Analysis Of In Vivo And In Vitro Metabolites Of Indazole Amide Synthetic Cannabinoids

Synthetic cannabinoids,a class of new psychoactive substances,are the most severely abused in the world.In order to avoid the attack,the structure of this new type of drug is rapidly updated.It has developed to the eighth generation of indazole amide synthetic cannabinoids from the emergence of the first generation of naphthoyl indole synthetic cannabinoids in 2006.Indazolamide synthetic cannabinoids are rapidly metabolized in the human body,so that a small amount of ingestion will produce anesthesia and excitement.At the current stage of drug control,there are two challenges in the determination of indazole amide synthetic cannabinoids: on the one hand,the lack of standard products to compare the retention time and peak area of the substances detected in actual cases;on the other hand,the lack of relevant research on the metabolic process of this type of substance,and there is still no reliable data to determine some drug-taking behaviors.It is difficult to establish a standard test method for this type of substance,while indazole amide synthetic cannabinoids have appeared in many domestic cases.Therefore,it is urgently necessary to study the metabolic mechanism of indazol amide synthetic cannabinoids,analyze the metabolites,infer the metabolic pathways and determine the metabolic markers,so as to provide a basis for the identification of indazolamide synthetic cannabinoids and the establishment of inspection methods in biological samples.It is of great significance to effectively monitor and combat the crimes involving indazole amide synthetic cannabinoid through the work of drug control.Based on 8 common indazole amides synthetic cannabinoids,the in vitro incubation model of human liver microsomes and zebrafish animal experimental model were established to simulate the metabolic process of the human body in this paper.Liquid chromatography Q Exacitive? HF combined quadrupole Orbitrap mass spectrometry technology(LC-QE-HF-MS)were used to detect azole amide synthetic cannabinoids in vivo and in vitro phase I and phase II metabolites,analyze and infer metabolic pathways,and screen metabolic markers based on the signal response intensity.The results provided data support for the establishment of inspection methods for such substances in biological samples.The main research contents are as follows:1.Indazole amide synthetic cannabinoids PX-2,MMB-FUBINACA,MDMBFUBINACA,APINACA,5F-APINACA,CUMYL-THPINACA,ADB-PINACA,ABPINACA standard product were detected by LC-QE-HF-MS.According to the mass spectrum,the lysis process and characteristic fragments of the original drug were analyzed and determined.It provided an experimental basis for the metabolite structure analysis of human liver microsomal and zebrafish.2.An in vitro incubation model of human liver microsomes was established,the human metabolic environment was simulated,and 8 kinds of indazole amides to synthesize cannabinoid metabolites were analyzed.The results showed that the indazole amide synthetic cannabinoids produced phase Ⅰand phase Ⅱmetabolites,respectively.The main metabolic pathways of phase Ⅰmetabolism were hydroxylation,carboxylation,dealkylation and carbonylation,and the pathway of phase metabolism was glucuronidation.By compariⅡ ng the signal response intensity of metabolites,the main metabolites were hydroxylated metabolites.3.A metabolic model of zebrafish was established to analyze and verify the metabolite structure of indazole amide synthetic cannabinoids in vivo.By comparing the signal response intensity,the metabolites produced by the human liver microsomes and zebrafish were analyzed and compared,and the metabolic pathways of indazole amides synthesize cannabinoids were inferred.We regarded as hydroxylated metabolites and de-N-indazole amide group metabolites as metabolic markers for indazole amide synthetic cannabinoids.