綠升麻中菠蘿蜜烷型三萜類成分研究

1、<p>　　綠升麻中菠蘿蜜烷型三萜類成分研究</p><p>　　作者：盧濱 范云雙 萬定榮 段宏泉</p><p>　　【摘要】 　　目的研究綠升麻中菠蘿蜜烷型三萜皂苷類化學(xué)成分。方法以硅膠柱層析、凝膠層析等多種柱層析分離，制備高效液相色譜純化，得到單體化合物，以各種有機(jī)波譜鑒定化合物結(jié)構(gòu)。結(jié)果分離得到15個(gè)菠蘿蜜烷型三萜類皂苷化合物，鑒定為asiaticoside A (1

2、)；asiaticoside B (2)；26-deoxycimicifugoside(3)；cimicifugoside(23S,4)；25-甲氧基升麻醇-3-O-b-D-木糖苷(25- O-methyl-cimigenol-3-O- b-D-xylopyranoside,5)；25, 2′-二乙酰升麻醇-3-O-β-D-木糖苷(25,2′-O-diacetylcimigenol-3-O-β-D-xylopyranoside,6)；2

3、4-乙酰升麻醇-3-O-β-D-木糖苷(24-O-acetylshengmanol-3-O-β-D-xylopyranoside(23R,24R,7)；24-O-acetyldahurinol-3-O-β-D-xylopyranoside(8)；23-乙酰升麻醇-3-O-β-D-木糖苷(23-O-a</p><p>　　【關(guān)鍵詞】 綠升麻； 醋酸乙酯提取物； 菠蘿蜜烷型三萜皂苷</p><p

4、>　　Abstract：ObjectiveTo isolate and elucidate the constituents of Actaea asiatiea. MethodsChemical constituents were isolated by repeated column chromatography (Toyopearl HW-40C and preparative HPLC) and their structu

5、res were elucidated on the basis of spectral data analysis.ResultsFifteen cycloartane triterpenes were isolated from Actaea asiatiea，asiaticoside A, asiaticoside B, 26-deoxycimicifugoside,Cimicifugoside(23S),25-O-methyl-

6、cimigenol-3-O- β-D-xylopyranoside,25,2′-O-diacetylcimigenol-3-O-β-</p><p>　　Key words：Actaea asiatiea; Ethyl acetated extraction; Cycloartane triterpenes</p><p>　　綠升麻為湖北省土家族傳統(tǒng)藥物，應(yīng)用歷史悠久，為毛茛科植

7、物類葉升麻Actaea asiatiea Hara的根莖，主要分布在我國東北、內(nèi)蒙古、河北、山西、陜西、甘肅、青海、湖北、四川、云南以及西藏東部。綠升麻味辛、微苦，性平，具有散風(fēng)熱、祛風(fēng)濕、透疹、解毒的作用，在湖北省鄂西地區(qū)土家族民間用于治療風(fēng)熱頭痛，咽喉腫痛、風(fēng)濕疼痛、風(fēng)疹塊、麻疹不透、百日咳、子宮脫垂、犬咬傷等［1，2］。本文運(yùn)用各種色譜分析手段，結(jié)合制備高效液相色譜，從綠升麻的醋酸乙酯提取物中分離得到15個(gè)菠蘿蜜烷型三萜皂苷類化合

8、物，通過各種有機(jī)波譜解析鑒定了各單體化合物的結(jié)構(gòu)。</p><p><b>　　1 器材</b></p><p>　　核磁共振儀: Bruker AVANCE 300 instrument（TMS內(nèi)標(biāo)）; 制備高效液相色譜儀：日本分光公司（JASCO）, PU-1580 (泵), RI-1530 和 UV-1575（檢測器）；制備色譜柱：YMC-Pack ODS-A

9、 SH-343-5 20X250mm（YMC），Econosphere silica 10μm 22 X250mm (Alltech); 液質(zhì)聯(lián)用色譜儀：Alliance 2695, Quattro Micro TM ESI (Waters); Toyopearl HW-40C（Tosoh, 凝膠滲透層析）；柱色譜和薄層色譜用硅膠均系青島海洋化工廠生產(chǎn)，所用試劑均為分析純。</p><p>　　綠升麻Actae

10、a asiatiea Hara 200409采自湖北省鶴峰縣，由中南民族大學(xué)生命科學(xué)院萬定榮教授鑒定，標(biāo)本（D20040901）存放于天津醫(yī)科大學(xué)藥學(xué)院。</p><p><b>　　2 提取分離</b></p><p>　　綠升麻2.6 kg，粉碎后用95%的乙醇加熱回流提取3次，6 h/次。提取液減壓濃縮至浸膏600 g，浸膏加水混懸后，分別用石油醚、醋酸乙酯

