高效液相色谱指纹图谱在中药芦根上的应用

利用现代分离分析技术如高效液相色谱(HPLC)与气相色谱(GC),能够获得中草药的色谱指纹图谱。本实验报道了高效液相指纹图谱应用于快速检测中草药提取物以及应用于研究、开发与生产中的质量控制体系中,利用HPLC建立芦根的色谱指纹图谱。实验条件为反相C18柱,流速为1．0mL／min,紫外检测器(λ=230nm)。通过分析比较色谱指纹图谱中的相对保留值α以及相对面积St对不同产地的芦根进行对比研究。这种基于HPLC的分析策略为中草药的精确鉴定和质量控制提供了有效的信息,并为HPLC在复杂组分样品中的应用开拓了新的领域。     

高效液相色谱指纹图谱在地黄研究中的应用(Ⅰ)

长期以来,中药有效成分的提取分离和质量控制一直是中药产业中特有和尚未很好解决的问题.随着天然产物化学和现代仪器分析技术等相关学科的迅速发展,国内外在植物药及其制剂质量控制研究方面的重点均已转向利用各种仪器进行品质评价方面.中药来源于自然,其化学成分的多样性和复杂性是中药发挥其疗效的物质基础,同时也是质量评价与控制的重点与难点.中药的化学成分复杂,所体现的是综合效应和整体疗效.然而多年以来在中药的研究靠中,采取了与化学药品相同的研究思路,结果将药材及制剂内部各成分的综合作用和相互关系彼此孤立.在中药材及复方制剂质量标准的研究制定中,也往往是采用各种分析检测手段测定其中某种有效成分,并以其含量多少来判断比较某种药材的质量.这种质量控制模式很难做到全面衡量中药及其制剂的质量、疗效和稳定性.单一成分含量达标并不能说明该中药材质量合格.指纹图谱质量控制模式能综合反映某一特定药材各主要组分及其相对含量.因此它比测定任何一种或几种成分所提供的信息都丰富和有用的多.中药提取物在给定的实验条件下所得到色谱指纹图谱反映了该中药的化学组成及其含量分布状况,其特征峰可作为鉴别中药的依据.本文研究结果表明,利用色谱指纹图谱的相关参数来对中药原药材的质量进行控制,进而使中成药的质量得到保证,此方法的运用是切实可行的.另外,在有效成分不明确的前提下,利用色谱指纹图谱来评价中药的质量是有其意义的.色谱指纹图谱作为控制中药质量的新手段将在推进中药现代化进程上起到积极的作用.本文利用高效液相色谱建立了地黄的色谱指纹图谱.采用反相C18柱(5μm,150 mm×4.6 mm)、流动相甲醇:水(V/V=5:95)、检测波长为280 nm、流速1.0 mL/min进行试验.根据相对保留值和相对面积值对色谱指纹图谱进行分析对比研究,建立了控制地黄药材质量的新方法.该法为中药样品的鉴定提供了较全面的信息.     

不同产地白芷药材高效液相色谱指纹图谱研究

本文采用高效液相色谱-二极管阵列检测器(HPLC-DAD)法建立中药白芷的指纹图谱.应用化学计量学中两种不同的模式识别方法(主成分分析法和系统聚类分析法)对实验数据进行处理,以找出来自三个不同产地30个中药白芷样品间的相似性及差异性.两种模式识别方法均能成功地按样品的来源将不同产地的样品正确分类.建立了不同产地中药白芷的识别方法,该方法能有效地控制中药白芷的质量,并能为其它中药产品的化学模式识别提供参考.     

穿心莲药材高效液相色谱指纹图谱研究及其应用

应用反相高效液相色谱(RP-HPLC)法建立穿心莲药材指纹图谱。首先以穿心莲对照药材为对象建立其乙醇提取物特征色谱图,再用5种不同来源穿心莲药材的乙醇提取物验证色谱图的指纹特性。在穿心莲药材乙醇提取物特征色谱图中有共有峰10个,各共有峰的相对保留时间与相对峰面积的相对标准偏差(RSD)均小于4%。对照药材中共有峰峰面积占总峰面积的98.5%以上。该指纹图谱用于清火栀麦胶囊中穿心莲的鉴别专属性良好。     

