川芎、高良姜等挥发油的水蒸汽蒸馏法和超临界流体萃取法比较研究

目的:确定川芎、高良姜、片姜黄和羌活的挥发油提取最佳条件.方法:用水蒸汽蒸馏法和超临界流体萃取的方法提取挥发油.结果:(1)水蒸汽蒸馏法提取挥发油的最佳条件确定为:川芎:浸泡时间12h,粉末粒度细,提取时间3h,加水比1:10;高良姜:浸泡时间0h,粉末粒度粗,提取时间3h,加水比1:10;片姜黄:浸泡时间24h,粉末粒度中,提取时间1.5h,加水比1:5;羌活:浸泡时间0h,粉末粒度粗,提取时间3h,加水比1:10.(2)超临界流体萃取法提取挥发油的最佳条件确定为:川芎:温度为60℃,粉末粒度细,压力25MPa,提取时间1.5h;高良姜:温度为60℃,粉末粒度中,压力25MPa,提取时间0.5h;片姜黄:温度为60℃,粉末粒度细,压力20MPa,时间1h;羌活:温度为60℃,粉末粒度细,压力25MPa,提取时间1h.结论:超临界流体萃取法要优于水蒸汽蒸馏法.     

Determination and pharmacokinetic study of p-hydroxyphenethyl anisate following intravenous and oral administration in rats by RP-HPLC method

In the present study, we developed a rapid and specific reversed-phase high-performance liquid chromatographic (RP-HPLC) method for the quantification of p-hydroxyphenethyl anisate (HPA), one of the main bioactive constituents of the roots and rhizomes ofNotopterygium incisumand N. franchetii, in rat plasma after an intravenous (20 mg/kg, i.v.) and an intragastrical (200 mg/kg,i.g.) administration to rats, respectively. The method involved a plasma clear-up step using liquid-liquid extraction by EtOAc, followed by RP-HPLC separation and detection. Separation of HPAwas performed on an analytical Diamonsil^TM ODS C₁₈ column with the mobile phase of MeOH–H₂O at ratios of 75:25 (v/v) for i.v. and 70:30 (v/v) for i.g. administration. The flow-rate was 1.0 mL/min, and UV detection was performed at 256 nm. The calibration curves were linear over the ranges of 0.05–5.0 μg/mL (r² = 0.9984) for i.v. and 0.5–10.0 μg/mL (r² = 0.9995) for i.g. administrationin rat plasma. The extraction recoveries were in the range of 82.01%–87.97%. The intra- and inter-day precisions were between 1.71% and 3.99%, with accuracies ranging from 91.22% to 110.5%. The absolute bioavailability of an orally administered HPA in rats was about 48.17%. The developed method was suitable for the determination and pharmacokinetic studyof HPA in rat plasma.     

羌活薄层色谱和HPLC指纹图谱结合多成分含量测定的质量评价方法研究

目的?建立薄层色谱、HPLC指纹图谱、多成分含量测定分析方法,评价18批不同产地羌活市售药材,为提高羌活质量标准提供技术支持。方法?采用高效薄层色谱法对羌活进行鉴别;采用HPLC-DAD法建立羌活指纹图谱,运用相似度软件对18批药材进行质量评价,建立HPLC法测定羌活中多个有效成分（阿魏酸苯乙醇酯、羌活醇、异欧前胡素、镰叶芹二醇）的含量。结果?建立了高效便捷的羌活薄层鉴别方法;所建立的指纹图谱出峰时间适中、方法稳定,共标定了14个共有峰,并通过对照品比对指认了6个色谱峰。18批市售药材指纹图谱相似度按共有峰计算在0.887~0.992;建立羌活多类型有效成分含量测定方法,阿魏酸苯乙醇酯、羌活醇、异欧前胡素、镰叶芹二醇在该色谱条件下分离良好,线性关系良好（r>0.999?8）,平均回收率分别为95.3%、99.5%、98.5%、98.5%。结论?建立的羌活薄层鉴别方法和HPLC指纹图谱稳定可靠,能有效区别羌活、宽叶羌活及其易混淆品;同时建立了羌活中多个有效成分的HPLC含量测定方法,为全面提高羌活质量控制方法提供指导。      

