朝鲜淫羊藿的色谱指纹图谱及液相色谱/电喷雾串联质谱分析

建立了长白山区朝鲜淫羊藿药材的高效液相色谱指纹图谱的分析方法. 确定了18批朝鲜淫羊藿药材的13个共有峰, 该指纹图谱的精密度、稳定性和重现性的相对标准偏差均低于3.0%. 结合液相色谱/电喷雾串联质谱对特征峰进行了结构确认, 并根据电喷雾串联质谱数据推测了13个特征化合物的结构. 结果表明采用高效液相色谱与质谱联用技术对朝鲜淫羊藿色谱指纹图谱中的特征峰进行结构确认, 使其色谱指纹图谱的特征性更强, 更适合于药材质量的鉴别与评价.     

基于超高效液相色谱-质谱红花药材指纹图谱的建立

以红花为研究对象，建立超高效液相色谱-质谱（UPLC-MS）指纹图谱分析方法，为红花药材质量鉴别提供借鉴.通过考察检测波长、梯度洗脱条件、提取溶剂、提取方法等因素，在最优条件下对20个产地的红花样品进行分离分析.色谱柱：Thermo Hypersil Gold C₁₈，100 mm×2.1 mm，1.9 μm，流动相：0.1%甲酸水溶液（A）-乙腈（B）；流速：0.3 mL/min；柱温：30℃；检测波长：270 nm；进样量：2 μL，质谱条件：鞘气流速：45 arb，辅助气流速15 arb，雾化电压：3.3 KV，离子传输管温度：350℃，辅助气加热温度：400℃.试验结果：共确定了67个共有峰，通过UPLC-MS指认了其中11个共有峰，对20批不同产地的红花药材指纹图谱进行了相似度评价，其相似度在0.687~0.971之间.建立的UPLC-MS指纹图谱可为红花药材的质量控制提供参考依据.     

沉香的高效液相色谱-四极杆-飞行时间质谱数字化指纹图谱研究

采用高效液相色谱-四极杆-飞行时间质谱联用(HPLC-Q-TOF-MS)技术,研究构建了一种沉香数字化色谱-质谱指纹图谱的新方法。沉香药材经乙醇提取后,采用HPLC-Q-TOF-MS测定,并同时采集HPLC-Q-TOF-MS及液相色谱-紫外数据,得到液相色谱-紫外检测(HPLC-UV)色谱图和高分辨飞行时间质谱(TOF-MS)总离子流色谱图。对色谱图中的各个色谱峰进行精确质量数识别,建立数字化指纹图谱,以精确质量数结合保留时间表征沉香中的化学成分,即为每个色谱峰给出具有唯一性的数字信息,以数字化的形式反映其化学成分,并根据精确质量及同位素推算出分子式,结合二级质谱及文献资料共鉴定出30个化学成分。该方法对沉香的每种化学成分给出了类似于身份认定的数字化信息,具有唯一性,能全面反映沉香的物质成分,可为沉香的药理、药效及质量标准研究提供科学的数据。     

丹参水溶性成分的电喷雾质谱行为及其特征图谱的初步研究

应用电喷雾-质谱(ESI-MS)技术研究了原儿茶醛和丹参素的ESI-MS规律,结合ESI-MS技术,对9种丹参水溶性成分的质谱行为进行了研究,并对其中5种成分进行了定性分析。同时利用选择性离子检测质谱技术,初步建立了丹参水溶性成分SIM-MS特征图谱,该图谱简单、易于解析,重现性与特征性较好,可以用于丹参药材及复方丹参的快速指纹鉴别。     

山麦冬、麦冬对照药材电喷雾-质谱(ESI-MS)特征图谱的比较研究

报道了采用电喷雾-质谱负离子全扫描法,分析山麦冬、麦冬对照药材的甲醇提取物。经分析发现,山麦冬、麦冬对照药材负离子的全扫描质谱图差异显著,从中选择16强峰建立山麦冬、麦冬对照药材甲醇提取物的特征图谱。研究表明,该分析方法有较好的重现性,图谱特征性强,可快速、准确鉴别山麦冬、麦冬药材。     

栀子药材的指纹图谱整体性分析

建立了药材栀子水提部分的高效液相色谱指纹图谱,脂溶性成分和挥发油的气相色谱．质谱联用指纹图谱,并分别计算了此3部分指纹图谱的相似系数和总体相似系数,同产地结果RSD≤5％。通过比较不同产地栀子相似系数的关系,以及对9个主要药效成分定量结果的分析,对不同产地栀子的指纹图谱做了整体性评价,结果表明,只有全面测定成分及整体性评价后,药材的质量才能得到有效评价。     