11、、正丁醇進(jìn)行萃取，得石油醚提取物41 g，醋酸乙酯提取物210 g，正丁醇提取物31g。醋酸乙酯提取物經(jīng)硅膠柱層析分離，大孔吸附樹脂分離，制備高效液相色譜純化得到化合物1～5；經(jīng)凝膠滲透色譜，制備高效液相色譜純化得到6～15。</p><p><b>　　3 結(jié)構(gòu)鑒定</b></p><p>　　化合物1：白色無定型粉末。ESI-MS m/z: 667 ［M+Na］

12、+ (分子式 C37H56O9)。1H-NMR ( 300 MHz, Pyr ) δ: 0.21, 0.62(each 1H, d, J= 4.0 Hz, 19-H), 0.91, 1.04, 1.14， 1.17， 1.19， 1.24, 1.30 (each 3H , s，7×CH3), 2.15(3H, s, COCH3 ), 4.82 (2H, d, J=2.8Hz), 6.25(2H, d, J=7.3Hz, 1′-

13、H)。13C-NMR數(shù)據(jù)見表1。化合物1經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［3］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為asiaticoside A。</p><p>　　化合物2：白色無定型粉末。ESI-MS m/z: 665 ［M+Na］+(分子式C37H54O9 )。1H-NMR ( 300 MHz, Pyr ) δ: 0.50, 0.95 (each 1H, d, J =3.6H

14、z,19-H)， 1.04, 1.10, 1.10, 1.14, 1.16, 1.27, 1.36, ( 3H, s,7×CH3), 2.20（3H，s, COCH3）, 4.86(1H, d, J =7.1Hz,11′-H),5.19(1H, d, J =6.6Hz,7-H), 5.41(1H，d,J =9.0Hz，12-H)，6.25（1H, s, 22-H）。13C-NMR數(shù)據(jù)見表1?；衔?經(jīng)酸水解, TLC Rf值對

15、照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［3］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為asiaticoside B。</p><p>　　化合物3：無色針晶。ESI-MS m/z: 658 ［M+Na］+( 分子式 C37H54O10)。1H-NMR (300 MHz, Pyr ) δ: 0.52，1.06 (each 1H, d, J =4.0Hz , 19-H)，1.02( 3H, d, J = 6.0 Hz, 21-

16、H), 1.05(3H, s )，1.17(3H, s), 1.47(3H, s), 1.49(3H, s),1.32(3H,s), 3.74(1H,s,24-H) 。13C-NMR數(shù)據(jù)見表1?；衔?經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［4］報(bào)道的數(shù)據(jù)基本一致鑒定化合物為26-deoxycimicifugoside。</p><p>　　化合物4：白色無定型粉末。ESI-MS m/

17、z: 659 ［M+H］+( 分子式 C37H54O10)。1H-NMR ( 300 MHz , Pyr ) δ: 0.57, 1.09(each 1H, d, J =3.6Hz, 19-H)，0.95( 3H, d, J =5.6 Hz, 21-H), 1.02, 1.02, 1.33, 1.42, 1.80 (each 3H, s, 5×CH3), 2.20 (3H, s, COCH3), 2.95(1H, dd, J =

18、16.0, 9.0Hz, 11H), 3.45(1H, dd, J =11.5, 3.8Hz, 3-H)，3.94 (1H，s，24－H)，4.72 (1H, q－like, J = 7.2Hz, 16-H)，4.85 (1H, d, J = 8.5 Hz, 11′-H)，5.07(1H, d, J =6.4 Hz, 7-H)，5.23 (1H, d, J =8.7 Hz, 12-H), 5.76 (1H，s, 27-H)。13C-NM

19、R數(shù)據(jù)見表1?；衔?經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文</p><p>　　化合物5：白色無定型粉末。ESI-MS m/z:637［M+H］+ (分子式C36H58O9)。1H-NMR ( Pyr ) δ: 0.30, 0.56(each 1H, d, J = 3.8Hz, 19-H)； 0.85 (3H, d, J = 6.2 Hz, 21-H)； 1.08, 1.16, 1.2

20、0, 1.27, 1.27, 1.34 (each 3H, s, 6×CH3), 3.21 (3H, s, COCH3), 3.53 ( 1H, dd, J = 12.0, 4.0 Hz, 3-H), 3.66 (1H, br s, 24-H), 4.26 (1H, s, 15-H), 4.63 (1H, br d, J =9.0 Hz, 23-H), 4.88 (1H, d, J = 7.3 Hz, 1′-H)。13C-NM

