35 1 2 0 1 7 1 DOI 10. 13193 /j. issn. 1673-7717. 2017. 01. 035 1 1 1 1 2 1. 150036 2. 150040 D101 DEAE Cellulose 52 Sephacryl S - 300 - - PJP - 1a PJP - 3a 1. 34 10 3 Da 1. 70 10 7 Da - PJP - 3a 4. 56 7. 53 1 2. 48 41. 4 17. 94 PJP - 1a PJP - 3a β PJP - 1a R284. 2 A 1673-7717 2017 01-0119-05 Extract Isolation Purification and Identification of Polysaccharides from Exocarp of Unripe Fruits of Juglans mandshurica YANG Hongxia 1 REN Xiaolei 1 HUO Jinhai 1 WANG Weiming 1 2 1. Insititute of Chinese Materia Medica Heilongjiang Academy of Traditional Chinese Medicine Harbin 150036 Heilongjiang China 2. Center of Forect Resources Research and Development of Heilongjiang Harbin 150040 Heilongjiang China Abstract The purpose of this study was to extract isolate and purify polysaccharide from exocarp of unripe fruits of Juglans mandshurica analyze the monosaccharide composition and preliminarily analyze its structure of the obtained homogeneous molecular weight polysaccharide. A purified polysaccharide was obtained by hot water extractionethanol 2016-09 - 16 2009ZX09102-138 2013RFQYJ056 1989-1964 - E-mail zyyjy@ 163. com 10.. M. 1991 19. M. 856-859. 2012 3. 11. M. 1992 41 20. M.. 125. 12. M. 440 482. 2008 13. M. 22. M. 1984 300. 1958 1049. 14. M. 1962 2007 296. 16. M. 1997 233. 2012 112-113. 2011 107-108. 21 2012 339.. M. 23. M. 1991 354. 1460. 24. M. 1983 15. M. 228. 25. M. 26. M. 1988 42. 17. M. 27. M. 1955 2011 88. 63. 18. M. 28. J. 2011 177. 2013 19 1 3. 119
35 1 2 0 1 7 1 precipitation deproteinization and decolor. The purified polysaccharide then was isolated with DEAE Cellulose 52 column chromatography and Sephacryl S - 300 column chromatography. The purity identification and molecular weight determination were adopted by HPGPC. The monosaccharide composition and structure were analyzed by GC - MS and IR. The result showed that both the two obtained polysaccharides were homogeneous polysaccharide. The molecular weights of PJP - 1a and PJP - 3a were 1. 34 103 Da and 1. 70 107 Da respectively. PJP - 3a was mainly composed of rhamnose arabinose xylose mannose glucose and galactose at a molar ratio of 4. 56 7. 53 1 2. 48 41. 4 17. 94. IR suggested that both PJP - 1a and PJP - 3a had pyran ring structure and glycosidic bonds of PJP - 1a and PJP - 3a were both β type and PJP - 1a had acetamido structure. Key words Juglans mandshurica polysaccharide isolate purify analysis 120 Jusland emandehurica Maxim. 1 land emandehurica Maxim. 2 > 200 U mg - 1 Jus- 3 D101 DE- AE Cellulose 52 Whatman 4 5 Sephacryl S - 300 Pharmacia 00892-100MG 00270-100MG 00271-100MG 00891 P - 012-110821 D - B - 079-140425 D - G - 042-130612 D - M - 066-110701 L - A - 012-6 - 7 8-12 13-14 140111 L - S - 037-1401267 > 15 98% 1. 