黄芪药材的指纹图谱研究方法的建立

用反相高效液相色谱－紫外－质谱联用技术对黄芪药材进行指纹图谱研究，为阐明不同产地药材的异同性，建立黄芪质量的国际统一标准奠定基础，黄芪的总提取物各类成分得到很好分离，紫外和质谱两种检测器可对不同特性化合物的检测进行互补，获得相对充分的指纹图谱信息。     

质谱差谱方法及其在中药复方研究中的应用

采用HPLC-ESI-MSn分析得到了复方合煎液中有效部分提取物的指纹图谱数据, 通过比对找出对复方指纹谱有贡献的有效成分峰. 再用质谱差谱方法处理多级质谱数据, 提取不同类型的中性丢失, 实现了对复方有效成分化合物的快速分类鉴定. 用该法研究了复方柴苓汤有效部分黄芩总黄酮和柴胡总皂苷.     

基于选择离子集成谱的中药质量评价新方法

采用液相色谱-质谱(LC-MS)联用技术建立了不同产地的双丹方(丹参和丹皮)煎剂的指纹图谱, 并提出一种将二维的LC-MS数据转化为一维的选择离子集成谱(ISIC)的方法. 通过主成分分析考察了ISIC与总离子流色谱图(TIC)和保留时间校正后的TIC对不同产地药材的区分能力. 结果表明, ISIC能全面反映不同产地的丹参和丹皮成分间的差异; 而TIC由于色谱峰重叠严重, 对不同产地的药材无区分能力, 保留时间校正后的TIC仅能反映不同产地丹参成分间的差异. 该方法运算速度快, 适于大批量样本分析, 生成的ISIC信息量大, 且不需校正保留时间, 可直接用于后续的统计分析.     

邓老凉茶颗粒的超高效液相色谱质谱联用指纹图谱研究

建立了适用于邓老凉茶颗粒质量控制的超高效液相色谱质谱联用指纹图谱分析方法。样品采用甲醇索氏萃取60 min,萃取液采用超高效液相色谱质谱法进行指纹图谱分析。色谱柱采用Waters ACQUITY HSST3 C18(150 mm×3.0 mm,1.8μm),以0.5%甲酸-乙腈为流动相进行梯度洗脱,流速为0.8 mL/min,柱温35℃。质谱采用负离子ESI模式,选择基峰离子流质量色谱图进行指纹图谱研究。32个共有峰在15 min内得到良好分离,其中15个共有峰通过对照品进行了确证。通过《中药色谱指纹图谱相似度评价系统2004A版》对邓老凉茶颗粒样品进行相似度分析,15个批次样品的相似度均达到0.960以上,表明邓老凉茶颗粒的产品质量稳定性很好。以32个共有峰的相对峰面积进行主成分分析,邓老凉茶颗粒样品之间的细微质量差异得到明显区分。该方法快速、高效、可靠,可有效地用于邓老凉茶颗粒的质量控制。     

Quality evaluation of cortex moutan by high performance liquid chromatography coupled with diode array detector and electrospary ionization tandem mass spectrometry

An high performance liquid chromatography (HPLC) coupled with diode array detector (DAD) and electrospray ionization tandem mass spectrometry (ESI/MS(n)) method was developed for quality evaluation of Cortex Moutan through identification of common constituents based on chromatographic fingerprints and determination of key pharmacological compounds. The representative chromatographic fingerprints of Cortex Moutan were obtained by analyzing 10 batches of samples under the optimized HPLC conditions and the results showed that the chromatographic profiles of the analyzed samples were very similar. Total of nineteen common peaks were detected and seventeen of them were identified rapidly by their characteristic UV profile and the information of molecular structure provided by ESI/MS(n) experiments. Simultaneously, five key pharmacological compounds, namely gallic acid, oxypaeoniflorin, paeoniflorin, benzoylpaeoniflorin and paeonol, were determined by the validated HPLC-DAD method. The linear calibration curves were acquired with correlation coefficients higher than 0.999. The precisions of intra-day and inter-day were not exceeding 3.1%, and the recoveries of five analytes were from 92.86 to 99.35%. This developed method that combined the chromatographic fingerprints and quantification assay ensured the phytoequivalence and pharmacological effects of Cortex Moutan and was successfully applied to the quality control of Cortex Moutan.     

Determination of moutan tannins by high‐performance liquid chromatography and capillary electrophoresis

Cortex Moutan (Radicis Cortex Moutan), the dried root bark of Paeonia moutan and P. spp., contains a series of water‐soluble tannins. With the eight components, 1 4,6‐di‐O‐GG (4,6‐di‐O‐galloyl‐D‐glucose), 2 1,2,3,6‐tetra‐O‐GG, 3 1,2,3,4,6‐penta‐O‐GG, 4 1,3,4,6‐tetra‐O‐GG, 5 3,4,6‐tri‐O‐GG, 6 1,3,6‐tri‐O‐GG, 7 3,6‐di‐O‐GG, and 8 1,2,6‐tri‐O‐GG, as marker substances, a rapid and efficient method of analysis based on HPLC and CE was developed. Using a phosphate eluent, a 5C18‐MS separating column, and a detection wavelength of 280 nm, HPLC was successfully used to analyze the eight constituents within 60 min. The analysis can be completed within 50 min, using the MEKC mode with a buffer composed of borate, SDS, and isopropanol, and a detection wavelength of 210 nm. The detection limit for the marker substances varied from 0.04 to 0.93 μg/mL for the HPLC method and 0.02 to 0.36 μg/mL for the CE method.     

