Synthesis and applications of epicatechin and epiafzelechin derivatives from proanthocyanidins

SYNTHESIS AND APPLICATIONS OF EPICATECHIN AND EPIAFZELECHIN DERIVATIVES FROM PROANTHOCYANIDINS FU CAILI NATIONAL UNIVERSITY OF SINGAPORE 2010 SYNTHESIS AND APPLICATIONS OF EPICATECHIN AND EPIAFZELECHIN DERIVATIVES FROM PROANTHOCYANIDINS FU CAILI B Eng China Agricultural University M Eng China Agricultural University A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF PHILOSOPHY DEPARTMENT OF CHEMISTRY NATIONAL UNIVERSITY OF SINGAPORE 2010 Acknowledgements The last four years have been one of the most important stages in my life The experience in my Ph.D period will benefit me for a lifetime I would like to take this opportunity to express my immense gratitude to all those who have kindly helped me and all those who have made my graduate life at NUS both productive and enjoyable At the very first, I am honored to express my deepest gratitude to my dedicated supervisor, Dr HUANG Dejian This thesis would not have been possible without his able supervision He has offered me a great many of invaluable ideas and great suggestions, profound knowledge, and rich research experience From him, I learn not only the knowledge, but also the professional ethics, both of which will stay with me for many years to come His encouragement, patience and kindness throughout all these years are greatly appreciated and I am very much obliged to his efforts of helping me finishing the dissertation My most sincere gratitude is expressed to the following people for their various contributions to this research effort: Ms Quek Yi Ling, Ms Chen Wei, Ms Tan Kheng Ling, Ms Xie Bingbing, Ms Ng Wei Ling, Ms Wendy Wen Yi Leong, Ms Amylia Bte Abdul Ghani and Mr Ni Runyan from the National University of Singapore (NUS) for their contribution in various experiments Dr Wang Shuhua, Dr Yao Wei, Dr Feng Shengbao and Ms Koh Lee Wah from NUS for their technique support Mdm Lee Chooi Lan, Ms Lew Huey Lee, Ms Jiang Xiaohui and Mr Abdul Rahman bin Mohd Noor for their generous assistance in the laboratories Mdm Wong Lai Kwai, Mdm Lai Hui Ngee, Mdm Han Yan Hui and Miss Tan Geok Kheng from the Chemical, Molecular and Materials Analysis Centre (CMMAC), NUS for their i assistance in mass spectrometry, nuclear magnetic resonance spectroscopy, and Single crystal X-Ray diffraction analysis Last, but certainly not the least, I would like to thank my family I want to express my gratitude to my dearest wife, for her unceasing love and continuous support I also want to thank my parents for their love and support all the way From the bottom of my heart, I thank all my friends whose name may not be mentioned one by one here, but had never hesitate to lend me their helping hands whenever I am in need Fu Caili August 2010 ii Table of Contents page Abstract viii List of Figures x List of Tables xiv List of Schemes xv List of Abbreviations .xvi Chapter Literature Review 1.1 Structures, sources and bioavailability of proanthocyanidins 1.1.1 Structure of proanthocyanidins 1.1.2 Analytical methods of proanthocyanidins 1.1.3 Sources and contents of proanthocyanidins .11 1.1.4 Absorption and bioavailability of proanthocyanidins 12 1.2 Bioactivities of proanthocyanidins 14 1.2.1 Antioxidant activities 14 1.2.2 Antibacterial activities 16 1.2.3 Prevention of cardiovascular diseases .19 1.2.4 Anticancer and antiinflammatory activities .20 1.2.5 Other bioactivities 21 1.3 Epicatechin derivatives from proanthocyanidins and their bioactivities 22 1.4 Stereochemistry of epicatechin derivatives 24 1.5 The aim of this research 26 iii Chapter Isolation, Purification and Characterization of Proanthocyanidins from Mangosteen Pericarps 2.1 Introduction .28 2.2 Results and discussion .28 2.2.1 Isolation of proanthocyanidins from mangosteen pericarps 28 2.2.2 Structural characterization .30 2.2.3 Thiolysis 35 Chapter Comparison among Different Proanthocyanidins 3.1 Introduction .38 3.2 Results and discussion .38 3.2.1 Extraction and NMR analysis of cocoa