Progress and prediction of multicomponent quantification in complex systems with practical LC-UV methods

Complex systems exist widely,including medicines from natural products,functional foods,and biological samples.The biological activity of complex systems is often the result of the synergistic effect of multiple components.In the quality evaluation of complex samples,multicomponent quantitative analysis(MCQA)is usually needed.To overcome the difficulty in obtaining standard products,scholars have proposed achieving MCQA through the“single standard to determine multiple components(SSDMC)”approach.This method has been used in the determination of multicomponent content in natural source drugs and the analysis of impurities in chemical drugs and has been included in the Chinese Pharmacopoeia.Depending on a convenient(ultra)high-performance liquid chromatography method,how can the repeatability and robustness of the MCQA method be improved?How can the chromatography conditions be optimized to improve the number of quantitative components?How can computer software technology be introduced to improve the efficiency of multicomponent analysis(MCA)?These are the key problems that remain to be solved in practical MCQA.First,this review article summarizes the calculation methods of relative correction factors in the SSDMC approach in the past five years,as well as the method robustness and accuracy evaluation.Second,it also summarizes methods to improve peak capacity and quantitative accuracy in MCA,including column selection and twodimensional chromatographic analysis technology.Finally,computer software technologies for predicting chromatographic conditions and analytical parameters are introduced,which provides an idea for intelligent method development in MCA.This paper aims to provide methodological ideas for the improvement of complex system analysis,especially MCQA.

双标多测法在枣仁安神颗粒6个成分含量测定中的应用

目的建立枣仁安神颗粒中斯皮诺素、6'''-阿魏酰斯皮诺素、迷迭香酸、五味子醇甲、五味子乙素和丹酚酸B 6个成分的双标多测法,验证该方法在枣仁安神颗粒质量评价中应用的准确性和可行性。方法采用HPLC法,以乙腈为流动相A,0.2%冰醋酸溶液为流动相B,梯度洗脱,流速为1.0 mL·min^(-1),切换波长320 nm(斯皮诺素、6'''-阿魏酰斯皮诺素、迷迭香酸、丹酚酸B)、250 nm(五味子醇甲、五味子乙素),柱温28℃。采用5根不同的C_(18)色谱柱,测定枣仁安神颗粒6个成分的实际保留时间,计算各成分在各色谱柱保留时间的平均值,得到各成分的标准保留时间,选取斯皮诺素(峰1)、五味子乙素(峰6)作为双标化合物,使用双标线性校正法准确定位各成分色谱峰,预测成分在未知色谱柱的保留时间并进行方法学验证。以五味子乙素为参照物,计算其余各成分的相对校正因子,对各成分进行定量。结果双标线性校正法能够准确地预测待测组分在未知色谱柱上的保留时间;斯皮诺素、6'''-阿魏酰斯皮诺素、迷迭香酸、五味子醇甲和丹酚酸B相对于五味子乙素的相对校正因子分别为0.7342、0.6063、0.5664、0.9975及0.7773,其RSD均小于1%;各成分含量计算值和实测值无显著差异。结论双标多测法同时测定枣仁安神颗粒中6个成分可行且准确。

Simultaneous Determination of Seven Components in Gamboge and Its Processed Products Using a Single Reference Standard

Objective To establish a quality control method for simultaneous determination of multiple components in gamboge. Methods A single reference standard for the determination of multiple components （SSDMC） with HPLC was proposed. Seven major components of gamboge including gambogenic acid （S）, β-morellic acid （C1）, 2R-30-hydroxygambogic acid （C2）, isogambogenic acid （C3）, gambogellic acid （C4）, 2R-gambogic acid （C5）, and 2S-gambogic acid （C6） were simultaneously analyzed using gambogenic acid as reference standard. The credibility and feasibility of SSDMC method were validated with respect to linearity, limits of detection and quantification, precision, stability, repeatability, accuracy, ruggedness, and robustness. The relative conversion factors （RCFs） of S and C1-6 were calculated. Twelve batches of gamboge including crude and processed products were successfully analyzed by applying the SSDMC and traditional external standard （ES） methods. Results The SSDMC method was credible and feasible. The RCFs of S and C1-6 were 1.000, 0.913, 0.864, 1.064, 0.777, 0.921, and 0.919, respectively. No significant difference was observed in the contents of the seven components between SSDMC and ES methods. The heat-processing technique caused a reduction in the seven components. Conclusion SSDMC is a simple, reliable, and effective method for the analysis of the complex multiple components in gamboge, and it is also a practical and economical approach.