高效液相色谱指纹图谱应用于地黄的研究

利用高效液相色谱建立了地黄的色谱指纹图谱。采用反相C1 8柱 (5 μm ,1 5 0mm× 4.6mm) ,流动相甲醇∶水 (V V =5∶95 ) ,检测波长为 2 80nm ,流速 1 .0mL min进行实验。根据相对保留值和相对面积值对色谱指纹图谱进行分析对比研究 ,建立了控制地黄药材质量的新方法。该法为中药样品的鉴定提供了较全面的信息 ,并为HPLC在复杂组分的样品分析领域开拓了新的应用途径。     

高效液相色谱指纹图谱应用于板蓝根的鉴定

利用高效液相色谱建立了板蓝根的色谱指纹图谱。采用反相C18柱，流动相为水：甲醇（V/V=96：4），检测波长为260nm；流速1.0mL/min进行试验。根据相对保留值a和相对面积Sr对色谱指纹图谱进行分析对比研究，建立了一个快速、简易鉴别板蓝根样品的色谱指纹图谱分析方法。该法为中药样品的鉴定提供了较全面的信息，并为HPLC在复杂组分的样品分析和鉴别领域开拓了新的应用。     

桂枝茯苓丸的高效液相色谱指纹图谱研究

应用高效液相色谱(HPLC)法建立桂枝茯苓丸的指纹图谱。采用ANGLA Promosil C18色谱柱(250×4.6mm,5μm),以乙腈-0.1%磷酸水溶液为流动相,梯度洗脱,230nm检测,优化条件下,桂枝茯苓丸各成分得到较好的分离。经方法学验证,方法具良好的精密度、重复性和稳定性。采用中药色谱指纹图谱相似度评价系统软件(2004A版)进行指纹图谱分析,11批桂枝茯苓丸样品相似度均在0.90以上。实验表明:桂枝茯苓丸的HPLC指纹图谱特征性和专属性强,可较系统地用于桂枝茯苓丸的质量控制。     

五加生化胶囊的高效液相色谱指纹图谱研究

应用高效液相色谱(HPLC)法建立五加生化胶囊的指纹图谱。采用ANGLAVenusil XBP-C18色谱柱(250×4.6mm,5μm),以甲醇-0.1%磷酸溶液为流动相,梯度洗脱,检测波长为327nm,五加生化胶囊多数峰达到基线分离。采用2004A中药色谱指纹图谱相似度评价系统对11批样品的指纹图谱进行峰匹配,确定12个共有峰,11批五加生化胶囊指纹图谱的相似度均在0.90以上。结果表明:五加生化胶囊的HPLC指纹图谱特征性和专属性强,可较系统地用于五加生化胶囊的质量控制。     

高效液相色谱法研究当归指纹图谱

目的:研究当归药材的指纹图谱.方法:高效液相色谱法,Hypersil ODS柱(4.6 mm×250 mm,5μm),Kromasil ODS保护柱(4.6 mm×10 mm,5μm).甲醇-1%醋酸梯度流动相,流速1.0 mL/min,柱温25℃.以当归对照药材为对照品,色谱峰光谱采集范围:190~400 nm;以阿魏酸为参照物并测定了阿魏酸的含量,检测波长:323nm,流动相:甲醇-1%醋酸(40∶60,V/V),流速0.7 mL/min.结果:找出了22个共有峰,其中5号峰为阿魏酸,11个样品与当归对照药材之间的相似度均在90%以上,平均相似度为96.77%.结论:样品处理方法简单,研究所得的当归指纹图谱稳定性、重复性好,可以作为当归极性部分的特征性指纹图谱.     

高效液相色谱法测定茯苓皮的指纹图谱

本文用正交设计优化茯苓皮的超声提取条件,并有效提取了茯苓皮药材内的主要成分三萜类物质,同时利用高效液相色谱法(HPLC)建立了提取物的液相指纹图谱。实验考察了不同提取溶剂、提取条件对提取工艺的影响;确立了最佳色谱条件,得到了茯苓皮中24种总提取物的指纹图;同时对10种不同产地的茯苓皮药材进行分析,标定了17个共有峰。研究结果表明,HPLC指纹图谱方法重现性好,稳定可靠,可用于控制茯苓皮中三萜成分的质量。     

