Objective To split the classic prescription Baoyuan Decoction (保元汤) into different phases, study the relieving effects of different phases on exercise fatigue in rats, and screen for their effective phases. Methods The gradient centrifugation dialysis method was used to separate Baoyuan Decoction into the nano phase (BD nano phase, BD-N), precipitation phase (BD-P) and solution phase (BD-S). The appearance was physically characterized and contents of main ingredients was determined. A rat swimming fatigue model was established, control group, model group, positive control (Dazhu Hongjingtian Capsule) group, Baoyuan Decoction group, BD-N group and BD-P group were set up, active phases was screened by measuring behavioral indicators, organ indices, serum biochemical indicators, and pathological changes in liver and muscle tissues. Based on combined analysis of gut microbiome, metabolomics and transcriptomics, the potential mechanism of Baoyuan Decoction and its effective phase in alleviating exercise fatigue in rats was explored. Four differentially expressed genes (Gck, Ackr3, Pde4b, Ppp1r3b) were randomly selected for qRT-PCR validation. Results The particle morphology of BD-N was spherical and Tyndall effect was significant. The particle size was in the order of BD-P > Baoyuan Decoction > BD-N. The polydispersity index (PDI) of each phase was in the order of Baoyuan Decoction > BD-N > BD-P. The absolute value of zeta potential for each phase was BD-P > BD-N > Baoyuan Decoction. Ginsenoside Rb₁, calycosin-7- O-β-D-glucoside, liquiritin, ammonium glycyrrhizinate, cinnamic acid, cinnamaldehyde and 6-gingerol were mainly distributed in BD-N. The comprehensive efficacy ranking was Baoyuan Decoction ≈ positive control drug > BD-N > BD-P. The ability of BD-N in regulating the abundance of gut microbiota, increasing the level of Lactobacillus genus and regulating 45 differential metabolites related to fatigue was similar to Baoyuan Decoction. The qRT-PCR results showed that the regulatory trends of BD-N and Baoyuan Decoction on four genes were consistent with the transcriptome sequencing results. Conclusion From the perspective of physics, chemistry and pharmacological effects, the effective phase of Baoyuan Decoction in relieving fatigue is screened, and BD-N is the effective phase of Baoyuan Decoction in relieving fatigue.

Baoyuan Decoction  /  phase state  /  exercise-induced fatigue  /  gut microbiota  /  metabolomics  /  transcriptomics  /  ginsenoside Rb₁  /  calycosin-7-O-β-D-glucoside  /  liquiritin  /  ammonium glycyrrhizinate  /  cinnamic acid  /  cinnamaldehyde  /  6-gingerol