HPLC法测定银州柴胡中五种皂苷的含量

首次建立HPLC法同时测定银州柴胡中柴胡皂苷a、c、d、e和f五种皂苷含量的方法。流动相为乙腈-水梯度洗脱,柱子为HibarRT RP-18(4.6 mm×250 mm,5μm),流速1.0 mL/min,柱温30℃,检测波长210 nm。柴胡皂苷a、c、d、e和f的线性范围及相关系数分别为:21.78～43.56μg(r=0.9993),3.94～9.85μg(r=0.9994),20.68～51.70μg(r=0.9998),1.67～5.57μg(r=0.9997),1.91～6.70μg(r=0.9992);柴胡皂苷a、d的加样回收率分别为101.5%和98.5%。该方法操作简便,结果准确,重现性好,为柴胡药材多指标质量分析方法的建立和银州柴胡药用提供了参考依据。     

北柴胡不定根培养及茉莉酸甲酯处理对柴胡皂苷含量的影响

目的:建立北柴胡不定根培养体系,明确茉莉酸甲酯(MeJA)处理对北柴胡不定根中柴胡皂苷含量积累的影响。方法:利用固体和液体相结合的组织培养技术培养北柴胡不定根;分别以不同浓度的MeJA处理不定根不同时间,利用HPLC测定处理后不定根中柴胡皂苷含量的积累变化。结果:培养了北柴胡不定根;MeJA处理对北柴胡不定根中柴胡皂苷含量的积累有明显促进作用,当MeJA浓度为200μmol/L时,柴胡皂苷含量最高,为0.45%;以200μmol/L MeJA处理北柴胡不定根26 d时,柴胡皂苷含量最高,为0.51%。结论:北柴胡不定根培养结合MeJA诱导,可做为柴胡皂苷次生代谢合成途径及其积累规律研究的有效技术体系。     

北柴胡营养器官中主要化学成分的组织化学定位及其含量比较

应用植物解剖学、组织化学定位和植物化学方法,研究了北柴胡各营养器官中柴胡皂苷和黄酮类化合物的积累分布状态及其含量变化。结果表明,柴胡皂苷在根中分布在中柱鞘和次生韧皮部中;在茎中主要分布在表皮、棱角处的厚角组织以及位于皮层和髓中的分泌道的上皮细胞中;在叶中,则分布在表皮细胞和整个叶肉组织中。而黄酮类化合物在茎中分布在表皮、棱角处的厚角组织、皮层、髓射线和髓鞘细胞中;在叶中,则主要分布在表皮和位于上下表皮内的厚角组织中。同时,北柴胡中柴胡总皂苷在根、茎、叶中的含量的变化规律为根>叶>茎;而总黄酮在根、茎、叶中的含量的变化规律为叶>茎>根;且在叶中含量相当高,从而为北柴胡的综合利用提供依据,对合理利用药材和保护北柴胡资源也有一定意义。     

UPLC-QTof-MS测定不同提取条件下柴胡皂苷a和柴胡皂苷d的转化规律

使用UPLC-QTof-MS研究了不同提取条件下柴胡皂苷a、d的转化规律。色谱柱为Acquity UPLC BEH C18(2.1 mm×50 mm,1.7μm);流动相为乙腈-0.01%甲酸水溶液;流速为0.45 mL/min。以四极杆串联飞行时间质谱(Q-Tof)为检测器,使用负离子检测模式,对经过12种不同提取方法提取的柴胡样品(1~12)进行检测。得到柴胡皂苷a、d不同提取方法下的转化规律:酸性常温条件下水解生成柴胡皂苷b1、b2,酸性加热情况下糖苷键水解生成皂苷元;中性、碱性条件下均较稳定。实验结果对柴胡皂苷a、d的提取、分离、分析提供了参考依据。     