Simultaneous Determination of Four Anthraquinones in Polygoni Multiflori Radix with Single Reference Standard by High Performance Liquid Chromatography

Objective: To establish a rapid, accurate and reliable analytical method for the simultaneous determination of four major anthraquinones in Polygoni Multiflori Radix(PMR) using single reference standard.Methods: The four components including emodin-8-O-β-D-(EMG), physcion-8-O-β-D-glucoside, emodin and physcion were separated on an ODS C18 column within 13 min and detected at 280 nm. Emodin was selected as the reference standard, and the response factor for each analyte with respect to emodin were calculated. Robustness were also tested including different columns, equipments, temperatures, detection wavelengths, and other chromatographic conditions which might influence stability of response factors.Results: The method was validated in terms of linearity(r^2> 0.9995), LOQs(0.820–3.05 ng/m L), LODs(0.180–0.920 ng/m L), precision,accuracy(95.8–103.6%, RSD < 2.80%) and stability. A total of 40 batches of PMR were analyzed and the results were found to have no statistically significant differences compared with those obtained using the external standard method.Conclusion: This work provided a single standard to determine multi-components method for quantitation of four anthraquinones in PMR,which could be applied in the quality control of this herbal drug.

猴耳环双标多测法及抗氧化、α-葡萄糖苷酶抑制活性成分快速筛选法的建立

目的:建立猴耳环中5个成分的双标多测法及其抗氧化、α-葡萄糖苷酶抑制活性成分的快速筛选法。方法:采用HPLC法,以双标线性校正法准确定位猴耳环中5个成分色谱峰,以没食子酸为参照物,计算没食子酸与其余4个成分的相对校正因子,对各成分进行定量,将所得结果与外标法进行比较。采用离线DPPH-HPLC法及亲和超滤液相联用法快速筛选猴耳环中具有抗氧化、α-葡萄糖苷酶抑制活性成分。结果:双标线性校正法相比于相对保留时间更能准确地预测待测组分在未知色谱柱上的保留时间;相对校正因子法和外标法结果的RSD<2.00%。采用离线DPPH-HPLC法及亲和超滤液相联用法可以快速筛选出猴耳环中7-没食子酰基特利色黄烷、杨梅苷等不同程度抗氧化活性和α-葡萄糖苷酶抑制活性的化合物。结论:该研究建立的双标多测同时测定猴耳环中5个成分的方法准确可行,采用离线DPPH-HPLC法及亲和超滤液相联用法能快速、灵敏、高通量筛选猴耳环中的抗氧化和α-葡萄糖苷酶抑制活性成分。

樟帮法甘草制吴茱萸指纹图谱的建立及生物碱类成分的含量测定

目的 建立樟帮法甘草制吴茱萸(以下简称“泡吴茱萸”)的HPLC指纹图谱,并同时测定吴茱萸碱、吴茱萸次碱、吴茱萸新碱的含量,为泡吴茱萸的质量控制及标准提高提供依据。方法 基于多波长切换法建立泡吴茱萸的指纹图谱,并进行相似度评价;以吴茱萸次碱为内参物,采用斜率校正法计算吴茱萸碱、吴茱萸新碱的相对校正因子,并比较一测多评(QAMS)法与外标法测定结果的差异。结果 建立了泡吴茱萸的指纹图谱,共有20个共有峰,与对照指纹图谱的相似度为0.970~0.998,共指认了7个共有峰;10批样品中QAMS法测定的吴茱萸碱、吴茱萸新碱的含量为1.754~7.542 mg/g和1.281~2.455mg/g,QAMS法与外标法两种检测方法所得结果相似。结论 所建立的HPLC法简单、可靠,分离度、重复性良好,可用于泡吴茱萸的质量评价。

基于一测多评法同时测定脑心舒通胶囊中6种成分的含量

目的:建立HPLC一测多评法同时测定脑心舒通胶囊中6种成分(丹参素、原儿茶醛、羟基红花黄色素A、迷迭香酸、天麻素、丹酚酸B),使用外标法验证此方法在脑心舒通胶囊中质量控制应用的可行性及适应性。方法:以丹参素为内标物,建立其与其他5种成分间的相对校正因子,并计算脑心舒通胶囊各成分含量,实现一测多评。结果:丹参素与其他五种成分的相对校正因子分别为1.025、2.157、0.760、1.820、6.224;9批脑心舒通胶囊中用一测多评法测定6种成分的含量与外标法测定结果无显著差异(相对误差<5%)。结论:该方法准确可靠,可用于脑心舒通胶囊的质量控制。

