Abstract: Background: The pathogenesis of depression is associated with the gut-brain axis, involving gut microbiota and their metabolites. The herb pair Radix bupleuri-Radix paeoniae Alba (BP) has been reported to exert antidepressant effects by ameliorating gut microbiota dysbiosis but; its mechanisms within the gut-brain axis remain incompletely understood. Objectives and Methods: A chronic restraint stress (CRS) mouse model of depression was used. Following determination of the optimal BP dosage, mice were allocated into four groups: Control, Model, BP-treated, and fluoxetine-treated. Antidepressant effects were evaluated through behavioural tests. Serum metabolites were profiled using untargeted metabolomics and pathway enrichment analysis. Gut microbiota composition was assessed via 16S ribosomal ribonucleic acid (rRNA) gene sequencing, with analysis of alpha and beta diversity, and linear discriminant analysis effect size (LEfSe). Multi-omics integration was applied to investigate correlations between metabolites and microbial communities. Key metabolites in the hippocampus, colon, and cecum were quantified using enzyme-linked immunosorbent assay (ELISA). Results: BP treatment was associated with improvements in CRS-induced depressive-like behaviours. Metabolomic analysis indicated systemic metabolic disruption, with glycerophospholipid metabolism identified as the most significantly altered pathway. BP treatment was associated with changes in phosphatidylcholine (20:4/20:5) [PC(20:4/20:5)] andwith restoration of CRS-related gut microbiota eubiosis, including changes in microbial diversity and composition. Integrated analysis revealed correlations between reshaped microbiota (such as increased Lactiplantibacillus and decreased Anaerotignum) and alterations in glycerophospholipid metabolism. Tissue measurements also showed a BP-associated pattern of lower phosphatidylcholine in the hippocampus and higher phosphatidylcholine in the colon. Conclusions: BP treatment was associated with ameliorated depressive-like behaviours, altered serum glycerophospholipid metabolism, normalisation of PC(20:4/20:5), restoration of gut microbial ecology, and tissue-specific differences in phosphatidylcholine levels along the microbiota-gut-brain axis. The observed PC redistribution was associated with antidepressant-like effects. These findings generate testable hypotheses; causal relationships remain to be validated in further intervention experiments.