Chromatographic fingerprinting and metabolomics for quality control of TCM

Chromatographic fingerprinting technique of traditional Chinese medicine (TCM) has proved to be a comprehensive strategy for assessing the intact quality of herbal medicine. In general, one could use the chromatographic techniques to obtain a relatively complete picture of herbal medicines, which are in common called chromatographic fingerprints of herbal medicines to represent the so-called phytoequivalence. Based on this, the features of chromatographic fingerprints of herbal medicines have been discussed in some detail. The technique based on chromatographic fingerprinting is essentially a kind of high-throughput and integral tools to explore the complexity of herbal medicines. In order to further control the comprehensive quality of TCMs, some new strategies are proposed to trace the chemical changes of chromatographic fingerprints both in product processing and/or after their administration by modern chromatographic techniques and chemometrics. Combined with metabolomics, it seems possible for one to reveal the working mechanism of TCMs and to further control their intrinsic quality. Finally, the intensive study of chromatographic fingerprinting coupled with multivariate analysis tools developed in bioinformatics and chemometrics are emphasized in order to achieve the aim to reveal the working mechanisms of TCMs and to further control and strengthen TCMs' intrinsic quality in a comprehensive manner.     

Chemometrics and modernization of traditional Chinese medicine

Development of chromatographic fingerprinting and its related chemometric methods in the research of quality control of traditional Chinese medicines(TCMs) are discussed. The quality control methods for guarantying the authentication and stability of products and semi-products of TCMs are firstly assessed. The technique based on chromatographic fingerprinting is essentially a kind of high-through put and integral tools to explore the complexity of herbal medicines. In order to further control the comprehensive quality of TCMs,confirmation and identification of their important chemical components are necessary. Some new strategies are proposed to trace the chemical changes of chromatographic fingerprints both in product processing and/or after their administration by modern chromatographic techniques and chemometrics. Combined with systems biology and bioinformatics,it seems possible for one to reveal the working mechanism of TCMs and to further control their intrinsic quality comprehensively.     

色谱指纹图谱与中药质量控制

本文综述了近年来色谱指纹图谱研究中多种色谱技术、相关化学计量学方法的发展及其在中药质量控制中的应用,并对中药质量稳定性和有效性的全面评价方法和可行性进行了初步探讨。提出以色谱指纹图谱为核心,依托现代色谱技术、化学计量学和系统生物学方法,建立中药化学成分信息与药效(毒性)信息相关关系的中药质量控制研究策略,深层次揭示中药的内在作用机理,期望真正实现中药质量的可控、安全和有效。     

中药分析新思路及其质量控制

随着国家对中药现代化的重视,为了更好地诠释中药药性的本质,研究者从不同角度对中药进行研究。分析化学、药学、生物学等多学科的多种不同的技术在这里融合,一些新的思路和方法正在不断涌现,中药的分析与质量控制正沿着综合且全面的方向在不断地发展和前进。该文对当前中药分析新思路及其质量控制进行了简要评述,在此基础上,对中药质量控制的色谱指纹图谱进行了分类说明与研究,表明采用现代分析手段,可以实现对中药质量的有效控制。     

Qualitative and quantitative analysis in quality control of traditional Chinese medicines

Separation techniques with high efficiency and sensitive detection have been widely used for quality control of traditional Chinese medicines (TCMs). High-performance liquid chromatography, gas chromatography, and capillary electrophoresis are commonly used to separate various components in TCMs. Ultraviolet detection, fluorescence detection, evaporative light-scattering detection, mass spectrometry and nuclear magnetic resonance can be applied to separation techniques for qualitative and quantitative analysis of TCMs. The development of quality control for TCMs based on quantitative and qualitative analysis from 2000 to 2007 are reviewed; the fingerprint technique is also discussed due to its broad application in the quality control of TCMs. Prospects for further research based on our primary results are also discussed.     