羌活中的香豆素类成分及其抑制脂多糖诱导的RAW 264.7细胞NO生成活性的研究

目的研究羌活Notopterygium incisum的香豆素类成分及其抗炎活性。方法采用硅胶、HPLC等柱色谱方法进行分离纯化,通过质谱、核磁共振波谱数据鉴定化合物的结构;采用脂多糖(LPS)诱导的小鼠巨噬细胞RAW 264.7炎症反应模型,考察羌活中香豆素类成分对炎症反应模型一氧化氮(NO)生成的影响。结果从羌活甲醇提取物分离得到24个香豆素类化合物,分别鉴定为异欧前胡素(1)、川白芷素(2)、补骨脂素(3)、香柑内酯(4)、茵陈素(5)、欧芹酚(6)、5-去氢羌活醇(7)、环氧脱水羌活醇(8)、7″-O-甲基异羌活醇(9)、佛手柑素(10)、7-异戊烯氧基-6-甲氧基-香豆素(11)、栓翅芹烯醇(12)、羌活醇(13)、去甲呋喃羽叶芸香素(14)、异羌活醇(15)、蛇床夫内酯(16)、6-异戊烯氧基伞形花内酯(17)、紫花前胡苷元(18)、异虎耳草素(19)、紫花前胡苷(20)、前胡苷V(21)、前胡苷I(22)、印枳苷元-11-O-β-D-吡喃葡萄糖基-(1→6)-β-D-吡喃葡萄糖苷(23)、羌活苷(24)。化合物7～10、13和15抑制LPS诱导的RAW 264.7细胞NO生成活性最强,最大半数抑制浓度(IC_(50))值为8.50～35.12μmol/L。结论化合物7为新的天然产物,化合物17为首次从羌活中分离得到;C-5位上具有多烯烃结构的香豆素抑制LPS诱导的RAW 264.7细胞NO生成活性较强。     

Quantitative analysis of chemical constituents in different commercial parts of Notopterygium incisum by HPLC–DAD–MS

Ethnopharmacological relevance

Notopterygium incisum used as an antirheumatic and analgesic medicine for the treatment of rheumatism, headache, cold, etc., is not only a traditional Qiang medicine, but also a traditional Tibetan remedy for thousands of years.

Aim of the study

In order to use this herbal medicine in a more effective way, in the present investigation, we analyzed the main chemical constituents of different commercial parts as well as fibrous roots of Notopterygium incisum.

Materials and methods

All crude Notopterygium incisum samples used in this study were collected in Sichuan province of China. We used HPLC-DAD-MS to analyze quantitatively the main chemical constituents of different commercial parts as well as fibrous roots of Notopterygium incisum.

Results

According to different commercial parts, 10 Notopterygium incisum samples collected from two main production areas have been investigated, and four main constituents have been quantitatively analyzed.

Conclusion

The quality of rhizomes is better than that of main roots. Unexpectedly, the content of two main constituents, notopterol and isoimperaorin, in fibrous roots is higher than any other parts of Notopterygium incisum. This result implied that the fibrous root may also have value, and we should study it to explore its potential.     

羌活化学成分研究

从羌活的乙醚提取部分分离鉴定了佛手柑内酯,异欧芹属素乙,去甲呋喃羽叶云香素,佛手酚,羌活醇,羌活酚,脱水羌活酚,花椒毒酚,佛手柑亭,苯乙基阿魏酸酯,对羟基间甲氧基苯甲酸,阿魏酸和娠烯醇酮等13个化合物。     

