Huangkui capsule (HKC), a proprietary traditional Chinese medicine, has been used for decades to clinically treat chronic kidney diseases. Gut microbiota may mediate its metabolism, yet inadequate research on HKC's in vivo metabolism hinders understanding of its pharmacodynamic basis. In this study, normal and pseudo-germ-free rats were orally administered HKC, with serum, urine, and fecal samples collected at serial time points. All samples were analyzed by UPLC-Q-TOF-MS/MS. Chemometrics tools were used for data processing, HKC-related xenobiotics (HRX) screening/identification, and dynamic profile characterization. Flora-related metabolites were screened via intergroup HRX comparison and validated by microbiota correlation analysis and literature evidence. As a result, a total of 112 HRX (20 prototypes and 92 metabolites) were identified and categorized into four structural types: 57 quercetin-related, 37 gossypetin-related, 7 myricetin-related, and 11 other HRX. HKC metabolism involved phase I biotransformations (mainly in quercetin-HRX) and phase II conjugations (mainly in gossypetin-HRX). Most HRX peaked in serum at 1 h post-administration, with urinary excretion occurring within 0-4 h and fecal excretion delayed to 12-24 h. Unique HRX showed distinct dynamics (e.g., serum double peaks), indicating the involvement of gut microbiota and enterohepatic circulation. The 39 differential HRX (DHRX) exhibited significant correlations with specific bacterial taxa such as Clostridium. Furthermore, based on three criteria, 63 of the 112 HRX that satisfied at least one criterion were classified as flora-related. Among these, 38 met at least two criteria and were designated as verified flora-related HRX. Finally, a total of 13 candidates were selected and proposed as PK markers. This study delineates the dynamic in vivo metabolite profiles of HKC in rats and clarifies the critical regulatory role of gut microbiota, and provide a scientific basis for the material basis of its nephroprotective effects.
Keywords: Gut microbiota; Huangkui capsule (HKC); Metabolite profiles; UPLC-Q-TOF-MS/MS.
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