Borinic Acid-Catalyzed Sulfation and Boronic Acid-Promoted Esterification of Carbohydrates

Yu Chen Lin

A thesis submitted in conformity with the requirements for the degree of Master of Science Department of Chemistry University of Toronto

Yu Chen Lin

Master of Science

Department of Chemistry University of Toronto

2017 Abstract

Carbohydrates and their O-sulfates play important roles in biological functions, including cellular recognition and adhesion, neural processes, fibrosis, growth factor regulation, cancer metastasis, and cellular entry of viruses. However, preparation of sulfated carbohydrates remains a synthetic challenge with conventional methods requiring lengthy protection and deprotection steps.

Described herein is our work toward the development of a method for the regioselective sulfation of fully unprotected carbohydrates using a borinic acid catalyst. Via an activated 1,2-cis-borinate intermediate, our method was shown to be robust in the sulfation of a range of substrates, including the synthesis of a sulfated galactosylceramide found in mammalian nervous systems. In addition, work on utilizing boronic acids as protective groups for the preparation of sugar fatty acid ester surfactants is also discussed.

Acknowledgments

I would like to thank my supervisor, Professor Mark Taylor, for the opportunity to join his lab, and his continued support in my studies, research, and pursuits.

I am also very grateful to all the Taylor lab members whom I’ve had the pleasure of meeting and working together with. You have all made me feel so at home here, and although it has only been a year, the great memories I’ve had here will stay with me long after. Furthermore, I’d like to give a shout out to my friends in and around the Department for every laughter and drink we’ve shared.

Lastly, I would like to thank my family for their unconditional love and support throughout the years and during my degree.

iii Table of Contents

Acknowledgments ...... iii

Table of Contents ...... iv

List of Tables ...... vi

List of Figures ...... vii

List of Abbreviations ...... ix

Chapter 1 Introduction ...... 1

1.1 Sulfated carbohydrates ...... 1

1.2 Direct, regioselective synthesis of sulfated carbohydrates ...... 4

1.3 Synthesis of sulfated carbohydrates via masked sulfates ...... 11

1.4 Organoboron compounds in carbohydrate chemistry ...... 18

1.5 Scope of Thesis ...... 20

Chapter 2 Regioselective, catalytic sulfation of unprotected carbohydrates ...... 21

2.1 Sulfation of carbohydrates with 2,2,2-trichloroethyl chlorosulfate ...... 21

2.2 Sulfation of carbohydrates with alkyl and aryl 1,2-dimethylimidazolium salt ...... 22

2.3 Sulfation of carbohydrates with sulfur trioxide amine complexes ...... 29

2.4 Summary and future work ...... 32

2.5 Experimental ...... 33

2.5.1. General Information ...... 33

2.5.2. General Procedure A ...... 34

2.5.3. Preparation of catalyst and carbohydrate substrates ...... 34

2.5.4. Synthesis and characterization of compounds ...... 36

Chapter 3 Boronic acid-promoted Fischer esterification ...... 47

3.1 Introduction ...... 47

3.2 Chemical synthesis of sugar fatty acid esters ...... 49

3.3 Summary ...... 51

3.4 Experimental ...... 51

3.4.1 General Information ...... 51

3.4.2 General Procedure B ...... 52

3.4.3 Synthesis and characterization of compounds ...... 52

Appendices ...... 58

A1. NMR spectra of reported compounds ...... 58

List of Tables

Table 01. Optimization of methyl 6-O-TBS-α-D-mannopyranoside sulfation with 2,2,2-TCE chlorosulfate ...... 22

Table 02. Optimization of methyl α-L-rhamnopyranoside sulfation with TCE-sulfuryl 1,2- dimethylimidazolium triflate ...... 24

Table 03. Solvent screen of methyl α-L-rhamnopyranoside sulfation with TCE-sulfuryl 1,2- dimethylimidazolium triflate ...... 25

Table 04. Boronic acid/Lewis base co-catalyst for sulfation of methyl α-L-rhamnopyranoside ....26

Table 05. Optimization of methyl α-L-rhamnopyranoside sulfation with arylsulfuryl 1,2- dimethylimidazolium triflate ...... 28

Table 06. Optimization of methyl α-D-mannopyranoside sulfation with sulfur trioxide amine complex ...... 30

Table 07. Optimization of n-octyl α-D-galactopyranoside sulfation with sulfur trioxide amine complex ...... 31

Table 08. Substrate scope of carbohydrate sulfation with SO3-Me3N ...... 32

Table 09. Substrate scope for fatty acid and sugar alcohol esterification ...... 50

List of Figures

Figure 01. Structure of select naturally occurring sulfated carbohydrates ...... 2

Figure 02. Structure of the major tri-sulfated disaccharide repeat unit in heparin ...... 3

Figure 03. Structure of heparin derivatives ...... 4

Figure 04. Conventional routes to access various patterns of carbohydrate sulfation ...... 5

Figure 05. Transient boronate ester protection in the regioselective sulfation of steroids ...... 6

Figure 06. Temperature-dependent regioselective sulfation of galactoside ...... 7

Figure 07. Effect of SO3-amine complexes in the sulfation of trimethylsilyl cellulose ...... 8

Figure 08. Regioselective sulfation with dibutyltin oxide of 1,2-cis-diol and 1,2-cis-dioxy ...... 9

Figure 09. Regioselective sulfation with dibutyltin oxide of 1,3-cis-diol ...... 10

Figure 10. Regioselective sulfation with dibutyltin oxide in the absence of 1,2-cis-diol ...... 10

Figure 11. Routes of nucleophilic attack on a carbohydrate sulfate diester ...... 11

Figure 12. Preparation and unmasking of phenyl sulfate diesters ...... 12

Figure 13. Preparation and unmasking of alkyl sulfate diesters, a) sulfation with iBu/nP chlorosulfate, b) unmasking of iBu sulfate diesters, c) unmasking of nP sulfate diesters ...... 13

Figure 14. Preparation and unmasking of trifluoroethyl sulfate diesters ...... 14

Figure 15. Stability of TFE-masked sulfate diester to further functionalizations ...... 15

Figure 16. Deprotection of TFE-masked sulfate diesters ...... 16

Figure 17. Preparation and unmasking of trichloroethyl sulfate diesters ...... 17

Figure 18. Regioselective TCE-masked sulfation of unprotected carbohydrates ...... 18

Figure 19. Tetracoordinate organoboron activation of diols for regioselective functionalization .19

vii

Figure 20. Synthesis of trichloroethyl chlorosulfate ...... 21

Figure 21. Synthesis of 2,2,2-trichloroethoxysulfuryl 1,2-dimethylimidazolium salts ...... 23

Figure 22. Sulfation with pre-formed cyclic arylboronate of methyl α-L-rhamnopyranoside ...... 26

Figure 23. Preparation of alkyl- and arylsulfuryl 1,2-dimethylimidazolium triflate ...... 27

Figure 24. Decomposition study of 2.08, 2.18, 2.19 in CD3CN; A: 2.08 in 1 equiv. 1,2- dimethylimidazole; B: 2.08 in 1 equiv. DIPEA; C: 2.18 in 1 equiv. DIPEA; D: 2.18 in 1 equiv. 3 MS-dried DIPEA; E: 2.19 in 1 equiv. DIPEA; F: 2.19 in 1 equiv. 3 MS-dried DIPEA ...... 29

Figure 25. Structure of select fatty acids and sugar alcohols ...... 47

Figure 26. Preparation of sugar fatty acid esters with lipases ...... 49

viii List of Abbreviations

Ac acetyl app apparent

Bn benzyl br broad Bu butyl Bz benzoyl calcd. calculated Cp cyclopentadienyl CSA (1S)-(+)-10-camphorsulphonic acid d doublet DAST (diethylamino)sulfur trifluoride DBU 1,8-diazabicyclo[5.4.0]undec-7-ene DCM dichloromethane dec. decomposed DIPEA N-N-diisopropylethylamine DMA N-N-dimethylacetamide DMAP 4-(dimethylamino)pyridine DMF N-N-dimethylformamide DMPU 1,3-dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone equiv equivalents ESI electrospray ionization Et ethyl

GML glycerol monolaurate h hour hept heptet HIV human immunodeficiency virus HMDS bis(trimethylsilyl)amide

HMPA hexamethylphosphoramide HRMS high-resolution mass spectra iBu isobutyl Im imidazole iPr isopropyl IR infrared m multiplet (NMR), medium (IR) m meta Me methyl min minute MP 4-methoxyphenyl Ms methanesulfonyl MS molecular sieve

NBS N-bromosuccinimide NMP N-methylpyrrolidine NMR nuclear magnetic resonance nP neopentyl o ortho p pentet p para Ph phenyl Piv pivaloyl PMB para-methoxybenzyl PMP 1,2,2,6,6-pentamethylpiperidine ppm parts per million q quartet rt room temperature s singlet (NMR), strong (IR) SFAE sugar fatty acid ester SIV simian immunodeficiency virus x

t triplet TBAF tetrabutylammonium fluoride TBS tert-butyldimethylsilyl tBu tert-butyl TCE 2,2,2-trichloroethyl TDS thexyl-dimethylsilyl Tf trifluoromethanesulfonyl TFA trifluoroacetic acid TFE 2,2,2-trifluoroethyl THF tetrahydrofuran TMS trimethylsilyl TREAT-HF triethylamine trihydrofluoride

UV ultraviolet w weak

Chapter 1 Introduction

Carbohydrates present a wealth of diverse structures derived from joining together monosaccharides, primarily in their pyranose or furanose forms. These structures can undergo further functionalization such as alkylation, macrocyclization, phosphorylation, and sulfation. Their structural diversity is mirrored by their broad range of biological functions and properties, from energy storage as starch and structural support as cellulose, to cellular recognition and other intercellular functions as glycolipids and glycoproteins.

The monosaccharides that make up complex carbohydrate structures contain several hydroxyl groups that differ in their stereochemical arrangements. Site selectivity among similar hydroxyl groups presents a difficult challenge for chemical modification. However, certain distinguishing features among monosaccharides have been exploited. Reactivity differences among amines, primary and secondary hydroxyl groups allow for site-selective modifications. cis-Diols present in mannose and galactose can be used to distinguish between two secondary hydroxyl groups. Differences between axial and equatorial hydroxyl groups can be used to influence stereochemical outcomes.

1.1 Sulfated carbohydrates Sulfated carbohydrates are common in nature and play key roles in biological functions (Figure 01). Dermatan sulfate 1.01, found in skin and blood vessels, plays a role in coagulation and fibrosis.1 Heparan sulfate 1.02, found on all cell surfaces, is a proteoglycan that functions in cell adhesion, blood coagulation, and growth factor regulation.1 Sulfated sialyl-Lewisx 1.03, found on cell surfaces, binds preferentially to lymphocyte cell-adhesion molecule L-selectin2 and has been

1 Capila, I.; Linhardt, R. J. Angew. Chem. Int. Ed. 2002, 41, 390-412. 2 Julien, S.; Ivetic, A.; Grigoriadis, A.; QiZe, D.; Burford, B.; Sproviero, D.; Picco, G.; Gillett, C.; Papp, S. L.; Schaffer, L. Cancer Res. 2011, 71, 7683-7693.

1 2 shown to play a role in bladder urothelial carcinoma metastasis.3 Sulfated galactosylceramide 1.04, found in the myelin sheath of nerve cells, play various functions in the nervous system. Abnormal expression of these sulfoglycolipids is associated with neurological disorders such as Alzheimer’s and Parkinson’s diseases.4 These sulfoglycolipids are also involved in the progression of other illnesses such as diabetes mellitus and the cellular entry of HIV-1.5

OH HOOC -O SO OH O O - 3 O O OSO3 O HO O O O OH AcHN O O O O O HO O HO -O SO OSO - NHAc OSO - 3 HOOC 3 HOOC 3 AcHN O Dermatan sulfate 1.01 Heparan sulfate 1.02

OH - HO COOH OSO3 OH OH O HO O OH O O OH AcHN O O HN fatty acid OH O OH OH O NHAc - O C13H27 O3SO O OH OH OH OH OH Sulfated galactosylceramide 1.04 6-Sulfo sialyl-Lewisx 1.03

Figure 01. Structure of select naturally occurring sulfated carbohydrates.

Perhaps the most well-known sulfated carbohydrates belong to the heparin polysaccharide family (Figure 02). Discovered in 1916, naturally occurring heparin contains on average 25 units of the disaccharide 1.05, giving a molecular weight of 5–40 kDa.1 It is the biological macromolecule with the highest density of negative charge due to its sulfate and carboxylate groups. Heparin

3 Taga, M.; Hoshino, H.; Low, S.; Imamura, Y.; Ito, H.; Yokoyama, O.; Kobayashi, M. Urol. Oncol.: Semin. Orig. Invest. 2015, 33, 496.e1-496.e9. 4 Eckhardt, M. Mol. Neurobiol. 2008, 37, 93-103. 5 Compostella, F.; Panza, L.; Ronchetti, F. C. R. Chim. 2012, 15, 37-45.

3 initiates the blood coagulation cascade by binding to enzyme antithrombin III, which then inactivates thrombin and other proteases.1

- OSO3

O O O O HO HO - OSO - - OOC 3 O3SHN O

Heparin 1.05

Figure 02. Structure of the major tri-sulfated disaccharide repeat unit in heparin.

First approved in 1939, heparin sodium salt is an anticoagulant drug administered through intravenous catheter or as an injection. It remains one of the oldest pharmaceutical drugs that is still in use. Pharmaceutically relevant effects aside from anticoagulation include anti-inflammatory properties for the treatment of ulcerative colitis and relief of obstructive pulmonary diseases like asthma.6 Most pharmaceutical-grade heparin on the market is isolated from animal tissues, particularly from porcine intestine or bovine lungs. However, the sulfation pattern of livestock- grown heparin is variable and difficult to control, and the extracted heparin is prone to viral and bacterial contamination. Recently, mammalian cell production of heparin using the Chinese hamster ovary cell lines have been shown to be a safer and more robust alternative.6,7

Due to the success of heparin as a drug, a number of derivatives and low-molecular weight analogues have been studied. Fondaparinux 1.06, a pentasaccharide marketed by GlaxoSmithKline, was approved in 2001 as an anticoagulant (Figure 03).8 Fondaparinux has advantages over heparin in that the former has a longer half-life, thus requiring a lower dosage.

6 Oduah, E. I.; Linhardt, R. J.; Sharfstein, S. T. Pharmaceuticals 2016, 9, 38. 7 Baik, J. Y.; Gasimli, L.; Yang, B.; Datta, P.; Zhang, F.; Glass, C. A.; Esko, J. D.; Linhardt, R. J.; Sharfstein, S. T. Metab. Eng. 2012, 14, 81-90. 8 Bauer, K. A.; Hawkins, D. W.; Peters, P. C.; Petitou, M.; Herbert, J. M.; Boeckel, C. A. A.; Meuleman, D. G. Cardiovasc. Ther. 2002, 20, 37-52.

Pentosan polysulfate 1.07, a plant-derived oligosaccharide, exhibits anti-HIV activity.9 Heparin tetrasaccharide 1.08, with increased oral bioavailability, has anti-allergic activity and is being studied for the treatment of asthma.10

- - OSO - OSO3 OSO3 3 -OOC O O O O O O HO O HO HO - HO O3SO HO - - - - O SHN - OH O SHN OOC OSO3 3 O3SHN 3 OMe O O Fondaparinux 1.06

- OSO3 OH O OH OSO - 3 OH O O OSO - 3 -OOC O OSO - O O 3 OH O - n HO O OSO3 - O - OOC O3SHN Pentosan polysulfate 1.07 OSO - OH 3 Heparin tetrasaccharide 1.08

Figure 03. Structure of heparin derivatives.

1.2 Direct, regioselective synthesis of sulfated carbohydrates

Synthesis of sulfated carbohydrates remains a challenge despite their long history of biologically relevant properties. Sulfation is commonly carried out with sulfur trioxide-amine complexes, together forming a Lewis acid-base adduct. These complexes are easier to handle than liquid sulfur trioxide, which requires distillation prior to use. The relative reactivities of the SO3 complexes

9 Baba, M.; Nakajima, M.; Schols, D.; Pauwels, R.; Balzarini, J.; De Clercq, E. Antiviral Res. 1988, 9, 335-343. 10 Ahmed, T.; Smith, G.; Abraham, W. M. Pulm. Pharmacol. Ther. 2013, 26, 180-188.

5 generally vary inversely with the strength of the Lewis base component. Common complexes used 11 are listed in order of decreasing basicity: Me3N ≈ Et3N > pyridine > DMF.

Conventional routes to sulfated carbohydrates consist of numerous protection and deprotection steps. As exemplified in Figure 04, the synthesis of galactopyranoside monosulfate requires multiple orthogonal protecting groups, functional group manipulation, and selective deprotection to reveal the free hydroxyl at the position of interest for sulfation.12

O OH OPiv OPiv O OH AcO HO O O O O O O O OR OR OR HO OR O OR HO BzO O OPiv OBz OH OH OH

OPiv OH SO3-amine SO3-amine SO3-amine O BzO OR OBz

SO3-amine

OSO - OPiv OPiv OPiv 3 - O O AcO O3SO O O O O OR OR O -O SO OR BzO OR O - 3 OH OSO3 OPiv OBz

6-O-monosulfate 2-O-monosulfate 3-O-monosulfate 4-O-monosulfate

Figure 04. Conventional routes to access various patterns of carbohydrate sulfation.12

To overcome lengthy protection/deprotection steps, methods for direct and regioselective sulfation have been studied.13 Boronate ester protection was utilized by McLeod to transiently mask the cis- diols of steroid 1.09 for selective functionalization at the remaining hydroxyl group.14 The

11 Gilbert, E. E. Chem. Rev. 1962, 62, 549-589. 12 Marinier, A.; Martel, A.; Banville, J.; Bachand, C.; Remillard, R.; Lapointe, P.; Turmel, B.; Menard, M.; Harte, W. E.; Wright, J. J. K. J. Med. Chem. 1997, 40, 3234-3247. 13 Al-Horani, R. A.; Desai, U. R. Tetrahedron 2010, 66, 2907-2918. 14 Hungerford, N. L.; McKinney, A. R.; Stenhouse, A. M.; McLeod, M. D. Org. Biomol. Chem. 2006, 4, 3951-3959.

6 boronate ester of 1.10 can then be cleaved to reveal the cis-diol in 1.11 for selective sulfation to give 1.12 (Figure 05). Although this route still requires protecting groups, it reduces the number of separate purification steps.

OH Ph OH O B 1) PhB(OH)2, DMF/CH2Cl2 H O , aq. NaOH OH 2) TBS-Cl, imidazole O 2 2 OH THF 73% over 3 steps HO TBSO TBSO H H H 1.11

1.09 1.10

SO3-pyridine DMF, pyridine 61%

OH OH

- + 80% AcOH/H O - + OSO3 Na 2 OSO3 Na

69% HO TBSO H H 1.13 1.12

Figure 05. Transient boronate ester protection in the regioselective sulfation of steroids.14

A temperature-dependent regioselective sulfation of galactoside 1.14 was described by Kondo 15 (Figure 06). Sulfation performed at room temperature with SO3-pyridine yielded the 3,4-O-bis- sulfate 1.15 while at 0 oC, the reaction afforded only the 4-O-sulfate 1.16 without observable 3-O- or bis-sulfate. The 3-O-sulfate 1.18, however, could only be prepared after first protecting the C4- hydroxyl group. This method was not shown to be generalizable to other sugar moieties or to be applicable in the presence of exposed primary or C2-hydroxyl groups.

