To investigate the effects of meal frequency on plasma lipid and carbohydrate metabolism, 19 healthy normocholesterolemic free-living men and women consumed their usual diet as three or nine meals per day in random order for 2 wk each. There was no significant difference in macronutrient intake. Compared with the three-meal/d diet, nine meals per day reduced fasting plasma total, low-density lipoprotein (LDL), and high-density lipoprotein (HDL) cholesterol by 6.5% (P < 0.005), 8.1% (P < 0.005), and 4.1% (P < 0.05), respectively. Body weight, fasting triglycerides, apolipoproteins A-I and B, and the LDL-HDL cholesterol ratios were not different for the two diets, as were 24-h urinary C peptide-creatinine ratios and insulin-glucose response to a glucose load. The insulin-glucose curve measured over 3 h in the evening after the evening meal was flatter for the nine meals, but the areas under the curves were not significantly different. Increasing meal frequency while maintaining a constant nutrient intake produces a small but significant decrease in LDL cholesterol in normolipidemic free-living subjects.