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OBJECTIVE To establish the fing erprint of Temurin- 5 powder，conduct chemical pattern recognition analysis ，and determine the contents of 4 components simultaneously. METHODS The fingerprints of 10 batches of Temurin- 5 powder were established and similarity evaluation was performed by using high performance liquid chromatography （HPLC）combined with the Similarity Evaluation System of Chromatographic Fingerprints of Traditional Chinese Medicine （2012 edition）；common peaks were identified by comparing with mixed substance control. The common peaks were analyzed by systematic cluster analysis and principal component analysis with SPSS 26.0 software. The HPLC method was used to determine the contents of gallic acid ， geniposide，chlorogenic acid and ellagic acid in 10 batches of samples. RESULTS A total of 15 common peaks were identified from the fingerprints of 10 batches of Temurin-5 powder，and the similarity was 0.997-0.999. It was identified that peak 1 was gallic acid ，peak 3 was geniposide ，peak 5 was chlorogenic acid and peak 12 was ellagic acid. Among the 10 batches of samples ， S4 and S 9 were grouped into one category ，S6-S8 were grouped into one category ，and the other batches of samples were grouped into one category. The accumulative variance contribution rate of first three principal components was 89.245％. The linear ranges of gallic acid ，geniposide，chlorogenic acid and ellagic acid were 5.55-177.5，15.98-511.5，2.56-82.0 and 13.48-431.5 μg/mL， respectively. RSDs of precision ，stability（24 h）and repeatability tests were all less than 2％（n＝6 or n＝7）. The average recoveries were 101.56％，102.21％，98.60％ and 96.62％，respectively，RSDs were 1.90％，1.61％，1.58％ and 1.73％（n＝6）. Average contents of above components were 5.03-5.64，10.38-12.16，1.40-1.69，6.47-7.11 mg/g，respectively. CONCLUSIONS The established fingerprint is stable and feasible ，and the content determination method meets the relevant regulations. Combined with chemical pattern recognition analysis ，it can be used for the quality control of Temurin- 5 powder.
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Objective:Based on pharmacokinetics， the antitussive and expectorant related quality markers （Q-marker） of Trichosanthis Fructus were screened from diosmetin-7-<italic>O</italic>-glucopyranoside， diosmetin， apigenin， vanillic acid and cinnamic acid， and the candidate Q-marker was evaluated by multivariate statistical method. Method:Six healthy rats were randomly selected and the 70% ethanol extract of Trichosanthis Fructus （dose of 20 g·kg<sup>-1</sup>） was given by intragastric administration. Blood was collected from the orbital vein at different time points， and the plasma concentrations of 5 components （diosmetin-7-<italic>O</italic>-glucopyranoside， diosmetin， apigenin， vanillic acid and cinnamic acid） from Trichosanthis Fructus were detected simultaneously by high performance liquid chromatography-triple quadrupole tandem mass spectrometry （HPLC-QqQ-MS/MS）. The main detection conditions were as following：mobile phase of 0.2% formic acid aqueous solution （A）-acetonitrile （B） for gradient elution （0-4 min， 6%-23%B； 4-5 min， 23%-59.5%B； 5-10 min， 59.5%-60%B）， flow rate of 0.5 mL·min<sup>-1</sup>， the detection wavelength at 254 nm， electrospray ionization （ESI）， positive ion mode detection， multiple reaction monitoring （MRM） mode scanning， scanning range of <italic>m</italic>/<italic>z</italic> 50-1 500. Diosmetin-7-<italic>O</italic>-glucopyranoside， diosmetin， apigenin and vanillic acid with clear pharmacokinetic behaviors were selected as candidate Q-marker about antitussive and expectorant of Trichosanthis Fructus. The contents of these components in 9 batches of medicinal materials were determined and the main detection conditions were the same as the pharmacokinetic study. SPSS 21.0 was used for cluster analysis and principal component analysis （PAC） based on the results of determination. Result:The pharmacokinetic results showed that the area under concentration-time curve （AUC<sub>0-</sub><italic><sub>t</sub></italic>） of 4 components （diosmetin-7-<italic>O</italic>-glucopyranoside， diosmetin， apigenin and vanillic acid） were （111.28±9.94）， （27.08±2.76）， （1 376.12±101.86）， （631.32±64.72） μg·h·L<sup>-1</sup>， respectively. The 9 batches of Trichosanthis Fructus samples were clustered into 3 groups by systematic cluster analysis. The clustering results were related to the variety of Trichosanthis Fructus and also affected by the origin. The PCA results showed that the comprehensive scores of Gaotang Trichosanthis Fructus， Shanxi Trichosanthis Fructus， Hebei Ben Trichosanthis Fructus were 1.919， 1.356 and 0.299， respectively， ranking in the top 3 among all samples. The comprehensive scores of Nongkeyuan No. 1， Hebei Trichosanthis Fructus and Nongkeyuan No. 2 were -0.804， -1.085， -1.120， respectively， which were in the last 3 positions among all samples. Conclusion:The pharmacokinetic characteristics and quality evaluation of diosmetin-7-<italic>O</italic>-glucopyranoside， diosmetin， apigenin and vanillic acid meet the requirements about antitussive and expectorant related Q-marker of Trichosanthis Fructus.