15 Tsukida, T.; Yoshida, M.; Kurokawa, K.; Nakai, Y.; Achiha, T.; Kiyoi, T.; Kondo, H. J. Org. Chem. 1997, 62, 6876-6881.

OBz OH O HO OR OBz 1.14 OBz AcO O HO OR OBz 1.17

SO3-pyridine SO3-pyridine SO3-pyridine DMF, rt DMF, 0 oC DMF, 0 oC 73% 73% 90%

OBz OBz OBz - - O3SO O3SO AcO O O O - OR OR - OR O3SO HO O3SO OBz OBz OBz 1.15 1.16 1.18

Figure 06. Temperature-dependent regioselective sulfation of galactoside.15

Following up on previous reports of sulfate insertion into O-Si bonds,16,17 Richter showed that the Lewis base used in the sulfate complex can be tuned to influence the regiochemical outcome 18 (Figure 07). Trimethylsilyl cellulose 1.19 with SO3-DMF preferentially yields the 6-O-sulfate

1.20 while SO3-Et3N preferentially yields the 2-O-sulfate 1.21. The authors rationalize this effect by stating that the electron-donation of Et3N polarizes the O-S bond of its SO3 complex, promoting insertion at the more polarized O-Si bond of the O-2 position. This electron-donating effect is absent in the DMF complex, which favors the more sterically accessible position at O-6.

16 Stein, A.; Wagenknecht, W.; Philipp, B.; Klemm, D.; Schnabelrauch, M., German Patent DD 299313, 1989. 17 Wagenknecht, W.; Nehls, I.; Stein, A.; Klemm, D.; Philipp, B. Acta Polym. 1992, 43, 266-269. 18 Richter, A.; Klemm, D. Cellulose 2003, 10, 133-138.

OTMS

O O TMSO OTMS 1.19

SO3-DMF SO3-Et3N THF THF

OTMS O Si O S O O O O TMSO O O S O O O O TMSO Si OTMS

NaOH NaOH MeOH MeOH

- OSO3 OH

O O O O HO HO - OH OSO3 major product, 1.20 major product, 1.21

18 Figure 07. Effect of SO3-amine complexes in the sulfation of trimethylsilyl cellulose.

Dibutyltin oxide was employed by Flitsch in 1994 to facilitate the regioselective sulfation of 1.22 (Figure 08).19,20 The dibutylstannylene acetal 1.23 was first formed at the cis-diol group with super-stoichiometric amount of dibutyltin oxide, followed by a solvent-switch and addition of SO3-

Me3N complex to afford the product of sulfation at the more sterically accessible position of the

19 Guilbert, B.; Davis, N. J.; Flitsch, S. L. Tetrahedron Lett. 1994, 35, 6563-6566. 20 Guilbert, B.; Davis, N. J.; Pearce, M.; Aplin, R. T.; Flitsch, S. L. Tetrahedron: Asymmetry 1994, 5, 2163-2178.

1,2-cis-diol. The galactosylceramide glycolipid 1.24, found in mammalian nervous systems, was synthesized in excellent yield. This strategy was also shown for disaccharides, including a lactoside 1.25 to give the 3′-O-sulfate 1.26 as the major product with 10% of the 3′,6′-O-bis-sulfate byproduct 1.27. In the absence of Bu2SnO, this reaction gave no observable 1.26. Maltosides 1.28 protected at the primary hydroxyl groups that do not possess a cis-diol were selectively 2′-O- sulfated to give 1.29 in decent yield. The authors rationalize that the regioselectivity is due to either the higher reactivity of the 2′-hydroxyl group or the C1′-C2′ cis-dioxy configuration. However, they did not perform the control reaction to show that the maltoside sulfation is only regioselective in the presence of Bu2SnO.

OH Bu OH OH O O OH Bu2SnO (1.5 equiv) SO3-Me3N (2 equiv) OH Sn O HN C8H17 O Bu O R = OR OR - OR C13H27 HO MeOH O THF O3SO OH reflux, 2 h OH OH rt, 4 h OH 1.22 1.23 1.24 97%

OH OH OR′ OH 1) Bu2SnO (1 equiv) HO MeOH, reflux, 2 h HO O O O O O SPh O SPh HO OH 2) SO3-Me3N (2 equiv) RO OH OH OH dioxane, rt, 30 h OH OH

- 1.25 1.26 R = SO3 , R′ = H 76% - - 1.27 R = SO3 , R′ = SO3 10% Ph Ph O O O 1) Bu2SnO (1 equiv) O O OTBS MeOH, reflux, 2 h O OTBS OH OH - OH O3SO O O 2) SO3-Me3N (2 equiv) O O OH O-allyl dioxane, rt, 93 h OH O-allyl OH OH 1.28 1.29 56%

Figure 08. Regioselective sulfation with dibutyltin oxide of 1,2-cis-diol and 1,2-cis-dioxy.20

In 2004, Gelb adapted the method for the synthesis of 1.32, a fluorescent probe used in the assay for screening newborns against Hunter syndrome, a disease caused by the deficiency of iduronate- 2-sulfatase (Figure 09).21 The dibutyltin oxide, in this case, binds to the 1,3-cis-diol of iduronic ester 1.30, delivering the sulfate to the more nucleophilic O-2 affording 1.31.13

21 Blanchard, S.; Turecek, F.; Gelb, M. H. Carbohydr. Res. 2009, 344, 1032-1033.

O O O H H H t-BuO N t-BuO N t-BuO N N N N H H H O O O 1) Bu2SnO (1.5 equiv) MeOH, reflux, 40 min NaOH

O O O O O O H O O O O OH 2) SO3-Me3N (1.5 equiv) OH 2 OH o MeOOC O DMF, 55 C, 24 h MeOOC O -OOC O

- - OH OH OH OSO3 OH OSO3 1.30 1.31 1.32 61% over 2 steps

Figure 09. Regioselective sulfation with dibutyltin oxide of 1,3-cis-diol.21

In 2009, Kosma demonstrated the regioselective sulfation of xylopyranosides and xylotriosides.22 Low-molecular weight xylans and their sulfates have been shown to possess antithrombin23 and antiasthmatic24 activities. With the dibutyltin oxide strategy, 1.33 and 1.35 gave the terminal 4-O- sulfated products 1.34 and 1.36, respectively (Figure 10).22 However, this selectivity for substrates without the presence of a cis-diol was not explained, and the remaining mass balance of the sulfation of 1.35 was unaccounted for.

1) Bu2SnO (1.08 equiv) O toluene, reflux, 15 h - O HO O3SO HO OMe HO OMe OH 2) SO3-Me3N (1.1 equiv) OH THF, rt, 48 h 1.34 1.33 74%

1) Bu2SnO (4 equiv) O O O toluene, reflux, 15 h - O O O HO O O O3SO O O HO HO HO OMe HO HO HO OMe OH OH OH 2) SO3-Me3N (3.2 equiv) OH OH OH THF, rt, 72 h 1.35 1.36 27%

Figure 10. Regioselective sulfation with dibutyltin oxide in the absence of 1,2-cis-diol.22

Although activation with dibutyltin oxide has shown to be highly selective for sulfating the equatorial hydroxyl of a cis-diol, it requires—at minimum—a stoichiometric amount of tin, long reaction times (93 h to afford 1.29), and a two-step activation/sulfation process. That being said,

22 Abad-Romero, B.; Mereiter, K.; Sixta, H.; Hofinger, A.; Kosma, P. Carbohydr. Res. 2009, 344, 21-28. 23 Yamagaki, T.; Tsuji, Y.; Maeda, M.; Nakanishi, H. Biosci. Biotechnol. Biochem. 1997, 61, 1281-1285. 24 Kuszmann, J.; Medgyes, G.; Boros, S. Carbohydr. Res. 2005, 340, 1739-1749.

11 direct sulfation of carbohydrates presents a concise and efficient alternative to traditional protecting group-based methods. However, this strategy is often only applicable as the final step in a complex synthesis. The highly polar sulfated products are insoluble in organic solvents, making them difficult to purify and manipulate for subsequent elaborations. To this end, masked sulfates have been devised.

1.3 Synthesis of sulfated carbohydrates via masked sulfates

Masked sulfates, or protected sulfate esters, provide a way to address the issues of sulfated products, allowing for chemical transformations of the molecule elsewhere. Following appropriate transformations, the sulfate can be unmasked to reveal the desired product. These masked sulfates must be stable to conventional purification techniques and survive a wide range of reaction conditions including, in particular, the deprotection conditions of other functional groups on the molecule.

Sulfate diesters have generally been used, providing the added benefit of a spectroscopic handle that is compatible with common characterization techniques. Sulfate diesters are prone to nucleophilic attack and substitution at one of three possible sites (Figure 11).25 Substitution via Route A leading to desulfation is generally slow, particularly on a carbohydrate’s secondary hydroxyl group. Design of sulfate diesters would ideally disfavor attack on the sulfur center (Route B), or premature deprotection of the ester (Route C).

O O R S carbohydrate O O B C A Nu

Figure 11. Routes of nucleophilic attack on a carbohydrate sulfate diester.25

25 Proud, A. D.; Prodger, J. C.; Flitsch, S. L. Tetrahedron Lett. 1997, 38, 7243-7246.

Perlin first introduced phenyl chlorosulfate as a masked sulfating group for carbohydrates.26 The stable phenyl chlorosulfate is synthesized from phenol, sodium hydroxide, and sulfuryl chloride in high yields. After reaction with 1.37 (Figure 12), unmasking of the sulfate diester 1.38 was achieved by catalytic hydrogenolysis of the phenyl group to a cyclohexyl group, followed by alkyl fission to give 1.39. Although yields for unmasking the phenyl group were not reported, the authors stated that about 10% desulfation had occurred. The phenyl-masked sulfate is stable to the

TFA/CHCl3 conditions used to deprotect the 5,6-O-isopropylidene group, and also stable to 1:1

Ac2O:H2SO4, which was used to deprotect both isopropylidene groups. However, this method is not compatible with functional groups sensitive to base and hydrogenolysis.

O O OSO - O S O 3 O O O OPh O O O O OH 1) NaH, THF, 30 min O K2CO3, PtO2, H2 O O 2) O EtOH, H2O, 20 h O O O PhO S Cl , 20 h O O 1.37 1.38 1.39 75%

Figure 12. Preparation and unmasking of phenyl sulfate diesters.26

Alkyl-masked sulfating groups have been described by Widlanski with particular emphasis on isobutyl (iBu) and neopentyl (nP) chlorosulfates.27 The carbohydrate 1.37 was first stirred in NaHMDS, followed by the addition of the masked chlorosulfate to give sulfated products 1.40 and 1.41 (Figure 13a). Sulfation with iBu chlorosulfate was slower and required large excess of the chlorosulfate, while sulfation with nP chlorosulfate required DMPU as a co-solvent. The stabilities of the sulfates were studied using phenyl iBu or nP sulfate diesters as model substrates. The iBu sulfate diester degraded completely in 6% piperidine after 24 h, but was relatively stable in 50% TFA (7.1% degradation after 24 h). No degradation of the nP sulfate diester was observed in 20% piperidine, versus 6.9% degradation in 50% TFA after 24 h. Both sulfate diesters were stable under

26 Penney, C. L.; Perlin, A. S. Carbohydr. Res. 1981, 93, 241-246. 27 Simpson, L. S.; Widlanski, T. S. J. Am. Chem. Soc. 2006, 128, 1605-1610.

conditions used to deprotect benzyl and isopropylidene groups (hydrogenolysis with Pd/C and H2, and acidification with aq. H2SO4/THF)

a) O O S O O O O OR O 1) NaHMDS, THF O OH O

O o 2) ClSO2OiBu (5–10 equiv), −15 C O O or O o 1.37 ClSO2OnP (1 equiv), DMPU, −75 C 1.40 R = iBu 95% 1.41 R = nP 95% b) OH OH NaI O O HO HO o - iBu-O3SO acetone, 55 C O3SO OH OH OH OH 1.42 1.43 97% c) OSO -nP O 3 O O NaI no reaction O acetone, 55 oC 1.44 O

- OSO3-nP OSO O O 3 O O O O NaN3 O DMF, 70 oC O O 1.44 1.39 O 98%

OH OH NaN O 3 O HO HO o nP-O3SO DMF, 70 C OH OH OH OH N3 1.45 1.46

Figure 13. Preparation and unmasking of alkyl sulfate diesters, a) sulfation with iBu/nP chlorosulfate, b) unmasking of iBu sulfate diesters, c) unmasking of nP sulfate diesters.27

Unmasking of the diester was examined through Route C (Figure 11) via the attack of a small nucleophile on the alkyl group to reveal the sulfated carbohydrate. iBu sulfate diester 1.42 was unmasked cleanly in the presence of NaI at elevated temperatures to give 1.43 (Figure 13b). nP sulfate diester 1.44, however, was more difficult to unmask due to its increased bulk. Under the same deprotection conditions for the iBu sulfate diester, the nP sulfate diester 1.44 yielded no

reaction (Figure 13c). NaN3 was found to be able to unmask the nP sulfate diester of the glucofuranose sulfate 1.44 to give 1.39, but led to the azide substitution product of the glucopyranose sulfate 1.45 to give 1.46. Although the method shown by Widlanski were not generally applicable and showed limited scope, it does offer a masked-sulfate approach with the potential for alkyl group manipulation to tune the reactivity of the masked sulfate.

Trihaloethyl-masked sulfates have also long been considered as an alternative to aryl-masked sulfates. Trifluoroethyl (TFE) and trichloroethyl (TCE) are thought to hinder Routes B and C (Figure 11) compared to alkyl and aryl masking groups due to both steric and electronic reasons. TCE esters have been used in the protection of phosphate and carboxyl groups, and removed selectively by Zn/AcOH. Flitsch sought to extend trihaloethyl masking to the sulfation of carbohydrates.25 Focusing initially on TCE-masked sulfates, the authors were unable to sulfate carbohydrates in sufficiently high yields for further studies, citing steric hindrance of the trichloro group. Switching direction to TFE-masked sulfates, the authors were unable to introduce the TFE- sulfate in one step from the TFE-chlorosulfate. Instead, the carbohydrate 1.47 had to be first sulfated with an SO3-amine complex, followed by TFE protection with 2,2,2-trifluorodiazoethane in moderate yields (Figure 14). The TFE-sulfate diester 1.48 was stable to conditions including hydrogenation, 20% TFA/EtOH, and NaOMe/MeOH used for isopropylidene deprotection. Harsh conditions (reflux in t-BuOK/t-BuOH) were required for attack on the sulfur center and elimination of TFE to give the unmasked sulfate 1.49. However, deprotection of TFE-sulfate diester on O-2 or O-3 positions resulted in sulfate migration to free, adjacent hydroxyl groups. Apart from difficult deprotection, this masked sulfate strategy was not widely adopted in carbohydrate synthesis due to the need for a two-step sulfation process and the potentially explosive nature of trifluorodiazoethane.

O OH O OSO3TFE - 1) 4:1 TFA:EtOH OSO3 O o O HO 1) SO3-pyridine, MeCN, 80 C rt, 2 h, 97% O O O O O 2) HO O F3C N2 O 2) t-BuOK, t-BuOH, reflux, 96% OH OH citric acid, MeCN, rt 1.47 1.48 51% 1.49

Figure 14. Preparation and unmasking of trifluoroethyl sulfate diesters.25

Linhardt then showed the versatility and stability of TFE-masked sulfate diesters in subsequent functionalizations. Sulfate diesters 1.50 and 1.52 were stable to either TBAF in AcOH or TREAT- HF (Figure 15), thus demonstrating orthogonality to silyl protective groups.28 The masked sulfate diesters were also stable to fluorination and glycosylation conditions, affording TFE-masked sulfate disaccharides 1.54 and 1.56 in decent yields.

OSO TFE O OSO3TFE O 3 1) TBAF, AcOH, THF, −25 oC Cl Cl O O O O OTDS BzO 2) DAST, DCM, , −30 oC BzO N3 N3 F 1.50 1.51 66%

OSO TFE BnO OSO3TFE BnO 3 1) TREAT-HF O O OTDS BzO 2) DAST, DCM, −30 oC BzO N3 N3 F 1.52 1.53 63%

BnO OSO3TFE O O OH BzO OSO TFE BnO 3 N3 O O O O AgClO4, Cp2ZrCl2 + O BzO O O N3 O 4 Å MS, DCM O F O O 1.53 1.47 1.54 64%

OBn TFEO3SO O TFEO SO OBn OH 3 O BzO O O N3 BF3ꙓEt2O O BzO O O N + 3 O O Cl C O O 4 Å MS, DCM 3 O O NH O 1.56 1.55 1.47 49%

Figure 15. Stability of TFE-masked sulfate diester to further functionalizations.28

28 Karst, N. A.; Islam, T. F.; Linhardt, R. J. Org. Lett. 2003, 5, 4839-4842.

Deprotection of TFE-masked sulfate diesters proved to be quite challenging.29 Previously reported t-BuOK deprotection conditions were effective on monosaccharide 1.57 to give 1.58, but led to the decomposition of disaccharide 1.59 (Figure 16). However, NaOMe/MeOH was found to be able to unmask the diester of disaccharide 1.59 in good yields to give 1.60 with minimal decomposition. Deprotection of TFE-masked bis-sulfate disaccharide 1.61 required a two step process. Unmasking with NaOMe/MeOH gave 1.62 in excellent yield, but the harsher t-BuOK reagent used in the second step gave 1.63 in only moderate yield, along with disaccharide decomposition, desilylation, and desulfonation.

Ph O t-BuOK, t-BuOH Ph O O O O O OMP OMP BnO BnO - OSO3TFE OSO3 1.58 1.57 82% TFEO SO OBn 3 - OBn O3SO O O O O BzO O N3 NaOMe, MeOH HO O O N3 O O O O O O O 1.59 1.60 70%

OBn OTBS OBn - OBn OTBS -O SO TFEO SO OTBS O3SO 3 3 NaOMe, MeOH t-BuOK, t-BuOH O O O O O O O O O OMP OH OMP HO BnO HO BnO BzO BnO N - N3 OSO TFE 3 OSO3 N3 OSO3TFE 3 1.62 1.63 1.61 90% 50%

Figure 16. Deprotection of TFE-masked sulfate diesters.29

In 2004, Taylor re-visited Flitsch’s attempts to develop a method for TCE-masked sulfate diesters and succeeded in sulfating aryl alcohols with TCE chlorosulfate.30 In follow-up reports of extending this methodology to the sulfation of carbohydrate 1.47, the authors managed to synthesize the sulfate diester 1.64 in approximately 50% yield, with the majority of the remaining mass balance being the chlorosugar byproduct 1.65 (Figure 17). This result, in contrary to Flitsch’s

29 Karst, N. A.; Islam, T. F.; Avci, F. Y.; Linhardt, R. J. Tetrahedron Lett. 2004, 45, 6433-6437. 30 Liu, Y.; Lien, I. F. F.; Ruttgaizer, S.; Dove, P.; Taylor, S. D. Org. Lett. 2004, 6, 209-212.