21、R數(shù)據(jù)見表1?；衔?經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［6］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為25-甲氧基升麻醇-3-O-β-D-木糖苷(25- O-methyl-cimigenol-3-O- β-D-xylopyranoside</p><p>　　化合物 6：白色無定型粉末。ESI-MS m/z:705［M+H］+ (分子式C39H60O11)。1H-NMR ( Pyr ) δ

22、: 0.26, 0.50(each 1H, d, J = 3.8Hz, 19-H), 0.87 (3H, d, J = 6.3 Hz, 21-H), 0.99, 1.11, 1.16, 1.20, 1.68, 1.70 (each 3H, s，6×CH3), 1.98, 2.17 (each 3H, s, 2×Ac), 3.40 (1H, dd, J= 12.0, 4.0 Hz, 3-H), 4.13 (1H, br

23、 s, 24-H), 4.30 (1H, s, 15-H), 4.61 (1H, br d, J=9.0 Hz, 23-H), 4.83 (1H, d, J= 8.0 Hz, 1′-H), 5.56 (1H, t, J= 8.0 Hz, 3′-H)。13C-NMR數(shù)據(jù)見表1?；衔?經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［7］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為25,2′-二乙酰升麻醇-3-O-β-D-木糖苷(2

24、5,2′</p><p>　　化合物7：白色無定型粉末。ESI-MS m/z: 681 ［M+H］+(分子式C37H60O11)。1H-NMR ( Pyr ) δ: 0.29, 0.58 (each 1H, d, J= 3.6 Hz, 19-H), 0.97 (3H, d, J= 5.9 Hz, 21-H), 1.07, 1.20, 1.23, 1.33, 1.48, 1.52 (each 3H, s，6

25、5;CH3), 2.13 (3H, s, Ac), 3.50(1H, dd, J= 11.5, 4.0, 3-H), 4.17 (1H, s, 15-H), 4.90 (1H, d,J=7.4 Hz, 1′-H), 5.78 (1H, d, J= 8.0 Hz, 24-H)。13C-NMR數(shù)據(jù)見表1?；衔?經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［4］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為24-乙酰升麻醇-3-O-

26、β-D-木糖苷(24-O-acetylshengmanol-3-O-β-D-xylopyranoside (23R,24R) )。</p><p>　　化合物8：白色無定型粉末。ESI-MS m/z: 664 ［M+H］+(分子式C37H58O10)。1H-NMR ( Pyr ) δ: 0.28, 0.51 (each 1H, d, J= 3.6 Hz, 19-H), 0.95 (3H, d, J= 6.0 Hz

27、, 21-H), 1.05, 1.09, 1.19, 1.33, 1.51, 1.51 (each 3H, s，6×CH3), 2.12 (3H, s, Ac), 3.53(dd, J = 11.0, 3.6 Hz, 3-H), 3.85(1H, d, J= 12.5 Hz, 16-H), 4.22(m), 4.38 (1H, dd, J= 11.3, 5.0 Hz, 5′-H), 4.88 (1H, d, J = 7.4 H

28、z, 1′-H), 5.42 (1H, d, J = 6.0 Hz, 24-H)。13C-NMR數(shù)據(jù)見表2?；衔?經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［8］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為24-O-acetyldahurinol-3-O-β-D-xylopyranoside</p><p>　　化合物9：白色無定型粉末。ESI-MS m/z: 663 ［M+H］+ (分子式C37H

29、58O10)。1H-NMR (CDCl3.) δ: 0.45, 0.66 (each 1H, d, J = 3.6 Hz, 19-H), 1.09 (3H, d, J= 6.4 Hz, 21-H), 0.86, 0.86, 0.98, 1.29, 1.33, 1.37 ( each 3H, s, 6×CH3), 2.09（3H, s, Ac）, 2.80（1H, d, J= 8.4 Hz, 24－H）, 3.24 (1H, d

30、d, J= 11.0, 4.0 Hz, 3-H), 3.99 (1H, s, 15-H), 4.95 (1H, br t, J= 8.4 Hz, 23-H), 4.51 (1H, d, J = 7.4 Hz,11′-H)。13C-NMR數(shù)據(jù)見表2?；衔?經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［9］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為23-乙酰升麻醇-3-O-β-D-木糖苷(23-O-acetylshengman

31、ol-3-O-β-D-xylopyrano</p><p>　　化合物10：白色無定型粉末。1H-NMR ( Pyr.) δ： 0.33, 0.59 (each 1H, d, J= 3.6 Hz, 19-H), 1.28 (3H, d, J= 6.4 Hz, 21-H), 1.08, 1.22, 1.27, 1.34, 1.39, 1.41 ( each 3H, s, 6×CH3), 1.99, 2.0