山西不同产地柴胡皂苷含量与生态因子的相关性研究

目的：研究山西不同产地柴胡皂苷含量与生态因子的相关性，为山西柴胡的规范化种植提供参考依据。方法：以山西产柴胡为研究对象，收集山西11个柴胡产地的经纬度、海拔、年均温、年降雨量、年均日照和无霜期7个生态因子信息，采用高效液相色谱法测定不同产地柴胡的皂苷含量，分析柴胡皂苷含量与生态因子的相关性。结果：经纬度、无霜期和年均温是影响柴胡皂苷含量的主要生态因子，柴胡皂苷含量与经纬度呈正相关，与无霜期和年均温呈负相关。结论：研究结果可为山西柴胡的生态区划种植、品质提升提供一定的参考依据。     

绛县产人工栽培与野生柴胡质量对比研究

目的:以绛县迴马岭人工栽培与野生柴胡为样本,对比人工栽培与野生柴胡根部性状及质量。方法:采用性状鉴别、显微鉴别并结合高效液相色谱法,以柴胡皂苷a、d含量为指标评价,对人工栽培与野生柴胡根部性状及质量进行比较研究。结果:人工栽培柴胡较野生柴胡顺直且色浅,药效成分含量高于野生柴胡。结论:在临床上,为人工栽培柴胡代替野生柴胡提供了理论依据。     

狭叶柴胡营养器官中柴胡皂苷和黄酮类化合物的积累部位及含量比较

采用植物解剖学、组织化学定位和紫外分光光度计分析方法,对狭叶柴胡各营养器官(根、茎、叶)的结构特征、营养器官中柴胡皂苷和黄酮类化合物的积累部位及其含量进行比较研究。结果表明:(1)狭叶柴胡根由周皮、中柱鞘薄壁组织和次生维管组织组成,茎由表皮、皮层和维管柱组成,叶为等面叶结构,其营养器官的结构均表现出对旱生环境的适应性。(2)组织化学定位显示,柴胡皂苷在狭叶柴胡根中主要分布在维管形成层和次生韧皮部及次生木质部靠近维管形成层的木薄壁细胞中,在茎中主要分布在表皮、皮层、维管形成层及韧皮薄壁细胞中,在叶中主要分布于表皮和叶肉中;狭叶柴胡的黄酮类化合物在茎中主要分布于表皮、棱角处的厚角组织及部分皮层细胞中,在叶中主要分布于表皮和位于叶缘及下表皮内的厚角组织中,在根内分布很少。(3)定量分析表明,柴胡总皂苷在狭叶柴胡根、茎、叶中的含量分别为2.635%、1.045%和0.981%;而黄酮类化合物在根、茎、叶中的含量分别为0.032%、1.212%和2.259%;定量分析结果与组织化学实验结果相符。研究表明,柴胡总皂苷在狭叶柴胡的根中含量最高,支持《中国药典》以根入药的结论;而且狭叶柴胡叶中的黄酮类化合物含量较高,建议对狭叶柴胡的叶进行综合开发利用。     

乙酸钠、水杨酸和Cu2+对北柴胡不定根培养合成皂苷的影响

通过添加柴胡皂苷等目的产物合成的外源前体物质和诱导子,探讨了北柴胡不定根培养高效合成柴胡皂苷的条件。结果表明,在1/2MS培养基中添加乙酸钠、水杨酸和Cu2+均有利于不定根合成柴胡皂苷。在培养基中添加0.8 g/L乙酸钠,有利于原产于山西万荣的北柴胡的不定根的皂苷合成;在培养基中添加0.6 g/L乙酸钠,有利于原产于山西左权的北柴胡的不定根的皂苷合成。在培养基中添加0.06 g/L水杨酸,有利于万荣、左权北柴胡不定根的皂苷合成。添加0.1 mg/L Cu2+,有利于原产于山西芮城的北柴胡的皂苷合成;添加0.25mg/L Cu2+,有利于万荣北柴胡不定根的皂苷合成。     