17 report in 1997,25 showed that TCE-sulfated carbohydrates could indeed be prepared in good yields. Encouraged by this finding, Taylor designed a sulfating reagent that did not release an effective nucleophilic species. With the optimized 1,2-dimethylimidazolium triflate salt 1.67 in hand, the authors achieved TFE-masked sulfation of carbohydrate 1.66 to give 1.68 in high yields, as well as deprotection of 1.68 under mild conditions with Zn or Pd/C and ammonium formate to give 1.58 (Figure 17).31 The sulfating reagent 1.67 was stable to prolonged storage at room temperature and the TCE-masked sulfate diester 1.68 was stable to a range of reaction conditions:

NaOMe/MeOH and ZnCl2/AcOH/Ac2O for debenzylation and deacetylation, NBS in acetone/water, DBU, and acidic conditions for benzylidene opening.

O OH O O OSO3TCE O Cl TCEO S Cl O O O O O O + O O O O O AgCN, Et3N, DMAP, THF O O

1.64 1.47 ~ 50% 1.65

O OTf TCEO S N N Zn or Pd/C O O Ph O 1.67 Ph HCO NH Ph O O O O O 2 4 O O OMP OMP OMP BnO BnO BnO 1,2-dimethylimidazole MeOH - OH OSO3TCE OSO3 DCM, 24 h 1.68 1.66 1.58 96% with Zn: 94%

Figure 17. Preparation and unmasking of trichloroethyl sulfate diesters.31,32

Masked sulfation provides products that can be manipulated for further synthetic transformations. However, despite these recent advances in the field of masked sulfation of carbohydrates, there remains a need for a reliable method toward the regioselective installation of a masked sulfate group, particularly on fully unprotected carbohydrates. In 2016, Kaji and Makino33 bridged that gap by using the transient boronate ester protection for the regioselective sulfation of steroids

31 Ingram, L. J.; Desoky, A.; Ali, A. M.; Taylor, S. D. J. Org. Chem. 2009, 74, 6479-6485. 32 Ingram, L. J.; Taylor, S. D. Angew. Chem. Int. Ed. 2006, 45, 3503-3506. 33 Fukuhara, K.; Shimada, N.; Nishino, T.; Kaji, E.; Makino, K. Eur. J. Org. Chem. 2016, 2016, 902-905.

18 described previously14 and the TCE-masked sulfate developed by Taylor.31 In one pot, unprotected carbohydrates 1.69 with 1,2-cis- or 4,6-diols are protected by phenylboronic acid, followed by TCE-masked sulfation at the remaining hydroxyl group of 1.70, and transesterification of the boronate ester with pinacol to reveal the TCE-masked sulfate carbohydrate 1.71 in good yields (Figure 18).

1) O OTf Ph TCEO S N N O PhB(OH)2 O O B O TCEO SO O 3 (HO)n O OR DCM 1,2-dimethylimidazole (HO)n OR rt, 24–48 h 1.69 (HO)n OR 4 Å MS, THF or DCM 1.70 20–24 h, 0 oC to rt 1.71 2) pinacol, DCM

Product: OMe OH OMe OH HO O O O O HO TCEO3SO OSO3TCE OMe O HO OH TCEO3SO OH OH OH OH TCEO3SO

76% 74% 66% 72%

Figure 18. Regioselective TCE-masked sulfation of unprotected carbohydrates.33

1.4 Organoboron compounds in carbohydrate chemistry

As discussed previously, organoboron compounds have been used in carbohydrate chemistry for functional group protection14,33 among other purposes.34 In addition, organoboron compounds in their tetracoordinate state have been used for the activation of diols. As first described by Aoyama, the phenylboronate ester of the 1,2-cis-diol of methyl α-L-fucopyranoside 1.72 can be activated by an amine base to give the tetracoordinate complex 1.73 (Figure 19).35 It can then undergo regioselective alkylation at the more accessible position to give 1.74. Other regioselective transformations following this strategy have also been explored, including the glycosidation of

34 McClary, C. A.; Taylor, M. S. Carbohydr. Res. 2013, 381, 112-122. 35 Oshima, K.; Kitazono, E.-I.; Aoyama, Y. Tetrahedron Lett. 1997, 38, 5001-5004.

19 unprotected carbohydrate 1.72, via the tetracoordinate complex 1.75, to give the disaccharide 1.76 (Figure 19).36

OMe OMe OMe O 1) PhB(OH) O 2 OH n-BuI O OH O OH OH 2) Ag2O, Et3N O On-Bu B OH benzene, reflux Ph OH 1.72 Et3N 1.74 1.73 50%

OAc O OMe B O OH AcO OMe OMe O AcO OH AcO O Br OH O O OAc OH O OH B O OH Ag2CO3 O O OH Et N+ I-, 4 Å MS AcO 4 AcO 1.72 AcO 1.76 1.75 93% H2 Ph N B (10 mol%) Ph O HO OTBS O OTBS HO OTBS Ph BzCl, iPr NEt O 2 B O O Ph HO MeCN O BzO HO HO HO OMe OMe OMe 1.77 1.79 1.78 95% NHBz BzO

Figure 19. Tetracoordinate organoboron activation of diols for regioselective functionalization.34

Expanding on the reactivity of tetracoordinate boronates, our group has developed catalytic versions employing borinic acid derivatives, avoiding the need for complexation with a Lewis

36 Oshima, K.; Aoyama, Y. J. Am. Chem. Soc. 1999, 121, 2315-2316.

20 base. Catalytic, regioselective acylation,37 sulfonylation,38 alkylation,39 and glycosylation40 of carbohydrates have since been accomplished. An example with 1.77 is shown in Figure 19, forming the tetracoordinate complex 1.78, to give the regioselectively benzoylated product 1.79.

1.5 Scope of Thesis

Carbohydrates and their O-sulfated derivatives have long been known to play important roles in biology. With a growing number of carbohydrate drugs being approved in recent years, it follows that concise and elegant approaches to the synthesis of carbohydrate derivatives, including O- sulfates, are also increasingly needed. Furthermore, organoboron compounds have shown to be powerful in imparting regiocontrol in carbohydrate functionalization, both as protecting and activating groups.

The remainder of this thesis discusses my research on expanding the scope of borinic acid catalysis and boronic ester protection in carbohydrate synthesis. Chapter 2 outlines the development of a method for the regioselective sulfation of unprotected sugars. In contrast to previously described sulfation methods, this synthesis employs catalytic borinic acid to effect direct sulfation with high selectivity. Chapter 3 describes the use of boronic acids in Fischer esterification between sugar alcohols and fatty acids for the preparation of surfactants. The esterification project outlined in this last chapter was conducted in conjunction with a fellow laboratory group member, Sanjay Manhas, and individual contributions to the joint work are delineated therein.

37 Lee, D.; Taylor, M. S. J. Am. Chem. Soc. 2011, 133, 3724-3727. 38 Lee, D.; Williamson, C. L.; Chan, L.; Taylor, M. S. J. Am. Chem. Soc. 2012, 134, 8260-8267. 39 Chan, L.; Taylor, M. S. Org. Lett. 2011, 13, 3090-3093. 40 Gouliaras, C.; Lee, D.; Chan, L.; Taylor, M. S. J. Am. Chem. Soc. 2011, 133, 13926-13929.

Chapter 2 Regioselective, catalytic sulfation of unprotected carbohydrates

As discussed in Chapter 1, although regioselective methods using stoichiometric amounts of additives have been developed for direct sulfation, they are only suitable as the final step in a synthesis due to the highly polar nature of the products. Masked sulfates, with their various protective ester groups, address some of the issues of sulfated products but no regioselective strategies have been developed for sulfation. My research began by employing the catalytic method developed in our lab using borinic acid-activation of cis-diols to prepare TCE-masked sulfates of carbohydrates.

2.1 Sulfation of carbohydrates with 2,2,2-trichloroethyl chlorosulfate

TCE chlorosulfate 2.02 was prepared according to literature41 in 84% yield from sulfuryl chloride 2.01 and 2,2,2-trichloroethanol (Figure 20).

O 2,2,2-trichloroethanol (1 equiv) Cl3C O Cl S Cl O S Cl pyridine (1 equiv) O O Et2O (0.5 M) (1 equiv) 1.5 h at −20 oC, 2.02 2.01 then 0.5 h at rt 84%

Figure 20. Synthesis of trichloroethyl chlorosulfate.

Initial attempts for TCE-masked sulfation of 6-O-TBS-α-D-mannopyranoside 2.03 with 2.02, catalyzed by the borinic ester pre-catalyst 2.04 yielded the cyclic sulfate 2.05 in low yields (Table 01). After the initial sulfation reaction, the sulfate diester presumably undergoes a rapid cyclization with the adjacent free hydroxyl group on the carbohydrate. The 2,2,2-trichloroethoxide generated in situ as a result of the cyclization may explain the intermolecular TBS migration product 2.06 observed under 1.5 equivalents of DIPEA or Et3N (Entries 1 and 5). However, the sugar without a TBS-group was not able to be isolated cleanly for characterization. Furthermore, the strongly basic

41 Pitts, A. K.; O'Hara, F.; Snell, R. H.; Gaunt, M. J. Angew. Chem. Int. Ed. 2015, 54, 5451-5455.

21 22 ethoxide could cause decomposition of the sulfating reagent 2.02, leading to low yields of sulfated products. Decreasing the reaction time (Entry 2), using DCM as solvent (Entry 3), or using 1.1 equivalents of DIPEA (Entry 4) also gave cyclic sulfates with none of the desired, uncyclized product observed. A preliminary base screen with Et3N and pyridine (Entries 5, 6) did not yield any productive results. 1,2,2,6,6-Pentamethylpiperidine (PMP) was used as a bulky base (Entry 7) in hopes of minimizing cyclic sulfate formation, but the TCE-masked sulfate diester was not observed.

Table 01. Optimization of methyl 6-O-TBS-α-D-mannopyranoside sulfation with 2,2,2-TCE chlorosulfate.

Cl3C O O S Cl O O TBSO OTBS OTBS O OH O O S O S Ph O B O (1.5 equiv) O O + O Ph N HO HO HO Base (1.5 equiv) TBSO H2 O O OMe 2.04 (10 mol %) 2.04 MeCN (0.2 M) OMe OMe 2.03 2-aminoethyl phenylborinate 24 h, rt 2.05 2.06

Entry Base 2.03 Yield (%)a 2.04 Yield (%)a

1 DIPEA 40 5 2b DIPEA 10 - 3c DIPEA 6 - 4d DIPEA 3 -

5 Et3N 6 1 6 pyridine 29 - 7 1,2,2,6,6-pentamethylpiperidine 19 -

aCrude 1H NMR yields were determined based on tetramethylsilane as an internal standard. b6 hour reaction time. cDCM used as solvent. d1.1 equivalents of 1.58 and DIPEA used.

2.2 Sulfation of carbohydrates with alkyl and aryl 1,2- dimethylimidazolium salt

Following Taylor’s success using the sulfating reagent in the form of an imidazolium salt,31 we synthesized the 2,2,2-trichloroethoxysulfuryl 1,2-dimethylimidazolium salts from the reaction of 2.02 with 2-methylimidazole to give 2.07, followed by methylation to give the dimethylimidazolium salts (Figure 21). MeOTf gave high yields of 2.08, but methylation with MeI or Me3OBF4 gave low conversion to 2.09 and 2.10, respectively, and were not pursued further.

MeOTf (1 equiv) O OTf TCEO S N N Et2O (0.2 M) 0 oC, 3 h O 2.08 82% HN N

O (3.6 equiv) O MeI (1 equiv) O I TCEO S Cl TCEO S N N TCEO S N N O THF (0.025 M) THF (0.2 M) 1 h at 0 oC, O 0 oC to rt, 4 d O 2.02 then 1 h at rt 2.07 2.09 76% low conversion

BF4 Me3OBF4 (1 equiv) O TCEO S N N THF (0.2 M) 0 oC, 24 h O 2.10 low conversion

Figure 21. Synthesis of 2,2,2-trichloroethoxysulfuryl 1,2-dimethylimidazolium salts.

We then investigated TCE-masked sulfation with the imidazolium triflate salt using methyl α-L- rhamnopyranoside as a model substrate for easier characterization compared to the TBS-protected sugars and to avoid the possibility of silyl migration (Table 02). Conditions A with a tertiary amine base for substrate/catalyst binding in MeCN were based on previously optimized conditions for catalyst activity in our group.37 Conditions B with 1,2-dimethylimidazole as base in DCM at 0 oC to room temperature were based on Taylor’s optimized conditions for TCE-sulfation.31 TCE- masked sulfated rhamnopyranosides were observed, and a clear pattern of regioselectivity became evident. With Conditions A, there is a slight preference for the 3-O-sulfated product 2.12 which presumably goes through a tetracoordinate borinate complex, delivering the TCE-sulfate to the more accessible, equatorial position at O-3. However, with Conditions B, background reaction in the absence of catalyst was very prominent, with a slight preference for the 2-O-sulfated product 2.13. The catalyst did not appear to have a significant effect for Conditions B, affording neither an increase in yield nor influence on regioselectivity. This may be due to the inability of 1,2- dimethylimidazole to promote effective substrate binding to the borinic acid catalyst.

Table 02. Optimization of methyl α-L-rhamnopyranoside sulfation with TCE-sulfuryl 1,2- dimethylimidazolium triflate.

OTf Cl C O 3 N (1.5 equiv) OMe O S N OMe OMe O O O + O HO HO HO Conditions A or Conditions B HO TCEO3SO HO OH OH OSO3TCE 2.11 2.12 2.13

a a Entry Catalyst (XX mol%) Equiv. of Base Yield (%) with 2.12:2.13 Ratio Yield (%) with 2.12:2.13 Ratio Conditions A with Conditions A Conditions B with Conditions B

1 no catalyst 1.5 15 1:1 60 1:2 2 2.04 (10 mol%) 1.5 24 1:1 54 1:2 3 2.14 (5 mol%) 1.5 39 1.5:1 50 1:2.3 4 2.15 (10 mol%) 1.5 37 1.8:1 - - 5 no catalyst 0.2 35 1:1 - - 6 2.04 (10 mol%) 0.2 4 3:1 41 1:1.5 aCrude 1H NMR yields were determined based on tetramethylsilane as an internal standard. Product ratio determined by 1H NMR. Conditions A: catalyst (XX mol%), DIPEA (YY equiv), MeCN (0.2 M), 24 h, rt. Conditions B: catalyst (XX mol%), 1,2- dimethylimidazole (YY equiv), DCM (0.2 M), 24 h, 0 oC to rt.

O O Ph Ph O Ph B B B Ph N B H Ph Ph 2 OH 2.04 2.14 2.15

Proceeding with the result that gave the most promising regioselectivity (Table 02, Entry 4, Condition A), a solvent screen quickly revealed that amide solvents were essential (Table 03). DMF and DMA gave the 3-O-sulfate 2.12 in improved regioselectivity (Entries 4, 5). A control reaction in DMA without catalyst gave trace yields (Entry 6), showing that the borinic acid is crucial. However, the switch to NMP did not prove to be productive (Entry 7). The use of solvent mixtures, DMA/MeCN and DMA/H2O, also did not prove to be beneficial (Entries 8, 10). In an attempt to improve yield, we increased the reaction temperature to 50 oC but were met with decreased yield, although complete regiocontrol (Entry 9). This result was puzzling at the time, and will be re-visited later on in the chapter.

Table 03. Solvent screen of methyl α-L-rhamnopyranoside sulfation with TCE-sulfuryl 1,2- dimethylimidazolium triflate.

OTf Cl C O 3 N (1.5 equiv) OMe O S N OMe OMe O O O + O HO HO HO 2.15 (10 mol%) HO TCEO3SO HO OH DIPEA (1.5 equiv) OH OSO TCE Solvent (0.2 M) 3 2.11 24 h, rt 2.12 2.13

Entry Solvent Yield (%)a 2.12:2.13 Ratio

1 MeCN 37 1.8:1 2 DCM 45 1.2:1 3 THF 54 1.2:1 4 DMF 15 4.3:1 5 DMA 41 4.3:1 6b DMA trace - 7 NMP trace 1:1 8 DMA/MeCN (1:1) 35 1.6:1 9c DMA/MeCN (1:1) 20 >20:1

10 DMA/H2O (20:1) - -

aCrude 1H NMR yields were determined based on tetramethylsilane as an internal standard. Product ratio determined by 1H NMR. bNo catalyst 2.15 used. cReaction performed at 50 oC.

After further optimization of the reaction conditions (base, catalyst, various methods of reagent addition) in DMA, we were unable to raise the yield of the desired 3-O-TCE-sulfated product 2.12 above 50% and achieved only modest regioselectivity. Turning to alternate modes of activation, we looked at complexation induced activation with boronic acid derivatives. The pre-formed cyclic arylboronate of methyl α-L-rhamnopyranoside 2.14 was hypothesized to be activated by an external Lewis base (Et3N) through a tetracoordinate intermediate for sulfation (Figure 22). However, the cyclic boronate acted as a protective group instead to give the 4-O-sulfate, followed by hydrolysis of the boronate group during work-up to give 2.15. This showed that the competing background sulfation reaction occurs very rapidly. In the presence of the relatively unreactive 4- hydroxyl group, the direct sulfation reaction is more favorable than sulfation through the activated boronate.

OMe OTf Cl C O O 3 N (1.5 equiv) HO O S N OMe O O O O B O O Et3N (6 equiv) S TCE O O HO MeCN (0.2 M) OH 24 h, rt; aqueous work-up F C 2.15 3 28% 2.14

Figure 22. Sulfation with pre-formed cyclic arylboronate of methyl α-L-rhamnopyranoside.

Revisiting previous work in our group of using electron-deficient boronic acids and Lewis base co-catalyst system for the silylation of pyranosides,42 we sought to adapt those conditions for regioselective sulfation. However, a brief screen of phosphine oxide and phosphoramide Lewis bases with 3,5-bis(trifluoromethyl)phenylboronic acid yielded no regioselectivity, though the 4- O-sulfate was not isolated in this case (Table 04).

Table 04. Boronic acid/Lewis base co-catalyst for sulfation of methyl α-L-rhamnopyranoside.

OMe OMe OMe 2.08 (1.5 equiv) O O + O HO HO HO F3C HO TCEO3SO HO OH OH OSO3TCE B(OH) 2.11 2 2.12 2.13

F3C (20 mol%) Lewis Base (20 mol%) DIPEA (1.5 equiv) MeCN (0.2 M) 24 h, rt

Entry Lewis Base Yield (%)a 2.12:2.13 Ratio

1 n-Bu3P=O 35 1 : 1.2 2 Ph2MeP=O 30 1.1 : 1 3 Ph3P=O 41 1 : 1.1 4 HMPA 33 1.2 : 1

aCrude 1H NMR yields were determined based on tetramethylsilane as an internal standard. Product ratio determined by 1H NMR.

