| Objective:(1)Studying the chemical components of Ophiopogon japonicus in Zhejiang to provide material basis for the formulation of quality standards and grade standards;(2)To establish the HPLC fingerprint of Ophiopogon japonicus in Zhejiang,and characterize the basic group of Ophiopogon japonicas;Establishing the method for the determination of HPLC multi-index components to provide the basis for quality control.(3)Studying the relationship between the appearance characteristics and the intrinsic quality of Ophiopogon japonicas to develop grade standards.Method:(1)The chemical composition of Ophiopogon japonicus in Zhejiang was analyzed to identify the relevant components.The target components were extracted,separated and purified by silica gel column chromatography and preparative liquid chromatography.The structures of the compounds were identified by spectral analysis.(2)High-performance liquid chromatography was used to optimize the chromatographic conditions and HPLC fingerprints of Ophiopogon japonicus were established.The chromatographic peaks of fingerprints were analyzed by reference substance control and LC/MS to clarify the chemical composition of Ophiopogon japonicus.The similarity evaluation of decoction pieces made of Ophiopogon japonicus in Zhejiang and Sichuan was performed and were compared to analyze the differences in fingerprints.The HPLC-ELSD method was established for the determination of the content of L-borneol 2-O-β-D-furosereose-(1→6)-β-Dglucopyranoside in Ophiopogon japonicus and HPLC method for simultaneous determination of p-hydroxybenzoic acid and other components,and the determination of Ophiopogon japonicus in Zhejiang was compared with Ophiopogon japonicus in Sichuan.(3)According to the requirements of 1984 "76 medicinal product specifications",connect the appearance grading index of Ophiopogon japonicas with the internal chemical composition such as fingerprint,p-hydroxybenzoic acid and other contents,To find the difference of the internal composition as a grading indicator,formulate the grading index of Ophiopogon japonicus,and classify the Ophiopogon japonicus by clustering and other methods.Results:(1)Three monomeric compounds were isolated from the ethanol extract and identified by spectral analysis and literature control.The structures were p-coumaric acid(I),p-hydroxybenzoic acid(II),(trans)-N-[2].-Hydroxy-2-(4-hydroxyphenyl)ethyl]-3-(3-hydroxy-4-methoxyphenyl)-acrylamide(III),Compound III was first isolated from Liliaceae.(2)The HPLC fingerprint of Ophiopogon japonicus was established and 13 common peaks were obtained and 5 of them were identified.The similarity measurement results showed that the similarity of Ophiopogon japonicus in Zhejiang was greater than 0.90.The similarity of Ophiopogon japonicus in Sichuan was between 0.59 and 0.87.A HPLC-ELSD method was established for the determination of L-borneol 2-O-β-D-furosereose-(1→6)-β-D-glucopyranoside in Ophiopogon japonicus.The method showed a good linear relationship in 0.3000-4.0000μg.The average recovery was 100.6%(RSD=4.2%).The HPLC method was established for the simultaneous determination of four components in Ophiopogon japonicus.The method of four components(P-hydroxybenzoic acid,(trans)-N-[2-hydroxy-2-(4-hydroxyphenyl)ethyl]-3-(3)-Hydroxy-4-methoxyphenyl)-acrylamide,Methylophiopogonanone A and Methylophiopogonanone B)exhibited good linear relationship in 0.00326-0.0652μg,,0.00457-0.0914μg,,0.0223-0.445μg,,0.0240-0.481μg.The recovery ratesas was100.6%(RSD=1.1%),99.7%(RSD=1.4%),101.0%(RSD=0.9%),and 100.8%(RSD= 1.0%).(3)The traditional graded traits indicators and internal indicators of chemical composition were measured and correlation analysis was performed using spss 22.0.The results showed that(trans)-N-[2-hydroxy-2-(4-hydroxyphenyl)ethyl]-3-(3-Hydroxy-4-methoxyphenyl)-acrylamide and Methylophiopogonanone B showed significant correlation with appearance traits,which can be used as evaluation indexes of Ophiopogon japonicaConclusion:(1)Three monomeric compounds were isolated from the ethanol extract,which provided the material basis for further research and improvement of the quality standard of Ophiopogon japonicas in Zhejiang.(2)HPLC fingerprints can be used to identify the origin of Ophiopogon japonicus,and can also be used to distinguish Ophiopogon japonicas in Sichuan.The establishment of Ophiopogon japonicus single index and multiple index determination methods can be used to provide evidence for Ophiopogon japonicus quality control.(3)The grade standard combines the relationship between appearance characteristics and intrinsic quality of Ophiopogon japonica in Zhejiang was determined which reflects the quality of Ophiopogon japonica to some extent and has certain rationality and feasibility. |
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