一测多评法同时测定车前草中4种活性成分的含量

目的:建立一测多评法(Quantitative Analysis of Multi-Components by Single Marker,QAMS)同时测定车前草中原儿茶酸、儿茶素、槲皮素和木犀草素的含量,并验证此方法的可行性及适应性。方法:以槲皮素为内标物,建立其与其他3种成分间的相对校正因子,并计算4种成分含量,实现一测多评。结果:槲皮素与其他3种成分的相对校正因子分别为1.199 2、0.861 3、1.606 9;采用一测多评法测定的10批次车前草中4种活性成分的含量与外标法测定结果无显著差异。结论:该方法准确稳定,可用于车前草多指标质量控制。

A feasible,economical,and accurate analytical method for simultaneous determination of six alkaloid markers in Aconiti Lateralis Radix Praeparata from different manufacturing sources and processing ways

Aconiti Lateralis Radix Praeparata(Fuzi) is a commonly used traditional Chinese medicine in clinic for its potency in restoring yang and rescuing from collapse. Aconiti alkaloids, mainly including monoester-diterpenoidaconitines(MDAs) and diester-diterpenoidaconitines(DDAs), are considered to act as both bioactive and toxic constituents. In the present study, a feasible, economical, and accurate HPLC method for simultaneous determination of six alkaloid markers using the Single Standard for Determination of Multi-Components(SSDMC) method was developed and fully validated. Benzoylmesaconine was used as the unique reference standard. This method was proven as accurate(recovery varying between 97.5%-101.8%, RSD < 3%), precise(RSD 0.63%-2.05%), and linear(R > 0.999 9) over the concentration ranges, and subsequently applied to quantitative evaluation of 62 batches of samples, among which 45 batches were from good manufacturing practice(GMP) facilities and 17 batches from the drug market. The contents were then analyzed by principal component analysis(PCA) and homogeneity test. The present study provided valuable information for improving the quality standard of Aconiti Lateralis Radix Praeparata. The developed method also has the potential in analysis of other Aconitum species, such as Aconitum carmichaelii(prepared parent root) and Aconitum kusnezoffii(prepared root).

骨髓炎颗粒HPLC指纹图谱及一测多评法测定7种成分含量

目的建立骨髓炎颗粒的HPLC指纹图谱及一测多评法(QAMS)测定骨髓炎颗粒中绿原酸、连翘酯苷、芦丁、黄芩素、黄芩苷、汉黄芩苷、党参炔苷等7种主要成分的含量。方法采用Diamonsil-C18色谱柱(150 mm×4.6 mm,4μm),以乙腈-0.1%的磷酸水溶液为流动相,梯度洗脱,流速为1.0 mL·mL^(-1),柱温为30℃,检测波长为280 nm,建立骨髓炎颗粒HPLC指纹图谱与测定绿原酸、连翘酯苷、芦丁、黄芩素、黄芩苷、汉黄芩苷、党参炔苷等7种主要成分含量。结果10批样品共标定了26个共有峰,指认了7个峰,相似度均大于0.995。以黄芩苷为内参物,分别建立绿原酸、芦丁、连翘酯苷、黄芩素、汉黄芩苷、党参炔苷的相对校正因子分别为2.3131、4.1377、2.1300、0.7248、1.9161、8.1718;其校正因子在不同色谱条件下的RSD均小于2%。该6种成分含量用外标法进行测定,与QAMS法比较成分含量结果无明显差异。结论建立的HPLC指纹图谱及7种成分QAMS法,科学合理、简便易行,可用于骨髓炎颗粒中有效成分含量的测定,为深入研究其质量标准奠定基础。

一测多评法同时测定金母颗粒中6种成分

目的建立一测多评法同时测定金母颗粒(金刚刺、土茯苓、火炭母等)中没食子酸、绿原酸、虎杖苷、落新妇苷、白藜芦醇、盐酸小檗碱等6种成分的含量。方法采用高效液相法分析,Agilent ZORBAX SB-C18色谱柱(250 mm×4.6 mm,5μm);流动相为乙腈-0.1%磷酸,梯度洗脱;体积流量1.0 mL·min^(-1);柱温35℃;检测波长291 nm;进样量10μL。以盐酸小檗碱为内标物,计算其他5种成分的相对校正因子,测定其含量。同时通过外标法和一测多评法测定11批金母颗粒中6个成分的含量,并比较两者的差异。结果6种成分在各自范围内线性关系良好(r>0.9994),平均加样回收率为88.74%~106.03%,RSD为0.41%~2.85%,以盐酸小檗碱为内标,没食子酸、绿原酸、虎杖苷、落新妇苷和白藜芦醇的相对校正因子分别为0.8666、0.7463、0.4663、0.6591、0.2683。一测多评法所得结果与外标法接近,相对平均偏差(RAD)<5.0%。结论该方法稳定,准确可靠,可为金母颗粒的质量控制和评价提供理论参考。