42 Lee, D.; Taylor, M. S. Org. Biomol. Chem. 2013, 11, 5409-5412.

To evaluate the effects of the sulfate ester substituent on reactivity and selectivity, we attempted to synthesize a series of alkyl- and arylsulfuryl 1,2-dimethylimidazolium triflate (Figure 23). The reaction of sulfuryl chloride with phenol and para-methoxyphenol gave 2.14 and 2.15, respectively, in good yields. However, the same reaction with benzyl alcohol, para-methoxybenzyl alcohol, or sec-butyl alcohol gave products in trace yields or decomposed upon work-up or storage under air at room temperature overnight. Previously attempts to prepare benzyl chlorosulfates were reported to lead to the formation of polybenzyl,43 and thus benzyl and alkyl chlorosulfates were not pursued further. Reaction of 2.14 and 2.15 with 2-methylimidazole gave 2.16 and 2.17, respectively, and methylation gave the triflate salts 2.18 and 2.19, respectively, in good overall yields.

O Cl S Cl HN N O MeOTf O OTf (1 equiv) (3.6 equiv) O (1 equiv) O R-OH O S Cl O S N N O S N N pyridine, Et O R THF Et O 2 O R O 2 R O 2.14 R = Ph 54% 2.16 R = Ph 82% 2.18 R = Ph 73% 2.15 R = p-OMePh 94% 2.17 R = p-OMePh 31% 2.19 R = p-OMePh 82% R = Bn decomposition R = PMB decomposition R = sec-Bu trace

Figure 23. Preparation of alkyl- and arylsulfuryl 1,2-dimethylimidazolium triflate.

Sulfation with 2.18 and 2.19 gave markedly better regioselectivity than with TCE-sulfuryl 1,2- dimethylimidazolium triflate (Table 05). Amide solvents such as DMF and DMA were, again, particularly effective (Entries 4-7 and 10) and the presence of catalyst was crucial (Entries 1, 3, 8). The optimal conditions achieved gave 52% yield with 13:1 selectivity of 3-O-sulfate:2-O-sulfate (Entry 6). However, this yield could not be improved further.

43 Gibbons, R. A.; Gibbons, M. N.; Wolfrom, M. L. J. Am. Chem. Soc. 1955, 77, 6374-6374.

Table 05. Optimization of methyl α-L-rhamnopyranoside sulfation with arylsulfuryl 1,2- dimethylimidazolium triflate.

OTf R O N O S N OMe OMe OMe O (1.5 equiv) O O + O HO HO HO Catalyst (10 mol%) HO DIPEA (1.5 equiv) TCEO3SO HO OH OH OSO TCE Solvent (0.2 M) 3 2.11 24 h, rt 2.12 2.13

Entry Catalyst Solvent Yield (%)a 2.12:2.13 Ratio

R = H, 2.18 1 no catalyst MeCN 29 2:1 2 2.15 MeCN 49 8:1 3 no catalyst DMF 0 - 4 2.15 DMF 30 >20:1 5 2.15 DMA 36 >20:1 6b 2.15 DMA/MeCN (1:1) 52 13:1 7 2.15 DMA/MeCN (1:1) 48 >20:1 R = p-OMe, 2.19 8 no catalyst MeCN 34 2:1 9 2.15 MeCN 48 8:1 10 2.15 DMA 20 >20:1 aCrude 1H NMR yields were determined based on tetramethylsilane as an internal standard. Product ratio determined by 1H NMR. b3 equiv of 2.18 and DIPEA used instead.

A closer examination of the sulfating reagents revealed that 2.08, 2.18, and 2.19 underwent appreciable base-mediated hydrolysis in deuterated acetonitrile, as monitored over time by 1H NMR spectroscopy (Figure 24). The triflate salts, in the absence of base, were stable in solution (<2% decomposition after 24 h, not shown in Figure 24 for clarity). 2.08 with 1 equiv. of 1,2- dimethylimidazole (Figure 24, A) or 1 equiv. of DIPEA (Figure 24, B) showed >50% decomposition after 24 h. 2.18 with 1 equiv. of DIPEA (Figure 24, C) showed >60% decomposition, while only 50% decomposition with 1 equiv. of DIPEA dried overnight with 3 molecular sieves (Figure 24, D). The greater disparity after 24 h was seen with 2.19 in the presence of DIPEA (Figure 24, E, 52% decomposition) and 3 MS-dried DIPEA (Figure 24, F, 28% decomposition). This base-mediated hydrolysis of the sulfuryl imidazolium triflates explains our

29 inability to increase sulfation yields by heating the reaction or increasing the concentration—both changes presumably accelerated the decomposition of sulfating reagents to a greater extent than they increased the rate of sulfation. Although rigorously drying the reaction mixture would, in theory, prevent hydrolysis and decomposition of the sulfating reagent, we decided to place this portion of the project on hold and focus instead on direct, regioselective sulfation without masking groups.

2.3 Sulfation of carbohydrates with sulfur trioxide amine complexes

We began studying direct sulfation using methyl α-D-mannopyranoside 2.20 as a model substrate (Table 06). In the absence of catalyst (Entry 1), the reaction mixture included unreacted starting material and a distribution of O-sulfate and bis-sulfates. A brief catalyst screen (Entries 2-4) revealed that the reaction is highly influenced by catalyst, with the product being predominantly the 3-O-sulfate 2.21. This is particularly promising given the presence of a more reactive primary hydroxyl group in the substrate. Replacing SO3-pyridine with SO3-Me3N (Entry 5) diminished conversion but afforded a cleaner reaction. Increasing the temperature to 60 oC (Entry 6) and the

30 equivalence of sulfating reagent (Entry 7) gave increased yield while maintaining the regioselectivity of sulfation. Further increasing the amount of sulfating reagent (Entry 8) was not beneficial and afforded more over-sulfated products than the desired mono-3-O-sulfate (3-

OSO3:over-sulfation 3:1).

Table 06. Optimization of methyl α-D-mannopyranoside sulfation with sulfur trioxide amine complex.

HO OH HO OH O Sulfating Reagent (XX equiv) O HO HO - HO Catalyst (10 mol%) O3SO OMe DIPEA OMe 2.20 MeCN (0.2 M) 2.21 3 h, Temperature

Sulfating Reagent Equiv. of Entry Catalyst Temperature (oC) Conversion (%)a Yield (%)a (XX equiv) DIPEA

1 SO3-pyridine (1.5 equiv) 1.5 no catalyst 40 79 28 2 SO3-pyridine (1.5 equiv) 1.5 2.14 40 48 31 3 SO3-pyridine (1.5 equiv) 1.5 2.15 40 82 61 4 SO3-pyridine (1.5 equiv) 1.5 2.16 40 79 66 5 SO3-Me3N (1.5 equiv) 1.5 2.15 40 27 27 6 SO3-Me3N (1.5 equiv) 1.5 2.15 60 49 49 7 SO3-Me3N (3 equiv) 3 2.15 60 90 80 8 SO3-Me3N (4.5 equiv) 4.5 2.15 60 89 67

aCrude 1H NMR yields were determined based on 1,3,5-trimethoxybenzene as an internal standard.

O S Ph O Ph B B Ph Ph B B OH OH 2.14 2.15 2.22

Further optimization was performed on n-octyl α-D-galactopyranoside to assess the efficiency of sulfation on a more soluble substrate with the standard conditions set out previously (Table 06, Entry 7). As discussed by Gilbert in 1962,11 the reactivities of sulfur trioxide amine complexes vary inversely with the strength of the Lewis base component. A survey of conventional sulfur trioxide amine complexes in our reaction, however, afforded the opposite trend (Table 07). SO3-

Et3N (Entry 2) gave higher conversion but the regioselectivity of sulfation suffered drastically (3-

OSO3:over-sulfation 0.6:1). Weaker Lewis base complexes (Entry 3, 4) gave low or no conversion.

To test the hypothesis that a highly Lewis basic additive could accelerate the sulfation, quinuclidine was added to the reaction with either SO3-Me3N (Entry 5) or SO3-DMF (Entry 6); however, both changes were not fruitful.

Table 07. Optimization of n-octyl α-D-galactopyranoside sulfation with sulfur trioxide amine complex.

OH OH OH OH O SO3-Me3N (3 equiv) O - HO 2.15 (10 mol%) O3SO OH DIPEA (3 equiv) OH O-Octyl MeCN (0.2 M) O-Octyl 2.23 3 h, 60 oC 2.24

Entry Changes to Above Conditions Conversion (%)a Yield (%)a

1 - 79 60 2 SO3-Et3N (3 equiv) instead of SO3-Me3N >95 36 3 SO3-pyridine (3 equiv) instead of SO3-Me3N 51 17 4 SO3-DMF (3 equiv) instead of SO3-Me3N 0 0 5 quinuclidine (3 equiv) as additive 1 1 6 SO3-DMF (3 equiv) instead of SO3-Me3N, quinuclidine (3 equiv) as additive 0 0 aCrude 1H NMR yields were determined based on 1,3,5-trimethoxybenzene as an internal standard.

With the optimized conditions in hand, we began to explore the substrate scope (Table 08). The sulfated products as a trialkylamine salt were exchanged with DOWEX Na+-resin to give the final product as a sodium salt. Substrates without primary hydroxyl groups gave sulfated products in high yields (2.25, 2.26). Sulfation could also be performed in good yields in the presence of primary hydroxyl groups for a wide range of manno- and galacto-derived pyranosides. No significant difference in reactivity between α- and β-anomers of methyl-D-galactopyranoside was observed (2.29 and 2.30). A known sulfated glycolipid 2.34 found in the mammalian nervous system was also synthesized in good yield.

Table 08. Substrate scope of carbohydrate sulfation with SO3-Me3N.

O O SO3-Me3N (3 equiv) + - Na O3SO (HO)n 2.15 (10 mol%) OR (HO) OR DIPEA (3 equiv) n MeCN (0.2 M) 3 h, 60 oC

OMe OMe HO OH HO OH O O O O HO HO HO OH + - + - - + Na O3SO Na O3SO OSO3 Na Na+ -O SO OH 3 OH OMe SPh

>99% 72% 65% 54% 2.25 2.26 2.27 2.28

OH OH OH OH OH OH O O O + - Na O3SO + - OMe + - O OH Na O3SO Na O3SO OH OH OMe 53% 57% 57% 2.29 2.30 2.31

O OH OH OH OH OH OH O HN C15H31 O O i + - O C H + - S Pr Na O3SO Na+ -O SO 13 27 Na O3SO AcHN 3 OH OMe OH OH 54% 46% 42%a 2.32 2.33 2.34

Reactions were performed at 0.1 mmol scale of substrate. Isolated yields are reported. aReaction was performed at 0.005 mmol scale of substrate with 6 equiv. SO3-Me3N, 40 mol% 2.15, 6 equiv. DIPEA, and 0.5 M MeCN.

2.4 Summary and future work

In summary, a catalytic, regioselective sulfation method was outlined that tolerated a range of fully unprotected carbohydrates. Our approach is similar to the one described by Flitsch in 199419,20 in that both strategies utilize cis-diol activation to selectively functionalize at the equatorial hydroxyl group. However, our method uses benign, organoboron compound in catalytic quantities as opposed to toxic, organotin reagent in stoichiometric amounts. Furthermore, our one-step

33 sulfation/activation is achieved in drastically shorter reaction time compared to those reported using the two-step process with dibutyltin oxide. There is still work to be done to understand the mechanism of sulfation and the reactivity of the sulfur trioxide amine complexes, particularly why the trend observed with various Lewis bases of the complex (Table 07) is opposite to that previously reported.11

2.5 Experimental

2.5.1. General Information

All reactions were performed using a Teflon-coated magnetic stir bar under argon. All solvents used were dried using the Pure Solv-MD solvent purification system (Innovative Technology) or previously dried overnight with 3 molecular sieves. All reagents and carbohydrates, unless otherwise stated, were purchased from Sigma-Aldrich or Carbosynth Ltd. Flash column chromatography was performed using silica gel (60 , 230-400 mesh) (SiliCycle). Analytical thin-plate chromatography was performed on aluminum-backed silica gel 60 F254 plates (EMD Milipore) and visualized under UV or with aqueous basic permanganate stain.

1H, 13C and 2D nuclear magnetic resonance (NMR) spectra were acquired on the Agilent DD2 600 MHz or Agilent DD2 500 MHz, both equipped with a OneNMR probe. Chemical shifts (δ) are reported in parts per million (ppm), calibrated to the residual protium in the deuterated solvent. Spectral features are tabulated as follows: chemical shift (δ, ppm); multiplicity (app = apparent, s = singlet, d = doublet, t = triplet, q = quartet, p = pentet, hept = heptet, m = multiplet, where the range of chemical shift is given); number of protons, coupling constants (J, Hz); assignment. Infrared (IR) spectra were acquired on the Fourier-transform Spectrum 100 spectrometer (PerkinElmer) equipped with a single-bounce diamond/ZnSe ATR accessory. Spectral features are tabulated as follows: wavenumber (cm-1); intensity (s = strong, m = medium, w = weak, br = broad). High-resolution mass spectra (HRMS) were acquired on the Agilent 6538 UHD Q-TOF for electrospray ionization, negative mode (ESI−). Optical rotations were acquired on the AUTOPOL IV (Rudolph Research Analytical) in a 0.6 dm polarimeter sample cell, at 589 nm wavelength, at 20 oC, and the sample concentrations are reported in g per 100 mL in methanol. Melting points were acquired on the Mel-Temp II (Laboratory Devices Inc.), and reported as a

34 range of melting or decomposing (denoted by dec.).

Presence of a single sulfate group is confirmed by HRMS. Assignment of sulfation position is based on change in NMR chemical shift between starting carbohydrate and sulfated product (1H: approx. +0.8 ppm, 13C: approx. +7 ppm).

2.5.2. General Procedure A

To a 2-dram vial equipped with a magnetic stir bar was added the carbohydrate (0.1 mmol, 1 equiv), sulfur trioxide trimethylamine complex (42 mg, 0.3 mmol, 3 equiv), and 2.15 (2 mg, 0.01 mmol, 0.1 equiv). The reaction vial was capped with a septum and purged with argon. Acetonitrile (0.5 mL, 0.2 M) was added to the vial, followed by N,N-diisopropylethylamine (0.06 mL, 0.3 mmol, 3 equiv). The septum was quickly replaced with a screw cap, sealed with Teflon tape, and the reaction was stirred at 60 oC for 3 hours. The mixture was then quenched with MeOH and the solvent was removed by rotary evaporation. The crude mixture was purified by flash chromatography on silica gel (2% to 15% MeOH in DCM). Fractions containing the product were combined and stirred with Dowex 50WX2 Na+-form (50–100 mesh) for 30 min. The resulting mixture was dried, filtered through Celite with DCM, then MeOH. The MeOH fraction was collected and dried to give the product as a solid.

2.5.3. Preparation of catalyst and carbohydrate substrates

10H-Dibenzo[b,e][1,4]oxaborinin-10-ol (2.15):

O O 1) n-BuLi, diphenyl ether, THF 2) tributyl borate B 3) 4N HCl OH

10H-Dibenzo[b,e][1,4]oxaborinin-10-ol was prepared according to literature procedure44 from diphenyl ether and tributyl borate (Sigma-Aldrich). Spectral features are in agreement with those previously reported.

Methyl-2-acetamido-2-deoxy-α-D-galactopyranoside:

OH OH OH OH Amberlyst H+ resin O O HO MeOH, reflux HO AcHN AcHN OH OMe

Methyl-2-acetamido-2-deoxy-α-D-galactopyranoside was prepared according to an adapted literature procedure45 from N-acetyl-D-galactosamine (Sigma-Aldrich). The ⍺-anomer was separated cleanly by flash chromatography on silica gel to give the desired product. Spectral features are in agreement with those previously reported.46

β-D-galactosyl N-palmitoyl-D-erythro-sphingosine:

1) Ms2O, PMP, CH2Cl2 2) O

HN C15H31 HO C13H27 O OPMB OH OPMB PMBO PMBO (Ph2B)2O, CH2Cl2 HN C H O O 15 31 O C13H27 PMBO 3) CF3COOH, anisole, CH2Cl2 PMBO PMBO OH PMBO OH

44 Dimitrijević, E.; Taylor, M. S. Chem. Sci. 2013, 4, 3298-3303. 45 Liang, H.; Grindley, T. B. J. Carbohydr. Chem. 2004, 23, 71-82. 46 Grönberg, G.; Nilsson, U.; Bock, K.; Magnusson, G. Carbohydr. Res. 1994, 257, 35-54.

β-D-galactosyl N-palmitoyl-D-erythro-sphingosine was prepared according to a literature procedure47 from 2,3,4,6-tetra-O-4-methoxybenzyl-D-galactopyranose and N-palmitoyl-D- sphingosine (Sigma-Aldrich). Spectral features are in agreement with those previously reported.

2.5.4. Synthesis and characterization of compounds

2,2,2-Trichloroethoxysulfuryl chloride (2.02):

2.02 was prepared according to a literature procedure.31 Cl3C O O S Cl Spectral features are in agreement with those previously O reported.

Chemical Formula: C2H2Cl4O3S Molecular Weight: 247.8950

Methyl 6-O-tert-butyldimethylsilyl-α-D-mannopyranoside 2,3-cyclic sulfate (2.05):

1 H NMR (399 MHz, CDCl3) δ 5.00 (app s, 1H, H-1), 4.94 O OTBS (dd, J = 5.3, 0.9 Hz, 1H, H-2), 4.89 (dd, J = 7.8, 5.3 Hz, 1H, O S O O H-3), 4.25 (dd, J = 9.8, 7.8 Hz, 1H, H-4), 3.96 (dd, J = 10.6, HO O 4.9 Hz, 1H, H-6a), 3.88 (dd, J = 10.6, 5.7 Hz, 1H, H-6b), 3.65 OMe (dddd, J = 9.8, 5.6, 4.9, 0.6 Hz, 1H, H-5), 3.42 (s, 3H, 1- Chemical Formula: C13H26O8SSi OCH ), 0.91 (s, 9H, TBS), 0.12 (d, J = 3.2 Hz, 6H, TBS); 13C Molecular Weight: 370.4880 3 NMR (100 MHz, CDCl3) δ 95.4 (C-1), 85.3 (C-3), 78.9 (C-

2), 69.1 (C-4), 67.8 (C-5), 64.0 (C-6), 55.3 (1-OCH3), 25.7 (TBS), 18.2 (TBS), -5.5 (TBS), -5.6 (TBS).

