Insulin resistance is commonly associated with obesity. The present study was performed to investigate the relative importance of total fat mass versus localization of adipose tissue in insulin-stimulated glucose disposal (Rd) and skeletal muscle glycogen synthase (GS) activity in obese individuals. Twenty obese women with an average body mass index (BMI) of 37.8 ± 1.3 kg/m2 and a waist to hip ratio (WHR) ranging from 0.78 to 1.02 were examined during basal conditions and following hyperinsulinemia (hyperinsulinemic euglycemic clamp). To accurately determine body composition, the following three methods were used: anthropometric measurements, dual-energy x-ray absorptiometry scanning (DEXA-scan), and bioelectric impedance measurements. In addition, indirect calorimetry and muscle biopsy were performed. Insulin-stimulated glucose Rd was negatively correlated with WHR (R = -.52, P < .025) whereas there were no correlations with BMI or percent fat (R = .16, NS and R = .16, NS, respectively). Furthermore, a negative correlation between WHR and insulin stimulation of GS activity in skeletal muscle was found (R = -.62, P < .005). In contrast, BMI and percent fat were not correlated with the insulin effect on GS activity in skeletal muscle (R = .34, NS and R = -.35, NS, respectively). The concentration of nonesterified fatty acids (NEFA) during hyperinsulineamia was strongly correlated with WHR and abdominal localization of adipose tissue (determined by DEXA-scan; R = .60, P < .005 and R = .60, P < .007, respectively). Moreover, lipid oxidation during hyperinsulinemia was positively correlated with WHR (R = .58, P < .007), whereas there was no correlation with BMI or percent fat (R = .31, NS and R = .29, NS, respectively). Finally, NEFA concentrations during hyperinsulinemia were negatively correlated with insulin-stimulated glucose Rd (R = .73, P < .0009). We conclude that in a group of massively obese women localization of adipose tissue in the abdominal region (assessed by various techniques) was accompanied by impaired insulin-stimulated glucose Rd, decreased capacity of insulin to stimulate GS activity in skeletal muscle, and higher circulating NEFA concentrations during the hyperinsulinemic period. In contrast, no correlations between BMI, percent fat, or other measurements of total amount of adipose tissue and these variables were found. Finally, the sophisticated measurements (DEXA-scan and bioelectric measurements) were not superior to BMI and WHR in predicting insulin resistance in this group of obese women.