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Objective::To establish a pre-column derivatization reverse-phase high-performance liquid chromatography (RP-HPLC) method for the simultaneous determination of 17 amino acids in Cynomorii Herba from different producing areas and conduct a multivariate statistical analysis. Method::RP-HPLC with pre-column derivatization was employed, with phenyl isothiocyanate (PITC) as derivatization reagent. Separation was performed on a WondaSil C18-WR column (4.6 mm×150 mm, 5 μm), with 0.05 mol·L-1 sodium acetate solution (pH 6.5) as mobile phase A, and acetonitrile-methanol-water (3∶1∶1) as mobile phase B for gradient elution at a flow rate of 0.8 mL·min-1. The detective wave length was set at 254 nm, and the column temperature was maintained at 35 ℃. Principal component analysis (PCA) and systematic cluster analysis (HCA) models were established for multivariate statistical analysis and quality evaluation. Result::17 Kinds of amino acid were detected in Cynomorii Herba, 7 of which were essential amino acids. The 17 amino acids showed good linearity in respective concentration range, r = 0.999 0-0.999 9.The average recoveries were between 98.03%-103.89%with RSD<3.5%. The results of PCA and HCA were basically the same, and both methods can be used to clearly distinguish Cynomorii Herba from 12 municipal producing areas into 6 regions. PCA can be used to classify Cynomorii Herba according to different municipal or provincial production areas, and HCA can be used to classify it according to provincial production areas. It showed that the amino acid contents in Cynomorii Herba from different municipal and provincial producing areas had differences, and the content distribution showed obvious geographical clustering characteristics. PCA showed that Cynomorii Herba from Gansu province and Inner Mongolia had higher amino acid contents and better quality as compared with other producing areas. Conclusion::The established method can be used for content determination of 17 amino acids in Cynomorii Herba from different producing areas, and provide a reference for its comprehensive quality evaluation.
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Objective: To study the fingerprints of Paeoniae Radix Rubra in different habitats, and determine the content of five chemical components (gallic acid, albiflorin, paeoniflorin, benzoic acid, and benzoyl paeoniflorin) and systematically cluster them. The relationship between origin and content was analyzed by grey correlation degree to provide reference for the quality evaluation of Paeoniae Radix Rubra. Methods: Fingerprints of Paeoniae Radix Rubra from 21 different producing areas were constructed by high performance liquid chromatography. The results were classified by principal component analysis and systematic cluster analysis. The gray correlation degree method was used to process the index components and their relative correlations were calculated. Results: HPLC fingerprints of Paeoniae Radix Rubra from 21 habitats were established, 11 common peaks were confirmed, and five of them (gallic acid, albiflorin, paeoniflorin, benzoic acid, and benzoyl paeoniflorin) were identified. The similarity of Paeonia lactiflora was greater than 0.9, and the similarity of Paeonia veitchii was less than 0.9. It was divided into two categories by principal component analysis combined with cluster analysis. The results of grey correlation analysis showed that the relative correlation (ri) was the largest in Gansu, followed by Ganzi in Sichuan. Conclusion: There is a big difference in the relative yield of Paeoniae Radix Rubra in different habitats. This experiment provides a scientific basis for the quality evaluation of Paeoniae Radix Rubra by fingerprint analysis, principal component analysis combined system cluster analysis and grey correlation analysis method.
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Objective: To discuss the phenotypic character and the HPLC fingerprints of radial striations from different germplasms Rehmanniae Radix. Method: The changes in the shape and column diameter of the radial striations of Rehmanniae Radix were observed and measured in the whole growth period. Besides,the HPLC fingerprints of the root,radial and un-radial striations were established to sign the chemical quality and analyzed by principal component analysis(PCA)and systematic cluster analysis. Result: There were significantly differences and regularities in the shape and proportion of the radial striations of different germplasms Rehmanniae Radix. The fingerprints showed the consistency between different types of chemical ingredients,and the differences in chemical quality characteristics mainly lay in the content of chemical compositions and theirs relative ratio. The results of PCA indicated that active ingredients, such as acteoside,catalpol,rehmaionoside D,rehmaionoside A and leonuride, were involved in the quality expression of different parts from various germplasms of Rehmanniae Radix,but each ingredient had a distinctive contribution rate to the differential quality expression between different parts from various germplasms of Rehmanniae Radix. However,the other components involved in the differential quality expression had different contribution rates in different germplasms.The systematic cluster analysis indicated that great differences in the chemical quality between the radial striations and un-radial striations of Beijing-1,Qinhuai,Qinhuai Zheng and 1706 germplasms,but with small differences in 85-5 and Baixuan germplasms. Conclusion: There are differences in phenotypic character of the radial striations and HPLC fingerprints between different germplasms Rehmanniae Radix.