47 D’Angelo, K. A.; Taylor, M. S. Chem. Commun. 2017, 53, 5978-5980.

Methyl 4,6-bis-O-(tert-butyldimethylsilyl)-α-D-mannopyranoside 2,3-cyclic sulfate (2.06):

1 H NMR (400 MHz, CDCl3) δ 4.99 (d, J = 0.8 Hz, 1H, H-1), O OTBS 4.94 (dd, J = 5.2, 0.9 Hz, 1H, H-2), 4.80 (dd, J = 7.7, 5.3 Hz, O S O 1H, H-3), 4.19 (dd, J = 9.9, 7.8 Hz, 1H, H-4), 3.88 (dd, J = O TBSO 11.5, 2.0 Hz, 1H, H-6a), 3.82 (dd, J = 11.5, 4.8 Hz, 1H, H- O OMe 6b), 3.55 (dddd, J = 9.9, 4.8, 2.1, 0.6 Hz, 1H, H-5), 3.40 (s,

Chemical Formula: C19H40O8SSi2 3H), 0.90 (s, 9H), 0.89 (s, 9H), 0.15 (d, J = 11.1 Hz, 6H), 0.08 Molecular Weight: 484.7510 13 (d, J = 3.3 Hz, 6H); C NMR (100 MHz, CDCl3) δ 95.3 (C-

1), 87.0 (C-3), 79.7 (C-2), 70.6 (C-5), 67.2 (C-4), 61.7 (C-6), 54.9 (1-OCH3), 25.8 (TBS), 25.7 (TBS), 18.3 (TBS), 18.0 (TBS), -4.7 (TBS), -5.2 (TBS), -5.3 (TBS), -5.4 (TBS).

1-(2,2,2-Trichloroethoxysulfuryl)-2-methyl imidazole (2.07):

2.07 was prepared according to a literature procedure.31 Cl C O 3 Spectral features are in agreement with those previously O S N N O reported.

Chemical Formula: C6H7Cl3N2O3S Molecular Weight: 293.5430

1-(2,2,2-Trichloroethoxysulfuryl)-2,3-dimethylimidazolium triflate (2.08):

2.08 was prepared according to a literature procedure.31 O OTf Cl3C Spectral features are in agreement with those O S N N O previously reported.

Chemical Formula: C8F3H10Cl3N2O6S2 Molecular Weight: 308.5775

Methyl 3-trichloroethylsulfo-α-L-rhamnopyranoside (2.12):

1 OMe H NMR (500 MHz, CDCl3) δ 4.85 (q, J = 10.8 Hz, 2H,

CH2CCl3), 4.85 (dd, J = 9.2, 3.1 Hz, 1H, H-3), 4.71 (d, J = O HO 2.0 Hz, 1H, H-1), 4.32 (dd, J = 3.2, 2.0 Hz, 1H, H-2), 3.78 O O OH S (app t, J = 9.3 Hz, 1H, H-4), 3.75–3.68 (m, 1H, H-5), 3.39 (s, O O 13 3H, 1-OCH3), 1.38 (d, J = 6.1 Hz, 3H, 5-CH3); C NMR (100 Cl3C MHz, CDCl3) δ 100.6 (C-1), 86.8 (C-3), 80.0 (CH2), 70.7 (C- Chemical Formula: C9H15Cl3O8S Molecular Weight: 389.6210 4), 69.3 (C-2), 68.3 (C-5), 55.3 (1-OCH3), 17.7 (5-CH3).

Methyl 2-trichloroethylsulfo-α-L-rhamnopyranoside (2.13):

1 OMe H NMR (399 MHz, CDCl3) δ 4.97 (d, J = 10.7 Hz, 1H,

CH2CCl3), 4.93 (d, J = 1.7 Hz, 1H, H-1), 4.89–4.82 (m, 1H, O HO H-2), 4.78 (s, 1H, CH2CCl3), 4.01 (dd, J = 9.5, 3.3 Hz, 1H, HO O O H-3), 3.71–3.61 (m, 1H, H-5), 3.48 (app t, J = 9.5 Hz, 1H, S O O H-4), 3.40 (s, 3H, 1-OCH3), 1.34 (d, J = 6.2 Hz, 3H, 5-CH3); Cl3C 13 C NMR (101 MHz, CDCl3) δ 97.7, 79.8, 77.9, 72.9, 69.4,

Chemical Formula: C9H15Cl3O8S 68.0, 55.3, 17.4. Molecular Weight: 389.6210

Benzenesulfonyl chloride (2.14):

2.14 was prepared according to a literature procedure.48 Spectral features are in agreement with those previously O O S Cl reported. O Chemical Formula: C6H5ClO3S Molecular Weight: 192.6130

48 DeBergh, J. R.; Niljianskul, N.; Buchwald, S. L. J. Am. Chem. Soc. 2013, 135, 10638-10641.

4-Methoxybenzenesulfonyl chloride (2.15):

2.15 was prepared according to a literature procedure.48 MeO Spectral features are in agreement with those previously O reported. O S Cl O 1 H NMR (400 MHz, CDCl3) δ 7.25 (d, J = 9.2 Hz, 2H, Ph), Chemical Formula: C H ClO S 7 7 4 13 Molecular Weight: 222.6390 6.89 (d, J = 9.2 Hz, 2H, Ph), 3.77 (s, 3H, CH3); C NMR (101

MHz, CDCl3) δ 159.5, 143.5, 122.8, 115.1, 55.8.

2-Methyl-1-(phenylsulfonyl) imidazole (2.16):

2.16 was prepared according to a literature procedure.49

O Spectral features are in agreement with those previously N O S N reported. O

Chemical Formula: C10H10N2O3S Molecular Weight: 238.2610

1-[(p-Methoxyphenyl)sulfonyl]-2-methyl imidazole (2.17):

2.17 was prepared according to an adapted literature MeO procedure.49 Spectral features are in agreement with those 50 O previously reported. N O S N O

Chemical Formula: C11H12N2O4S Molecular Weight: 268.2870

49 Desoky, A. Y.; Hendel, J.; Ingram, L.; Taylor, S. D. Tetrahedron 2011, 67, 1281-1287. 50 Reuillon, T.; Bertoli, A.; Griffin, R. J.; Miller, D. C.; Golding, B. T. Org. Biomol. Chem. 2012, 10, 7610-7617.

1,2-Dimethyl-3-(phenylsulfonyl)-imidazolium triflate (2.18):

2.18 was prepared according to a literature procedure.49 OTf O Spectral features are in agreement with those previously N O S N reported. O

Chemical Formula: C12F3H13N2O6S2 Molecular Weight: 253.2955

1-[(p-Methoxyphenyl)sulfonyl]-2,3-dimethylimidazolium triflate (2.19):

2.19 was prepared according to an adapted literature MeO procedure.49 OTf O N 1 O S N H NMR (400 MHz, CDCl3) δ 7.51 (d, J = 2.4 Hz, 1H, O Im), 7.33 (d, J = 2.4 Hz, 1H, Im), 7.21 (d, J = 9.2 Hz, 2H,

Chemical Formula: C13F3H15N2O7S2 Molecular Weight: 283.3215 Ph), 6.96 (d, J = 9.2 Hz, 2H, Ph), 4.09 (s, 3H, 3-CH3), 13 3.80 (s, 3H, OCH3), 2.93 (s, 3H, 2-CH3); C NMR (101

MHz, CDCl3) δ 160.0, 147.9, 142.5, 123.9, 122.5, 120.6, 115.9, 55.8 (OCH3), 37.3 (3-CH3), 11.9

(2-CH3).

Methyl α-L-fucopyranoside 3-(sodium sulfate) (2.25):

2.25 was prepared according to General Procedure A from OMe methyl α-L-fucopyranoside (18 mg, 0.1 mmol, 1 equiv) and O OH purified by flash chromatography on silica gel (5% to 15% - + OSO3 Na OH MeOH in DCM) to give a beige solid in >95% yield (28 mg). Chemical Formula: C7H13NaO8S Molecular Weight: 280.2228

$% 51 '( [α]# = −20.2° (c 0.1, MeOH) (lit. : [α]& = −157.4° (c 1.3, H2O)); m.p.: 212–214 (dec.) 51 o 1 (lit. , monohydrate: 161–162 C); H NMR (600 MHz, MeOD-d4) δ 4.68 (d, J = 3.9 Hz, 1H, H- 1), 4.47 (dd, J = 10.2, 3.2 Hz, 1H, H-3), 4.07 (dd, J = 3.2, 1.2 Hz, 1H, H-4), 3.97–3.92 (m, 2H, H- 13 2, H-5), 3.39 (s, 3H, 1-OCH3), 1.23 (d, J = 6.5 Hz, 3H, 5-CH3); C NMR (151 MHz, MeOD-d4)

δ 100.0 (C-1), 78.0 (C-3), 70.3 (C-4), 66.6 (C-2), 65.8 (C-5), 54.1 (1-OCH3), 15.1 (5-CH3); IR (thin film, cm-1): 3438 (br, m), 2946 (w), 1650 (br, m), 1216 (s), 1046 (s), 987 (s), 839 (s), 748 − − (m); HRMS (ESI ): calcd for C7H13O8S [M − Na] 257.0337 m/z; found 257.0341 m/z.

Methyl α-L-rhamnopyranoside 3-(sodium sulfate) (2.26):

2.26 was prepared according to General Procedure A from OMe methyl α-L-rhamnopyranoside (18 mg, 0.1 mmol, 1 equiv) O HO and purified by flash chromatography on silica gel (5% to + - Na O3SO OH 15% MeOH in DCM) to give a white solid in 79% yield (22 Chemical Formula: C7H13NaO8S Molecular Weight: 280.2228 mg).

$% 1 [α]# = −94.3° (c 0.27, MeOH); m.p.: 201–202 (dec.); H NMR (600 MHz, MeOD-d4) δ 4.58 (d, J = 1.8 Hz, 1H, H-1), 4.42 (dd, J = 9.5, 3.3 Hz, 1H, H-3), 4.16 (dd, J = 3.3, 1.8 Hz, 1H, H-2),

3.65–3.60 (m, 1H, H-5), 3.55 (app t, J = 9.5 Hz, 1H, H-4), 3.36 (s, 3H,1-OCH3), 1.28 (d, J = 6.2 13 Hz, 3H, 5-CH3); C NMR (151 MHz, MeOD-d4) δ 101.2 (C-1), 78.8 (C-3), 70.61 (C-4), 69.2 (C- -1 2), 68.3 (C-5), 53.7 (1-OCH3), 16.6 (5-CH3); IR (thin film, cm ): 3407 (br, m), 2943 (w), 2532 (br, m), 1653 (br, m), 1217 (s), 1049 (s), 970 (s), 840 (s), 782 (m); HRMS (ESI−): calcd for − C7H13O8S [M − Na] 257.0337 m/z; found 257.0340 m/z.

Methyl α-D-mannopyranoside 3-(sodium sulfate) (2.27):

51 Leder, I. G. J. Carbohydr. Chem. 1993, 12, 95-103.

2.27 was prepared according to General Procedure A from HO OH Methyl α-D-mannopyranoside (19 mg, 0.1 mmol, 1 equiv) O HO + - and purified by flash chromatography on silica gel (5% to Na O3SO OMe 15% MeOH in DCM) to give a beige solid in 65% yield (19 Chemical Formula: C7H13NaO9S Molecular Weight: 296.2218 mg).

$% 52 '( 52 [α]# = +9.1° (c 0.21, MeOH) (lit. : [α]& = +55° (c 0.9, MeOH)); m.p.: 232–234 (dec.) (lit. : o 1 246–248 C); H NMR (600 MHz, MeOD-d4) δ 4.67 (d, J = 1.8 Hz, 1H, H-1), 4.48 (dd, J = 9.6, 3.3 Hz, 1H, H-3), 4.17 (dd, J = 3.3, 1.9 Hz, 1H, H-2), 3.84 (dd, J = 12.0, 2.4 Hz, 1H, H-6a), 3.82 (app t, J = 3.6 Hz, 1H, H-4), 3.73 (dd, J = 11.8, 5.7 Hz, 1H, H-6b), 3.57 (ddd, J = 10.0, 5.6, 2.3 13 Hz, 1H, H-5), 3.39 (s, 3H, 1-OCH3); C NMR (151 MHz, MeOD-d4) δ 101.1 (C-1), 79.0 (C-3), -1 73.1 (C-5), 69.0 (C-2), 65.3 (C-4), 61.3 (C-6), 53.8 (1-OCH3); IR (thin film, cm ): 3461 (br, m), − 2690 (br, m), 1201 (s), 1045 (s), 964 (s), 839 (s), 788 (m); HRMS (ESI ): calcd for C7H13O9S [M − Na]− 273.0286 m/z; found 273.0285 m/z.

Phenyl α-D-thiomannopyranoside 3-(sodium sulfate) (2.28):

2.28 was prepared according to General Procedure A from HO OH O phenyl α-D-thiomannopyranoside (27 mg, 0.1 mmol, 1 HO + - Na O3SO equiv) and purified by flash chromatography on silica gel SPh (5% to 15% MeOH in DCM) to give a white solid in 54% Chemical Formula: C12H15NaO8S2 Molecular Weight: 374.3538 yield (20 mg).

$% 1 [α]# = +30.4° (c 0.25, MeOH); m.p.: 181–184 (dec.); H NMR (600 MHz, MeOD-d4) δ 7.55– 7.52 (m, 2H, 1-SPh), 7.33–7.30 (m, 2H, 1-SPh), 7.29–7.25 (m, 1H, 1-SPh), 5.44 (d, J = 1.2 Hz, 1H, H-1), 4.49 (dd, J = 9.4, 3.2 Hz, 1H, H-3), 4.47 (dd, J = 3.2, 1.6 Hz, 1H, H-2), 4.11 (ddd, J = 10.2, 5.1, 2.7 Hz, 1H, H-5), 3.92 (app t, J = 9.6 Hz, 1H, H-4), 3.82–3.76 (m, 2H, H-6a,b); 13C

NMR (151 MHz, MeOD-d4) δ 134.1 (1-SPh), 131.7 (1-SPh), 128.7 (1-SPh), 127.2 (1-SPh), 88.6 (C-1), 79.0 (C-3), 74.3 (C-5), 70.5 (C-2), 65.4 (C-4), 61.0 (C-6); IR (thin film, cm-1): 3391 (br,

52 Contreras, R. R.; Kamerling, J. P.; Vliegenthart, J. F. G. Recl. Trav. Chim. Pays-Bas 1991, 110, 85-88.

m), 2935 (w), 2499 (br, m), 1638 (br, w), 1220 (s), 1056 (s), 995 (s), 738 (s); HRMS (ESI−): calcd − for C12H15O8S2 [M − Na] 351.0214 m/z; found 351.0207 m/z.

Methyl α-D-galactopyranoside 3-(sodium sulfate) (2.29):

2.29 was prepared according to General Procedure A from OH OH methyl α-D-galactopyranoside (19 mg, 0.1 mmol, 1 equiv) O + - and purified by flash chromatography on silica gel (5% to Na O3SO OH OMe 15% MeOH in DCM) to give a white solid in 50% yield (15

Chemical Formula: C7H13NaO9S mg). Molecular Weight: 296.2218

$% 53 '( 53 [α]# = +23.3° (c 0.25, MeOH) (lit. : [α]& = +146 (c 1.1, H2O)); m.p.: 219–221 (dec.) (lit. , o 1 recrystallized: 166–167 C); H NMR (600 MHz, MeOD-d4) δ 4.76 (d, J = 3.8 Hz, 1H, H-1), 4.47 (dd, J = 10.2, 3.2 Hz, 1H, H-3), 4.31 (dd, J = 3.2, 1.2 Hz, 1H, H-4), 3.97 (dd, J = 10.2, 3.9 Hz, 1H, 13 H-2), 3.82–3.79 (m, 1H, H-5), 3.76–3.67 (m, 2H, H-6a,b), 3.42 (s, 3H, 1-OCH3); C NMR (151

MHz, MeOD-d4) δ 99.9 (C-1), 77.8 (C-3), 70.7 (C-5), 67.9 (C-4), 66.8 (C-2), 61.3 (C-6), 54.1 (1- -1 OCH3); IR (thin film, cm ): 3413 (br, m), 2938 (m), 2538 (br, m), 1644 (m), 1211 (s), 1001 (s), − − 972 (s), 859 (s); HRMS (ESI ): calcd for C7H13O9S [M − Na] 273.0286 m/z; found 270.0288 m/z.

Methyl β-D-galactopyranoside 3-(sodium sulfate) (2.30):

2.30 was prepared according to General Procedure A from OH OH methyl β-D-galactopyranoside (19 mg, 0.1 mmol, 1 equiv) O + - OMe and purified by flash chromatography on silica gel (5% to Na O3SO OH 15% MeOH in DCM) to give a white solid in 57% yield (17 Chemical Formula: C7H13NaO9S Molecular Weight: 296.2218 mg).

53 Contreras, R. R.; Kamerling, J. P.; Breg, J.; Vliegenthart, J. F. G. Carbohydr. Res. 1988, 179, 411-418.

$% 20 '( 1 [α]# = +1.2° (c 0.25, MeOH) (lit. : [α]& = +8.3° (c 3.6, MeOH)); m.p.: 228–230 (dec.); H

NMR (500 MHz, MeOD-d4) δ 4.25–4.24 (m, 2H, H-1, H-4), 4.22 (dd, J = 9.0, 3.6 Hz, 1H, H-3), 3.79–3.71 (m, 2H, H-6), 3.68 (dd, J = 9.4, 7.8 Hz, 1H, H-2), 3.56 (ddd, J = 6.7, 5.5, 1.1 Hz, 1H, 13 H-5), 3.53 (s, 3H, 1-OCH3); C NMR (126 MHz, MeOD-d4) δ 104.3 (C-1), 80.6 (C-3), 74.9 (C- -1 5), 69.3 (C-2), 67.2 (C-4), 61.0 (C-6), 55.8 (1-OCH3); IR (thin film, cm ): 3387 (br, m), 2939 (w), 2849 (w), 1643 (br, m), 1383 (br, m), 1216 (s), 996 (s), 811 (m); HRMS (ESI−): calcd for − C7H13O9S [M − Na] 273.0286 m/z; found 273.0291 m/z.

n-Octyl β-D-galactopyranoside 3-(sodium sulfate) (2.31):

2.31 was prepared according to General OH OH Procedure A from n-octyl β-D-galacto- O + - O pyranoside (29 mg, 0.1 mmol, 1 equiv) and Na O3SO OH purified by flash chromatography on silica gel Chemical Formula: C14H27NaO9S Molecular Weight: 394.4108 (5% to 15% MeOH in DCM) to give a white solid in 57% yield (22 mg).

$% 1 [α]# = −0.8° (c 0.27, MeOH); m.p.: 215–216 (dec.); H NMR (500 MHz, MeOD-d4) δ 4.31 (d, J = 7.8 Hz, 1H, H-1), 4.24 (dd, J = 3.3, 1.1 Hz, 1H, H-4), 4.22 (dd, J = 9.6, 3.3 Hz, 1H, H-3),

3.89 (dt, J = 9.5, 6.9 Hz, 1H, O-CH2), 3.75–3.72 (m, 2H, H-6a,b), 3.68 (dd, J = 7.5, 5.0 Hz, 1H,

H-2), 3.58–3.52 (m, 2H, H-5, O-CH2), 1.65–1.58 (m, 2H, CH2), 1.41–1.34 (m, 2H, CH2), 1.34– 13 1.27 (m, 8H), 0.91–0.87 (m, 3H, CH3); C NMR (126 MHz, MeOD-d4) δ 103.3 (C-1), 80.6 (C-

3), 74.8 (C-5), 69.5 (O-CH2), 69.3 (C-2), 67.2 (C-4), 60.9 (C-6), 31.6 (CH2), 29.4 (CH2), 29.2 -1 (CH2), 29.0 (CH2), 25.7 (CH2), 22.3 (CH2), 13.0 (CH3); IR (thin film, cm ): 3422 (br, m), 2926 (m), 2856 (m), 2544 (br, w), 1647 (w), 1376 (w), 1220 (s), 1058 (s), 987 (s), 811 (s); HRMS − − (ESI ): calcd for C14H27O9S [M − Na] 371.1381 m/z; found 371.1380 m/z.

