Oxfenicine is a carnitine-palmitoyl transferase 1b (CPT-1b)-specific inhibitor that has been shown to improve whole-body insulin sensitivity while suppressing fatty acid (FA) oxidation and increasing circulating FA. Since the white adipose tissue (WAT) is an organ that stores and releases FAs, this study investigated whether oxfenicine-induced inhibition of FA oxidation affected adiposity and WAT metabolism in rats fed either low (LF) or high-fat (HF) diets. Following 8 weeks of dietary intervention, male Sprague-Dawley rats were given a daily i.p. injection of oxfenicine (150 mg/kg body weight) or vehicle (PBS) for 3 weeks. Oxfenicine treatment reduced whole-body fat oxidation, body weight, and adiposity and improved insulin sensitivity in HF-fed rats. All these effects occurred without alterations in food intake, energy expenditure, and ambulatory activity. In vivo oxfenicine treatment reduced FA oxidation and lipolysis in subcutaneous inguinal (SC Ing) adipocytes, whereas glucose incorporation into lipids (lipogenesis) was significantly reduced in both SC Ing and epididymal (Epid) adipocytes. In summary, our results show that oxfenicine-induced inhibition of CPT-1b markedly affects WAT metabolism, leading to reduced adiposity through a mechanism that involves reduced lipogenesis in the Sc Ing and Epid fat depots of rats.