模拟干旱胁迫处理下北柴胡种苗MYC2基因的调控研究

该研究以北柴胡一年生种苗为材料,使用质量分数为10%、20%的PEG6000营养液进行模拟干旱胁迫实验,检测北柴胡根内源信号物质OPR、JA含量,转录因子BcMYC2和柴胡皂苷生物合成途径中4个关键酶基因HMGR、IPPI、FPS、β AS表达量,以及柴胡皂苷a、d含量,探究模拟干旱胁迫下茉莉酸信号通路调控BcMYC2进而影响柴胡皂苷生物合成的机制。结果表明：(1) PEG6000模拟干旱胁迫处理北柴胡种苗后,20% PEG6000处理组根内OPR含量在2 h时出现峰值,10% PEG6000处理组的根内OPR含量在6 h时出现峰值;两个胁迫组内源JA含量均在2 h时出现峰值。(2)两个胁迫处理组北柴胡根内BcMYC2相对表达量均在2 h处出现峰值,之后的2~4 h时间段内大幅度下降,且20% PEG6000处理组的BcMYC2相对表达量高于10% PEG6000处理组;其余4个柴胡皂苷生物合成途径关键酶基因HMGR、IPPI、FPS、β AS的相对表达量均在4 h时出现峰值,晚于BcMYC2相对表达量出现峰值的时间。(3)经模拟干旱胁迫处理后,北柴胡根内的柴胡皂苷含量在36 d内逐渐上升;在处理36 d时,20% PEG6000处理组的柴胡皂苷含量略高于10% PEG6000处理组,且两处理组均显著高于对照组。研究发现,经PEG6000模拟干旱处理后,北柴胡根内信号物质OPR及JA含量提高,促进BcMYC2的表达,进而提高柴胡皂苷生物合成途径中的关键酶基因的表达,最终显著提高了根内柴胡皂苷的含量。     

柴胡属植物中柴胡皂甙的资源利用

本文用高效液相色谱法(HPLC),分析了国产19种柴胡及市售商品柴胡中柴胡皂甙a、c、d(Saikosaponin a、c、d)的含量。结果表明19种柴胡根中均含上述三种成分,但44个样品的含量不同,其中北柴胡以河南信阳的含量较高,南柴胡以陕西延安的为好,多种柴胡的茎叶含有上述三种成分。但含量较根部低,少数种未检出。     

柴胡类药材的柴胡皂甙分析

柴胡类药材的柴胡皂甙分析李光慧罗燕燕王瑛袁昌齐王年鹤（北京临床药学研究所,北京１０００３５）（江苏省中国科学院植物研究所,南京２１００１４）ＡｎａｌｙｓｉｓｏｆｓａｉｋｏｓａｐｏｎｉｎｓｉｎｍｅｄｉｃｉｎａｌＢｕｐｌｅｕｒｕｍｓｐｐ．ＬｉＧｕａｎｇ...     

Localization and dynamic change of saikosaponin in root of Bupleurum chinense

Anatomical,histochemlcal and phytochemical methods were used to investigate the structure,the localization and content changes of total saikosaponin and saikosaponin-a of the roots of Bupleurum chinense DC.at different developmental stages.Results showed that saikosaponin was mainly distributed in pericycle and primary phloem in the young root;but In the mature root,it was mainly distributed in vascular camblum and secondary phloem.During the whole growth period from the pre-blossom,blossom,fruit,and fruit mature periods until the pre-withering period,it was in the fruit mature period that both the total saikosaponin content and the saikosaponin-a content reached the highest level.So the last 20 d of October was considered as the right collecting season for the drug of B.chinense.In addition,the quality of 1-year-old drug was better than that of 2-year-old drug due to its higher saikosaponin content.On the other hand,judging from the external characteristics of the drug,the one with an acerose taproot and more lateral roots was of better quality.The results offered theoretical bases for selecting medicinal material of high quality and determining the most appropriate harvesting stage and part of B.chinense.     

Two New Species of the Genus Bupleurum (Umbelliferae) from China

Bupleurum is a genus largely distributed in temperate regions of the Northern Hemisphere. In China 36 species, 17 varieties and 7 forms have hitherto been reported and most of them are used as Chinese traditional drugs under the name of Chai-Hu. Chai-Hu is one of the most popular drugs used ever since the ancient time and is prescribed principally in the treatment of fevers and influenza. In this article, the authors report two new species, B. kunmingense Y. Li et S. L. Pan and B. polyclonum Y. Li et S. L. Pan which were discovered in Yunnan Province based on morphological studies and preliminary phytochemical tests. The ultraviolet spectrums and thin layer chromatograms of the essential oil and the crude saikosaponins of the two new species are similar to those of B. chinense DC., the standard material medica of Chai-Hu. Moreover, two new saikosaponin spots located between saikosaponin a and c in TLC were discovered in the two new species, and this could be served as a chemical evidence for identification purpose. In the histochemistry examination, the reaction of saikosaponin with color developing agent in parenchyma of the roots of the two new species is obviously more significant than that of B. chinense DC. and this phenomenon has also been proved to be true by TLC of the crude saikosaponin extracts of these two new species. It is suggested that the two new species be used as a substitute ofhigh quality for Chai-Hu.     

