Omega-3 fatty acids (FAs) reduce postprandial triacylglycerol (TG) concentrations. This study was undertaken to determine whether this effect was due to reduced production or increased clearance of chylomicrons. Healthy subjects (n = 33) began with a 4-week, olive oil placebo (4 g/d) run-in period. After a 4-week wash-out period, subjects were randomized to supplementation with 4 g/d of ethyl esters of either safflower oil (SAF), eicosapentaenoic acid (EPA), or docosahexaenoic acid (DHA) for 4 weeks. Results for EPA and DHA were similar, and therefore the data were combined into one omega-3 FA group. Omega-3 FA supplementation reduced the postprandial TG and apolipoprotein B (apo B)-48 and apoB-100 concentrations by 16% (P = 0.08), 28% (P < 0.001), and 24% (P < 0.01), respectively. Chylomicron TG half-lives in the fed state were reduced after omega-3 FA treatment (6.0 +/- 0.5 vs. 5.1 +/- 0.4 min; P < 0.05), but not after SAF (6.9 +/- 0.7 vs. 7.1 +/- 0.7 min). Omega-3 FA supplementation decreased chylomicron particle sizes (mean diameter; 293 +/- 44 vs. 175 +/- 25 nm; P < 0.01) and increased preheparin lipoprotein lipase (LPL; 0.6 +/- 0.1 vs. 0.9 +/- 0.1 micromol/h/ml; P < 0.05) activity during the fed state, but had no effect on postheparin LPL or hepatic lipase activities. The results suggest that omega-3 FA supplementation accelerates chylomicron TG clearance by increasing LPL activity, and that EPA and DHA are equally effective.