牛奶中12种磺胺类药物残留的高效液相色谱测定方法

建立了高效液相色谱(HPLC)检测牛奶中12种磺胺类药物(磺胺二甲基嘧啶、磺胺间甲氧嘧啶、磺胺甲噁唑、磺胺对甲氧嘧啶、磺胺喹噁啉、磺胺氯哒嗪、磺胺氯吡嗪、磺胺嘧啶、磺胺噻唑、磺胺甲基嘧啶、磺胺甲噻二唑、磺胺地索辛)残留的方法,考察了样品的提取、净化及色谱分析条件。牛奶样经乙酸乙酯提取、固相萃取净化后上机分析,11种磺胺药物标准曲线在10~800μg/L,SM2在5~200μg/L质量浓度范围内相关系数r>0.999,回收率为76%~106%,相对标准偏差小于15.9%。检出限为5~8μg/L,定量下限为14~27μg/L。     

On-line fluorescence detection in HPLC: A high sensitive technique for the measurement of low enzyme activities: The case of tryptophan-hydroxylase

Summary The use of on-line fluorescence detection after high-performance liquid chromatographic separation permits the direct quantitative analysis of the reaction mixture of enzymes with low activity. The application to the determination of tryptophan-hydroxylase in vitro as well as in vivo is described. The whole system has a detection limit of 0.1 g 5-hydroxytryptophan formed/hour/gram of brains.     

Comparison of microemulsion electrokinetic chromatography with high-performance liquid chromatography for fingerprint analysis of resina draconis

Microemulsion electrokinetic chromatography (MEEKC) has been developed for fingerprint analysis of resina draconis, a substitute for sanguis draconis in the Chinese market. The microemulsion as the running buffer was made up of 3.3% (w/v) sodium dodecyl sulfate (SDS), 6.6% (w/v) n-butanol, 0.8% (w/v) n-octane, and 10 mmol/L sodium tetraborate buffer (pH 9.2), which was also used as the solvent for ultrasonic extraction of both water- and fat-soluble compounds in the traditional Chinese medicine samples. Four batches of resina draconis obtained from different pharmaceutical factories located in different geographic regions were used to establish the electrophoretic fingerprint. MEEKC was performed using a Beckman PACE/MDQ system equipped with a diode-array detector and with monitoring at 280 nm. The fingerprint of resina draconis comprised 27 common peaks within 100 min. The relative standard deviations of the relative migration time of these common peaks were less than 2.1%. Through repetitive injection of the sample solution six times in 24 h, all relative standard deviations of the migration time and peak area of loureirin A and loureirin B were less than 2.5 and 3.8%, which demonstrated that the method had good stability and reproducibility. The relative peak areas of these common peaks in the electropherograms of four batches of resina draconis were processed with two mathematical methods, the correlation coefficient and the interangle cosine, to valuate the similarity. The values of the similarity degree of all samples were more than 0.91, which showed resina draconis samples from different origins were consistent. On the other hand, high-performance liquid chromatography (HPLC) coupled with photodiode-array detection was also applied to establish the fingerprint of resina draconis. The samples were separated with a LiChrospher C₁₈ column using acetonitrile (solvent A) and water containing 0.1% H₃PO₄ (solvent B) as the mobile phase in linear gradient elution mode at a flow rate of 0.6 mL/min and detection was at 280 nm. There were only 20 common peaks in the HPLC fingerprint, and the values of the similarity degree of all samples were also more than 0.91. Though the similarity results of fingerprint analysis seemed to be the same, MEEKC resulted in more common peaks and higher separation efficiency for a variety of polarities of the components than HPLC. So, MEEKC was more suitable for development of the fingerprint of resina draconis.     

高效液相色谱－荧光检测法测定蒽和荧蒽

采用高效液相色谱－荧光检测法测定多环芳烃化合物蒽和荧蒽。在Ｓｈｉｍ－ｐａｃｋＣＬＣ－ｐｈｅｎｙｌ柱上，以５０％的乙腈水溶液作流动相，１２ｍｉｎ内实现了两组分的同时分离测定。线性范围为０～８０μｇ／ｍＬ，检测限低于５０ｎｇ／ｍＬ，相对标准偏差小于１％。     