宽叶羌活化学成分研究

从宽叶羌活的根及根茎中分离并鉴定了９个化合物，其中珊瑚菜内酯为首次从该属植物中分得，其余８个化合物的理化数据和光谱数据与文献报道一致。     

羌活水溶性部分的化学成分

目的对羌活(Notopterygium incisumTing ex H.T.Chang)的水溶性部分化学成分进行研究。方法采用开放ODS柱色谱和凝胶柱色谱分离纯化,NMR、MS等波谱技术分析鉴定结构。结果分离并鉴定了5个化合物,分别为绿原酸(chlorogenic acid,1)、紫花前胡苷(nodakenin,2)、香柑酚OβD吡喃葡萄糖苷(bergaptolOβDglucopyranoside,3)、前胡苷Ⅴ(decurosideⅤ,4)和阿魏酸(fer-ulic acid,5)。结论其中绿原酸为首次从羌活中分离得到。     

狭叶羌活根茎和根的挥发油成分的GC-MS分析

目的分析狭叶羌活(Notopterygium incisumTing ex H. T. Chang)根茎和根的挥发油的化学成分,为其质量评价提供科学依据。方法采用水蒸气蒸馏法提取狭叶羌活根茎和根的挥发油,用GC毛细管柱进行分析,归一化法测定其相对含量,并用GC-MS法鉴定化学成分。结果检出242个色谱峰,初步鉴定出83个化合物,占挥发油总量的75·77%。结论狭叶羌活根茎和根的挥发油中的化学成分主要为单萜和倍半萜类化合物,两者分别占总油中化学成分的13·63 %和67.93 %。(1S)-β-蒎烯(1·67%)、3-蒈烯(1·05%)、柠檬烯(1·22%)和1S-endo-醋酸冰片酯(1·68%)为单萜类化合物中的主要成分;( )-β-榄香烯(6·78%)、苜蓿烯(1·54%)、α-石竹烯(2·64%)、异大香叶烯D (1·67%)、桉烷-4 (14),11-二烯(2·36%)、α-芹子烯(2·42%)、δ-杜松烯(1·55%)、3,7,11-三甲基-2,6,10-十二烷三烯-1-醇(1·03%)、(±)-榄香烯(5·18)、(-)-匙叶桉油烯醇(1·40%)、愈创醇(3·81%)、去羟基异菖蒲烯二醇(1·06%)、γ-桉醇(1·05%)、α-桉醇(7·97%)、异愈创木醇(3·09%)和胡萝卜醇(2·30%)为倍半萜类化合物中的主要成分。总挥发油中,反式肉桂酸异丙酯占11·3%,为所有化合物中含量最高者。     

羌活醇提物预处理对大鼠HIRI模型的防护作用

目的探讨羌活醇提物对大鼠肝缺血再灌注损伤（hepatic ischemia reperfusion injury,HIRI）模型肝功能和抗氧化酶的影响。方法将60只Wistar大鼠分为空白组（6只）、模型组（18只）、丹参预处理组（DS组,18只）和羌活预处理组（QH组,18只）,后三组组内再分为缺血45min组（145）、缺血45min再灌注30rain组（145R30）和缺血45min再灌注60min组（145R60）3小组,每组6只,造模后测定大鼠各时点即刻的血清丙氨酸氨基转移酶（alanine amiotransferase,ALT）、门冬氨酸氨基转移酶（aspartate aminotransferase,AST）水平以及大鼠肝组织中丙二醛（malondiadehyde,MDA）含量、超氧化物歧化酶（superoxide dismutase,SOD）活力水平。结果MDA：羌活组145R30min即刻点MDA含量明显高于145min即刻点（P〈0．01）;145min即刻点,羌活组MDA含量高于模型组（P〈0．05）;145R30min即刻点,丹参组与羌活组MDA含量均明显高于模型组（P均〈0．01）;SOD：羌活组145R30min和145R60min时点的肝组织SOD活力均高于145min时点（P均〈0．01）;145R30min即刻点模型组肝组织SOD活力均低于丹参组和羌活组（P均〈0．001）。结论羌活醇提物可能是通过提高HIRI大鼠肝组织的总SOD活力,增强其对氧自由基的清除能力,减轻脂质过氧化损伤,从而起到保护肝脏的作用。