基于UPLC-QAMS同时测定参鹿酒中7种人参皂苷含量

该研究旨在建立超高效液相色谱(UPLC)-一测多评(QAMS)法同时测定参鹿酒中7种人参皂苷(Rg1、Re、Rf、Rb1、Rc、Rb2及Rd)的含量。首先采用UPLC测定参鹿酒中7种人参皂苷含量,并进行方法学考察。然后以人参皂苷Rg1为参照物,计算其余6种人参皂苷的相对校正因子及含量,并考察QAMS法的耐用性。最后与外标法相比,验证QAMS的可行性。结果表明,7种人参皂苷在各自范围内线性关系良好(R^(2)>0.999),精密度、重复性及稳定性试验结果的相对标准偏差(RSD)均<3%,加样回收率为100.13%~101.73%。6种人参皂苷的相对校正因子的RSD值均<5%,重现性良好。采用相对校正因子计算得到的13批参鹿酒样品中7种人参皂苷的含量与外标法实测值的相对误差(RE)均<5%,说明所建立的UPLC-QAMS法可有效、快速的评价参鹿酒的质量。

一测多评法测定无花果叶中4种成分的含量

目的:建立无花果叶中绿原酸、芦丁、补骨脂素和佛手柑内酯4种成分含量测定的一测多评方法,并验证此方法的可行性与准确度。方法:采用高效液相色谱法,以芦丁为内参物,建立绿原酸、补骨脂素和佛手柑内酯的相对校正因子(f_(s/i)),并利用f_(s/i)计算无花果叶样品中各成分的含量,同时用外标法测定不同产地无花果叶中各成分的含量,比较一测多评法和外标法所得结果的差异。结果:一测多评法得到的绿原酸、补骨脂素和佛手柑内酯f_(s/i)重现性较好,且通过一测多评法测得的不同产地的无花果叶各成分含量与外标法测得的结果无显著差异。结论:本研究建立的一测多评法准确可行,可用于无花果叶的定量分析和质量评价。

一测多评法同时测定刺五加配方颗粒中6种成分的含量

目的利用一测多评(QAMS)法同时测定刺五加配方颗粒中原儿茶酸、新绿原酸、绿原酸、隐绿原酸、紫丁香苷和刺五加苷E的含量。方法采用UPLC法,以紫丁香苷作为内参物,计算原儿茶酸、新绿原酸、绿原酸、隐绿原酸、刺五加苷E的相对校正因子,并考察各成分相对校正因子的耐用性;同时利用相对校正因子计算各成分含量并与外标一点法(ESM)实测值进行比较。结果在一定线性范围内,原儿茶酸、新绿原酸、绿原酸、隐绿原酸、刺五加苷E的相对校正因子分别为0.788、2.358、2.229、2.718、2.558,重复性较好。QAMS法计算所得12批刺五加配方颗粒中原儿茶酸、新绿原酸、绿原酸、隐绿原酸、刺五加苷E的含量与ESM法实测值无显著性差异。结论该方法准确可行,可节约对照品及检测成本,同时测定刺五加配方颗粒中6种成分的含量,为刺五加配方颗粒的多指标成分质量控制提供参考。

“一测多评”法与外标法测定三黄片中4种黄芩黄酮类成分

目的采用"一测多评"法(QAMS)与外标法(ESM)同步测定三黄片(大黄、盐酸小檗碱及黄芩浸膏)中的4种黄芩黄酮类成分含有量并进行比较,以验证"一测多评"法在三黄片中应用的可行性。方法以黄芩苷为内参物,采用XtimateTMC18(4.6 mm×250 mm,5μm)色谱柱,流动相为甲醇-0.2%磷酸水溶液,梯度洗脱,体积流量为1 mL/min,柱温30℃,检测波长274 nm。结果 2种方法测得的含有量无显著性差异,RSD<5%。结论 "一测多评"法可用于三黄片中4种黄芩黄酮类成分的质量评价。

