Gypenosides are the main bioactive components in Gynostemma pentaphyllum. This study investigated whether gypenosides diet intervention could reduce the development of overweight and regulate the gut microbiota in C57BL/6J mice， which were fed with a high-fat diet for 12 weeks. The results showed that gypenosides diet intervention at 600 mg/kg bw/d significantly reduced body weight gain， body fat percentage and adipocyte size in mice， compared with the high-fat diet group. Meanwhile， it also reduced low-density lipoprotein cholesterol， alanine aminotransferase and leptin，and increased high-density lipoprotein cholesterol. Moreover， the results of 16S rRNA high-throughput sequencing showed that gypenosides diet intervention increased the α diversity and relative abundance of Oscillospiraceae， Alloprevotella， Parabacteroides， Lachnospiraceae DW59 and Burkholderiales YL45 at genus level.