2 3 kg 14 1 h 14 1 h 4 95 % 4 4000 r /min 15 min 16-17 18 1. 3 1. 3. 1 Viskase - 100MG 00983-100MG SIGMA - ALDRICH D - 0. 01 g ml - 1 19-24 ph 6 3% 60 2 h 5 min 25-1. 3. 2 D101 1 D101 1 h 2 BV h - 1 1. 1 UV - 160-1. 4 KD - 2KY 1. 4. 1 DEAE Cellulose 52 RE - 52AA JM1812 30 ml 3 ml DEAE Cellulose 52-1 0 0. 1 0. 2 0. 3 0. 5 mol L - 1 NaCl LC - 10AS 2. 5 ml min - 1 5 ml / - Alltech ELSD 2000ES 620 nm PL aquagel - OH MIXED - H 8 μm GPC Spectrum Two 3500 Da 12 h Perkin Elmer Clarus 680 - SQ8T - 12 h PJP - 1 PJP - Perkin Elmer 3500 Da 2 PJP - 3 PJP - 4 PJP - 5 1. 4. 2 Sephacryl S - 300 2014 8 35 mg 2. 5 ml Sephacryl S - 300
35 1 2 0 1 7 1 0. 3 ml min - 1 min 25 1 42 3 ml / - 620 nm 1. 2 ml min - 1 2 EI 70 ev 250 250 m /z 43 ~ 500 2. 5 scan s - 1 PJP - 1a PJP - 3a 1. 7 1. 5 1 mg 200 mg KBr 1. 5. 1-400 ~ 4000 cm - 1 HPLC - GPC 2 mg 1 ml 2 mg 2 ml - 1 0. 45 μm 2. 1 DEAE Cellulose 52 LC - 10AS HPLC aquagel - OH MIXED - H 8 μm GPC Alltech ELSD 2000ES 5 PJP - 1 2 mg ml - 1 10 μl 0. 3 0. 1 mol L - 1 NaCl PJP - 2 0. 2 mol L - 1 ml min - 1 115 1. 8 L min - 1 NaCl PJP - 3 0. 3 mol L - 1 NaCl PJP - 4 1 ml 2 mg ml - 1 0. 45 μm 10 μl 2 mg ml - 1 T - PJP - 1 PJP - 3 logmw 2. 2 Sephacryl S - 300 1. 5. 1 PJP - 1 PJP - 3 Sephacryl S - 300 1. 6 PJP - 1a PJP - 3a 1. 6. 1 10 mg 4. 0 ml 2. 0 mol L - 1 TFA 110 2. 3 PJP - 1a PJP - 3a 3 h 4000 r /min 10 min 2. 3. 1 PJP - 1a PJP - 3a TFA 110 - HPLC - GPC PJP - 1a PJP - 3a 15 min 2 PJP - 1a PJP - 1. 6. 2 2. 0 ml 3a 60 mg 3 h 2. 3. 2 PJP - 1a PJP - 3a 110 y = 0. 256 3x + 14. 293 R 2 = 0. 996 15 min 1. 6. 3 3. 0 ml 1. 0 ml 100 2 h 1. 34 10 3 Da 1. 70 10 7 Da 5. 0 ml 3 2. 4 PJP - 3a 0. 22 μm GC - MS PJP - 3a L - 22. 90 min L - 23. 21 min D - 1. 6. 4 GC - MS 1 DB - 5 30 m 0. 25 mm 0. 25 μm min D - 28. 24 min 6 80 1 min5 min - 1 200 2 min - 1 220 10 min - 1 270 1 DEAE Cellulose 52 1. 5. 2 2 mg 0. 5 mol L - 1 NaCl PJP - 5 1 PJP - 1 PJP - 3 PJP - 1a PJP - 3a HPLC - GPC 31. 340 min 19. 823 min 23. 51 min D - 27. 82 min D - 27. 93 4. 56 7. 53 1 2. 48 41. 4 17. 94 PJP - 3a GC - MS 3 ~ 4 121 1 DEAE Cellulose 52
35 1 2 0 1 7 1 2 PJP - 1a PJP - 3a HPLC - GPC 3 GC - MS 122 2. 5 4 PJP - 3a GC - MS PJP - 1a PJP - 3a 5 PJP 3-1a PJP - 3a 3400 cm - 1 Sephadex G100 Sephacryl S - 300 2935 cm - 1 - OH C - H PJP - 1a 1615 cm - 1 - NH 2 COCH 3 C = O 27 PJP - 1a PJP - 3a - 1600 cm - 1 PJP - 1a 2 mg ml - 1 10 μl 115 PJP - 3a 0. 2 mol L - 1 NaCl 1. 8 L min - 1 0. 3 ml min - 1 C = O 1600 cm - 1 PJP - 1a 892. 61 cm - 1 β - 764. 85 cm - 1 28 PJP GC - MS IR - 3a 1 099. 52 cm - 1 C - O - C 29 PJP - 3a 891. 30 cm - 1 β - 4. 56 7. 53 1 PJP - 1a PJP - 3a β - 2. 48 41. 4 17. 94 PJP - 1a PJP - 3a
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