In this study, we evaluated the antidiabetic effect of a chloroform fraction of a methanol extract of Vigna nakashimae (designated VN) and compared it with that of a water fraction. Both fractions were administrated to eight-week old db/db mice for two weeks, after which the plasma glucose, triglyceride, and total cholesterol levels were measured. The chloroform fraction (VN-C) lowered the fasting glucose and blood glycated hemoglobin in the db/db mice more effectively than those of the water fraction (VN-W). VN-C also improved the glucose tolerance and led to a significant decrease in the plasma levels of free fatty acids and triglycerides. VN-C enhanced the phosphorylation of AMP-activated protein kinase (AMPK) and increased the expression of carnitine palmitoyltransferase-1 (CPT-1) in HepG2 and C2C12 cells more significantly than VN-W. Consistent with AMPK activation, VN-C inhibited cAMP/Dex-stimulated expression of gluconeogenic genes and increased glucose uptake in C2C12. Collectively, these results suggest that VN-C has an antidiabetic effect, which is exerted via AMPK activation, and that this effect is stronger than that of VN-W.