Solidification of floating organic drop liquid‐phase microextraction cell fishing with gas chromatography–mass spectrometry for screening bioactive components from Amomum villosum Lour

In this study, a novel solidification of floating organic drop liquid‐phase microextraction cell fishing with gas chromatography–mass spectrometry (SFOD‐LPME‐CF‐GC‐MS) method was established and used to screen, isolate and analyze bioactive components from Amomum villosum Lour. extract. Through comparision of its effect on the models of normal cell and inflammatory cells, anti‐inflammatory active components of essential oil from A. villosum Lour. were readily screened, and the components obtained are in agreement with related pharmacological articles. SFOD‐LPME‐CF‐GC‐MS was used to analyze the interaction of A. villosum Lour. extracts with normal and lipopolysaccharide‐stimulated RAW264.7 macrophage cells. The effect of A. villosum Lour. essential oil extracts in the LPS‐stimulated RAW264.7 model were also assessed in terms of cytotoxicity and nitric oxide production as an indication of bioactivity. Three potentially bioactive components were identified, demonstrating that SFOD‐LPME‐CF‐GC‐MS can be used successfully in the drug‐screening process. This approach avoids the requirement for protein precipitation, but more importantly, generates a high concentration ratio, allowing analysis of trace components in traditional Chinese medicines. SFOD‐LPME‐CF‐GC‐MS is a simple, fast, effective and reliable method for the screening and analysis of bioactive components, and it can be extended to screen other bioactive components from TCMs. Copyright © 2014 John Wiley & Sons, Ltd.     

Multi‐component determination and chemometric analysis of Paris polyphylla by ultra high performance liquid chromatography with photodiode array detection

Multi‐source analysis of traditional Chinese medicine is key to ensuring its safety and efficacy. Compared with traditional experimental differentiation, chemometric analysis is a simpler strategy to identify traditional Chinese medicines. Multi‐component analysis plays an increasingly vital role in the quality control of traditional Chinese medicines. A novel strategy, based on chemometric analysis and quantitative analysis of multiple components, was proposed to easily and effectively control the quality of traditional Chinese medicines such as Chonglou. Ultra high performance liquid chromatography was more convenient and efficient. Five species of Chonglou were distinguished by chemometric analysis and nine saponins, including Chonglou saponins I, II, V, VI, VII, D, and H, as well as dioscin and gracillin, were determined in 18 min. The method is feasible and credible, and enables to improve quality control of traditional Chinese medicines and natural products.     

中药物质基础的高效液相色谱分离分析方法研究

高效液相色谱方法已成为中药物质基础研究的重要手段．在中药质量控制、中药标准品制备、活性化合物发现等方面发挥着不可替代的作用．然而我们的实验数据表明,一个中药材可能包含上万个化合物．中药物质基础的复杂性对高效液相色谱方法提出了巨大挑战．本文针对液相色谱方法在中药物质基础分离分析中存在的问题与难点,结合红花、黄连、姜黄三味药材样品,从亲水色谱分离模式、新型色谱固定相、二维液相色谱分离系统和液相色谱质谱联用技术等方面讨论了液相色谱的分离分析方法和发展方向．结果表明,高效液相色谱新技术新方法在中药复杂体系的分离分析中具有很大的发展潜力和应用前景．     

Fingerprint analysis of Ligusticum chuanxiong using hydrophilic interaction chromatography and reversed-phase liquid chromatography

Fingerprint analysis is considered one of the most powerful approaches to quality control in traditional Chinese medicines (TCMs). In this study, a binary chromatographic fingerprint analysis was developed using hydrophilic interaction chromatography (HILIC) and reversed-phase liquid chromatography (RPLC) to gain more chemical information about polar compounds and weakly polar compounds. This method was used to construct a chromatographic fingerprint of Ligusticum chuanxiong. The two chromatographic methods demonstrated good precision, reproducibility, and stability, with relative standard deviations of <2% for retention time and 7% for peak area for both HILIC and RPLC separations. Data from the analysis of 14 samples by HILIC and RPLC were processed with similarity analysis, with correlation coefficients and congruence coefficients. This binary fingerprint analysis, using two chromatographic modes, is a powerful tool for characterizing the quality of samples, and can be used for the comprehensive quality control of TCMs.     

