[Mechanism for enhancing efficacy through processing of Ziziphi Spinosae Semen based on in vivo and in vitro metabolic transformation].

This study aimed to elucidate the mechanism for enhancing efficacy through processing of Ziziphi Spinosae Semen(ZSS) by integrating in vitro intestinal microbiota transformation and in vivo pharmacokinetic studies to clarify the differences in metabolic transformation characteristics between raw and processed ZSS. High-performance liquid chromatography was employed to analyze the metabolic rates of the two major components, spinosin and jujuboside A, in raw ZSS and fried Ziziphi Spinosae Semen(FZSS) mediated by the intestinal microbiota. High-throughput 16S rRNA sequencing was used to investigate the regulatory effects of ZSS and FZSS on the intestinal microbiota in rats with insomnia. A UPLC-Q-TOF-MS/MS method was developed and validated for the quantification of spinosin and jujuboside A in rat plasma and subsequently applied to a comparative pharmacokinetic study of ZSS and FZSS in a rat model of insomnia induced by DL-4-chlorophenylalanine(PCPA). The results showed that the metabolic rates of spinosin and jujuboside A mediated by the intestinal microbiota were significantly accelerated in the FZSS group compared to the ZSS group. Relative to the non-medicated control group, both ZSS and FZSS groups significantly increased the relative abundance of beneficial bacteria, such as Firmicutes and Lactobacillus, while decreasing the relative abundance of harmful bacteria, such as Proteobacteria and Clostridium. The maximum plasma concentration(C_(max)) of spinosin in the FZSS group was 20.01 ng·mL~(-1), which was significantly lower than that in the ZSS group(45.41 ng·mL~(-1)). Similarly, the C_(max) of jujuboside A was significantly lower in the FZSS group(12.32 ng·mL~(-1)) compared to the ZSS group(16.31 ng·mL~(-1)). The clearance rate of spinosin and jujuboside A was significantly faster in the FZSS group(38 058.83,28 141.27 L·h~(-1)·kg~(-1), respectively) than in the ZSS group. Stir-frying process increased the content of amino acids in FZSS, promoted the growth of Lactobacillus and Bifidobacterium, accelerated the decomposition rate of spinosin and jujuboside A in the intestine, and consequently reduced their plasma concentration in rats. Building upon our previous research, the stir-frying process accelerated the intestinal decomposition of spinosin and jujuboside A, promoted the production of their secondary metabolites, and enhanced the sedative and hypnotic effects. This study elucidates the metabolic characteristics of spinosin and jujuboside A in ZSS and FZSS from both in vivo and in vitro metabolic perspectives, preliminarily revealing the significant impact of processing on the bioavailability of these components. These findings provide a reference for elucidating the processing mechanism underlying "enhancing efficacy through processing" of ZSS.