三叶木通活性成分的TLC鉴别及含量测定
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摘要
目的:建立三叶木通药材中皂苷PH、皂苷PJ1的TLC鉴别和皂苷PH、akemisaponin E、皂苷PJ1和scheffoleoside A的HPLC含量测定方法。方法:TLC法,展开剂为三氯甲烷-甲醇-甲酸(20∶10∶1),显色剂为10%硫酸乙醇溶液;HPLC法,采用依利特C_(18)色谱柱(250 mm×4.6 mm,5μm),以乙腈-水(21∶79)为流动相,流速1.0 mL·min~(-1),检测波长200 nm,柱温30℃。结果:三叶木通药材中皂苷PH、皂苷PJ1的TLC分离效果均较好;HPLC法测得皂苷PH、akemisaponin E、皂苷PJ1和scheffoleoside A的进样量分别在0.201～5.025μg(r=0.999 9)、0.076～1.888μg(r=0.999 9)、0.224～5.608μg(r=1.000 0)和0.041～1.035μg(r=0.999 9)范围内与峰面积呈现良好的线性关系;平均回收率(n=3)分别为99.8%、98.2%、99.5%、98.9%,RSD分别为0.46%、1.2%、0.40%和1.0%;5批样品中皂苷PH、akemisaponin E、皂苷PJ1和scheffoleoside A的含量范围分别为2.094～3.928、0.956～1.418、2.958～4.472和0.526～0.797 mg·g~(-1)。结论:此方法简便易行,重复性好,可作为三叶木通药材质量控制指标。
Objective:To establish TLC identification method of saponin PH and saponin PJ1,and HPLC determination method of saponin PH,akemisaponin E,saponin PJ1 and scheffoleoside A.in Akebia trifoliata(Thunb.)Koidz. Methods:TLC:Silica-G-CMCNa plate was used with chloroform-methanol-formic acid(20 ∶10 ∶1)as the developing solvent and 10% sulfuric acid ethanol solution as the colorimetric agent. HPLC:Separation was performed on an ELITE C_(18)(250 mm×4.6 mm,5 μm)column and the mobile phase composed of acetonitrilewater(21 ∶79)at a flow rate of 1.0 mL·min~(-1). The detection wavelength was 200 nm,and the column temperature was set at 30 ℃. Results:Saponin PH and saponin PJ1 in A. trifoliata(Thunb.)Koidz could be separated by TLC.The linear ranges of saponin PH,akemisaponin E,saponin PJ1 and scheffoleoside A were 0.201-5.025 μg(r=0.999 9),0.076-1.888 μg(r=0.999 9),0.224-5.608 μg(r=1.000 0)and 0.041-1.035 μg(r=0.999 9),respectively. The average recoveries(n=3)were 99.8%(RSD=0.46%),98.2%(RSD=1.2%),99.5%(RSD=0.40%)and 98.9%(RSD=1.0%). The contents of saponin PH,akemisaponin E,saponin PJ1 and scheffoleoside A in five batches of samples were 2.094-3.928,0.956-1.418,2.958-4.472 and 0.526-0.797 mg·g~(-1). Conclusion:This method is specific,accurate and reproducible,and can be used in control the quality of A. trifoliata(Thunb.)Koidz.
Objective:To establish TLC identification method of saponin PH and saponin PJ1,and HPLC determination method of saponin PH,akemisaponin E,saponin PJ1 and scheffoleoside A.in Akebia trifoliata(Thunb.)Koidz. Methods:TLC:Silica-G-CMCNa plate was used with chloroform-methanol-formic acid(20 ∶10 ∶1)as the developing solvent and 10% sulfuric acid ethanol solution as the colorimetric agent. HPLC:Separation was performed on an ELITE C_(18)(250 mm×4.6 mm,5 μm)column and the mobile phase composed of acetonitrilewater(21 ∶79)at a flow rate of 1.0 mL·min~(-1). The detection wavelength was 200 nm,and the column temperature was set at 30 ℃. Results:Saponin PH and saponin PJ1 in A. trifoliata(Thunb.)Koidz could be separated by TLC.The linear ranges of saponin PH,akemisaponin E,saponin PJ1 and scheffoleoside A were 0.201-5.025 μg(r=0.999 9),0.076-1.888 μg(r=0.999 9),0.224-5.608 μg(r=1.000 0)and 0.041-1.035 μg(r=0.999 9),respectively. The average recoveries(n=3)were 99.8%(RSD=0.46%),98.2%(RSD=1.2%),99.5%(RSD=0.40%)and 98.9%(RSD=1.0%). The contents of saponin PH,akemisaponin E,saponin PJ1 and scheffoleoside A in five batches of samples were 2.094-3.928,0.956-1.418,2.958-4.472 and 0.526-0.797 mg·g~(-1). Conclusion:This method is specific,accurate and reproducible,and can be used in control the quality of A. trifoliata(Thunb.)Koidz.
