Metabolic Profile and Dynamic Characteristics of Ziziphi Spinosae Semen Flavonoids During Biotransformation and Their Effect on Human Gut Microbiota In Vitro.

The Ziziphi spinosae semen flavonoids (ZSSF) possess significant nutritional value and exhibit a wide range of health benefits, exerting positive effects on diseases related to the central nervous system. Given the excellent bioactivity and limited absorption of ZSSF, elucidating the underlying mechanism of ZSSF is imperative. In this study, in vitro fecal microbial culture fermentation was performed to investigate the metabolism of ZSSF and its effect on the gut microbiota. A total of 45 chemical constituents were identified by Ultra-performance liquid chromatography-quadrupole time-of-flight-tandem mass spectrometry (UPLC-Q-TOF-MS/MS), and possible biotransformation pathways were postulated. The degradation rates of spinosin and 6‴-feruloylspinosin were 2.53 ± 0.07 μg/mL/h and 0.79 ± 0.06 μg/mL/h, respectively, with swertisin accumulating to 55.00 ± 3.55 μg/mL at 24 h before declining. The abundance of Firmicutes increased from 8.42% at 0 h to 48.26% in the ZSSF fermentation group at 48 h, while Actinobacteria and Proteobacteria decreased from 60.02% and 30.25% to 23.46% and 20.02%, respectively. Furthermore, gas chromatography-mass spectrometry (GC-MS) was used to analyze short-chain fatty acids (SCFAs), revealing that ZSSF was associated with higher butyrate concentration at 48 h under the tested batch-fermentation conditions, with significantly higher levels observed in the ZSSF fermentation group compared to the control (1.34 ± 0.03 vs. 1.10 ± 0.05 mmol/L, p < 0.05). This study provides the first integrated analysis of ZSSF metabolism, gut microbiota modulation, and SCFAs production in a single in vitro fermentation model, offering a comprehensive understanding of the gut microbiota-mediated biotransformation of ZSSF and its metabolic consequences. Notably, these findings are derived from an in vitro batch fermentation model, and therefore, in vivo validation is required to confirm the physiological relevance of these observations. Collectively, this study provides insight into the exploration of the metabolic functions and mechanisms of ZSSF.