Species discrimination of Radix Bupleuri through the simultaneous determination of ten saikosaponins by high performance liquid chromatography with evaporative light scattering detection and electrospray ionization mass spectrometry

A simple, rapid and robust high performance liquid chromatography-evaporative light scattering detection (HPLC-ELSD) method was established for the species discrimination and quality evaluation of Radix Bupleuri through the simultaneous determination of ten saikosaponins, namely saikosaponin-a, -b(1), -b(2), -b(3), -b(4), -c, -d, -g, -h, and -i. These compounds were chromatographed on an Ascentis(?) Express C18 column with a gradient elution of acetonitrile and water containing 0.1% acetic acid at a flow rate of 1.0 mL/min. Saikosaponins were monitored by ELSD, which was operated at a 50°C drift tube temperature and 3.0 bar nebulizer gas (N(2)) pressure. The developed method was validated with respect to linearity, intra- and inter-day accuracy and precision, limit of quantification (LOQ), recovery, robustness and stability, thereby showing good precision and accuracy, with intra- and inter-assay coefficients of variation less than 15% at all concentrations. Furthermore, a high performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-ESI-MS) method was developed to certify the existence of ten saikosaponins, as well as to confirm the reliability of ELSD. The extraction conditions of saikosaponins from Radix Bupleuri were also optimized by investigating the effect of extraction methods (sonication, reflux and maceration) and various solvents on the extraction efficiencies for saikosaponins. Sonication with 70% methanol for 40 min was found to be simple and effective for extraction of major saikosaponins. This analytical method was applied to determine saikosaponin profiles in 20 real samples consisting of four Bupleurum species, namely B. falcatum, B. chinense, B. sibiricum and the poisonous B. longiradiatum. It was found that three major saikosaponin-a, -c and -d were the major constituents in B. falcatum, B. chinense, and B. longiradiatum, while one major saikosaponin (saikosaponin-c) was not identified from B. sibiricum. In addition, no saikosaponin-b(3) was detected in B. longiradiatum samples, indicating that the toxic B. longiradiatum may be tentatively distinguished from officially listed Bupleurum species (B. falcatum and B. chinense) based on their saikosaponin profiles. Overall the simultaneous determination of ten saikosaponins in Radix Bupleuri was shown to be a promising tool to adopt for the discrimination and quality control of closely related Bupleurum species.     

A New Species of the Genus Bupleurum (Umbelliferae) from China

Chai-Hu is one of the most popular Chinese traditional drugs used ever since the ancient time and is prescribed principally in the treatment of fevers and influenza. In this article, the authors report a new species, Bupleurum luxieuse Y. Li et S. L. Pan which was discovered in Yunnan Province and used as Chinese drug under the name of Chai-Hu. The morphology was analysed and preliminary phytochemical tests of B. luxiense were carried out. The thin layer chromatograms and gas chromatograms of the essential oil and the qualitative analysis of the saikosaponin of this new species are similar to those of Bupleurum chinense DC., a standard material medica of Chai-Hu. Moreover, the roots of B. luxiense is discovered to have saikoside 2 times more than B. chinense. The results suggest that the new species be used as a substitute of high quality for Chai-Hu.     