Recent analytical approaches in quality control of traditional Chinese medicines—A review

Traditional Chinese medicines (TCMs) are gaining more and more attention all over the world, due to their specific theory and long historical clinical practice. But the uncontrollable quality is a bottleneck for its modernization and globalization. This paper reviewed the recent analytical methods in the quality control of TCMs, including screening strategies of bioactive markers from TCMs through biochromatographic methods, the traditional chromatographic methods, DNA methods, as well as the spectroscopic methods, including FT-IR, NIR and NMR. The comprehensive methods, such as fingerprint and multi-component quantification are emphasized; hyphenated techniques, like HPLC-MS, GC-MS, CE-MS, LC-NMR, chemometric methods, and combination of chemical and biological methods, such as biofingerprint, metabolic fingerprint are now more and more widely used in TCMs. In a few word, the analysis and quality control of TCMs are moving towards an integrative and comprehensive direction, in order to better address the inherent holistic nature of TCMs.     

Recent applications of capillary electrophoresis in the analysis of traditional Chinese medicine

Traditional Chinese medicines (TCMs) are generally mixtures of herbal plants or extracts which comprise hundreds of different constituents with widely difference in the content and physiochemical properties. In order to analyze bioactive compounds in TCMs and control the quality, a large number of analytical tools have been developed, among which capillary electrophoresis (CE) has become a powerful technique with increasing importance. Some formats of CE, including capillary zone electrophoresis, micellar electrokinetic chromatography, non-aqueous CE and capillary electrochromatography have been widely employed for the analysis of TCMs. The general characteristics of these formats are briefly described, and their applications to the analysis of TCMs during the past five years are summarized.     

中药质量控制技术发展展望

本文从中药产业需求、现代化需求、技术需求、机遇与挑战等方面概括了中药质量控制技术发展的背景;讨论了中药质量控制技术对于提高中药药效和安全性、推动产业发展和推进中药国际化的意义;综述了中药质量控制技术的现状,分析了在过程控制、安全性控制、标准品和对照品制备、指纹图谱技术等方面的不足;提出了中药质量控制技术应重点发展以分离和表征技术为主的中药质量控制关键技术、中药安全性控制技术、中药质量控制标准体系、中药质量控制原创性技术和中药标准品、对照品生产技术,制定技术标准,建立具有中药特色的过程控制和产品质量控制标准。     

Screening and quantifying the quality markers of DuHuo by fingerprint modeling

Abstract

With environmental health and safety becoming a concern for people, supercritical fluid chromatography has been regarded as a “green chemistry” method that gradually has peaked people’s interest. The purpose of this study was to establish a method to screen and quantify the quality markers of Duhuo (DH) through an environmentally friendly ultra-performance convergence chromatography (UPC²) method. Firstly, the fingerprint of different batches of DH samples was established by UPC². Then, the quality markers that can represent the whole information of DH was screened through chemometrics. Finally, the content of the selected quality markers was determined using the UPC² method. From the fingerprint analysis, 24 common peaks were identified. The quantitative analysis results showed that the four compounds could be used as quality markers to represent the overall information and to monitor the quality of DH. This is the first time a traditional Chinese medicine (TCM) fingerprint has been established by UPC² and also the first time to screen the quality markers in DH. The method established in this study can be used for a comprehensive and overall analysis of DH or other natural medicines, and it could serve as an efficient method to contribute to the expansion of the global market for TCMs.     

