| Bupleurum is a traditional Chinese medicine,with a long history.In the 2015 edition of the China Pharmacopoeia contained two original Bupleurum,namely Bupleurum chinense DC.and Bupleurum scorzonerifolium Willd..Bupleurum has bitter acrid cold with many activities such as antipyretic,antifebrile,antibacterial,antiviral and relieve chest pain effects.Bupleurum species are widely distributed in our country,in addition to Hainan have not been reported,the remaining provinces are have Bupleurum species distribution,in recent years,Guangyuan and Mianyang have gradually become the main origins of Bupleurum.In this study,we analyzed the main chemical constituents of saikosaponins,flavonoids and polysaccharides in Zhongchai No.2 of Bupleurum produced in Sichuan,and established HPLC and GC fingerprints of Zhongchai No.2,Which provided the scientific basis for the quality evaluation,the identification and the medicine of Zhongchai No.2.The main results are as follows:Determine the HPLC method for determination of saponins and flavonoids in Zhongchai No.2 of Sichuan Bupleurum by the optimization of pretreatment conditions,detection conditions and elution conditions,the selection of mobile phase and column of Zhongchai No.2.HPLC experiments were performed on Hypersil GOLD column(4.6mm×250 mm,5.0 ?m),eluted with acetonitrile(A)-0.1% phosphoric acid(B)as the mobile phases in gradient mode(0-5 min,10%A-30%A;5-25 min,30%A-70%A)at a flow rate of 0.8 mL·min-1,The column temperature is room temperature detection(30oC)at the injection volume of 10 ?L,the running time of 25 min,the wavelength of 210 nm.Determination of saikosaponins and flavonoids in Zhongchai No.2 by HPLC showed that the underground part contains saikosaponin a,saikosaponin c,saikosaponin d,and the total contents of saikosaponin a and saikosaponin d were more than 0.30%,higher than the minimum requirement of Bupleurum medicinal content on 2015 volune of Chinese Pharmacopoeia,up to 1.01%,the underground part of Zhongchai No.2contains little flavonoids.The upper part of Zhongchai No.2 contains little saikosaponin a and saikosaponin d,and its up to 0.10%,the upper part of Zhongchai No.2 contains more flavonoids,up to 0.68%.Determination of total saponins,total flavonoids and total polysaccharides in Zhongchai No.2 by UV showed that the total saponins content of the upper part was 0.26%-0.82%,the total flavonoids was 0.81%-2.76%,the soluble polysaccharide was 0.996-3.859 mg·g-1,the crude polysaccharide was 6.032-10.632mg·g-1 and the total polysaccharides up to 12.837 mg·g-1;the total saponins content of underground part was 1.05%-2.12%,the total flavonoids was 0.18%-0.46%,thesoluble polysaccharide was 2.118-13.809 mg·g-1,the crude polysaccharide was7.153-20.257 mg·g-1 and the total polysaccharides up to 28.992 mg·g-1.The total saponin content of upper part of Zhongchai No.2 reached the highest value in August and September and decreased in October and November,the total saponins content of underground part of Zhongchai No.2 reached the highest value in July and August and decreased in September,October and November;The total flavonoids content of upper part of Zhongchai No.2 reached the highest value in August and September and decreased in October and November,the total flavonoids content of underground part of Zhongchai No.2 reached the highest value in July and August and decreased in September,October and November;The content of total polysaccharides in upper part and underground part of Zhongchai No.2 were increasing from July to November.Import the HPLC and GC chromatograms of 10 batches Zhongchai No.2 samples into the Chinese medicine chromatography fingerprint similarity evaluation system,HPLC fingerprints matched 10 common peaks and identified three active ingredients of saikosaponin a,saikosaponin c and saikosaponin d,The relative retention time RSDs of common peaks were 0.18%-0.70%,The relative peak area RSDs of common peaks were 20.95%-101.05%;GC fingerprints matched 12 common peaks and the relative retention time RSDs of common peaks were 0.04%-0.32%,the relative peak area RSDs of common peaks 20.67%-88.92%.The results showed that the main components of the samples were basically identical,but the content of each component was quite different,which indicated that each chemical composition of same varieties of the different areas in Sichuan Bupleurum significantly different.The similarity of HPLC fingerprints of Zhongchai No.2 was showed that the similarity between each batch of samples and control fingerprints was 0.39%-0.97%,and the similarity among the samples was 0.05%-0.95%;The similarity of GC fingerprints of Zhongchai No.2 was showed that the similarity between each batch of samples and control fingerprints was 0.76%-0.99%,and the similarity among the samples was 0.33%-0.98%.Although the Bupleurum samples with the same base source,but because of the different growth environments,the fingerprint similarity was also different.By the research on the fingerprints of 10 batches of Zhongchai No.2 produced in Sichuan,this paper established HPLC and GC fingerprint analysis method of Zhongchai No.2,the common peak informations of saikosaponins and volatile oils were respectively shown in GC and HPLC chromatograms.More comprehensive scientificevaluation of the quality of herbs.The method is simple,reproducible,which can provides a scientific basis for the quality control of Bupleurum produced in Sichuan. |
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