32、7（3H, s, 2×Ac）, 3.05（1H, d, J = 8.4 Hz, 24－H）, 3.50 (1H, dd, J= 11.5, 4.2 Hz, 3-H), 4.38 (1H, s, 15-H), 5.40 (1H, br t, J= 8.4 Hz, 23-H), 4.89 (1H, d, J= 7.5 Hz, 11′-H), 5.42(1H, m, 4′-H )。13C-NMR數(shù)據(jù)見表2。化合物10經(jīng)酸水解, TL

33、C Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［10］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為Cimiracemoside M。</p><p>　　化合物11：白色無定型粉末。ESI-MS m/z: 579［M+H］+(分子式 C33H48O9 )。1H-NMR ( Pyr ) δ: 0.50, 1.02 (each 1H, d, J= 3.6 Hz, 19-H), 0.98(3H, d, J= 6.3 Hz, 2

34、1-H), 1.03, 1.06, 1.28 , 1.34 ( each 3H, s，4×CH3), 2.14 (3H, s, Ac), 2.90 (1H, dd, J= 16.0, 9.0 Hz, 11-H), 3.44 (1H, dd, J= 11.6, 3.8 Hz, 3-H), 4.85 (1H, d, J= 7.5 Hz, 1′-H), 4.90 (1H, d, J= 7.5 Hz,16-H), 5.12 (1H,

35、d, J=8.7 Hz, 7-H), 5.14 (1H, d, J= 8.7 Hz, 12-H)。13C-NMR數(shù)據(jù)見表2?；衔?1經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［11］報(bào)道的數(shù)據(jù)一致，鑒定化合物為12β-acetoxy-3β-hydroxy-24,25,26,</p><p>　　化合物12：白色無定型粉末。ESI-MS m/z: 591 ［M+H］+ (分子式 C33H5

36、0O9 )。1H-NMR ( Pyr ) δ: 0.20, 0.56 (each 1H, d, J= 3.9 Hz, 19-H), 0.98(3H, d, J= 6.2 Hz, 21-H), 0.85, 1.02, 1.24 , 1.32 ( each 3H, s, 4×CH3), 2.14 (3H, s, Ac), 2.70(1H, dd, J= 16.0, 9.0 Hz, 11-H), 3.47 (1H, dd, J= 1

37、1.6, 3.8 Hz, 3-H), 4.84 (1H, m, 16-H), 4.86 (1H, d, J= 7.5 Hz, 1′-H), 5.07 (1H, dd, J= 3.1, 8.7 Hz, 12-H)。13C-NMR數(shù)據(jù)見表2?；衔?2經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［11］報(bào)道的數(shù)據(jù)基本一致，鑒定化合12b-acetoxy-3β-hydroxy-24,25,26,27-tetranor-c

38、ycloartan-23,16β-o</p><p>　　化合物13：白色無定型粉末。ESI-MS m/z: 679 ［M+H］+(分子式 C37H58O11)。1H-NMR ( Pyr ) δ: 0.23, 0.56 (each 1H, d, J = 3.6 Hz, 19-H), 1.22 (3H, d, J= 6.2 Hz, 21-H), 0.87, 1.02, 1.28, 1.33, 1.71, 1.78(

39、 each 3H, s, 6×CH3), 2.09(3H, s, Ac), 2.73（1H, dd, J= 16.0, 9.0 Hz, 11-H）, 3.48(1H, dd, J= 11.2, 4.0 Hz, 3-H), 3.90(1H, d, J= 10.5Hz, 22-H), 4.23(1H, s, 24-H), 4.87 (1H, d, J= 7.5 Hz, 1′-H), 5.00(1H, q-like, J= 8.1H

40、z, 16-H), 5.16 (1H, dd, J= 3.6, 9.0 Hz, 12-H)。13C-NMR數(shù)據(jù)見表2。化合物13經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［12］報(bào)道的數(shù)據(jù)</p><p>　　化合物14：白色無定型粉末。ESI-MS m/z: 663 ［M+H］+(分子式 C37H58O10)。1H-NMR ( Pyr ) δ: 0.31, 0.54(d, J= 3.6

41、Hz, 19-H), 0.99(d, J= 5.7Hz, 21H), 1.08, 1.14, 1.20, 1.27, 1.40, 1.78(3H, s, 6×CH3), 2.10(3H, s, Ac), 3.52(1H,dd, J = 11.7, 4.2Hz, 3-H), 4.84(d, J= 7.4Hz, 1′-H), 5.19(1H, d, J= 7.6Hz, 24-H)。13C-NMR數(shù)據(jù)見表2?；衔?4經(jīng)酸水解, T

