 2010 Ranjan Banerjee and S. Bruce King. “Synthesis of Cyclic Hydroxamic Acids through – NOH Insertion of Ketones” Oral presentation at 61st Southeast Regional Meeting of the American Chemical Society, San Juan, PR, October 21-24, 2009 Ranjan Banerjee and S. Bruce King. “New Synthetic Approaches Towards The Natural Hydroxamic Acid Cobactin Core” Presented at 41st National Organic Symposium, Boulder, CO, June 7-11, 2009 Ranjan Banerjee and S. Bruce King. “A New Synthetic Route to Cyclic Hydroxamic Acids” Presented at 40th National Organic Symposium, Durham, NC, June 3-7, 2007

PEER-REVIEWED PUBLICATIONS

R. Banerjee and S. Bruce King “Synthesis of Cyclic Hydroxamic Acids through –NOH Insertion of Ketones” Org. Lett. 2009, 11 (20), 4580-4583

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