Simultaneous determination of wogonin,oroxylin a,schisandrin, paeoniflorin and emodin in rat serum by HPLC–MS/MS and application to pharmacokinetic studies

Wogonin and oroxylin A in Scutellariae Radix, schisandrin in Chinensis Fructus, paeoniflorin in Moutan Cortex and emodin in Polygoni Cuspidate Rhizome et Radix are anti‐inflammatory active compounds. A method for simultaneous determination of the five compounds in rat was developed and validated using high‐performance liquid chromatography with tandem mass spectrometry (HPLC–MS/MS). The separation was performed on a Symmetry C₁₈ column (4.6 × 50 mm, 3.5 μm) with acetonitrile and 0.1% formic acid aqueous solution as the mobile phases. The detection was performed using multiple‐reaction monitoring with electrospray ionization source in positive–negative ion mode. The calibration curves showed good linearity (r ≥ 0.9955). The lower limit of quantification (LLOQ) was 5 ng/mL for wogonin and schisandrin, 10 ng/mL for oroxylin A and emodin, and 15 ng/mL for paeoniflorin, respectively. The relative standard deviations of intraday and interday precisions were <11.49 and 14.28%, respectively. The extraction recoveries and matrix effects were acceptable. The analytes were stable under the experiment conditions. The validated method has been successfully applied to pharmacokinetic studies of the five compounds in rats after oral administration of Hu‐gan‐kan‐kang‐yuan capsule. This paper would be a valuable reference for pharmacokinetic studies of Chinese medicine preparations containing the five compounds.     

加压毛细管电色谱在建立中药黄柏指纹图谱中的方法学研究

以加压毛细管电色谱（pCEC）为技术平台,对其在建立中药黄柏指纹图谱中的方法学进行了研究．通过对提取溶剂、流动相中有机相种类、盐溶液等条件的优化,发现1％盐酸的甲醇溶液为提取溶液,20mmol／LNH4C1溶液．乙腈为流动相梯度洗脱,紫外检测波长为230nm时对其分离效果最好．并通过在色谱柱上施加不同的电压,详细地阐明了pCEC的双重分离机制对分离选择性的影响,发现黄柏中的主要成分药根碱、巴马汀和小檗碱在pCEC模式中随电压的不同,有不同的出峰顺序．当电压为0—4kV时出峰顺序为药根碱、巴马汀和小檗碱,当电压为8—14kV时出峰顺序为药根碱、小檗碱和巴马汀．对此原因进行了详细讨论,同时与微径液相色谱模式进行了比较,说明pCEC可以为复杂样品的分离提供更多更好的分离途径．     

Nine absorbed components pharmacokinetic of raw and processed Moutan Cortex in normal and blood-heat and hemorrhage syndrome model rats

Raw Moutan Cortex (RMC) and Processed Moutan Cortex (PMC) have a long history of use in China and other Asian countries. In this study, a rapid and accurate ultra-high-pressure liquid chromatography coupled with diode array detector (UHPLC–DAD) method was developed and validated for the simultaneous determination of nine absorbed compounds of RMC/PMC. After extraction by protein precipitation with methanol from plasma, the analytes were separated on an Acquity UPLC® BEH Shield RP₁₈ column (2.1 × 100 mm, 1.7 μm, Waters, USA). Acetonitrile (A) and 0.1% (v/v) formic acid in water (B) were selected as the mobile phase to perform gradient elution. The linearity of nine analytes was >0.9915. The intra- and inter-assay precision (RSD) values were within 11.18%, and accuracy ranged from 91.32 to 101.29%. Suitable stability, matrix effect and extraction recoveries were also obtained. The validated method was applied to compare the pharmacokinetics of RMC and PMC in Blood-Heat and Hemorrhage Syndrome Model and normal rats. The results revealed that processing and the pathological state could influence the pharmacokinetic characteristics of compounds in RMC/PMC. The study willbe useful for further studies on pharmacokinetics and clinical application of raw and processed Moutan Cortex.     

