AIMS/HYPOTHESIS: Insulin resistance usually precedes the diagnosis of Type II (non-insulin-dependent) diabetes mellitus. However, in most patients, the clinical expression of the disease could be prevented by dietary and lifestyle changes. We investigated the effects of a diet enriched in monounsaturated fatty acids (Mediterranean diet) and a low fat, high-carbohydrate diet on in vivo and in vitro glucose metabolism in 59 young subjects (30 men and 29 women). METHODS: We carried out an intervention dietary study with a saturated fat phase and two randomized-crossover dietary periods: a high-carbohydrate diet and a Mediterranean diet for 28 days each. We analysed the plasma lipoproteins fractions, free fatty acids, insulin sensitivity and glucose uptake in isolated monocytes at the end of the three dietary periods. RESULTS: In comparison to the saturated fat diet, the CHO and Mediterranean diets induced a decrease of LDL-cholesterol (p < 0.001) and HDL-cholesterol (p < 0.001). Steady-state plasma glucose decreased (p = 0.023) and basal and insulin-stimulated 2-deoxiglucose uptake in peripheral monocytes increased in both diets (CHO and Mediterranean), (p = 0.007) indicating an improvement in insulin sensitivity. Fasting free fatty acids plasma values were correlated positively with steady state plasma glucose (r = 0.45; p < 0.0001). In addition, there was an inverse correlation between the mean glucose of the steady state plasma glucose period and logarithmic values of basal (r = -0.34; p = 0.003) and insulin stimulated glucose uptake in monocytes (r = -0.32; p = 0.006). CONCLUSION/INTERPRETATION: Isocaloric substitution of carbohydrates and monounsaturated fatty acids for saturated fatty acids improved insulin sensitivity in vivo and in vitro, with an increase in glucose disposal. Both diets are an adequate alternatives for improving glucose metabolism in healthy young men and women.