Optimization Of Preparation Process And Application Of Naphthopyrone Reference Extracts From Cassia See

Cassiae Semen(CS)is a dry and mature seed of leguminous plants or small decoctions,with an effects of clear eyesight and active.At present,the quality control methods of CS medicinal materials are mostly controlled by the Chrysophanol and Aurantio-obtusin.However,these compounds are not specific ingredients in CS,and the content is relatively low,it is difficult to fully and scientifically reflect the quality of CS.The literature and previous studies have shown that naphthopyrones are a class of specific components in CS,which are significantly higher in content than Anthraquinone and are important pharmacodynamic material in CS.Therefore,incorporating it into the quality control method of CS is a beneficial supplement to the quality standard of CS.However,there is no relevant commercial references.Therefore,in order to solve the above problems,the laboratory has established a preparation process of a CS standard extract in the early stage,and controlled the quality of CS based on the standard extract.However,it was found that the preparation process of the standard extract in the earlier period was relatively complicated and could not be prepared in large quantities,and it was even more difficult to convert from the laboratory to industrial production.Therefore,this article intends to use a series of methods and techniques to separate the chemical constituents of naphthopyranone compounds and prepare the corresponding chemical reference materials,which will lay the material foundation for the study of preparation technology of subsequent control extracts.Secondly,on the basis of the original process,we further optimized the establishment of a simple and rapid preparation process for the standard extract,which were verified could be produce by a large scale.The standard extract was subjected to chemical composition characterization and calibration.Finally,a method for determination of CS based on a standard extract was established to enrich and improve CS quality control methods,provide a scientific basis for the comprehensive evaluation of CS quality,and provide for the development and application of standard extract.Separation of Naphthopyranones from Cassiae SemenThe research group uses a variety of isolation and structural identification technique to research chemical constituents of CS.Seven naphthopyrones are isolated and identified,including Nor-rubrofusarin-6-O-β-D-gentiobioside,cassiaside,Rubrofusarin triglucoside,cassiaside B2,cassiaside C2,rubrofusarin-6-O-β-D-gentiobioside and cassiaside C.The last four naphthopyrones are calculated by peak area normalization method.It is showed that the purity of the three compounds is more than 98%.So the three compounds can be used as a chemical reference to optimization cassia standard extract.The research of the standard extract from Cassiae SemenAn HPLC method is established for determinating of four naphthopyrones including cassiaside B2,cassiaside C2,rubrofusarin6-O-β-D-gentiobioside and cassiaside C simultaneously in CS.The results show that the method is simple,well repeatable and reproducible.The research group optimizes preparation method of cassia standard extract,with cassiaside B2,cassiaside C2,rubrofusarin-6-O-β-D-gentiobioside and cassiaside C as indexes.The prepared route of cassia standard extract that can rapidly produce is established for the first time.After the purified preparation,the total content of four main components reaches above 60%.The preparation method is stable,reproducible,and provides research base for the development of Chinese medicine standard extracts.The research group uses qualitative analysis,quantitative analysis,characteristic chromatogram and determination of moisture content to research cassia standard extract.(1)Qualitative analysis:the research group uses UPLC-MS method to study cassia standard extract.The results show that the cassias tandard extract containes cassiaside B2,cassiaside C2,rubrofusarin-6-O-β-D-gentiobioside and cassiaside C.(2)Quantitative analysis:HPLC method is used to determine the four main components of cassia standard extract.The results show that the method is well repeatable and reproducible.(3)Characteristic chromatogram:the research group establishs characteristic chromatogram of ten batches of cassia standard extract.The results show that the similarity of ten batches of cassia standard extract is above 0.9.(4)The homogeneity test was performed on uniformity cassia standard extract,and the results showed that the uniformity of cassia standard extract were well-distributed.(5)The stability of the cassia standard extract was tested,and the results showed that had good stability in 90 days,indicating that the prepared standard extract met the basic requirements for application and long-term storage.(6)Other inspections:the research group determines the water content and the hygroscopicityof cassia standard extract.The average moisture content achieved relevant standard.And the same time it has hygroscopic,which means it should be kept in closed storage.A HPLC method is used to determine the four main components and to study certification as well as uncertainty of cassia standard extract.The results of certification and uncertainty of three main components are(11.40±0.44%,(11.68±0.66)%,(16.60±0.42)%,(28.8±0.66)%.The uncertainties of the four components are all less than 1%.This suggests that uncertainty is normal and reliable.The cassia standard extract has good reproducibility between different laboratories and different instruments,which shows the feasibility of promotion of cassia standard extract.The application of the standard extract from Cassiae SemenThe research group establishes the qualitative identification method of CS and chinese herbal compound with standard extractas index,which make an important progress and development to the CS qualitative identification.The research group establishes the content determination method of cassia and chinese herbal compound with standard extract as index.Comparing the determination results of the method using standard extractas index and the method using reference substance as index,it shows no significant difference between the results of two quantitative methods.The result illustrates that standard extract can replace reference substance in cassia and chinese herbal compound quality control.The research group explores quality control standards of CS based on the content determination method using standard extract as index.Four limits of quality evaluation for CS are initially established,which provide some suggestions for the improvement of cassia quality evaluation system.Innnovation1.The evaluation method of Semen Cassiae was established,for the first time,by using the indexes of cassiaside B2,cassiaside C2,rubrofusarin-6-O-β-D-gentiobioside and cassiaside C.2.A industrial method producing cassia standard extract with cassiaside B2,cassiaside C2,rubrofusarin-6-O-β-D-gentiobioside and cassiaside C as indexes was estabilished for the first time.3.The evaluation method of Semen Cassiae and compounds was established for the first time,based on cassia naphthopyrone standard extract with cassiaside B2,cassiaside C2,rubrofusarin-6-O-β-D-gentiobioside and cassiaside C.