Quality assessment of cortex cinnamomi by HPLC chemical fingerprint, principle component analysis and cluster analysis

HPLC fingerprint analysis, principle component analysis (PCA), and cluster analysis were introduced for quality assessment of Cortex cinnamomi (CC). The fingerprint of CC was developed and validated by analyzing 30 samples of CC from different species and geographic locations. Seventeen chromatographic peaks were selected as characteristic peaks and their relative peak areas (RPA) were calculated for quantitative expression of the HPLC fingerprints. The correlation coefficients of similarity in chromatograms were higher than 0.95 for the same species while much lower than 0.6 for different species. Besides, two principal components (PCs) have been extracted by PCA. PC1 separated Cinnamomum cassia from other species, capturing 56.75% of variance while PC2 contributed for their further separation, capturing 19.08% variance. The scores of the samples showed that the samples could be clustered reasonably into different groups corresponding to different species and different regions. The scores and loading plots together revealed different chemical properties of each group clearly. The cluster analysis confirmed the results of PCA analysis. Therefore, HPLC fingerprint in combination with chemometric techniques provide a very flexible and reliable method for quality assessment of traditional Chinese medicines.     

Multi-component HPLC fingerprinting of Radix Salviae Miltiorrhizae and its LC-MS-MS identification

The multi-component fingerprinting method of Radix Salviae Miltiorrhizae (Root of Salvia miltiorrhiza BGE.), an important and popular medicinal herb in traditional Chinese medicine, was studied using reverse-phase HPLC and LC-MS-MS. Extract containing both the water-soluble phenolic compounds and nonpolar diterpenoid compounds known to be the herb's main bioactive components was prepared by a two-step extractive procedure. An HPLC fingerprinting method which can simultaneously separate these two types of compounds was established with gradient elution mode and photodiode array detection at 280 nm. Eighteen peaks in this HPLC fingerprint were structurally identified by employing LC-MS-MS techniques. The electrospray ionization (ESI) MS-MS spectra of most salvianolic acids displayed a characteristic behavior of loss of danshensu and caffeic acid moieties, while those of tanshinones showed a particular behavior of loss of H2O which is quite different from the fragmentation pattern in electron ionization mass spectrometry (EI-MS). The HPLC fingerprints of 7 batches of crude drugs showed similar separation pattern and provided much chemical information of the pharmacologically-active compounds in the crude drugs, which is useful for the authentication and quality evaluation of this medicinal herb.     

基于标准制剂控制模式和定量指纹图谱评价复方甘草片的质量一致性

通过建立复方甘草片标准制剂（SP）控制模式和定量高效液相色谱指纹图谱,结合5个质量标志物的精准定量评价了9个厂家共145批复方甘草片质量一致性。首先建立了复方甘草片标准制剂的标准指纹图谱（SP-RFP）,然后以SP-RFP作为评价标准,采用系统指纹定量法对145批复方甘草片进行整体定性和整体定量评价。用双标校正法校正定量指纹图谱的系统误差,结果表明所检样品质量均合格。此外,在统一化色谱条件下测定各原料药和模拟样品,对制剂指纹进行归属相关度和准确度评判,得到原料指纹与制剂指纹的相关性,从而实现智能预测制剂或原料药质量和阻止低劣原料入药。同时用紫外全指纹溶出度法测定5个厂家的复方甘草片的溶出度曲线,用以评价制剂工艺的合理性。以上方法可行且准确度高,实现了对复方甘草片质量和工艺的一致性评价。该文为中药质量一致性评价提供了基础评价模式和基本操作思路以及具体实例。     

A simple and effective method for identification of Fraxini Cortex from different sources by multi-mode fingerprint combined with chemometrics

Fraxini Cortex has a long history of being used as a medicinal plant in traditional Chinese medicine. However, it is challenging to differentiate and make quality evaluations for Fraxini Cortex from different origins due to their similarities in morphological features, as well as general chemical composition using traditional chemical analytical methods. In this study, a simple and effective method was developed to identify Fraxini Cortex from different origins by multi-mode fingerprint combined with chemometrics. Digital images of the high-performance thin-layer chromatography profiles were converted to grayscale intensity, and the common patterns of high-performance thin-layer chromatography fingerprints were generated with ChemPattern software. Authentication and quality assessment were analyzed by similarity analysis, hierarchical cluster analysis, principal component analysis, and multivariate analysis of variance. The ultra-high-performance liquid chromatography fingerprints were analyzed by similarity analysis, principal component analysis, and orthogonal partial least square-discriminant analysis. When combined with chemometrics, high-performance thin-layer chromatography and ultra-high-performance liquid chromatography fingerprint provided a simple and effective method to evaluate the comprehensive quality of Fraxini Cortex, and to distinguish its two original medicinal materials (Fraxinus chinensis Roxb. and Fraxinus rhynchophylla Hance.) recorded in the Chinese Pharmacopeia and its three adulterants (Fraxinus mandschurica Rupr., Fraxinus pennsylvanica Marsh., and Juglans mandshurica Maxim.). A similar workflow may be applied to establish a differentiation method for other medicinal and economic plants.