Lycium barbarum polysaccharide (LBP) has been shown to ameliorate insulin resistance, but the identification of compounds from LBP and the mechanisms have not been clarified. In this study, LBP-4a was purified from Lycium barbarum by DEAE cellulose and Sephadex G-100 column chromatography, and the effects of LBP-4a on insulin resistance were investigated. The results indicated that LBP-4a caused translocation of the glucose transporter isoform 4 (GLUT4) to the cell surface, which in turn stimulated glucose uptake, and the effect was sensitive to wortmannin, an inhibitor of phosphoinositol 3-kinase (PI3-K), and SB203580, an inhibitor of p38 mitogen activated protein kinase (p38 MAPK (α, β)). Furthermore, the effects of LBP-4a on p38 MAPK activities were abrogated by pretreatment of rat adipocytes using SB203580. In summary, LBP-4a improved insulin resistance via translocation and activation of GLUT4 in OLETF rats, and the activation of PI3-K and p38 MAPK contributed to these effects.