Analysis of global components in Ganoderma using liquid chromatography system with multiple columns and detectors

In present study, a multiple columns and detectors liquid chromatography system for analysis of global components in traditional Chinese medicines was developed. The liquid chromatography system was consist of three columns, including size exclusion chromatography column, hydrophilic interaction chromatography column, and reversed phase chromato‐graphy column, and three detectors, such as diode array detector, evaporative light scattering detector, and mass spectrometry detector, based on column switching technique. The developed multiple columns and detectors liquid chromatography system was successfully applied to the analysis of global components, including macromolecular (polysaccharides), high (nucleosides and sugars)‐, and low (triterpenes)‐polarity small molecular compounds in Ganoderma, a well‐known Chinese medicinal mushroom. As a result, one macromolecular chromatographic peak was found in two Ganoderma species, 19 components were identified in Ganoderma lucidum (two sugars, three nucleosides, and 14 triterpenes), and four components (two sugars and two nucleosides) were identified in Ganoderma sinense. The developed multiple columns and detectors liquid chromatography system was helpful to understand comprehensive chemical characters in TCMs.     

Chemical constituents and quality control of two Dracocephalum species based on high‐performance liquid chromatographic fingerprints coupled with tandem mass spectrometry and chemometrics

Two similar Dracocephalum species, namely, Dracocephalum tanguticum Maxim and Dracocephalum moldavica L, are commonly used as ethnic medicines in China. Here we describe a strategy of combining high‐performance liquid chromatography with quadrupole time‐of‐flight mass spectrometry, as well as fingerprints and chemometrics for characterization and discrimination of chemical constituents on the two herbs. A total of 49 compounds including 33 flavonoids, 5 phenylethanol glycosides, 1 coumarin glycoside, 8 organic acids, and 2 other types of compounds were unambiguously or tentatively identified from the two Dracocephalum species. Among the compounds identified, 26 were characterized for the first time and 4 compounds, rosmarinic acid ( 7 ), salvianolic acid B ( 10 ), luteoloside ( 22 ), diosmetin‐7‐O‐glucoside ( 28 ), were inferred as common constituents for the two herbs. Flavonoids featured in these two Dracocephalum species while their types presented significant differences. Acacetin ( 45 ) and acacetin glycosides (acatetin‐7‐O‐glucuronide ( 30 ), acacetin‐7‐O‐(6”‐O‐malonyl) glucoside ( 33 ), buddleoside ( 34 ), tilianin ( 35 ), and agastachoside ( 42 )) were detected only in D. moldavica, which can be used to discriminate two herbal medicines. In addition, six characteristic constitutes in D. tanguticum were simultaneously quantified. Moreover, the induced chemometrics methods including similarity analysis and hierarchical clustering analysis were successfully applied for origin discrimination and quality evaluation of D. tanguticum and D. moldavica.     

Development of microwave-assisted extraction followed by headspace single-drop microextraction for fast determination of paeonol in traditional Chinese medicines

Paeonol is an important active component present in traditional Chinese medicines (TCMs), which was used for the treatment of many diseases such as eczema. In this work, microwave-assisted extraction (MAE) was firstly combined with headspace single-drop microextraction (HS-SDME), and applied to rapid determination of paeonol in two TCMs of Cynanchum paniculatum and Paeonia suffruticosa. In the proposed method, paeonol in TCMs was isolated by using MAE, followed by extraction and concentration by HS-SDME, and detected by gas chromatography-mass spectrometry (GC-MS). The experiment parameters of MAE and HS-SDME were discussed, and the method precision, recovery and detection limit were also studied. To further demonstrate the reliability of the quantification, both the proposed method and a standard method of steam distillation (SD) were simultaneously applied to quantitative analysis of paeonol in TCM samples from different growing areas. The experimental results show that MAE-HS-SDME is a simple and rapid method for the quantitative analysis of paeonol in TCMs, and is also a potential and alternative tool for quality monitoring for the two TCMs of C. paniculatum and P. suffruticosa.     

