鹿茸饮片浸出物提取方法的建立及多成分测定在质量评价中的应用

Establishment of extraction method and quantitative analysis of multicomponents in evaluating different Cervi Cornu Pantotrichum decoction pieces

目的比较不同提取方式、不同溶剂、不同料液比、不同提取时间对鹿茸浸出物得率的影响,对15个产地养殖的60批鹿茸饮片中7种核苷类主要成分(尿嘧啶、鸟嘌呤、次黄嘌呤、黄嘌呤、尿苷、肌苷和鸟苷)进行分析,比较不同部位鹿茸饮片(蜡片、粉片、纱片和骨片)的差异。方法参照《中国药典》2015年版四部通则相关方法优化鹿茸浸出物的提取方法,建立鹿茸浸出物最佳提取工艺,采用UPLC法对60批不同批次鹿茸饮片7种核苷类成分进行测定,结合主成分分析(PCA)和正交偏最小二乘-判别分析(OPLS-DA)化学计量学方法对鹿茸饮片进行区分与比较。结果热浸法提取条件下水溶性浸出物含量显著高于冷浸法,同一批次来源不同部位鹿茸饮片的浸出物含量,骨片低于蜡片、粉片、纱片(P<0.05)。不同部位饮片中7种核苷类成分总量在0.95~5.20 mg/g。通过OPLS-DA得出5个主要差异性成分尿嘧啶、鸟嘌呤、次黄嘌呤、尿苷和鸟苷为不同部位鹿茸饮片之间含量变化差异较为显著的成分。结论建立的鹿茸浸出物提取方法简便高效,结合7种核苷类成分含量测定可将纱片、骨片、蜡片与粉片之间进行初步区分,从而用于鹿茸饮片的质量控制与评价,为进一步完善鹿茸质量标准提供科学依据。

Objective To study the effect of different extraction methods,solvent,material liquid ratio and time on the yield of extract Cervi Cornu Pantotrichum(CCP).To analyze the content of seven chemical constituents(uracil,guanine,hypoxanthine,xanthine,uridine,inosine,and guanosine)from the 60 batches of CCP pieces in 15 habitats,and compare the differences of different parts of CCP pieces(wax slices,powder slices,gauze slices,and bone slices).Methods The CCP extraction method was optimized according to Pharmacopoeia of the People’s Republic of China(2015 edition,volume IV),the optimal extraction method of CCP was determined.A total of 60 batches of CCP pieces from different origins were determined by UPLC.Principal component analysis(PCA)and orthogonal partial least squares-discriminant analysis(OPLS-DA)chemometrics were used to distinguish and compare CCP pieces.Results The water-soluble extract,hot dip method was better than the cold dip method.Water-soluble extracts under hot dip methods could be used to significantly distinguish bone slices with wax slices,powder slices,gauze slices from the same batch of CCP pieces(P<0.05).The total content of seven nucleotides were between 0.95-5.20 mg/g in the decoction pieces of CCP.And uracil,guanine,hypoxanthine,uridine,and guanine were five main components among different parts of CCP pieces by orthogonal partial least squares discriminant analysis(OPLS-DA).Conclusion The method is simple and efficient for the extraction of CCP.It is suggested that the method of water-soluble extract of CCP and the determination of seven nucleosides and nucleobases should be combined to evaluate the quality of CCP,which can provide scientific basis for further improving the quality standard.