Genetic Diversity And Quality Evaluation Of Bupleurum Chinense In Qinling Mountains

Objective: Genetic analysis was conducted on the samples of Bupleurum chinense from Qinling area.A quality evaluation method for B.chinense was established and the highquality B.chinense producing areas in Qinling region were summarized based on the data collected in this study.Methods: The genomic total DNA of samples was extracted by CTAB method,and the universal primer mat K pairs were used for PCR amplification.The PCR products were sequenced to analyze the genetic diversity of B.chinense from different habitats.The content of saikosaponin in samples from different producing areas was determined by UPLC.Gradient elution was carried out with acetonitrile-0.1% phosphoric acid solution as mobile phase.The detection wavelength was 210 nm with column temperature 30 ? and flow rate 0.2 m L·min-1.Analysis on the difference and variation trend of saponin contents among samples of B.chinense from different provenances.To establish alcohol-soluable fingerprint for B.chinense sampled from distinct areas,UPLC method was used with acetonitrile-0.1% phosphoric acid water as mobile phase detection wavelength set as 210 nm,column temperature 30 ?,and flow rate 0.2 m L·min-1.Cluster analysis and principal component analysis of sample data were carried out with SPSS 23.0 and SIMCA 14.1 software.With the help of UPLC approach,gradient elution was performed with acetonitrile-0.1% phosphoric acid solution as mobile phase.Detection wavelength was 254 nm,column temperature 30 ?,and flow rate 0.2 m L·min-1.Finally,water-soluble fingerprint of B.chinense from different producing areas was established.Cluster analysis as well as principal component analysis were used to analyze the data.Results: The length of mat K sequence was 508 bp.34 samples were divided into 8 haplotypes,and the base mutation rate was 1.57%.According to the results of phylogenetic tree clustering,most of the cultivated products and wild products of B.chinense in Gansu province could be clustered into one category,and some of the cultivated products of B.chinense in Shaanxi province and Shanxi province were clustered together with the samples of B.chinense from Gansu province.Regarding the content of saponins,three kinds of saikosaponin in Gansu B.chinense(2 years old and above)was higher than the average value of 9.8705 mg·g-1,and the content of saponins in Shanxi was close to the mean,while the content of saponins of samples from Shanxi is below average and has large volatility.The results of alcohol-soluble fingerprint of B.chinense from different producing areas showed that 14 common peaks were identified.The similarity analysis showed that all the samples except S17 were higher than 0.902.The cluster analysis showed that the clusters of B.chinense in Gansu were concentrated.Four principal components were extracted by principal component analysis(PCA): peak 5(SSa),peak 8(SSd),and two unknown components,peak 10 and peak 14,respectively.In the water-soluble fingerprints of B.chinense from different habitats,17 common peaks were calibrated and the similarity analysis of the samples showed that all the samples except S17 were above 0.905.The cluster analysis showed that among the samples from the three provinces of B.chinense,the clusters of samples produced in Shanxi province were more dispersed,compared with closer clustering distance by analyzing samples collected in Gansu province.Five principal components,peak 11(SSb2)and 4 unknown compounds were extracted by PCA,three of which correspond to peak 5,peak 13 and peak 16.Conclusion: Based on the analysis of genetic diversity of B.chinense in different habitats in Qinling Mountains,it was found that the genetic distance of B.chinense in different producing areas is relatively small and closely related.It can be considered that the difference in plant morphology of B.chinense is mainly caused by different growth environments.The contents of saponins,alcohol-soluble fingerprints and watersoluble fingerprints of B.chinense were studied.It was found that the content of saponins in Gansu province was higher and the quality was stable in Qinling area.The quality of samples grown in Shaanxi was second.The content of saponins of B.chinense in Shanxi province was the lowest with greater fluctuation in terms of quality.