防晒类化妆品中2种帕地马酯的高效液相色谱检测及质谱确证

建立了同时测定防晒类化妆品中帕地马酯A和帕地马酯O的高效液相色谱分析方法及质谱确证方法。化妆品样品以甲醇为提取溶剂进行超声提取,提取液离心处理,取上清液经微孔滤膜过滤后测定。采用XTerra MS C18柱(250×4.6mm,5μm)分离,外标法定量。结果表明防晒类化妆品中帕地马酯A和帕地马酯O的方法定量限均为0.5mg/kg,在低、中、高3个添加水平范围内平均回收率为89.1%~112.5%,相对标准偏差为2.0%~7.7%。对于疑似阳性样品,进一步采用液相色谱-串联质谱法进行确证分析。该方法准确、快速、灵敏度高,适用于防晒类化妆品的实际检验工作。     

Development of a novel method combining HPLC fingerprint and multi-ingredients quantitative analysis for quality evaluation of traditional Chinese medicine preparation

A novel method combining high performance liquid chromatography (HPLC) fingerprint and simultanous quantitative analysis of multiple acitve components was developed and validated for quality evaluation of one type of traditional Chinese medicine preparations: Shuang-huang-lian (SHL) oral liquid formulation. For fingerprint analysis, 45 peaks were selected as the common peaks to evaluate the similarities among several different SHL oral liquid preparations collected from manufacturers. Additionally, simultanous quantification of eleven markers, including chlorogenic acid, caffeic acid, rutin, forsythiaside, scutellarin, baicalin, forsythin, luteoloside, apigenin, baicalein and wogonin, was performed. Statistical analysis of the obtained data demonstrated that our method has achieved desired linearity, precision and accuracy. Finally, concentrations of these eleven markers in SHL oral liquid prepared by different manufacturers in China were determined. These results demonstrated that the combination of HPLC chromatographic fingerprint and simultaneous quantification of multi-ingredients offers an efficient and reliable approach for quality evaluation of SHL oral liquid preparations.     

Multiple compounds determination and fingerprint analysis of Lidanpaishi tablet and keli by high-performance liquid chromatography

The objective of this paper was to establish an efficient method for quality control of Lidanpaishi (LDPS) tablet and keli, two famous traditional Chinese medicines. A simple and reliable high-performance liquid chromatography (HPLC) coupled with photodiode array detector (DAD) method was developed both for fingerprint analysis (FA) and quantitative determination. In quantitative analysis, linear regressions, limit of detection (LOD) and quantification (LOQ), intra-day and inter-day precisions, recovery, repeatability and stability were all tested and good results were obtained to simultaneously determine the 15 marker compounds, namely chlorogenic acid, rhaponticin, 6,7-dimethoxycoumarin, naringin, hesperidin, neohesperidin, baicalin, wogonoside, baicalein, wogonin, chrysin, honokiol, magnolol, emodin, and chrysophanol in the herbal drugs. In fingerprint analysis, 34 peaks were selected as the characteristic peaks to evaluate the similarities of different samples collected from different pharmaceutical companies in China according to the State Food and Drug Administration (SFDA) requirements, and two kinds of data, relative retention time (RRT) and relative peak area (RPA) were used to identify the common peaks in 15 samples for investigation. Furthermore, hierarchical cluster analysis (HCA) was also performed to evaluate the variation of the herbal drugs. The present approach, i.e. HPLC coupled with multiple compounds determination (MCD) and FA is a powerful and meaningful tool to comprehensively conduct the quality control of traditional Chinese medicines.     

Dispersion measurements on conventional and miniaturised HPLC systems

Summary The volume dispersion produced by a number of commercial detector/injector (DI) system and of laboratory designed detector/injector systems has been measured by three different techniques: (A) by direct connection of injector to detector using a "zero-length" column; (B) by measuring the plate number N as a function of capacity ratio, k′; (C) by placing a 2m × 100μm bore straight open tube between injector and detector, and subtracting the contribution from the tube according to the Taylor-Aris equation. Setting as a criterion of adequate performance that there should be less than a 10% loss in plate efficiency due to the DI system for an unretained solute peak, it is found that in general the commercial systems examined show excessive dispersion in relation to the types of column for which they are claimed to have been designed. A 1μl flow cell designed in our laboratories offers a standard deviation below 1μl and is a possible candidate for use with miniaturised systems. However, extreme precautions to avoid extra-column dispersion must be taken. The column must enter into the detector cell and injection must be by flow splitter. It is considered unlikely that current HPLC technology can be adapted for use with high efficiency columns of bore below 2mm. Presented at the 15th International Symposium on Chromatography, Nürnberg, October 1984