Improvement Study Of Quality Standard Of XZK Based On Quality Markers

The prescription of Xingbei Zhike Keli(XZK)consists of nine kinds of herbs,which have the effects of relieving cough and removing phlegm.At present,the basic research of XZK is relatively weak,and the current quality standard control components are relatively simple.It is necessary to carry out researches on quality standards.This study intends to use UPLC-Q-TOF-MS/MS to analyze the chemical constituents of XZK,through the analysis of the source and specificity of the main ingredients of the prescriptions,the source and specificity of the chemical constituents of XZK were determined.The combination of the absorbed components in rat plasma and mouse ammonia water cough model were used to identify the components related to antitussive activity.Based on the above research results,the quality markers of XZK were determined to provide a basis for the improvement of quality standards.Based on quality markers,thin layer identification,content determination and fingerprinting were used to improve the quality standard of XZK.The specific research contents are as follows:1.Determination of quality markers of XZK1.1 Identification of chemicals of XZKTo analyze and identify the chemical components in XZK by using ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry(UPLC-Q-TOF-MS/MS).As a result,92 compounds were successfully identified and predicted,including alkaloids,flavonoids,coumarins and saponins.The main components were analyzed by source route and specificity to provide a basis for the screening of quality markers.1.2 Discovery and determination of quality markers related to antitussive activity of XZKThe absorbed components in SD rat plasma after oral administration of XZK were analyzed by UPLC-Q-TOF-MS/MS technique,compared with the blank plasma and chemical composition.A total of 28 components were detected,including 18 prototype components and 10 probable metabolites.Based on the chemical composition spectrum,the chemical source pathway and specificity analysis,and absorbed components in rat,17 components were initially screened out as the quality markers of XZK.In this experiment,the mouse ammonia cough model was used to screen for active ingredients in 5 compounds,including baicalein,chrysin,peimisine,isoliquiritin and oroxin A.The results showed that baicalein,chrysin,peimisine and isoliquiritin have antitussive activity.Screening results of antitussive effect on other 12 components in the early stage of the research group,it was determined that 1.1 compounds are the quality markers for XZK,including.ephedrine,pseudoephedrine,amygdalin,baicalin,(±)-praeruptorin A,praeruptorin B,praeruptorin C,peimine,peiminine,glycyrrhizic acid,and baicalein.2.Study on the quality standard of XZK2.1 Thin layer identification studyWe have optimized the original six thin layer chromatography(TLC)methods and have established the TLC method of Bitter Almond.Durability investigation results showed that the seven TLC methods were durable and had no negative interference.2.2 Content determination(1)Ephedrine and pseudoephedrine were detected by HPLC-UV,peimine and peiminine was detected by HPLC-ELSD.As a result,the two methods showed good precision,repeatability and accuracy.It can be applied to the quality evaluation of ephedrine,pseudoephedrine,peimine and peiminine in XZK.(2)Ephedrine,pseudoephedrine,amygdalin,baicalin,(±)-praeruptorin A,praeruptorin B,praeruptorin C,glycyrrhizic acid,baicalein,oroxin A and oroxin B were detected by two-dimensional ultra performance liquid chromatography(2D-UPLC).Method validation meeted requirements.As an in-depth study,this experiment is expected to provide guidamce for XZK quality standards.2.3 Fingerprint studyHPLC-UV was used to establish the fingerprint method of Xingbei Zhike Granules.A total of 26 common peaks were calibrated with 10 batches of Xingbei Zhike Granules.The 26 common peaks were identified by combining prescription drugs and UPLC-Q-TOF-MS.The results showed that the method could be applied to the overall quality evaluation of XZK.PC A and OPLS-DA were carried out on the fingerprints of 10 batches of XZK.The results showed that there were 5 main components affecting batch quality,which were mainly attributed to Stemona,Peucedanum praeruptorum Dunn and Oroxylum indicum.It is suggested that the quality of these three herbs should be paid attention to the production,and the batch quality of XZK can be controlled more scientifically and efficiently.