The human undergoing rapid and sustained weight loss by very low calorie dieting (VLCD) derives the majority of daily energy needs from adipose fatty acids. To evaluate the rates of metabolic utilization of individual fatty acids in humans, two groups of adult women outpatients were studied during major weight loss by VLCD. The diets used were either food or formula, providing the recommended dietary allowance for minerals and vitamins, with fat contents of 2-20 g/d. Group 1 consisted of 10 subjects [initial body mass index (BMI) 32.7, 157% of ideal body weight (IBW)] with a mean loss of 17.7 kg in 3-5 months. Group 2 consisted of 14 subjects (initial BMI 36.7, 167% of IBW) with a mean loss of 25.6 kg in 4-5 months. Adipose tissue biopsies were obtained by needle aspiration from Group 1 before and after weight loss and from Group 2 before, at the midpoint, and after weight loss. With weight loss in Group 1, the adipose tissue content of 18:1-omega-9, 18:2-omega-6, and 20:4-omega-6 did not change, but 18:3-omega-3 fell (0.67 to 0.56 wt%, p < 0.0001) as did 20:5-omega-3 (0.08 to 0.05, p < 0.01). Adipose tissue 22:6-omega-3 rose from 0.03 to 0.07 (p < 0.01). In Group 2, only 18:3-omega-3 showed a change, falling from 0.71 to 0.69 to 0.59 wt% across weight loss (p = 0.03 by analysis of variance). We conclude that the major fatty acids are oxidized in proportion to their composition in adipose triglyceride. The significant reduction in the concentration of 18:3-omega-3 during weight loss is unique among fatty acids. Its accelerated removal from adipose tissue indicates either a preferential step in beta-oxidation or a defined need during supplemented fasting which exceeds its rate of provision from adipose stores.