引文
[1]中华人民共和国药典2015年版.一部[S].2015:63ChP 2015.VolⅠ[S].2015:63
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[12]IWANAGA S,WARASHINA T,MIYASE T.Triterpene saponins from the pericarps of Akebia trifoliata[J].Chem Pharm Bull,2012,60(10):1264
[13]张宁.三叶木通藤茎HPLC指纹图谱和果实预知子有效成分的研究[D].西安:陕西师范大学,2015:13ZHANG N.Study on HPLC Fingerprint of Stems and Active Constituents of Fruits from Akebia trifoliata(Thunb.)Koidz.[D].Xi’an:Shaanxi Normal University,2015:13
[14]CHOI JW,JUNG HJ,LEE KT,et al.Antinociceptive and antiinflammatory effects of the saponin and sapogenins obtained from the stem of Akebia quinata[J].Med Food,2005,8(1):78
[15]TSEN T.Pharmacological studies on the components of Akebia longeracemosa,especially on the chemical and pharmacological properties of Akebia saponins[J].Shikoku Igaku Zasshi,1973,29(1):65
[2]李丽,陈绪中,姚小洪,等.三种木通属植物的地理分布与资源调查[J].武汉植物学研究,2010,28(4):498LI L,CHEN XZ,YAO XH,et al.Geographic distribution and resource status of three important Akebia species[J].J Wuhan Bot Res,2010,28(4):498
[3]刘桂艳,王晔,马双成,等.木通属植物木通化学成分及药理活性研究概况[J].中国药学杂志,2004,39(5):330LIU GY,WANG Y,MA SC,et al.Overview of research on chemical constituents and pharmacological activities of Akebia Dence.[J].Chin Pharm J,2004,39(5):330
[4]刘岩庭,侯雄军,谢月,等.木通属植物化学成分及药理活性研究进展[J].江西中医学院学报,2012,24(4):87LIU YT,HOU XJ,XIE Y,et al.Overview of research on chemical constituents and pharmacological activities of Akebia Dence.[J].JJiangxi Univ TCM,2012,24(4):87
[5]刘卫国,杨文珏,汪晓辉,等.高效液相色谱法测定三叶木通中齐墩果酸的量[J].时珍国医国药,2005,16(8):747LIU WG,YANG WJ,WANG XH et al.Determination of oleanolic acid in Akebia trifoliate by HPLC[J].LiShizhen Med Mater Med Res,2005,16(8):747
[6]成晓霞.三叶木通有效成分含量测定及指纹图谱研究[D].西安:陕西师范大学,2008:34CHENG XX.The Study on Fingerprint and Determination by HPLCof the Active Constituents from Akebia trifoliata(Thunb.)Koidz.[D].Xi’an:Shaanxi Normal University,2008:34
[7]欧金梅,储晓琴,张虹.高效液相色谱法测定安徽产木通药材中3种有效成分[J].安徽中医药大学学报,2015,34(1):70OU JM,CHU XQ,ZHANG H.Determination of three active constituents of Akebia Dence.in Anhui province by HPLC[J].JAnhui Univ Chin Med,2015,34(1):70
[8]高慧敏,王智民,傅雪涛,等.RP-HPLC测定木通药材中皂苷类成分的含量[J].中国中药杂志,2007,42(1):20GAO HM,WANG ZM,FU XT,et al.Determination of triterpenoid saponins in Caulis Akebiae by RP-HPLC[J].China J Chin Mater Med,2007,42(1):20
[9]高伟,幸伟年,敖婉初,等.高效液相色谱法测定木通属植物苯乙醇苷B含量[J].南方林业科学,2015,43(5):48GAO W,XING WN,AO WC,et al.Phenylethanoid glycosides Bcontent of Akebia plants determined by HPLC[J].South China Forest Sci,2015,43(5):48
[10]高慧敏,王智民.木通属药用植物研究进展[J].中国中药杂志,2006,31(1):10GAO HM,WANG ZM.Overview of research on chemical constituents of Akebia Dence.[J].China J Chin Mater Med,2006,31(1):10
[11]MATSUZAKI K,MURANO K,ENDO Y,et al.Nortriterpene saponins from Akebia trifoliate[J].Nat Prod Commun,2014,9(12):1695
[12]IWANAGA S,WARASHINA T,MIYASE T.Triterpene saponins from the pericarps of Akebia trifoliata[J].Chem Pharm Bull,2012,60(10):1264
[13]张宁.三叶木通藤茎HPLC指纹图谱和果实预知子有效成分的研究[D].西安:陕西师范大学,2015:13ZHANG N.Study on HPLC Fingerprint of Stems and Active Constituents of Fruits from Akebia trifoliata(Thunb.)Koidz.[D].Xi’an:Shaanxi Normal University,2015:13
[14]CHOI JW,JUNG HJ,LEE KT,et al.Antinociceptive and antiinflammatory effects of the saponin and sapogenins obtained from the stem of Akebia quinata[J].Med Food,2005,8(1):78
[15]TSEN T.Pharmacological studies on the components of Akebia longeracemosa,especially on the chemical and pharmacological properties of Akebia saponins[J].Shikoku Igaku Zasshi,1973,29(1):65