It is known that the fatty acid (FA) composition in serum cholesteryl esters to a certain extent mirrors not only the FA composition of dietary fat, but also the endogenous FA synthesis, where desaturases play an important part. A surrogate measure of delta9-, delta6- and delta5-desaturase activity can be calculated as a [product:precursor] fatty acid ratio. Delta9-desaturase activity is known to be high in conditions like diabetes, atherosclerosis and obesity. The aim of the present study was to relate the proportions of individual fatty acids in serum cholesteryl esters, as well as estimated desaturase ratios to markers of obesity and lifestyle variables (smoking, physical activity and dietary fat). We also studied gender differences. These relationships were studied in a reference population consisting of men (n=554) and women (n=295) who took part in a health survey concerning coronary heart disease in Sweden. We found positive and significant correlations between markers of obesity and the proportions of 16:0, 16:1 (n-7), 18:0, 18:3 (n-6), 20:3 (n-6), 20:4 (n-6), 20:5 (n-3), delta9 and delta6 activities, and an inverse correlation to delta5 activity and 18:2 (n-6). These relationships were independent of age and physical activity and in some cases of body mass index (BMI). For each standard deviation (SD) increase of delta9 and delta6 activities, the risk of being overweight was increased by about 60%, whereas the risk was reduced to about 30% for every SD increase of delta5 activity. Women were found to have significantly higher levels of delta9 and lower levels of delta6 desaturase activities than men. In conclusion, this study shows that a changed FA profile in serum cholesteryl esters and estimated desaturase activities are associated with obesity and lifestyle factors in men and women.