中药材一测多评中相对校正因子的规律性文献调查和分析

目的：探寻相对校正因子（RCF）在中药材一测多评中应用的规律性。方法：以中国知网（CNKI）文献数据库为来源,采用文献检索方法收集大量有关中药材一测多评的文献,提取文献中通过一测多评法（QAMS）所得含量的计算值与外标法所得含量的实测值,并通过SPSS 19.0软件计算其计算值与实测值间相对标准偏差的百分值（RSD,%）,绘制散点图分析RSD与RCF的相关性。结果：RCF值在〉0.62-1.53区间分布较多,占全部数据的76.7%,且在此区间时,计算值与实测值的相似度较大,而RSD〈3.0%的数据占此区间全部数据的88.8%,可信度较好。与此同时,计算值与实测值的相似程度还受到其他待测成分与内参物之间母核结构和理化性质差异的影响,母核结构相似时,计算值与实测值的相似度较大;母核结构差异大时,计算值与实测值的相似度较小;而母核结构相似、理化性质差异大时,计算值与实测值的相似度也较小。结论：QAMS法中RCF会影响中药材待测成分含量测定结果的可信度,有必要建立中药材多成分QAMS法测定中RCF、RSD等相关参数数据库。

光度分析中协同系数补偿方法的研究──三次标准加入法同时测定钼、钨和钛

本文在两次标准加入法[1]的基础上，提出了三组份同时测定的三次标准加入法，并应用于钼、钨和钛的流动注射同时测定。测定条件：［水杨基荧光酮（SAF）］＝1．25 ×10-4mol／L，［溴化＋六烷基三甲基铵（CTMAB）」＝2．5×10－3mol／L，pH1．35，25℃，采样、进样时间分别为10s和18s，进样频率128次／h。为消除组份间协同作用引起的吸光度加合性的偏离，定义了三组份协同系数。将其引入三组份同时测定方法，补偿了吸光度对线性的偏离。补偿后钼、钨和钛的测定范围分别为0．05～0．35，0．25～1．00，0．025～0．30mg／L。测定了钢样及模拟样品中钼、钨和钛含量，结果较好。

通腑泻热灌肠液中大黄素及大黄酚含量测定

目的建立通腑泻热灌肠液的质量标准及活性成分含量测定方法。方法采用薄层色谱（TLC）法对通腑泻热灌肠液中所含大黄、栀子、忍冬藤进行定性鉴别;并用高效液相色谱（HPLC）法测定通腑泻热灌肠液中大黄的2个有效成分含量。结果薄层色谱鉴别方法能定性检出大黄、栀子、忍冬藤,斑点清晰,分离度好,专属性强,且阴性对照无干扰;大黄素、大黄酚分别在0.0726~0.726μg（r=0.9998）、0.0071~0.071μg（r=0.9999）范围内与峰面积均呈现良好的线性关系,平均加样回收率分别为99.13%、103.16%,RSD（n=6）分别为2.30%、1.50%。结论所建立的定性、定量分析方法可行,专属性强,准确可靠,能有效控制通腑泻热灌肠液的质量。

主成分线性回归分光光度法同时测定复方制剂中苯酚和间苯二酚

目的:对吸收峰严重重叠的苯酚和间苯二酚二组份体系同时测定方法进行研究.方法:在波长 250～290 nm 范围内,每隔 0.5 nm 测定苯酚和间苯二酚二组份体系的吸光度,建立了主成分线性回归法测定二组份.按正交设计表L25(56)配置25组标准混合溶液,对苯酚和间苯二酚二组份进行线性回归;同时测定了模拟混合样的苯酚和间苯二酚二组份含量.结果:苯酚和间苯二酚二组份在模拟混合样和模拟复方雷锁辛涂剂处方配比中个别误差稍大,绝大部分分析结果相对误差小于 5%.结论:主成分线性回归法测定苯酚和间苯二酚二组份,测量体系稳定,结果可靠,提供了一种新的测量途径.

一测多评法测定洋金花叶中醉茄内酯类成分的含量

目的:建立同时测定药材洋金花叶中多种醉茄内酯类成分的一测多评质量控制方法。方法:以Daturataturin A为内标物,考察不同色谱柱、流动相系统、HPLC仪器条件下,Daturataturin A与Baimantuoluoside B、daturametelin I之间的相对校正因子,实现一测多评,同时采用外标法测定3种醉茄内酯的含量,比较计算值与实测值的差异,验证该质量控制方法的准确性和可行性。结果:Baimantuoluoside B(BB)、daturametelin I(dI)与Daturataturin A(DA)间的相对校正因子分别为0.664,1.136,相对校正因子重现性良好。洋金花叶中醉茄内酯成分采用校正因子计算的含量值与外标法实测值之间无显著性差异。结论:该方法可用于洋金花叶醉茄内酯类成分的定量分析。