To investigate causes of increased triglyceride concentrations in subjects with normal glucose tolerance (determined by oral glucose tolerance testing using World Health Organization criteria) 883 healthy subjects (389 men and 494 women) between 40 and 65 years of age were studied. Subjects were divided by gender into four groups according to 120-min glucose concentrations. Individuals in the highest quartile of glucose concentration had the highest mean triglyceride concentrations (p < 0.0001) and highest mean non-esterified fatty acid (NEFA) concentrations (p < 0.0001). There was also a clustering of cardiovacular risk factors normally associated with the insulin resistance syndrome in subjects in this group. Regression analysis showed that the most important determinants of triglyceride levels were smoking (men p = 0.001, women p = 0.005), waist:hip ratio (men p = 0.01, women p < 0.001) and NEFA suppression (men p = 0.02, women p = 0.005). NEFAs suppressed 16.7% in women compared to 2.4% in men during the first 30 min of the oral glucose tolerance test (p < 0.001). These results show that a clustering of cardiovascular risk factors associated with decreased NEFA suppression occurs in a sub-group of subjects with normal glucose tolerance and that the pattern of NEFA suppression differs between men and women.