This investigation was undertaken to study whether the risk to develop non-insulin-dependent diabetes mellitus (NIDDM) among 50-year-old men during a 10-year follow-up period was related to the fatty acid composition of their serum cholesterol esters. There were highly significant differences in the initial health survey between the fatty acid composition in serum in subjects who remained normoglycemic (w = 1,753) and in those who later developed NIDDM (n = 75). The main differences were that the latter had higher proportions of saturated fatty acids and palmitoleic acid (16:ω-7), a low proportion of linoleic acid (18:2ω-6), and a relatively high content of γ-linolenic (18:3ω-6) and dihomo-γ-linolenic (20:3ω-6) acids in the serum cholesterol esters. The picture was similar also after adjusting for differences in body mass index. In a logistic model, a high proportion of dihomo-γ-linolenic acid remained a significant contributor to the development of diabetes, along with the height of the insulin index, the blood glucose concentration at 60 min, and the fasting insulin concentration. The increased risk to develop NIDDM related to the serum cholesterol ester fatty acid composition may be mediated by diet and/or genetic factors.