Isopropyl β-D-thiogalactopyranoside 3-(sodium sulfate) (2.32):

2.32 was prepared according to General Procedure A from OH OH isopropyl β-D-thiogalactopyranoside (24 mg, 0.1 mmol, 1 O i equiv) and purified by flash chromatography on silica gel + - S Pr Na O3SO OH (5% to 15% MeOH in DCM) to give a white solid in 54% Chemical Formula: C9H17NaO8S2 Molecular Weight: 340.3368 yield (18 mg).

$% 1 [α]# = −4.0° (c 0.24, MeOH); m.p.: 216–217 (dec.); H NMR (600 MHz, MeOD-d4) δ 4.50 (d, J = 9.7 Hz, 1H, H-1), 4.29 (dd, J = 3.3, 1.1 Hz, 1H, H-4), 4.25 (dd, J = 9.3, 3.2 Hz, 1H, H-3), 3.75–3.68 (m, 3H, H-2, 6a,b), 3.57 (ddd, J = 6.6, 5.4, 1.1 Hz, 1H, H-5), 3.26 (app hept, J = 6.7 Hz, 13 1H, S-CH), 1.31 (dd, J = 10.7, 6.8 Hz, 6H, CH3); C NMR (151 MHz, MeOD-d4) δ 85.4 (C-1),

81.8 (C-3), 78.8 (C-5), 68.4 (C-2), 67.4 (C-4), 61.1 (C-6), 34.3 (S-CH), 23.0 (CH3), 22.9 (CH3); IR (thin film, cm-1): 3401 (br, m), 2965 (m), 2536 (br, m), 1644 (br, w), 1219 (s), 1056 (s), 879 − − (s), 798 (s); HRMS (ESI ): calcd for C9H17O8S2 [M − Na] 317.0370 m/z; found 317.0370 m/z.

Methyl-2-acetamido-2-deoxy-α-D-galactopyranoside 3-(sodium sulfate) (2.33):

2.33 was prepared according to General Procedure A from OH OH methyl-2-acetamido-2-deoxy-α-D-galactopyranoside (24 O + - mg, 0.1 mmol, 1 equiv) and purified by flash Na O3SO AcHN OMe chromatography on silica gel (5% to 15% MeOH in DCM)

Chemical Formula: C9H16NNaO9S to give a white solid in 46% yield (16 mg). Molecular Weight: 337.2748

$% 1 [α]# = +19.7° (c 0.25, MeOH); m.p.: 195–197 (dec.); H NMR (600 MHz, MeOD-d4) δ 4.79 (d, J = 3.5 Hz, 1H, H-1), 4.51 (dd, J = 11.3, 2.9 Hz, 1H, H-3), 4.42 (dd, J = 11.3, 3.5 Hz, 1H, H- 2), 4.29 (dd, J = 2.9, 1.2 Hz, 1H, H-4), 3.84–3.78 (m, 1H, H-5), 3.76–3.69 (m, 2H, H-6a,b), 3.39 13 (s, 3H, 1-OCH3), 1.97 (s, 3H, Ac); C NMR (151 MHz, MeOD-d4) δ 172.2 (C-NHAc), 98.6 (C-

1), 74.9 (C-3), 70.9 (C-5), 67.3 (C-4), 61.3 (C-6), 54.2 (1-OCH3), 48.5 (C-2), 21.3 (Ac); IR (thin film, cm-1): 3408 (br, m), 2942 (w), 2443 (br, w), 1635 (s), 1219 (s), 1109 (s), 990 (s), 845 (s); − − HRMS (ESI ): calcd for C9H16NO9S [M − Na] 314.0551 m/z; found 314.0552 m/z.

(2S,3R,4E)-2-(Hexadecanoylamino)-3-hydroxy-1-[[3-O-(sodiumoxysulfonyl)-beta-D- galactopyranosyl]oxy]-4-octadecene (2.34):

To a 1/2-dram vial equipped with a O OH OH magnetic stir bar was added β-D- HN C H O 15 31 galactosyl N-palmitoyl-D-erythro- + - O C13H27 Na O3SO OH sphingosine (3.5 mg, 0.005 mmol, 1 OH equiv), sulfur trioxide trimethylamine Chemical Formula: C40H76NNaO11S Molecular Weight: 802.0938 complex (4.2 mg, 0.003 mmol, 6 equiv), and 2.15 (0.4 mg, 0.002 mmol, 0.4 equiv). The reaction vial was capped with a septum and purged with argon. Acetonitrile (0.1 mL, 0.05 M) was added to the vial, followed by N,N-diisopropylethylamine (5.2 µL, 0.03 mmol, 6 equiv). The septum was quickly replaced with a screw cap, sealed with Teflon tape, and the reaction was stirred at 60 oC for 3 hours. The mixture was then quenched with MeOH and the solvent was removed by rotary evaporation. The crude mixture was purified by flash chromatography on silica gel (2% to 10% MeOH in DCM) in a Pasteur pipette. Fractions containing the product were combined and stirred with Dowex 50WX2 Na+-form (50–100 mesh) for 30 min. The resulting mixture was dried, filtered through Celite with DCM, then MeOH. The MeOH fraction was collected and dried to give the product as a white solid in 42% yield (1.7 mg).

1 H NMR (500 MHz, MeOD-d4) δ 5.73–5.63 (m, 1H, H-5), 5.44 (ddq, J = 15.2, 7.8, 1.4 Hz, 1H, H-4), 4.34 (d, J = 7.8 Hz, 1H, H-1ʹ), 4.28–4.22 (m, 2H, H-1a, H-4ʹ), 4.25 (dd, J = 8.0, 3.5 Hz, 1H, H-3ʹ), 4.11 (t, J = 8.2 Hz, 1H, H-3), 3.94 (dt, J = 8.6, 3.5 Hz, 1H, H-2), 3.79–3.71 (m, 3H, H-2ʹ, H-6ʹa,b), 3.60–3.53 (m, 2H, H-1b, H-5ʹ), 2.17 (t, J = 7.8 Hz, 1H, α-C=O), 2.05–2.00 (m, 2H), 1.58

(s, 1H, β-C=O), 1.41–1.36 (m, 2H, H-7), 1.29 (s, 44H, aliphatic CH2), 0.92–0.88 (m, 6H, CH3); 13 C NMR (126 MHz, MeOD-d4) δ 175.9, 134.9, 131.4, 105.1, 81.9, 76.5, 72.8, 71.1, 69.8, 68.5, 62.4, 54.7, 37.4, 33.5, 33.1, 33.1 (2C), 30.9–30.4 (18C), 27.2, 23.8 (2C), 14.5 (2C); HRMS (ESI−): − calcd for C40H76NO11S [M − Na] 778.5145 m/z; found 778.5139 m/z.

Chapter 3 Boronic acid-promoted Fischer esterification

Fischer esterification, first described by Emil Fischer and Arthur Speier in 1895,54 is the condensation between a carboxylic acid and alcohol in the presence of an acid catalyst, typically at high temperatures. Due to the reversibility of this reaction, the water produced as the byproduct must be continuously removed, often via a Dean-Stark apparatus. The method described in this chapter focuses on the esterification between a fatty acid and sugar polyol promoted by boronic acid as an efficient way to synthesize non-ionic surfactants in a regioselective manner.

3.1 Introduction

Surfactants are amphiphilic molecules containing both a hydrophobic group (fatty acid chain) and a hydrophilic group (free hydroxyl groups of the sugar alcohol). Interest in the condensation of fatty acids and sugar alcohols arises from the advantage that both components are derived from renewable starting materials that can be used to generate a large pool of chiral products. For example, fatty acids of various chain lengths 3.01–3.05 (Figure 25) are isolated from plants or from the saponification of animal and vegetable oils. Sugar alcohols 3.06–3.09 (Figure 25) are derived from the reduction of monosaccharide sugars or, in the case of glycerol 3.06, isolated from triglycerides from plant and animal sources.

HO n OH OH OH HO OH Acyl chain length: HO OH HO OH HO OH OH 10 Capric acid 3.01 OH OH OH OH OH OH 12 Lauric acid 3.02 14 Myristic acid 3.03 3.06 3.07 3.08 3.09 16 Palmitic acid 3.04 Glycerol Xylitol Ribitol L-(−)-Arabitol 18 Stearic acid 3.05

Figure 25. Structure of select fatty acids and sugar alcohols.

54 Fischer, E.; Speier, A. Eur. J. Inorg. Chem. 1895, 28, 3252-3258.

47 48

Surfactants are commonly found as soap and detergent, emulsifiers and adhesives, and as additives in food, cosmetics, and agrochemicals. Surfactants of sugar fatty acid esters (SFAE) in particular are biodegradable, non-toxic, and non-hazardous to the environment.55 Glycerol monolaurate (GML), a well-studied example of SFAE, has many applications and biological activities. GML was shown to demonstrate antiviral properties against HIV-1 and SIV (simian immunodeficiency virus that infects non-human primates) by preventing mucosal transmission,56 and against herpes simplex virus 2.57 GML was also effective against a range of infectious bacteria while exhibiting significantly reduced drug resistance compared to common antibiotics.58

Synthesis of SFAE via chemical methods have been described;59 however, high temperatures are required, often leading to discoloration of products and the formation of byproducts through acyl migration, cyclization of sugar alcohols, or decomposition.55 High-boiling point solvents needed for these reactions are also difficult to remove completely. Therefore, SFAE are typically prepared from unprotected sugars via enzymatic methods with lipases (Figure 26).60 However, long reaction times are still required to afford 3.11, and in some cases, only giving the SFAE 3.13 in low yields. Furthermore, enzymatic methods require specialized equipment and technology, are generally more expensive than chemical synthesis, and are not accessible to all research laboratories.

55 Ducret, A.; Giroux, A.; Trani, M.; Lortie, R. Biotechnol. Bioeng. 1995, 48, 214-221. 56 Li, Q.; Estes, J. D.; Schlievert, P. M.; Duan, L.; Brosnahan, A. J.; Southern, P. J.; Reilly, C. S.; Peterson, M. L.; Schultz-Darken, N.; Brunner, K. G. Nature 2009, 458, 1034. 57 Sands, J.; Auperin, D.; Snipes, W. Antimicrob. Agents Chemother. 1979, 15, 67-73. 58 Carpo, B. G.; Verallo-Rowell, V. M.; Kabara, J. J. Drugs Dermatol. 2007, 6, 991-998. 59 Queneau, Y.; Chambert, S.; Besset, C.; Cheaib, R. Carbohydr. Res. 2008, 343, 1999-2009. 60 Gumel, A. M.; Annuar, M. S. M.; Heidelberg, T.; Chisti, Y. Process Biochem. 2011, 46, 2079-2090.

O OH O OH O HO Novozyme 435 + OH O o 14 OH 14 OH 60 C, 72 h OH OH O OH OH 3.04 3.10 3.11 78%

OH OH Mucor miehei O OH OH O Lipozyme TM 20 + O O HO 16 O 16 OH 46 h OH OH OH OH 3.05 3.12 3.13 9%

Figure 26. Preparation of sugar fatty acid esters with lipases.61,62

3.2 Chemical synthesis of sugar fatty acid esters

This project was pioneered by a fellow laboratory group member Sanjay Manhas who established the optimized reaction conditions. I, then, joined the project for the completion of the substrate scope, which will be the main focus of this section. Four sugar alcohols and four fatty acids were examined for the substrate scope (Table 09). Glycerol in the presence of phenylboronic acid formed a 1,2-cyclic boronate ester intermediate, revealing a free, terminal hydroxyl for esterification catalyzed by (1S)-(+)-10-camphorsulphonic acid (CSA) (Entries 1, 2). The reaction was performed in a Dean-Stark apparatus at reflux in toluene for the continuous removal of the water byproduct. Following completion of the reaction, the boronate ester was transesterified with sorbitol upon work-up and the SFAE product was purified by recrystallization. Xylitol (Entries 3- 6), ribitol (Entries 7-10), and L-arabitol (Entries 11, 12) can form several binding modes with phenylboronic ester, either through the 1,2-cis-diol or 1,3-diol. C3 and C5 sugar alcohols were chosen so that after boronate ester formation, only a single hydroxyl group remains for esterification. Yields with xylitol were noticeably better than those of ribitol and L-arabitol. This is because both 1,2- and 1,3-boronate esters formed with xylitol permit a free terminal hydroxyl group for esterification. For ribitol, only 1,3-boronate esters would lead to esterification at the

61 Šabeder, S.; Habulin, M.; Knez, Ž. J. Food Eng. 2006, 77, 880-886. 62 Oguntimein, G. B.; Erdmann, H.; Schmid, R. D. Biotechnol. Lett. 1993, 15, 175-180.

50 terminal hydroxyl group. For L-arabitol, esterification at the terminal hydroxyl group gave rise to two diastereomers (1:1 crude ratio as determined by 1H NMR) separable by recrystallization, and the yields reflect the isolation of a single diastereomer. Our method allows not only high regioselectivity for the terminal alcohol but also prevents over-esterification, a challenge commonly found with conventional chemical synthesis of these polyol surfactants.

Table 09. Substrate scope for fatty acid and sugar alcohol esterification.

O O HO OH O R O HO O R OH B O OH CSA (25 mol%) Ar O PhB(OH)2 (1.2 or 2.4 equiv) or + or or HO R toluene (0.2 M) Ar reflux, 24 h OH (1 equiv) O B OH O O HO OH O O HO O R OH OH B O OH OH O R (1.2 equiv) Ar

Entry Sugar Alcohol Acyl Chain Length Product Yield (%)a

3.14b 1 HO OH 10 61 Glycerol 3.06 2 OH 18 3.15 55

b 3 OH 10 3.16 61 4 12 3.17 64 HO OH Xylitol 3.07 5 OH OH 14 3.18 77 b 6 18 3.19 81

b 7 OH 10 3.20 47 8 HO OH Ribitol 3.08 12 3.21 45 OH OH 9 14 3.22 41 10 18 3.23 44

b 11 OH 10 3.24 38

12 HO OH L-(−)-Arabitol 3.09 18 3.25 33 OH OH

Reactions were performed at 2 or 8 mmol scale of the fatty acid. aIsolated yields are reported. bReaction performed and product characterized by Sanjay Manhas.

The effect of acyl chain length (C10, C12, C14, C18) on the efficiency of esterification was examined for two sugars (xylitol and ribitol). Reaction yield of xylitol exhibits a positive correlation with chain length, but reaction yield of ribitol did not seem to vary significantly with chain length.

3.3 Summary

In summary, an efficient method toward the preparation of SFAE surfactants was described for the esterification of sugar alcohols and fatty acids. The protective boronate ester intermediate not only affords high regiocontrol for esterification at the terminal hydroxyl group but also prevents over- esterification. In contrast to enzymatic methods, this strategy avoids the need for expensive lipases while affording high product yields in a relatively short reaction time.

3.4 Experimental

3.4.1 General Information

1H, 13C and 2D nuclear magnetic resonance (NMR) spectra were acquired on the Bruker Avance III 400 MHz equipped with a BBFO probe, Varian Mercury 400 MHz equipped with a ATB probe, or Agilent DD2 500 MHz equipped with a OneNMR probe. Chemical shifts (δ) are reported in parts per million (ppm), calibrated to the residual protium in the deuterated solvent. Spectral features are tabulated as follows: chemical shift (δ, ppm); multiplicity (app = apparent, s = singlet, d = doublet, t = triplet, q = quartet, p = pentet, h = heptet, m = multiplet, where the range of chemical shift is given); number of protons, coupling constants (J, Hz); assignment. Infrared (IR) spectra were acquired on the Fourier-transform Spectrum 100 spectrometer (PerkinElmer) equipped with a single-bounce diamond/ZnSe ATR accessory. Spectral features are tabulated as follows: wavenumber (cm-1); intensity (s = strong, m = medium, w = weak, br = broad). High- resolution mass spectra (HRMS) were acquired on the JEOL AccuTOF for direct analysis in real time (DART) ion source, positive mode (DART+).

3.4.2 General Procedure B

To a 25 mL or 50 mL round-bottom flask equipped with a magnetic stir bar was added the sugar alcohol (1.2 equiv), fatty acid (1 equiv), (1S)-(+)-10-camphorsulphonic acid (CSA) (25 mol%), and phenylboronic acid (1.2 equiv or 2.4 equiv depending on the sugar alcohol). Toluene (0.2 M) was added to the flask and the reaction flask was fitted to a Dean-Stark apparatus equipped with a reflux condenser. The joints were greased, sealed with Teflon tape, and the reaction setup was purged with argon. The reaction was stirred at reflux for 24 h. The solvent was removed by rotary evaporation and the residue was dissolved in Et2O (~150 mL). The mixture was transferred to a separatory funnel to which was added basic sorbitol solution (1 M Na2CO3/1 M D-sorbitol in H2O; 50 mL for a 2 mmol scale reaction, 200 mL for an 8 mmol scale reaction). The mixture was shaken vigorously for 5 min and the aqueous layer was collected. To the remaining organic layer was added another portion of the basic sorbitol solution and shaken vigorously for 5 min. The organic layer was collected and the combined aqueous layers were back-extracted with Et2O (~150 mL ×

3). The combined organic layers were dried over MgSO4, filtered, and concentrated under reduced pressure. The residue was purified by recrystallization in hexanes or EtOH.

3.4.3 Synthesis and characterization of compounds

(±)-Glycerol monostearate (3.15):

3.15 was prepared according to O General Procedure B from HO O OH glycerol (221 mg, 2.4 mmol,

Chemical Formula: C21H42O4 1.2 equiv), stearic acid (569 Molecular Weight: 358.5630 mg, 2 mmol, 1 equiv), CSA (116 mg, 0.5 mmol, 25 mol%), phenylboronic acid (293 mg, 1.2 mmol, 2.4 equiv), and toluene (10 mL, 0.2 M). The crude residue was purified by recrystallization in EtOH to give a white solid in 55% yield (394 mg).

1 H NMR (500 MHz, DMSO-d6) δ 4.85 (d, J = 5.1 Hz, 1H, O-H), 4.61 (app t, J = 5.6 Hz, 1H, O- H), 4.03 (dd, J = 11.1, 4.2 Hz, 1H, H-1a), 3.89 (dd, J = 11.1, 6.5 Hz, 1H, H-1b), 3.62 (m, 1H, H-

2), 3.39–3.27 (m, 2H, H-3a, H-3b), 2.28 (t, J = 7.4 Hz, 2H), 1.51 (app p, J = 7.0 Hz, 2H), 1.23 (m, 13 28H), 0.89–0.82 (m, 3H); C NMR (126 MHz, DMSO-d6) δ 173.4, 69.7, 65.9, 63.1, 33.9, 31.7, 29.5, 29.5, 29.5, 29.5, 29.4, 29.3, 29.2, 29.2, 28.9, 24.9, 22.5, 14.4; IR (thin film, cm-1): 3295 (br, m), 3236 (br, m), 2916 (s), 2849 (s), 1730 (s), 1471 (m), 1176 (s), 1048 (m), 720 (m); HRMS + + (DART ): calcd for C21H46NO4 [M + NH4] 376.3427 m/z; found 376.3418 m/z.