42、LC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［13］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為Neocimicigenosides B。</p><p>　　化合物15：白色無定型粉末。ESI-MS m/z: 667 ［M+Na］+ (分子式 C37H56O9 )。1H-NMR ( Pyr ) δ: 0.37, 0.62(d, 3.6 Hz, 19-H), 0.89(d, J = 6.4Hz, 21-H), 0.8

43、0, 0.94, 0.94, 1.09(3H, s, 5×CH3), 2.13(3H, s, Ac), 4.24(d, J= 8.4Hz, 23H), 4.86, 5.00(each 1H, s,26-H), 4.58(d, J= 5.7Hz, 1′-H)。13C-NMR數(shù)據(jù)見表2?；衔?5經(jīng)酸水解, TLC Rf值對照，鑒定為D－木糖。以上光譜數(shù)據(jù)與文獻(xiàn)［8］報(bào)道的數(shù)據(jù)基本一致，鑒定化合物為2′-O-乙酰基-25-脫水升麻

44、醇-3-O-β-D-木糖苷( 2′-O-acetyl-25-anhydrocimigenol-3-O-β-D-xylopyranoside)。</p><p>　　表1 化合物1～7的13C-NMR (Pyr)（略） </p><p>　　表2 化合物8～15的13C-NMR（略） </p><p><b>　　【參考文獻(xiàn)】</b><

45、/p><p>　?。?］ 萬定榮. 湖北省土家族常用植物藥（毛茛科）［J］.中藥材，1990，13（3）：13.</p><p>　?。?］ 《中華本草》編委會.中華本草［M］.上海： 上?？茖W(xué)技術(shù)出版社, 1999.</p><p>　　［3］ Jingchun Gao, Feng Huang, Jinchao Zhang, et al. Cytotoxic cycl

46、oartane triterpene saponins from Actaea asiatica［J］.J. Nat. Prod， 2006：1500.</p><p>　?。?］ Kusano A, Takahira M , Shibano M , et al. Studies on the constituents of cimicifuga species. XXVI..Twelve new cyclola

47、nostanol glycosides from the underground part of Cimicifuga simplex wormsk［J］.Chem. Pharm. Bull. , 1999, 47(4):511.</p><p>　?。?］ Kusano A, Takahira M , Shibano M , et al. Studies on the constituents of cimic

48、ifuga species. XX. Absolute stereostructures of Cimicifugoside and Actein from Cimicifu simplex wormsk［J］.Chem. Pharm. Bull. ,1998,46(3):467.</p><p>　?。?］ Erdal Bedir,Ikhlas A.Khan, Cimiracemoside A: a new c

49、yclolanostanol xyloside from the rhizome of Cimicifuga racemosa［J］.Chem. Pharm. Bull.,2000,48(3):425.</p><p>　　［7］ Liang Zhou, JunShan Yang, JianHua Zou, et al. Three new cycloartane triterpene glycosides fr

50、om Souliea vaginata［J］. Chem. Pharm. Bull., 2004,52(5):622.</p><p>　?。?］ Liang Zhou, JunShan Yang, GuangZhong Tu, et al. Cyclolanostane triterpene glycosides from Souliea vaginata［J］.Chem. Pharm. Bull.,2006,

51、54(6):823.</p><p>　　［9］ 潘瑞樂，陳迪華， 斯建勇，等.升麻地上部分皂苷類成分研究［J］.化學(xué)學(xué)報(bào), 2002,37(2):117.</p><p>　?。?0］ Shao-Nong Chen, Daniel S. Fabricant, et al. Cimiracemosides I-P, New 9,19-Cyclolanostane triterpene gl

52、ycosides from Cimicifuga racemosa［J］.J. Nat. Prod.,2002,65:1391.</p><p>　?。?1］ Makiko Nishida, Hitoshi Yoshimitsu, Toshihiro Nohara. Two new tetranor-cycloartane glycosides from Cimicifuga Rhizome［J］.Chem. P

53、harm. Bull.,2003,51(9):1117.</p><p>　?。?2］ Yu Shao,Amanda Harris, Mingfu Wang, et al.Triterpene glycosides from Cimicifuga racemosa［J］.J. Nat. Prod.,2000,63:905.</p><p>　?。?3］ Yoshihiro Mimaki,

54、Isao Nadaoka, Masaaki Yasue, et al.Neocimicigenosides A and B, cycloartane glycosides from the rhizomes of Cimicifuga racemosa and their effects on CRF-Stimulated ACTH secretion from AtT-20 cells［J］.J. Nat. Prod.,2006,69