Study On Quality Consistency Of Salvia Yunnanensis C.H.Wright And Salvia Miltiorrhiza Bge.based On Biological Potency

Salvia yunnanensis C.H.Wright(ZDS)is a common Chinese herbal medicine in southwest minority areas of China.Its clinical effects include promoting blood circulation and removing stasis,detoxification and swelling,cooling blood and carbuncle,etc.ZDS has a long medicinal history in Yunnan,Guizhou and other regions.It was first recorded in the Ming Dynasty《Materia medica of Southern Yunnan》: "the blood tonifying effect of this medicine is better than the four kinds of medicine".There are more than ten kinds of patent medicines with purple Salvia miltiorrhiza as preparation,such as Danshen Yixin Capsule,Zidan Huoxue tablet,zideng capsule,etc.,which are mainly used to treat irregular menstruation,chest pain,bone injury,blood stasis,swelling and pain and other diseases.Now it has been included in the "Standard of traditional Chinese medicine in Yunnan Province" and "Quality standard of traditional Chinese medicine in Guizhou Province" and other local regulations.Salvia miltiorrhiza Bge.(DS)is also a traditional medicine widely used in clinic,which has the effect of promoting blood circulation,cooling blood and carbuncle,etc.Because both ZDS and DS are medicinal plants belonging to Salvia of Labiatae family,their basic sources and clinical effects are similar,so they are often confused in Yunnan and Guizhou areas of China.However,there is still a lack of scientific basis for the rationality of this mixed use.In order to solve the quality consistency problem of ZDS and DS,this study starts from the clinical efficacy of ZDS and DS,and selects anti-platelet aggregation activity,inhibition of COX-2 enzyme activity and inhibition of NLRP3 inflammasome activity as indicators to evaluate their biological activity consistency.Multi-component chemical characterization of ZDS and DS were carried out by UPLC,and their quality consistency was comprehensively evaluated by chemical fingerprint and biological activity evaluation.The main research contents include the following parts:1.Quality consistency evaluation of ZDS and DS based on multi-component chemical characterization.Ultra performance liquid chromatography(UPLC)was used to establish a method for the determination of 10 components in ZDS and DS.Their chemical quality consistency was evaluated by fingerprint similarity analysis.The results showed that the similarity of 12 batches of ZDS was between 0.831-0.990,and that of 10 batches of DS was between 0.990-0.999,both of which were very high,meeting the requirements of fingerprint for similarity;in terms of chemical composition,salvianolic acids were the main components of purple Salvia miltiorrhiza,and tanshinones were the main components of Salvia miltiorrhiza,with significant differences.Principal component analysis of chromatographic peak area also showed that ZDS and DS were obviously divided into two categories according to the chemical content,with different chemical characteristics.There was no consistency in the chemical composition of ZDS and DS.2.Quality consistency evaluation of ZDS and DS based on the biological activity of anti platelet aggregation.In this study,the maximum platelet aggregation rate was used as the measurement index,cryptotanshinone was selected as the standard reference,and the anti-platelet aggregation method of ZDS and DS was established by investigating the solvent DMSO volume fraction,plasma centrifugal force,drug concentration and other factors.The results showed that the average blood activating potency of ZDS was 667.25U/μg,and that of DS was 1240.7U/μg,which was about twice of that of ZDS.Further analysis of variance also showed that there was a very significant difference in the biological potency of anti-platelet aggregation between ZDS and DS(P<0.0001),there was no consistency in the biological activity of anti-platelet aggregation between ZDS and DS.3.Quality consistency evaluation of ZDS and DS based on inhibition of COX-2activity.COX-2 was selected as the activity target,and the inhibition rate of COX-2 activity was used as the evaluation index to investigate the inhibition effect of different concentrations of ZDS and DS on COX-2 activity,so as to establish a biological activity evaluation method combined with the enzyme level of clinical indications.The biological activity of different batches of ZDS and DS was determined by this method.It was found that both of them had good inhibition effect on COX-2 activity.The average inhibitory potency of ZDS on COX-2 was 23.962U/mg,and that of DS was28.202U/mg.The variance analysis also showed that there was no significant difference in the inhibition of COX-2 bio-availability between the two groups(P=0.1641).That is,ZDS and DS had the same biological activity in inhibiting COX-2.4.Quality consistency evaluation of ZDS and DS based on inhibition of NLRP3 inflammasome activity.Caspase-1 was selected as the active target to establish a stable NLRP3 inflammasome model.The number of Caspase-1 secreted by NLRP3 inflammasome model was used as an evaluation index to investigate the biological activity of different batches of ZDS and DS on NLRP3 inflammasome.It was found that both of them had a good inhibitory effect on the expression of Caspase-1.The average inhibitory potency of ZDS on caspase-1 was 15.245 U/mg,and that of DS was 42.823 U/mg.The results of variance analysis showed that the biological potency of Caspase-1 was significantly different between ZDS and DS(P<0.05).That is,there was no consistency between ZDS and DS in inhibiting the activity of NLRP3 inflammasome.To sum up,this study took the quality consistency of ZDS and DS as the breakthrough point,constructed the quality evaluation method related to clinical efficacy,explored the main active components of them in playing different curative effects,and evaluated the quality consistency of ZDS and DS from multiple biological activity perspectives through the correlation between fingerprint and biological potency.This method provides a new idea and technical reference for the quality consistency evaluation of traditional Chinese medicine with similar clinical efficacy.In the selected three biological activities,the degree and activity of ZDS and DS are different,which indicates that there is insufficient evidence of consistency between the two groups.