(±)-Xylitol monolaurate (3.17):

3.17 was prepared according to General OH O Procedure B from xylitol (1.46 g, 9.6 HO O OH OH mmol, 1.2 equiv) and lauric acid (1.60 g,

Chemical Formula: C17H34O6 8 mmol, 1 equiv), CSA (465 mg, 2 mmol, Molecular Weight: 334.4530 25 mol%), phenylboronic acid (2.34 g, 19.2 mmol, 2.4 equiv), and toluene (40 mL, 0.2 M). The crude residue was purified by recrystallization in EtOH to give a white solid in 64% yield (1.71 g).

1 H NMR (500 MHz, DMSO-d6) δ 4.78 (d, J = 5.4 Hz, 1H, O-H), 4.47 (app t, J = 5.6 Hz, 1H, O- H), 4.43 (d, J = 5.4 Hz, 1H, O-H), 4.37 (d, J = 6.3 Hz, 1H, O-H), 4.05 (dd, J = 11.2, 4.4 Hz, 1H, H-1a), 4.00 (dd, J = 11.2, 7.2 Hz, 1H, H-1b), 3.77–3.69 (m, 1H, H-2), 3.55–3.50 (m, 1H, H-4), 3.44–3.40 (m, 2H, H-3, H-5a), 3.38–3.34 (m, 1H, H-5b), 2.27 (t, J = 7.4 Hz, 2H), 1.51 (app p, J = 13 7.2 Hz, 2H), 1.24 (m, 16H), 0.89–0.82 (m, 3H); C NMR (100 MHz, DMSO-d6) δ 173.4, 72.2 (C-4), 70.9 (C-3), 70.0 (C-2), 66.2 (C-1), 63.0 (C-5), 34.0, 31.7, 29.5, 29.4, 29.3, 29.2, 29.2, 28.9, 24.9, 22.5, 14.4; IR (thin film, cm-1): 3403 (br, m), 3289 (br, m), 2916 (s), 2850 (s), 1728 (s), 1471 (m), 1174 (s), 1028 (m), 719 (m).

(±)-Xylitol monomyristate (3.18):

3.18 was prepared according to OH O General Procedure B from xylitol HO O (1.46 g, 9.6 mmol, 1.2 equiv) and OH OH

Chemical Formula: C19H38O6 myristic acid (1.83 g, 8 mmol, 1 Molecular Weight: 362.5070 equiv), CSA (465 mg, 2 mmol, 25

54 mol%), phenylboronic acid (2.34 g, 19.2 mmol, 2.4 equiv), and toluene (40 mL, 0.2 M). The crude residue was purified by recrystallization in EtOH to give a white solid in 77% yield (2.23 g).

1 H NMR (400 MHz, DMSO-d6) δ 4.79 (d, J = 5.4 Hz, 1H, O-H), 4.48 (t, J = 5.5 Hz, 1H, O-H), 4.44 (d, J = 5.1 Hz, 1H, O-H), 4.38 (d, J = 6.4 Hz, 1H, O-H), 4.05 (dd, J = 11.2, 4.4 Hz, 1H, H- 1a), 4.00 (dd, J = 11.2, 7.1 Hz, 1H, H-1b), 3.78–3.68 (m, 1H, H-2), 3.56–3.47 (m, 1H, H-4), 3.48– 3.38 (m, 2H, H-3, H-5a), 3.40–3.32 (m, 1H, H-5b), 2.27 (t, J = 7.4 Hz, 2H), 1.51 (app p, J = 7.2 13 Hz, 2H), 1.24 (m, 20H), 0.89–0.81 (m, 3H); C NMR (101 MHz, DMSO-d6) δ 172.9, 71.7 (C- 4), 70.5 (C-3), 69.5 (C-2), 65.7 (C-1), 62.5 (C-5), 33.5, 31.3, 29.0, 29.0, 29.0, 28.9, 28.7, 28.7, 28.5, 24.4, 22.1, 13.9; IR (thin film, cm-1): 3406 (br, m), 3305 (br, m), 2919 (s), 2851 (s), 1727 (s), 1472 (m), 1173 (s), 1144 (s), 1028 (s), 719 (m).

(±)-Ribitol monolaurate (3.21):

3.21 was prepared according to General OH O Procedure B from ribitol (365 mg, 2.4 HO O mmol, 1.2 equiv) and lauric acid (401 OH OH mg, 2 mmol, 1 equiv), CSA (116 mg, 0.5 Chemical Formula: C17H34O6 Molecular Weight: 334.4530 mmol, 25 mol%), phenylboronic acid (585 mg, 4.8 mmol, 2.4 equiv), and toluene (10 mL, 0.2 M). The crude residue was purified by recrystallization in EtOH to give a white solid in 45% yield (301 mg).

1 H NMR (399 MHz, DMSO-d6) δ 4.82 (d, J = 5.5 Hz, 1H, O-H), 4.70 (d, J = 5.6 Hz, 1H, O-H), 4.60 (d, J = 5.1 Hz, 1H, O-H), 4.36 (t, J = 5.6 Hz, 1H, O-H), 4.14 (dd, J = 11.3, 2.8 Hz, 1H, H- 1a), 3.98 (dd, J = 11.3, 7.5 Hz, 1H, H-1b), 3.82–3.71 (m, 1H, H-2), 3.56 (ddd, J = 10.6, 5.6, 3.4 Hz, 1H, H-5a), 3.54–3.45 (m, 1H, H-4), 3.44–3.33 (m, 2H, H-3, H-5b), 2.28 (t, J = 7.4 Hz, 2H), 13 1.59–1.45 (m, 2H), 1.24 (m, 16H), 0.88–0.83 (m, 3H); C NMR (100 MHz, DMSO-d6) δ 173.5, 73.1 (C-3), 72.9 (C-4), 70.3 (C-2), 66.3 (C-1), 63.5 (C-5), 34.0, 31.7, 29.5, 29.4, 29.3, 29.2, 29.2, 28.9, 14.4; IR (thin film, cm-1): 3397 (br, m), 3291 (br, m), 2917 (s), 2850 (s), 1728 (s), 1472 (m), 1174 (s), 1028 (s), 719 (m).

(±)-Ribitol monomyristate (3.22):

3.22 was prepared according to OH O General Procedure B from ribitol HO O OH OH (365 mg, 2.4 mmol, 1.2 equiv) and

Chemical Formula: C19H38O6 myristic acid (457 mg, 2 mmol, 1 Molecular Weight: 362.5070 equiv), CSA (116 mg, 0.5 mmol, 25 mol%), phenylboronic acid (585 mg, 4.8 mmol, 2.4 equiv), and toluene (10 mL, 0.2 M). The crude residue was purified by recrystallization in EtOH to give a white solid in 41% yield (297 mg).

1 H NMR (399 MHz, DMSO-d6) δ 4.83 (d, J = 5.4 Hz, 1H, O-H), 4.70 (d, J = 5.6 Hz, 1H, O- H), 4.60 (d, J = 5.1 Hz, 1H, O-H), 4.37 (app t, J = 5.6 Hz, 1H, O-H), 4.14 (dd, J = 11.3, 2.8 Hz, 1H, H-1a), 3.98 (dd, J = 11.3, 7.5 Hz, 1H, H-1b), 3.80–3.73 (m, 1H, H-2), 3.56 (ddd, J = 10.6, 5.3, 3.4 Hz, 1H, H-5a), 3.52–3.47 (m, 1H, H-4), 3.42–3.34 (m, 2H, H-3, H-5b), 2.28 (t, J = 7.4 Hz, 2H), 1.51 (app p, J = 7.3 Hz, 2H), 1.24 (m, 20H), 0.85 (t, J = 6.7 Hz, 3H); 13C NMR (100 MHz,

DMSO-d6) δ 173.5, 73.1 (C-3), 72.9 (C-4), 70.3 (C-2), 66.3 (C-1), 63.5 (C-5), 34.0, 31.7, 29.5, 29.5, 29.5, 29.4, 29.3, 29.2, 29.2, 28.9, 24.9, 22.5, 14.4; IR (thin film, cm-1): 3350 (br, m), 3228 (br, m), 2916 (s) 2849 (s), 1723 (s), 1176 (s), 1038 (s), 887 (m), 718 (m), 611 (m).

(±)-Ribitol monostearate (3.23):

To a 25 mL or 50 mL OH O round-bottom flask HO O equipped with a magnetic OH OH

Chemical Formula: C23H46O6 stir bar was added ribitol Molecular Weight: 418.6150 (365 mg, 2.4 mmol, 1.2 equiv), stearic acid (569 mg, 2 mmol, 1 equiv), CSA (116 mg, 0.5 mmol, 25 mol%), and phenylboronic acid (585 mg, 4.8 mmol, 2.4 equiv). Toluene was added to the flask (10 mL, 0.2 M) and the reaction flask was fitted to a Dean-Stark apparatus equipped with a reflux condenser. The joints were greased, sealed with Teflon tape, and the reaction setup was purged with argon. The reaction was stirred at reflux for 24 h. The solvent was removed by rotary evaporation and the residue was transferred to a separatory funnel. The transesterification work-up with sorbitol described in General Procedure B was ineffective due to the formation of an emulsion; therefore, the procedure described below was performed. H2O (~200 mL) was added to the separatory funnel

56 and extracted with hexanes (~150 mL × 3). The combined organic layers were back-extracted with brine (~150 mL), collected, dried over MgSO4, filtered, and concentrated under reduced pressure. To a 50 mL round-bottom flask was added the crude residue and pinacol (2.36 g, 20 mmol, 10 equiv). Toluene was added to the flask (25 mL, 0.1 M) and the reaction flask was fitted to a reflux condenser. The joints were greased, sealed with Teflon tape, and the reaction setup was purged with argon. The reaction was stirred at reflux overnight. The solvent was removed by rotary evaporation and the residue was purified by recrystallization in EtOH to give a white solid in 44% yield (368 mg).

1 H NMR (500 MHz, DMSO-d6) δ 4.84 (d, J = 5.6 Hz, 1H, O-H), 4.72 (d, J = 5.6 Hz, 1H, O-H), 4.62 (d, J = 5.2 Hz, 1H, O-H), 4.39 (app t, J = 5.6 Hz, 1H, O-H), 4.13 (dd, J = 11.3, 2.8 Hz, 1H, H-1a), 3.98 (dd, J = 11.3, 7.6 Hz, 1H, H-1b), 3.80–3.72 (m, 1H, H-2), 3.55 (ddd, J = 10.9, 5.8, 3.4 Hz, 1H, H-5a), 3.52–3.47 (m, 1H, H-4), 3.42–3.34 (m, 2H, H-3, H-5b), 2.28 (t, J = 7.4 Hz, 2H), 1.51 (app p, J = 7.6 Hz, 2H), 1.23 (m, 28H), 0.89–0.81 (m, 3H); 13C NMR (126 MHz, DMSO- d6) δ 173.5, 73.1 (C-3), 72.9 (C-4), 70.3 (C-2), 66.3 (C-1), 63.5 (C-5), 34.0, 31.7, 29.5, 29.4, 29.4, 29.2, 29.1, 29.0, 24.9, 22.5, 14.4; IR (thin film, cm-1): 3294 (br, m), 3233 (br, m), 2916 (s), 2849 + (s), 1730 (s), 1471 (m), 1176 (s), 1048 (m), 720 (m); HRMS (DART ): calcd for C23H47O6 [M + H]+ 419.3373 m/z; found 419.3379 m/z.

L-Arabitol monostearate (3.25):

3.25 was prepared OH OH according to General HO O Procedure B from L-(−)- OH O

Chemical Formula: C23H46O6 arabitol (365 mg, 2.4 Molecular Weight: 418.6150 mmol, 1.2 equiv) and stearic acid (569 mg, 2 mmol, 1 equiv), CSA (116 mg, 0.5 mmol, 25 mol%), phenylboronic acid (585 mg, 4.8 mmol, 2.4 equiv), and toluene (10 mL, 0.2 M). The crude residue was purified by recrystallization in EtOH to give a white solid in 33% yield (276 mg).

1 H NMR (500 MHz, DMSO-d6) δ 4.79 (d, J = 6.0 Hz, 1H, O-H), 4.45 (dd, J = 6.1, 5.2 Hz, 1H, O-H), 4.26 (d, J = 7.7 Hz, 1H, O-H), 4.25 (dd, J = 11.2, 2.5 Hz, 1H, H-1a), 4.21 (d, J = 6.5 Hz,

1H, O-H), 3.94 (dd, J = 11.3, 6.9 Hz, 1H, H-1b), 3.72–3.63 (m, 2H, H-2, H-4), 3.44–3.36 (m, 2H, H-5a,b), 3.35–3.29 (m, 1H, H-2), 2.28 (t, J = 7.4 Hz, 2H), 1.51 (app p, J = 7.7 Hz, 2H), 1.23 (m, 13 28H), 0.89–0.82 (m, 3H); C NMR (126 MHz, DMSO-d6) δ 173.5, 70.7, 70.2, 68.9, 67.2, 63.2, 34.0, 31.7, 29.5, 29.5, 29.4, 29.4, 29.2, 29.2, 29.0, 24.9, 22.5, 14.4; IR (thin film, cm-1): 3399 (br, m), 3293 (br, m), 2919 (s), 2851 (s), 1727 (s), 1472 (m), 1172 (s), 1144 (s), 1028 (s), 719 (m); + + HRMS (DART ): calcd for C23H47O6 [M + H] 418.3373 m/z; found 418.3529 m/z.

Appendices A1. NMR spectra of reported compounds 0 7 4 9 3 1 5 7 9 9 0 7 0 4 2 8 2 9 0 2 0 9 0 0 1 1 1 0 2 1 ...... 1 0 1 1 1 1 1 3 9 6

9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A01. H NMR (399 MHz, CDCl3) of 2.05.

58 59 9 2 5 9 8 2 1 4 7 3 7 3 3 8 0 7 0 3 7 1 5 5 ...... 5 5 8 9 7 4 5 5 8 5 5 9 8 7 6 6 6 5 2 1 - -

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A02. C NMR (100 MHz, CDCl3) of 2.05.

3.1 3.2 3.3 3.4 3.5 3.6 3.7 3.8 3.9 4.0 )

4.1 m p p (

4.2 1 f 4.3 4.4 4.5 4.6 4.7 4.8 4.9 5.0 5.1 5.2 5.2 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 f2 (ppm)

Figure A03. 2D COSY NMR (399 MHz, CDCl3) of 2.05.

70 ) m p p (

75 1 f

100

5.3 5.2 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 3.0 2.9 2.8 2.7 2.6 f2 (ppm)

Figure A04. 2D HSQC NMR (399 MHz, CDCl3) of 2.05.

62 0 1 4 0 2 4 0 9 1 4 6 5 0 1 3 1 2 9 4 8 1 9 5 2 0 0 0 0 1 9 0 9 1 9 1 0 ...... 1 1 1 1 1 0 1 2 9 8 6 6

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A05. H NMR (399 MHz, CDCl3) of 2.06.

63 0 7 4 9 6 9 0 0 8 4 9 2 0 6 4 3 9 7 5 1 6 9 8 6 3 9 7 2 2 4 ...... 5 6 9 0 7 1 4 5 5 8 7 4 5 5 5 9 8 7 7 6 6 5 2 2 1 1 - - - -

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A06. C NMR (100 MHz, CDCl3) of 2.06.

3.3 3.4 3.5 3.6 3.7 3.8 3.9 4.0

4.1 ) m p

4.2 p (

1 4.3 f 4.4 4.5 4.6 4.7 4.8 4.9 5.0 5.1 5.2 5.2 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A07. 2D COSY NMR (399 MHz, CDCl3) of 2.06.

70 ) m p

75 p (

1 f

100 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 f2 (ppm)

Figure A08. 2D HSQC NMR (399 MHz, CDCl3) of 2.06.

66 2 6 9 2 4 2 0 9 0 8 9 9 7 0 ...... 2 1 0 0 0 2 3

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A09. H NMR (500 MHz, CDCl3) of 2.12.

67 0 5 9 1 2 8 7 3 9 2 0 7 6 5 3 4 8 6 3 9 8 8 5 1 9 . 8 9 6 2 2 3 6 0 ...... 0 6 9 0 9 8 5 7 1 8 7 7 6 6 5 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A10. C NMR (100 MHz, CDCl3) of 2.12.

2.9 3.0 3.1 3.2 3.3 3.4 3.5 3.6 3.7 3.8

3.9 ) m p

4.0 p (

4.1 f 4.2 4.3 4.4 4.5 4.6 4.7 4.8 4.9 5.0 5.1 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 3.0 2.9 f2 (ppm)

Figure A11. 2D COSY NMR (500 MHz, CDCl3) of 2.12.

75 ) m p

80 p (

1 f 85

100

105

5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A12. 2D HSQC NMR (500 MHz, CDCl3) of 2.12.

70 0 9 8 5 6 5 7 0 2 0 9 6 7 4 0 9 0 9 0 1 0 ...... 1 0 2 1 3 3 3

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A13. H NMR (400 MHz, CDCl3) of 2.19.

71 0 9 5 9 5 6 9 ...... 8 3 8 0 7 2 3 2 0 5 . . . 6 4 4 2 2 2 1 5 7 1 1 1 1 1 1 1 1 5 3 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A14. C NMR (101 MHz, CDCl3) of 2.19.

6.9

7.0

7.1

7.2 ) m p p ( 7.3 1 f

7.4

7.5

7.6

7.7

7.65 7.60 7.55 7.50 7.45 7.40 7.35 7.30 7.25 7.20 7.15 7.10 7.05 7.00 6.95 6.90 6.85 f2 (ppm)

Figure A15. 2D COSY NMR (400 MHz, CDCl3) of 2.19.

60 ) m p p (

70 f

100

110

120

7.6 7.4 7.2 7.0 6.8 6.6 6.4 6.2 6.0 5.8 5.6 5.4 5.2 5.0 4.8 4.6 4.4 4.2 4.0 3.8 3.6 3.4 3.2 3.0 2.8 f2 (ppm)

Figure A16. 2D HSQC NMR (400 MHz, CDCl3) of 2.19.

74 5 1 6 5 9 9 0 2 1 1 4 7 0 0 0 1 9 0 ...... 1 1 1 2 2 3

14 13 12 11 10 9 8 7 6 5 4 3 2 1 0 -1 -2 f1 (ppm)

1 Figure A17. H NMR (600 MHz, MeOD-d4) of 2.25.

75 0 . 0 3 6 8 1 1 0 ...... 0 8 0 6 5 4 5 1 7 7 6 6 5 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A18. C NMR (151 MHz, MeOD-d4) of 2.25.

3.2

3.3

3.4

3.5

3.6

3.7

3.8

3.9 ) m p

4.0 p (

1 f 4.1

4.2

4.3

4.4

4.5

4.6

4.7

4.8 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A19. 2D COSY NMR (600 MHz, MeOD-d4) of 2.25.

