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AIM To establish an UPLC-MS/MS method for simultaneous content determination of protocatechuic acid,epicatechin,chlorogenic acid,quercitrin,gaultherin and gaultheroside A in Gaultheria leucocarpa var.yunnanensis.METHODS The analysis was performed on a 40℃thermostatic Waters BEH C18 column(100 mm×2.1 mm,1.7 μm),with the mobile phase comprising of water(containing 0.1%formic acid)-acetonitrile(containing 0.1%formic acid)flowing at 0.3 mL/min in a gradient elution manner,and electron spray inoization source was adopted in positive and negative ion scanning with multiple reaction monitoring(MRM)mode.Hierarchical cluster analysis(HCA)and principal component analysis(PCA)was used to screen important components that affect the quality of medicinal materials.RESULTS Six constituents showed good linear relationships within their own ranges(R2≥0.998 2),whose average recoveries were 98.76%-101.88%with the RSDs of 1.0%-2.5%.The constituents of G.leucocarpa in the roots and aerial parts were quite different.Gaultherin,epicatechin and protocatechuic acid may be the quality mark constituents of G.leucocarpa.CONCLUSION This accurate and efficient method can be used for the quality control of G.leucocarpa.
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ObjectiveTo establish the identification method of Dalbergiae Odoriferae Lignum（DOL） and its counterfeits by nuclear magnetic resonance hydrogen spectrum（1H-NMR） combined with multivariate statistical analysis. Method1H-NMR spectra of DOL and its counterfeits were obtained by NMR， and the full composition information was established and transformed into a data matrix， and the detection conditions were as follows：taking dimethyl sulfoxide-d6（DMSO-d6） containing 0.03% tetramethylsilane（TMS） as the solvent， the constant temperature at 298 K（1 K=-272.15 ℃）， pulse interval of 1.00 s， spectrum width of 12 019.23 Hz， the scanning number of 16 times， and the sampling time of 1.08 s. Similarity examination and hierarchical cluster analysis（HCA） were performed on the data matrix of DOL and its counterfeits， and orthogonal partial least squares-discriminant analysis（OPLS-DA） was used to analyze the data matrix and identify the differential components between them. In the established OPLS-DA category variable value model， the category variable value of DOL was set as 1， and the category variable value of the counterfeits was set as 0， and the threshold was set as ±0.3， in order to identify the commercially available DOL. The OPLS-DA score plot was used to determine the types of counterfeits in commercially available DOL， and it was verified by thin layer chromatography（TLC）. ResultThe results of similarity analysis and HCA showed that there was a significant difference between DOL and its counterfeits. OPLS-DA found that the differential component between DOL and its counterfeits was trans-nerolidol. The established category variable value model could successfully identify the authenticity of the commercially available DOL. The results of the OPLS-DA score plot showed that there were heartwood of Dalbergia pinnata and D. cochinchinensis in the commercially available DOL， and were consistent with the TLC verification results. ConclusionThere is a phenomenon that heartwood of D. pinnata and D. cochinchinensis are sold as DOL in the market. 1H-NMR combined with multivariate statistical analysis can effectively distinguish DOL and its counterfeits， which can provide a reference for the identification of them.
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Objective: Poria cocos and Polyporus umbellatus are similar medicinal fungi in traditional Chinese medicines. A method for fingerprint analysis of monosaccharide composition of polysaccharides by HPLC combined with chemometrics methods has been developed for characterization and discrimination of them in this research. Methods: The polysaccharides were extracted by decocting in water, and then completely hydrolyzed with hydrochloride. Monosaccharides in the hydrolyzates were derivatized with 1-phenyl-3-methyl-5-pyrazolone (PMP) for HPLC analysis. More than 20 batches of P. cocos and P. umbellatus from different regions were analyzed. Results: The fingerprints of P. cocos showed five common characteristic peaks, which were identified by comparing with the reference substances. The five peaks corresponded to the derivatives of mannose, ribose, glucose, galactose, and fucose. At the same time, the fingerprints of P. umbellatus showed eight common characteristic peaks, of which seven were identified as the derivatives of mannose, ribose, rhamnose, glucose, galactose, xylose, and fucose. Moreover, the similarity of their fingerprints was respectively calculated by the Similarity Evaluation System for Chromatographic Fingerprint of TCM published by China Pharmacopoeia Committee (Version 2004A). And the data were further processed by hierarchical cluster analysis (HCA) and principal component analysis (PCA). The similarity evaluation and HCA indicated that there were no significant difference in P. cocos or P. umbellatus samples from different geographical regions, but PCA was performed to characterize the difference in monosaccharide constituents between P. cocos and P. umbellatus, and linear discriminant analysis (LDA) showed the overall correct classification rate was 100%. Conclusion: The fingerprint analysis method of monosaccharide composition of water-soluble polysaccharides can distinguish P. cocos and P. umbellatus, and can be applied for the authentication or quality control for P. cocos and P. umbellatus.
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Lipopeptides are currently re-emerging as an interesting subgroup in the peptide research field, having historical applications as antibacterial and antifungal agents and new potential applications as antiviral, antitumor, immune-modulating and cell-penetrating compounds. However, due to their specific structure, chromatographic analysis often requires special buffer systems or the use of trifluoroacetic acid, limiting mass spectrometry detection. Therefore, we used a traditional aqueous/acetonitrile based gradient system, containing 0.1% (m/v) formic acid, to separate four pharmaceutically relevant lipopeptides (polymyxin B1, caspofungin, daptomycin and gramicidin A1), which were selected based upon hierarchical cluster analysis (HCA) and principal component analysis (PCA). In total, the performance of four different C18 columns, including one UPLC column, were evaluated using two parallel approaches. First, a Derringer desirability function was used, whereby six single and multiple chromatographic response values were rescaled into one overall D-value per column. Using this approach, the YMC Pack Pro C18 column was ranked as the best column for general MS-compatible lipopeptide separation. Secondly, the kinetic plot approach was used to compare the different columns at different flow rate ranges. As the optimal kinetic column performance is obtained at its maximal pressure, the length elongation factorλ(Pmax/Pexp) was used to transform the obtained experimental data (retention times and peak capacities) and construct kinetic performance limit (KPL) curves, allowing a direct visual and unbiased comparison of the selected columns, whereby the YMC Triart C18 UPLC and ACE C18 columns performed as best. Finally, differences in column performance and the (dis)advantages of both approaches are discussed.
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Objective To establish a new identification method of Ophiopogonis japonicus from different habitats.Methods Using 1H-NMR to get the all component information on O.japonicus,and using pattern recognitions,such as principal component analysis(PCA),partial least squares-discriiminate analysis(PLS-DA),and hierarchical cluster analysis(HCA) to analyze the data from the 1H-NMR spectra.Results The 1H-NMR-pattern recognition(PR) method could identify the samples of O.japonicus from different habitats successively.Conclusion The 1H-NMR-PR is an useful method for identification of O.japonicus from different habitats and could be used for the quality control of traditional Chinese medicinal materials.