This project belongs to the new methods and technologies in the study of traditional Chinese medicine (TCM), and is an applied basic research field in the area of TCM pharmaceutical technology and in vivo process studies. After oral administration, not only are the active components of herbal decoctions absorbed by the body as drug prototypes, but some components also need to be transformed into active compounds under the action of intestinal flora before being absorbed. They then undergo phase I and phase II metabolism in the liver, thereby exerting pharmacological effects or producing certain toxic side effects. These in vivo metabolic processes are influenced by individual differences in metabolic enzymes and transporters within the body under different pathological and physiological states. Our research group has focused on systematic studies of the in vivo metabolic transformation patterns of active components in TCM for nearly ten years, and has adopted in vitro enzymatic hydrolysis technology to simulate the metabolic activation mechanisms of drugs in the intestinal tract of the body. We have explored targeted preparation processes for active components in TCM and screened out a batch of candidate compounds with stronger pharmacological activities. Lay the foundation for the research of new traditional Chinese medicine (TCM) drugs. The following series of achievements have been obtained: (1) Constructed research methods for the intestinal transformation and in vivo metabolism of active components in TCM. Using UHPLC-Q/TOF technology combined with pharmacokinetic research methods, studies on the in vivo metabolism of active components in more than 10 types of TCMs such as Gardenia, Gentian, and Anemarrhena were completed. Preliminary elucidation was made on the intestinal enzymatic activation and in vivo metabolic patterns of various glycoside components in TCMs such as Gentiopicroside, Icariin, and Anemarrhena saponins. This technology was applied to the study of the in vivo metabolic activation mechanism of iridoid glycosides in heat-clearing and bile-relieving TCMs. By comprehensively using analytical chemistry and computational chemistry techniques, it was first proven that after the removal of sugars from iridoid glycosides in the intestine, they need to undergo secondary metabolism or rearrangement to produce a pharmacologically active structure, laying theoretical foundations for the discovery of in vivo pharmacological structures of active components in TCMs. (2) A method for the determination of the content of phase II metabolites of active components in traditional Chinese medicine has been established. A bioenzymatic hydrolysis combined with HPLC-MS/MS detection technology was used to establish methods for determining the content of phase II metabolites in Gardenia jasminoides glycoside, Epimedium sagittatum saponin, and the active components in the traditional Chinese medicine compound Yin Chen Hao decoction. This was the first time that the pharmacokinetics of these phase II metabolites in vivo were completed, solving a technical problem in the field and providing new technologies and methods for related research by domestic and international peers. (3) Based on metabolic transformation rules and in vitro bioenzymatic hydrolysis technology, a targeted preparation process for active components of traditional Chinese medicine has been constructed. By simulating the in vivo metabolic transformation process of glycoside components in traditional Chinese medicine and using in vitro bioenzymatic hydrolysis and biphasic immobilized enzyme technology, a targeted preparation of secondary glycosides (Anemarrhena asphodeloides saponin AIII, Baohuo glycoside I) and aglycones (Anemarrhena asphodeloides saponin aglycone, Epimedium sagittatum saponin aglycone) and other active compounds has been achieved. Compared to conventional acid-base hydrolysis processes, it has a high degree of directionality and specificity. The preparation process and activity screening research for the aforementioned traditional Chinese medicine extracts and monomer compounds have been completed, meeting the requirements of pilot production, and a series of candidate drugs with highly active pharmacological effects such as liver protection, bile secretion promotion, antifungal, and antitumor have been screened out. (4) Research on the correlation between the in vivo metabolism of active components in traditional Chinese medicine and bilirubin metabolic enzymes and transporters clarified the mechanism of action of the traditional Chinese medicine compound Yin Chen Hao Tang in promoting bile secretion. Based on the laws of in vivo metabolism, combined with metabolomics and molecular informationomics technologies, by studying the relationship between the phase II metabolism of active components in traditional Chinese medicine and endogenous substances (bilirubin metabolic enzymes and transporters, etc.), we have for the first time proven that the mechanism by which Yin Chen Hao Tang promotes bilirubin metabolism is related to the upregulation of liver metabolic enzymes Ugt1a1 and transporters Mrp2, Oct1, The protein levels of Ntcp are related, and its main component, gardenoside, and its phase II metabolite may be the pharmacodynamic basis for its choleretic effect. This research elucidates the mechanism of its choleretic action from the perspective of drug metabolism. The research results have been applied in the study of new Chinese medicine drugs. Based on the study of the in vivo pharmacodynamic substances of traditional Chinese medicine compound prescriptions, the preparation process of the compound was optimized, and the quality standards of traditional Chinese medicine were improved.