70 ) m p

75 p (

1 f 80

100

4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A20. 2D HSQC NMR (600 MHz, MeOD-d4) of 2.25.

78 0 8 2 1 0 5 2 0 1 1 1 2 3 9 0 0 0 1 0 0 9 ...... 1 1 1 1 1 3 2

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A21. H NMR (600 MHz, MeOD-d4) of 2.26.

79 2 . 8 6 1 3 7 6 1 ...... 0 8 0 9 8 3 6 1 7 7 6 6 5 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A22. C NMR (151 MHz, MeOD-d4) of 2.26.

3.2

3.3

3.4

3.5

3.6

3.7

3.8

3.9 ) m p

4.0 p (

1 f 4.1

4.2

4.3

4.4

4.5

4.6

4.7

4.8 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A23. 2D COSY NMR (600 MHz, MeOD-d4) of 2.26.

70 ) m p

75 p (

1 f 80

100

4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A24. 2D HSQC NMR (600 MHz, MeOD-d4) of 2.26.

82 0 2 2 2 0 2 0 0 3 1 1 2 2 8 0 0 0 1 0 1 9 ...... 1 1 1 2 1 1 2

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A25. H NMR (600 MHz, MeOD-d4) of 2.27.

83 1 . 0 1 0 3 3 8 1 ...... 0 9 3 9 5 1 3 1 7 7 6 6 6 5

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A26. C NMR (151 MHz, MeOD-d4) of 2.27.

3.4

3.5

3.6

3.7

3.8

3.9

4.0 ) m p

4.1 p (

1 f 4.2

4.3

4.4

4.5

4.6

4.7

4.8 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 f2 (ppm)

Figure A27. 2D COSY NMR (600 MHz, MeOD-d4) of 2.27.

75 ) m p p (

1 80 f

100

4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 f2 (ppm)

Figure A28. 2D HSQC NMR (600 MHz, MeOD-d4) of 2.27.

3200

3000

2800

2600

2400

2200

2000

1800

1600

1400

1200

1000

800

600

400

200

0 7 8 7 0 1 4 0 4 1 5 3 8 0 1 5 5 1 4 9 9 9 0 0 9 0 0 1 ...... -200 1 1 0 1 1 0 1 1 2

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A29. H NMR (600 MHz, MeOD-d4) of 2.28.

87 1 7 7 2 . . . . 6 0 3 5 4 0 4 1 8 7 ...... 3 3 2 2 8 9 4 0 5 1 1 1 1 1 8 7 7 7 6 6

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A30. C NMR (151 MHz, MeOD-d4) of 2.28.

3.6 3.7 3.8 3.9 4.0 4.1 4.2 4.3

4.4 ) m p

4.5 p (

1 4.6 f 4.7 4.8 4.9 5.0 5.1 5.2 5.3 5.4 5.5 5.5 5.4 5.3 5.2 5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 f2 (ppm)

Figure A31. 2D COSY NMR (600 MHz, MeOD-d4) of 2.28.

90 ) m p

95 p (

1 f 100

105

110

115

120

125

130

135 7.6 7.4 7.2 7.0 6.8 6.6 6.4 6.2 6.0 5.8 5.6 5.4 5.2 5.0 4.8 4.6 4.4 4.2 4.0 3.8 3.6 f2 (ppm)

Figure A32. 2D HSQC NMR (600 MHz, MeOD-d4) of 2.28.

90 0 0 7 2 2 8 0 0 7 9 6 8 9 1 0 9 9 9 9 0 9 ...... 1 0 0 0 0 2 2

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A33. H NMR (600 MHz, MeOD-d4) of 2.29.

9 8 7 9 8 3 1 ...... 9 7 0 7 6 1 4 9 7 7 6 6 6 5

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A34. C NMR (151 MHz, MeOD-d4) of 2.29.

3.6

3.7

3.8

3.9

4.0

4.1 ) m p p (

4.2 f

4.3

4.4

4.5

4.6

4.7

4.8 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 f2 (ppm)

Figure A35. 2D COSY NMR (600 MHz, MeOD-d4) of 2.29.

75 ) m p p (

1 80 f

100

4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 f2 (ppm)

Figure A36. 2D HSQC NMR (600 MHz, MeOD-d4) of 2.29.

94 5 2 0 5 5 9 7 3 1 0 9 6 6 9 9 9 9 7 ...... 1 0 1 0 0 2

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A37. H NMR (500 MHz, MeOD-d4) of 2.30.

95 5 2 0 0 9 5 5 8 . 6 9 2 1 9 7 4 ...... 0 0 4 9 7 0 5 1 8 7 6 6 6 5

220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A38. C NMR (126 MHz, MeOD-d4) of 2.30.

3.4

3.5

3.6

3.7

3.8

3.9 ) m p p (

1 4.0 f

4.1

4.2

4.3

4.4

4.5 4.50 4.45 4.40 4.35 4.30 4.25 4.20 4.15 4.10 4.05 4.00 3.95 3.90 3.85 3.80 3.75 3.70 3.65 3.60 3.55 3.50 3.45 3.40 f2 (ppm)

Figure A39. 2D COSY NMR (500 MHz, MeOD-d4) of 2.30.

20170704_vnmrs_500_YCL-3-089-columnF2-Na-2-MeOHplug_MeOD_d4-gHSQC_01 — YCL-3-089-columnF2-Na-2-MeOHplug_MeOD_d4 —

75 ) m p

80 p (

1 f 85

100

105

110 4.50 4.45 4.40 4.35 4.30 4.25 4.20 4.15 4.10 4.05 4.00 3.95 3.90 3.85 3.80 3.75 3.70 3.65 3.60 3.55 3.50 3.45 3.40 f2 (ppm)

Figure A40. 2D HSQC NMR (500 MHz, MeOD-d4) of 2.30.

98 8 7 9 4 1 6 3 4 2 8 8 4 9 4 1 3 4 5 8 9 6 7 9 8 9 0 8 1 0 9 8 9 9 ...... 0 0 0 1 1 2 2 1 1 7 2

14 13 12 11 10 9 8 7 6 5 4 3 2 1 0 -1 -2 1H (ppm)

1 Figure A41. H NMR (500 MHz, MeOD-d4) of 2.31.

99 3 . 6 8 5 3 2 9 6 4 2 0 7 3 0 3 ...... 0 0 4 9 9 7 0 1 9 9 9 5 2 3 1 8 7 6 6 6 6 3 2 2 2 2 2 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 13C (ppm)

13 Figure A42. C NMR (126 MHz, MeOD-d4) of 2.31.

100

1.0

1.5

2.0 )

2.5 m p p (

H 1 3.0

3.5

4.0

4.5

4.6 4.4 4.2 4.0 3.8 3.6 3.4 3.2 3.0 2.8 2.6 2.4 2.2 2.0 1.8 1.6 1.4 1.2 1.0 0.8 1H (ppm)

Figure A43. 2D COSY NMR (500 MHz, MeOD-d4) of 2.31.

101

50 ) m p p (

C 3

60 1

100

110 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 1H (ppm)

Figure A44. 2D HSQC NMR (500 MHz, MeOD-d4) of 2.31.

102 0 0 9 1 1 6 7 0 9 9 9 6 0 3 0 9 9 0 0 0 1 ...... 1 0 0 3 1 1 6

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A45. H NMR (600 MHz, MeOD-d4) of 2.32.

103 4 8 8 4 4 1 3 0 9 ...... 5 1 8 8 7 1 4 3 2 8 8 7 6 6 6 3 2 2

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A46. C NMR (151 MHz, MeOD-d4) of 2.32.

104

3.1

3.2

3.3

3.4

3.5

3.6

3.7 )

3.8 m p p (

3.9 1 f

4.0

4.1

4.2

4.3

4.4

4.5

4.6 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 f2 (ppm)

Figure A47. 2D COSY NMR (600 MHz MeOD-d4) of 2.32.

105

72 ) m p p (

74 f

4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 f2 (ppm)

Figure A48. 2D HSQC NMR (600 MHz, MeOD-d4) of 2.32.

106 0 1 2 0 0 0 1 6 0 0 0 9 4 5 9 1 0 0 0 9 0 0 9 9 ...... 1 1 1 0 1 2 2 2

9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A49. H NMR (600 MHz, MeOD-d4) of 2.33.

107 2 . 6 9 9 3 3 2 5 3 2 ...... 7 8 4 0 7 1 4 8 1 1 9 7 7 6 6 5 4 2

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A50. C NMR (151 MHz, MeOD-d4) of 2.33.

108

3.2

3.3

3.4

3.5

3.6

3.7

3.8

3.9 )

4.0 m p p (

4.1 1 f 4.2

4.3

4.4

4.5

4.6

4.7

4.8

4.9 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A51. 2D COSY NMR (600 MHz, MeOD-d4) of 2.33.

109

70 ) m p

75 p (

1 f 80

100

4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A52. 2D HSQC NMR (600 MHz, MeOD-d4) of 2.33.

110 0 0 6 7 1 1 8 4 8 1 3 1 8 4 3 0 0 1 7 6 7 4 8 9 5 9 5 0 9 . 0 0 1 0 8 9 0 8 0 0 0 2 1 ...... 4 . 1 1 1 3 0 0 3 1 2 2 2 2 4 6

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A53. H NMR (500 MHz, MeOD-d4) of 2.34.

111 9 9 4 1 . . . . 9 5 8 1 8 5 4 7 4 5 1 1 9 4 2 8 5 5 4 1 5 ...... 7 3 3 0 1 6 2 1 9 8 2 4 7 3 3 3 0 0 7 3 4 1 1 1 1 8 7 7 7 6 6 6 5 3 3 3 3 3 3 2 2 1

220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A54. C NMR (126 MHz, MeOD-d4) of 2.34.

112

-0.5

0.0

0.5

1.0

1.5

2.0

2.5 ) m p p (

3.0 1 f 3.5

4.0

4.5

5.0

5.5

6.0

6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 f2 (ppm)

Figure A55. 2D COSY NMR (600 MHz, MeOD-d4) of 2.34.

113 0 0 9 5 5 3 4 6 4 9 3 . 0 9 0 0 0 8 0 1 1 ...... 7 . 1 0 1 1 1 2 2 2 2 3

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A56. H NMR (500 MHz, CDCl3) of 3.15.

114 4 . 7 9 1 9 7 5 5 5 4 4 3 2 1 9 9 5 4 3 ...... 7 9 5 3 3 1 9 9 9 9 9 9 9 9 8 4 2 4 1 6 6 6 3 3 2 2 2 2 2 2 2 2 2 2 2 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A57. C NMR (126 MHz, CDCl3) of 3.15.

115

3.1

3.2

3.3

3.4

3.5

3.6 ) m p

3.7 p (

1 f 3.8

3.9

4.0

4.1

4.2

4.3 4.20 4.10 4.00 3.90 3.80 3.70 3.60 3.50 3.40 3.30 3.20 3.10 f2 (ppm)

Figure A58. 2D COSY NMR (500 MHz, CDCl3) of 3.15.

116

55 56 57 58 59 60 61 62 63 64

65 ) m

66 p p (

67 1 f 68 69 70 71 72 73 74 75 76 77

5.1 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A59. 2D HSQC NMR (500 MHz, CDCl3) of 3.15.

117 9 0 4 2 0 5 7 0 3 3 9 0 4 9 1 . 0 0 1 9 0 0 1 1 9 0 1 1 1 ...... 5 . 1 1 1 0 1 1 1 1 1 1 2 2 1 3

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A60. H NMR (500 MHz, CDCl3) of 3.17.

118 4 . 2 9 0 2 0 0 7 5 4 3 2 2 9 9 5 4 3 ...... 7 2 0 0 6 3 4 1 9 9 9 9 9 8 4 2 4 1 7 7 7 6 6 3 3 2 2 2 2 2 2 2 2 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A61. C NMR (100 MHz, CDCl3) of 3.17.

119

3.2

3.3

3.4

3.5

3.6

3.7

3.8

3.9

4.0 ) m p

4.1 p (

1 4.2 f

4.3

4.4

4.5

4.6

4.7

4.8

4.9

5.0 5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A62. 2D COSY NMR (500 MHz, CDCl3) of 3.17.

120

66 ) m p p (

68 1 f 70

5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A63. 2D HSQC NMR (500 MHz, CDCl3) of 3.17.

121 5 0 6 9 0 5 9 3 3 5 7 9 1 1 . 0 1 9 0 1 0 0 5 1 0 1 1 ...... 0 . 1 1 0 1 2 1 1 2 1 2 2 2 3

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A64. H NMR (400 MHz, CDCl3) of 3.18.

122 9 . 7 5 5 7 5 5 3 0 0 0 9 7 7 5 4 1 9 2 ...... 7 1 0 9 5 2 3 1 9 9 9 8 8 8 8 4 2 3 1 7 7 6 6 6 3 3 2 2 2 2 2 2 2 2 2 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A65. C NMR (101 MHz, CDCl3) of 3.18.

123

3.2

3.3

3.4

3.5

3.6 ) m p

3.7 p (

1 f 3.8

3.9

4.0

4.1

4.2

4.3 4.25 4.15 4.05 3.95 3.85 3.75 3.65 3.55 3.45 3.35 3.25 3.15 f2 (ppm)

Figure A66. 2D COSY NMR (400 MHz, CDCl3) of 3.18.

124

65 ) m p p (

1 70 f

4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 f2 (ppm)

Figure A67. 2D HSQC NMR (400 MHz, CDCl3) of 3.18.

125 9 8 5 8 1 5 2 4 9 7 2 7 5 7 0 . 0 0 9 0 0 0 0 1 9 1 0 0 0 ...... 5 . 1 1 0 1 1 1 1 1 0 2 2 2 1 3 10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A68. H NMR (399 MHz, CDCl3) of 3.21.

126 5 . 1 9 3 3 5 0 7 5 4 3 2 2 9 9 5 4 3 ...... 7 3 2 0 6 3 4 1 9 9 9 9 9 8 4 2 4 1 7 7 7 6 6 3 3 2 2 2 2 2 2 2 2 1

230 220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A69. C NMR (100 MHz, CDCl3) of 3.21.

127

3.2

3.3

3.4

3.5

3.6 )

3.7 m p p (

1 f 3.8

3.9

4.0

4.1

4.2

4.25 4.20 4.15 4.10 4.05 4.00 3.95 3.90 3.85 3.80 3.75 3.70 3.65 3.60 3.55 3.50 3.45 3.40 3.35 3.30 3.25 f2 (ppm)

Figure A70. 2D COSY NMR (399 MHz, CDCl3) of 3.21.

128

57 58 59 60 61 62 63 64 65 66 )

67 m p p ( 68 1 f 69 70 71 72 73 74 75 76 77 78

5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 f2 (ppm)

Figure A71. 2D HSQC NMR (399 MHz, CDCl3) of 3.21.

129 3 0 6 3 2 4 2 1 2 2 7 1 7 0 5 . 0 9 9 9 0 0 0 1 0 0 1 0 0 ...... 0 . 1 0 0 0 1 1 1 1 1 2 2 2 2 3 10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A72. H NMR (399 MHz, CDCl3) of 3.22.

130 9 7 1 1 4 5 0 8 2 7 2 2 2 6 9 5 6 7 1 3 3 3 1 8 4 3 2 7 9 9 7 7 9 1 1 5 8 7 7 5 2 0 5 1 1 0 3 9 7 5 4 4 8 5 4 9 3 9 . 1 9 2 3 5 0 7 4 4 4 4 3 1 1 9 8 5 3 3 ...... 7 3 2 0 6 3 4 1 9 9 9 9 9 9 9 8 4 2 4 1 7 7 7 6 6 3 3 2 2 2 2 2 2 2 2 2 2 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A73. C NMR (100 MHz, CDCl3) of 3.22.

131

3.2

3.3

3.4

3.5

3.6 ) m p

3.7 p (

1 f 3.8

3.9

4.0

4.1

4.2

4.25 4.20 4.15 4.10 4.05 4.00 3.95 3.90 3.85 3.80 3.75 3.70 3.65 3.60 3.55 3.50 3.45 3.40 3.35 3.30 3.25 f2 (ppm)

Figure A74. 2D COSY NMR (399 MHz, CDCl3) of 3.22.

132

65 ) m p p (

1 70 f

5.0 4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A75. 2D HSQC NMR (399 MHz, CDCl3) of 3.22.

133 7 0 0 1 9 9 9 0 2 5 4 9 6 1 8 . 0 0 0 9 9 9 0 0 0 0 9 0 9 ...... 8 . 1 1 1 0 0 0 1 1 1 2 1 2 2 2

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A76. H NMR (500 MHz, CDCl3) of 3.23.

134 1 6 1 4 9 0 5 5 6 2 1 4 5 5 6 2 3 8 3 5 9 0 3 3 0 4 6 4 5 9 4 1 2 8 4 5 1 1 5 2 1 0 3 7 4 5 8 4 5 0 3 0 . 1 9 2 3 5 0 7 4 4 3 1 1 9 9 5 4 3 ...... 7 3 2 0 6 3 4 1 9 9 9 9 9 8 4 2 4 1 7 7 7 6 6 3 3 2 2 2 2 2 2 2 2 1

220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A77. C NMR (126 MHz, CDCl3) of 3.23.

135

3.1

3.2

3.3

3.4

3.5

3.6 ) m

3.7 p p (

1 f 3.8

3.9

4.0

4.1

4.2

4.3

4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A78. 2D COSY NMR (500 MHz, CDCl3) of 3.23.

136

65 ) m p p (

1 f

4.9 4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 f2 (ppm)

Figure A79. 2D HSQC NMR (500 MHz, CDCl3) of 3.23.

137 0 0 4 9 8 4 0 1 6 4 2 3 . 0 0 9 8 0 1 1 1 2 1 ...... 8 . 1 1 1 0 1 2 2 2 2 2 3

10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0 -0.5 -1.0 f1 (ppm)

1 Figure A80. H NMR (500 MHz, CDCl3) of 3.25.

138 5 . 7 2 9 2 2 0 7 5 5 4 4 2 2 0 9 5 4 3 ...... 7 0 0 8 7 3 4 1 9 9 9 9 9 9 9 4 2 4 1 7 7 6 6 6 3 3 2 2 2 2 2 2 2 2 2 1

200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0 -10 f1 (ppm)

13 Figure A81. C NMR (126 MHz, CDCl3) of 3.25.

139

3.2

3.3

3.4

3.5

3.6

3.7 ) m p p (

3.8 1 f

3.9

4.0

4.1

4.2

4.3

4.30 4.20 4.10 4.00 3.90 3.80 3.70 3.60 3.50 3.40 3.30 3.20 f2 (ppm)

Figure A82. 2D COSY NMR (500 MHz, CDCl3) of 3.25.

140

66 ) m p p ( 68 1 f

4.8 4.7 4.6 4.5 4.4 4.3 4.2 4.1 4.0 3.9 3.8 3.7 3.6 3.5 3.4 3.3 3.2 3.1 f2 (ppm)

Figure A83. 2D HSQC NMR (500 MHz, CDCl3) of 3.25.