Recent studies have shown that polysaccharides from Anoectochilus roxburghii (Wall.) Lindl. (ARPs) can reduce blood glucose levels, ameliorate oxidative stress and inflammation. However, whether ARPs have a beneficial effect on diet-induced obesity remain to be determined.This study aims to investigate the effect and mechanism of ARPs in improving obesity and metabolic disorders induced by high-fat diet (HFD).In this study, 6-week-old male mice were fed with HFD or chow diet for 13 weeks, and a dietary supplementation with ARPs was carried out. Glucose tolerance test and insulin tolerance test were performed to measure the glucose tolerance and insulin sensitivity. Adipose tissue and liver were isolated for analysis by qRT-PCR, Western blotting, hematoxylin-eosin staining and immunostaining.At week 13, body weight and fat mass were significantly increased by HFD, but ARPs supplementation abolished these phenotypes. Compared with HFD group, thermogenic genes including Ucp-1, Pgc-1α, Prdm16 and Dio2 in adipose tissue were up-regulated in ARPs-treated mice. In addition, ARPs decreased liver lipid accumulation by reducing lipid synthesis and increasing oxidation. Meanwhile, dyslipidemia and insulin resistance induced by HFD were improved by ARPs. Mechanistically, ARPs can promote fat thermogenesis via AMPK/SIRT1/PGC-1α signaling pathway.Dietary supplementation of ARPs can protect mice against diet-induced obesity, fatty liver and insulin resistance. Our study reveals a potential therapeutic effect for ARPs in regulating energy homeostasis.Copyright © 2022. Published by Elsevier GmbH.