A randomised crossover study of eight overweight or obese men (aged 24–49 years, BMI 25.5–31.3 kg/m2), who followed two diets for 4 weeks each, was performed to determine whether substitution of saturated fat with monounsaturated fat affects body weight and composition. Subjects were provided with all food and beverages as modules (selected ad libitum) of constant macronutrient composition, but differing energy content. The % total energy from saturated fat, monounsaturated fat and polyunsaturated fat was 24, 13 and 3% respectively on the saturated fatty acid (SFA)-rich diet and 11, 22 and 7% respectively on the monounsaturated fatty acid (MUFA)-rich diet. MUFA accounted for about 80% of the unsaturated fats consumed on both diets. Body composition, blood pressure, energy expenditure (resting and postprandial metabolic rates, substrate oxidation rate, physical activity), serum lipids, the fatty acid profile of serum cholesteryl esters and plasma glucose and insulin concentrations were measured before and after each diet period. Significant (P≤0·05) differences in total cholesterol and the fatty acid composition of serum cholesteryl esters provided evidence of dietary adherence. The men had a lower weight (-2·1 (se 0·4) kg, P=0·0015) and fat mass (-2·6 (se 0·6) kg, P= 0·0034) at the end of the MUFA-rich diet as compared with values at the end of the SFA-rich diet. No significant differences were detected in energy or fat intake, energy expenditure, substrate oxidation rates or self-reported physical activity. Substituting dietary saturated with unsaturated fat, predominantly MUFA, can induce a small but significant loss of body weight and fat mass without a significant change in total energy or fat intake.