赶黄草总黄酮树脂精制工艺与检测方法

本研究以总黄酮吸附量和解析率为检测指标,结合动态洗脱考察结果,考察11种树脂对赶黄草总黄酮的富集精制能力。优化树脂类型,选定了对总黄酮进行富集纯化较好的树脂HPD450;采用单因素及正交实验,确定最佳工艺条件:上样量10 mL,以6 mL/min的流速8 BV水、6 BV 80%甲醇溶液洗脱,pH值为6~7。以此方法得到90%以上的赶黄草总黄酮,实验结果良好,总黄酮精制工艺成效显著。用HPLC法精确测定赶黄草中槲皮苷、PGHG和ThA三个指标性黄酮成分,最佳色谱条件为:C18色谱柱,检测波长280 nm,进样量5μL,流动相:乙腈-0.05%磷酸溶液(0~25 min:12%~45%乙腈;25~40 min:45%~70%乙腈)梯度洗脱;精确测定赶黄草全草中3种黄酮成分的含量。采用分光光度法检测,指导优化总黄酮精制工艺,并以HPLC法精确测定三种黄酮代表成分。比较两种方法测定结果相一致,互为补充;故在工业生产时,可以分光光度法指导生产,以HPLC法精确定量测定。     

东北羊草草地锰、铜、锌含量特征的研究

 本文研究了羊草(Aneurolepidium chinense)草地割草场3种植物必需的微量元素锰、铜、锌的含量特征。结果表明,在生长季各时期,优势种羊草各器官元素含量有很大变化,总的趋势是：根>根茎>茎>叶>穗(Zn：叶>茎),各器官之间元素含量差异显著(P<0.05)。其它种类植物根中锰、铜、锌含量也显著高于地上各器官。羊草地上部锰、铜、锌积累量在生长季中的变化近似于“S”型曲线,但各元素曲线最高点出现的时间不同。寸草苔(Carex duriuscula)和针蔺(Heleocharis acicularis)地上部锰、铜、锌积累量变化与生物量变化相似,基本为双峰型曲线。三种元素在群落中的分布规律是：根>茎>叶>穗,与生物量分布规律相同。但锰、铜在地上各器宫中积累量占植物总积累量的比率低于相应的生物量比率,而锌这两者的比率则与生物量比率相近。群落中锰、铜、锌积累量与根层土壤中锰、铜、锌总量之比分别为0.10%,0.33%和0.09%。     

中药柴胡不同采收期的皂甙含量

中药柴胡不同采收期的皂甙含量潘泽惠庄体德周雪林林湘（江苏省中国科学院植物研究所,南京２１００１４）（黑龙江双鸭山矿务局师范学校,双鸭山１５５１２５）ＯｎｓｅａｓｏｎａｌｃｈａｎｇｅｓｏｆｔｈｅｔｏｔａｌｓａｐｏｎｉｎｓｉｎＣｈｉｎｅｓｅｔｒａｄｉｔ...     

不同提取方法对豫产白花蛇舌草多糖及抗氧化活性的影响

为评价不同提取方法对河南地区白花蛇舌草粗多糖的得率、总碳水化合物含量、杂质含量和抗氧化活性的影响,本实验分别以传统热水提取法、超声波辅助提取法、纤维素酶提取法从该药材中提取多糖。同时测定了粗多糖中总碳水化合物含量、杂质含量(总蛋白和总酚含量)以及羟基自由基清除能力、DPPH自由基清能力、还原力等抗氧化能力。实验结果显示三种提取法所得粗多糖相对于原药材的得率分别为6.77±0.09%、6.37±0.01%和4.81±0.90%;粗多糖中的总碳水化合物含量分别为289.14±1.33、246.83±2.33和278.92±2.92mg/g。粗多糖中总蛋白含量分别为28.59±2.21、23.26±2.43和4.96±0.18mg/g,总酚类成分含量分别为5.9±0.08、5.31±0.40和2.82±0.07mg/g。抗氧化活性实验结果表明,三种粗多糖的羟基自由基清除能力呈现出浓度依赖性,且均在3.0mg/mL时达到最大,对应最大清除率分别为89.13%、85.87%和74.71%。另外,三种粗多糖的DPPH自由基清除率分别在0.75~3.0mg/mL的浓度区间内达到最大并保持稳定,对应清除率的稳定区间分别为77.02%~77.90%、77.06%~77.96%和83.16%~85.73%。综上所述,以上三种不同提取方法对白花蛇舌草多糖的品质影响较大,可以为豫产白花蛇舌草的品质鉴别、多糖质量控制和开发利用提供依据。