Gut microbiota regulate the activation of adipose browning, which promote energy dissipation and combat diet-induced obesity. Pomegranate peel polyphenols (PPPs) have been shown to reduce obesity, regulate lipid metabolism in adipose tissue, and modulate the composition of gut microbiota in animal fed high-fat diet (HFD). However, the role of gut microbiota in the improvement of obesity by PPPs has not been elucidated. In current study, we applied antibiotics to inhibit gut microbiota in mice fed HFD and treated with PPPs. The results showed that the inhibition of gut microbiota impair the effect of PPPs on reducing obesity and promoting adipose browning, and change the fecal metabolomic profiles in respond to PPPs. Moreover, the inhibition of gut microbiota supressed the promotive effects of PPPs on the levels of Akkermansia and microbiota-related metabolites, such as urolithin A, short-chain fatty acids (SCFAs), and bile acids (BAs), which were associated with activating adipose browning. Therefore, our results suggested that the presence of gut microbiota is essential for PPPs to ameliorate HFD-induced obesity. The related bacteria or metabolites generated by the interaction between PPPs and microbiota promote adipose browning and facilitate the beneficial effects of PPPs.