proanthocyanidins .38 3.2.2 Synthesis of (2R,3R,4S)-2-(3,4-dihydroxyphenyl)-4-(2-hydroxy-4,6dimethoxyphenyl)-3,4-dihydro-2H-chromene-3,5,7-triol, .40 3.2.3 Optimization of the depolymerizing condition for carbon nucleophile, 3,5dimethoxyphenol 41 3.2.4 Depolymerization of MPPs by methyl thioglycolate .42 3.2.5 Comparison among proanthocyanidins from four sources 44 Chapter Synthesis of Epicatechin Derivatives from Mangosteen Pericarp Proanthocyanidins 4.1 Introduction .48 4.2 Results and discussion .48 4.2.1 Regioselective protecting of catechins 49 4.2.2 Synthesis of epicatechin derivatives from MPPs .51 Chapter Isolation, Identification and modification of Proanthocyanidins from Rhizomes of Selliguea feei iv 5.1 Introduction 63 5.2 Results and discussion .64 5.2.1 Isolation and characterization of proanthocyanidins from the rhizomes of Selliguea feei .65 5.2.2 Synthesis of four epiafzelechin derivatives .71 Chapter Applications of Sulfurcontaining Epicatechin Derivatives for Proanthocyanidins Synthesis 6.1 Introduction .76 6.2 Results and discussion .77 6.2.1 Kinetics of the dethiolation of three sulfur-containing epicatechin derivatives 77 6.2.2 Synthesis of procyanidin B2 via base-catalyzed condensation .79 6.2.3 Synthesis of epicatechin alkaloids 81 Chapter Conclusions and Suggestions for Future Work 7.1 Conclusions .85 7.2 Suggestions for future work 87 Chapter Experimental procedures 8.1 Instrument and reagents 88 8.2 Characterization of mangosteen pericarp proanthocyanidins 90 8.2.1 Isolation and identification of mangosteen pericarp proanthocyanidins 90 8.2.2 Thiolysis of mangosteen pericarp proanthocyanidins .91 8.3 Comparison among different proanthocyanidins 91 8.3.1 Extraction and characterization of cocoa proanthocyanidins 91 8.3.2 Depolymerization of MPPs with 3,5-dimethoxyphenol 92 v 8.3.3 Synthesis of 2-((2R,3S,4S)-2-(3,4-dihydroxyphenyl)-3,5,7trihydroxychroman-4-ylthio)acetate from MPPs 92 8.3.4 Gram-scale synthesis of from mangosteen pericarps directly 94 8.4 Screening regioselective protecting reagents of catechin derivatives .94 8.4.1 Reaction between (+)-catechin and phenylboronic acid 94 8.4.2 Reaction between (+)-catechin and p-methoxyphenylboronic acid 95 8.4.3 Reaction between epicatechin and methyl propiolate 95 8.4.4 Selective protection of catecholic group in with methyl propiolate .96 8.5 Synthesis of epicatechin derivatives 97 8.5.1 General procedure for the acid mediated depolymerization of proanthocyanidins in the presence of carbon and sulfur nucleophiles .97 8.5.2 General procedure for selective protection of the ortho-dihydroxyl groups in and 103 8.5.3 General procedure the preparation of Schiff base of compounds 7, 8, and 13 105 8.6 Characterization proanthocyanidins from the rhizomes of Selliguea feei and synthesis of epiafzelechin derivatives 110 8.6.1 Extraction and characterization of proanthocyanidins from the rhizomes of Selliguea feei 110 8.6.2 General procedure for the acid depolymerization of proanthocyanidins from rhizomes of Selliguea feei 113 8.6.3 Synthesis of epiafzelechin Schiff base, 26 .116 8.7 Applications of proanthocyanidins and the derivatives 117 8.7.1 Kinetics of the dethiolation of three sulfur-containing epicatechin derivatives 117 vi 8.7.2 Reaction of with epicatechin and carbon nucleophiles 117 Reference 118 List of Publications and Patent .130 Appendix 131 vii Abstract Proanthocyanidins (PAs), one of the most ubiquitous groups of plant polyphenols, are the natural polymer widespread throughout the plant kingdom but their application as raw materials for fine chemicals are largely unexplored This thesis documented the results on the effort towards chemical conversion of PAs into epicatechin and epiafzelechin derivatives with potential application such as multidentate chiral ligands Mangosteen pericarps proanthocyanidins (MPPs) were extracted with 0.66% yield (dry matter) from mangosteen peels, agricultural waste in South East Asian countries 13C{1H} and 1H NMR signals showed the presence of predominantly