Omega-3 fatty acids (n-3) can improve cardiovascular (CV) function. This study examined the effects of n-3 on endurance performance, recovery and CV risk factors in elite Australian Rules football players. 25 players were randomised, double-blind, to 6 g/day of docosahexaenoic acid (DHA)-rich fish oil (FO; n=12) or sunflower oil (SO; n=13) during 5 weeks of training. At baseline erythrocyte n-3 content, resting blood pressure (BP), fasting serum triglycerides (TG) and heart rate (HR) during treadmill running at 10 km/h were assessed. Two treadmill runs (T1 and T2) to exhaustion, separated by 5 min, were then performed at the average speed for a recent 2200 m time-trial. After 5 weeks, erythrocyte n-3 increased (FO 3.8+/-0.6%, SO 0.6+/-0.3%; P<0.001) while TG (FO -0.32+/-0.09 mmol l(-1), SO 0.08+/-0.05 mmol l(-1); P<0.001), diastolic BP (FO 1.3+/-1.3 mmHg, SO 6.8+/-1.7 mmHg; P=0.04) and HR during submaximal exercise (FO -7.8+/-2.3 beats min(-1), SO -1.9+/-1.9 beats min(-1); P=0.03) decreased in FO compared with SO. Time to exhaustion (TTE) during T1 increased by Week 5 (FO 10.2+/-2.2%, SO 17.3+/-4.3%; P<0.001 for time). Recovery (TTE for T2 as % T1) decreased in both groups (FO, -3.4+/-4.5%, SO -8.8+/-3.9%; P=0.05 for time). We conclude that 5 weeks of supplementation with FO improved CV function and reduced CV risk factors, but did not improve endurance performance or recovery in elite Australian Rules footballers.