An Expeditious Method Combining Multi-Components Determination and Fingerprinting Based on Ultra Performance Liquid Chromatography-Photo Diode Array-Tandem Mass Spectrometry and Chemometrics: Application for Authentication and Quality Evaluation of Cornus Officinalis Sieb. et Zucc

Selective and efficient analytical methods for authentication and quality evaluation of herbal medicines are significant and necessary. An expeditious method combining multi-components determination and fingerprinting based on ultra-performance liquid chromatography-photo diode array-tandem mass spectrometry (UPLC-PDA -MS/MS) and chemometrics for authentication and quality evaluation of Cornus Officinalis Sieb. et Zucc was developed. Tandem mass spectrometer operating in multiple reaction monitoring (MRM) was used for determination of three characteristic constituents (morroniside, sweroside, and loganin) in C. Officinalis. Meanwhile, UPLC fingerprint of C. Officinalis was established and the data set was submitted for classification to a suite of chemometrics method. Combing main biologically active components content level and chemometrics analysis, the effects of cultivation area, harvesting, and storage time on the quality of C. Officinalis were investigated. The study reveals that multi-components determination coupled with fingerprinting could be applied for authentication and quality evaluation of C. Officinalis which is accurate, efficient, and reliable.     

Micellar electrokinetic chromatography fingerprinting combined with chemometrics as an efficient strategy for evaluating the quality consistency and predicting the antioxidant activity of Lianqiao Baidu pills

An approach combining micellar electrokinetic chromatography fingerprinting with chemometrics was developed to evaluate the quality consistency of Lianqiao Baidu pills, which are traditional Chinese patent medicines composed of 19 herbs used mainly to treat skin ulcers, common cold, rheumatism, herpes, and constipation. The triangle optimization method was employed to choose a satisfactory background electrolyte, with the information index, I , as an objective function for assessing the capillary electrophoresis conditions. Then, under the optimal conditions, the micellar electrokinetic chromatography fingerprints of 28 batches of samples were established, and five marker compounds were quantitatively determined simultaneously. A limited‐ratio quantified fingerprint method was introduced to evaluate the chromatographic fingerprints both qualitatively and quantitatively. Principle component analysis revealed that the 28 batches of samples can be clustered according to different manufacturers. Moreover, the relationship between the fingerprint and the antioxidant activity was explored by orthogonal partial least‐squares regression, which provided critical medicinal efficacy information for quality control. The present study establishes a powerful and reliable method for monitoring the quality consistency of Lianqiao Baidu pill.     

色谱及相关技术在中药质量评价中的应用

对中药物质基础的研究方法——化学物质组学,以及中药质量评价的关键技术——指纹图谱技术做了介绍,并在此基础上综述了中药化学信息获取的关键技术——色谱及相关技术在中药质量评价中的应用。论述的结果表明,以化学物质组学为指导,结合色谱及相关技术获取指纹图谱信息及多指标成分定量信息的研究模式,对中药质量评价体系的建立具有指导意义。     

Efficient purification of high‐purity compounds from the stem of Lonicera japonica Thunb using two‐dimensional preparative chromatography

Purification of high‐purity compounds from traditional Chinese medicines (TCMs) plays an important role in investigating their bioactivity. Nevertheless, it is often quite difficult to isolate compounds with high purity because of the complexity of TCMs in chemical composition. In this work, a two‐dimensional preparation method was successfully developed for the preparation of high‐purity compounds from the stem of Lonicera japonica Thunb, based on two novel polar copolymerized RP stationary phases, XAqua C3 and XAqua C18. An XAqua C3 prep column was used to separate the sample in the first‐dimensional preparation, and 14 g of sample was fractionated into eight fractions with a recovery of 82%. An XAqua C18 prep column was selected to prepare high‐purity compounds in the second‐dimensional preparation for its good orthogonality with the XAqua C3 stationary phase. As a result, major compounds in the sample were isolated with more than 99% purity. This method is a potent method to realize the efficient purification of compounds with high purity from the stem of L. japonica Thunb and it shows great potential in the separation of high‐purity compounds from complex samples.