procyanidins together with a few prodelphinidin units along with small amounts of stereoisomers of afzelechin/epiafzelechin and gallocatechin/ epigallocatechin Depolymerization of MPPs with benzylmercaptan indicated that the mean degree of polymerization (mDP) is 6.6 The electrospray ionization–mass spectrometry (ESIMS) and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectra revealed the dominant B type oligomers with mainly epicatechin units and with a small amount of A-type oligomers MPPs and CPs extracted in our lab were compared with commercial pine bark proanthocyanidins (PBPs) and grape seed proanthocyanidins (GSPs) via MALDITOF-MS and thiolysis analysis Both thiolysis and MALDI-TOF-MS results indicated that MPPs were ideal for synthesis of the new chiral ligands due to the higher mDP and ratio of epicatechin units MPPs were depolymerized with selective carbon and sulfur nucleophiles The products were purified by column chromatography and characterized with ESI- viii Appendix H NMR spectrum of 16 16 174 Appendix 13 C NMR spectrum of 16 16 175 Appendix 176 Appendix 177 Appendix 178 Appendix 179 Appendix H NMR spectrum of 21 21 180 Appendix H NMR spectrum of 21 181 Appendix H NMR spectrum of 22 22 182 220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30.5000 39.8240 36.9293 52.3481 70.7416 76.0727 97.1863 96.5244 103.1647 116.2125 131.9440 129.8639 158.6651 158.5851 158.3596 157.6832 13 173.8221 Appendix C NMR spectrum of 22 de22fcl-345 22 (ppm) 30 20 10 183 7.8 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 1.0539 3.4293 1.0013 0.9392 1.4983 0.9860 1.0000 0.9539 0.9262 0.8510 4.4401 2.1318 2.0443 3.6 3.4 3.2 3.0 2.0862 2.0500 2.9446 3.5845 3.5297 3.7440 4.3702 4.3586 4.2891 4.1861 4.1735 4.1422 4.1368 5.3579 5.6171 6.1720 6.0904 6.0827 5.9896 5.9847 5.9819 6.9440 6.9149 6.8859 6.8563 7.6983 7.6699 7.5712 7.5422 Integral Appendix H NMR spectrum of 24 1H normal range AC300 24 (ppm) 2.8 2.6 2.4 2.2 2.0 1.8 1.6 1.4 1.2 184 220 210 200 190 180 170 160 150 140 130 120 110 100 90 80 70 60 50 40 30.5000 34.8638 45.1188 53.6864 61.2139 68.0215 71.1271 78.0801 104.2556 102.0010 100.7137 98.7500 98.1609 96.9317 107.6812 116.5325 115.9361 132.1331 131.3476 130.2203 130.0458 159.1815 158.6505 157.8941 157.6323 154.5631 154.3813 152.2503 13 173.3857 Appendix C NMR spectrum of 24 de22fcl-647 24 (ppm) 30 20 10 185 7.8 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 1.1930 1.0375 1.0526 0.9542 1.0000 1.0429 0.9469 0.9454 0.9261 1.0587 1.2077 2.1772 2.1421 1.0370 1.0352 2.0922 2.0251 3.3498 3.3145 3.3107 3.3082 4.1075 4.1049 4.1012 3.9612 4.2789 4.4743 4.4668 4.8261 5.7250 6.0855 6.0818 6.0364 6.0339 6.0288 6.1725 6.6062 6.5924 6.5772 7.3576 7.3425 7.1269 7.1118 7.0967 6.9063 6.9038 6.8899 6.8521 6.8344 6.8079 6.7916 7.6312 7.6148 7.5291 7.5266 7.5114 Integral Appendix H NMR spectrum of nv29fcl-fatp15 25 (ppm) 3.8 3.6 3.4 3.2 186 160 155 150 145 140 135 130 125 120 115 110 105 160.0 100 95 90 159.0 85 80 157.2506 157.7680 158.2126 158.4530 158.9194 158.0 75 70 157.0 65 60 55 50 45 40 35 29.4006 49.8563 49.6377 49.5211 49.4628 49.3462 49.2952 49.1786 49.0110 48.8361 48.6685 48.5009 45.6369 68.0529 70.4796 77.2205 101.6332 100.3215 98.3102 96.9912 96.6268 103.9142 107.5360 118.2922 116.7254 116.2663 116.1206 115.5667 131.7739 130.5205 130.1051 129.4346 138.3981 13 158.4530 158.2126 157.7680 157.2506 154.1899 154.0806 152.6741 151.8725 Appendix C NMR spectrum of 25 nv29fcl-fatp15 (ppm) 156.0 25 (ppm) 30 25 187 Appendix H NMR spectrum of 26 26 188 ... protecting of catechins 49 4.2.2 Synthesis of epicatechin derivatives from MPPs .51 Chapter Isolation, Identification and modification of Proanthocyanidins from Rhizomes of Selliguea... 8.6.3 Synthesis of epiafzelechin Schiff base, 26 .116 8.7 Applications of proanthocyanidins and the derivatives 117 8.7.1 Kinetics of the dethiolation of three sulfur-containing epicatechin. . .SYNTHESIS AND APPLICATIONS OF EPICATECHIN AND EPIAFZELECHIN DERIVATIVES FROM PROANTHOCYANIDINS FU CAILI B Eng China Agricultural University