Integrated approach toward absorption, distribution, metabolism, and excretion of Xiaoke pills in zebrafish based on UPLC-HRMS and DESI-MS techniques.

Traditional Chinese medicine (TCM) has been used in clinical settings for over 2000 years in China. The study of the absorption, distribution, metabolism, and excretion (ADME) of TCM in vivo could be beneficial for the discovery of the active components in TCM. However, the conventional strategies used for ADME research are based on rodent models and have the characteristics of lengthy experimental periods, complex processes, and extensive data processing, which make it difficult to perform rapid analyses and high-throughput ADME screening of the medicinal components of TCM. In this study, an integrated high-throughput research strategy for the in vivo ADME analysis of TCM was established based on a zebrafish model. Accordingly, a combination of ultra-performance liquid chromatography-high-resolution mass spectrometry (UPLC-HRMS), desorption electrospray ionization-mass spectrometry (DESI-MS) imaging, and in-house non-targeted precise-and-thorough background-subtraction (PATBS) data post-processing techniques were successfully applied for the analysis of the metabolism of zebrafish exposed to Xiaoke pills. A total of 49 compounds related to Xiaoke pills (including 13 prototypical components and 36 metabolites) were detected in zebrafish. In total, 32 of them, including puerarin, daidzein, deoxyschizandrin, formononetin, and glibenclamide, which have been identified to have hypoglycemic activity in our previous studies and are phase I and phase II metabolites resulting from the hydroxylation, demethylation, glucuronidation, sulfation, and glycosylation of the prototypical components in vivo, were found in rats treated with Xiaoke pills. Furthermore, the overall distribution of the known compounds in zebrafish exposed to Xiaoke pills was explored using DESI-MS. In summary, this study provides a practical approach for the high-throughput screening of the active components of TCM using a zebrafish model.

Quantitative Analysis of Twelve Active Components Combined With Chromatographic Fingerprint for Comprehensive Evaluation of Qinma Prescription by Ultra-Performance Liquid Chromatography Coupled With Diode Array Detection.

A combination method of ultra-performance liquid chromatography (UPLC) coupled with diode array detection has been developed for quality evaluation of Qinma prescription (QMP), based on chromatographic fingerprint technology with the similarity analysis (SA) and the quantitative analysis of 12 components by hierarchical cluster analysis (HCA). The established method has been validated by linearity, precision, repeatability, stability and recovery tests. The UPLC fingerprints with 17 common peaks of 5 QMP samples prepared by different extraction methods including water decoction extraction, water extraction-ethanol precipitation method, ethanol reflux extraction, ethanol extraction-water precipitation method and methanol ultrasonic extraction were obtained, and the SA results indicated that similarity index was greatly influenced by the large peak. The similarity index ranged from 0.816 to 0.999 basing on 17 peaks, which has been decreased to 0.683-0.999 basing on 16 peaks without the large peak of baicalin (BA). The results of simultaneous quantification of 12 components in these 5 QMP samples proved that BA, gallic acid (GA), wogonoside (WOG) and gentiopicroside (GEN) were the major ingredients in QMP with high contents >1.44 (mg/g), indicating that ethanol reflux was the most effective extraction method. Integrating fingerprint analysis, simultaneous determination and HCA, the established method is rapid, sensitive, accurate and readily applicable. All the results indicated that the combination method can control the quality of QMP and its related traditional Chinese medicinal compounds more comprehensively and scientifically.