Chronic consumption of fish and fish oil high in (n-3) PUFA reduces triacylglycerols (TG) but may increase oxidative stress, whereas consumption of soy isoflavones may reduce oxidative stress. Elevated serum TG and oxidative stress are considered cardiovascular disease (CVD) risk factors, but the effects of acute (n-3) PUFA and soy isoflavones on these CVD risk factors are unknown. The purpose of the study was to determine the effects of acutely supplementing a high-fat, high-fructose meal with fish oil and isoflavone placebo (FO) and fish oil placebo and soy isoflavones (ISO). In a randomized, double-blind, placebo-controlled, crossover study, 10 overweight or obese men consumed a high-fat, high-fructose meal with 4 dietary supplement combinations: fish oil placebo and isoflavone placebo (placebo); fish oil and isoflavone placebo (FO); fish oil placebo and isoflavones (ISO); and fish oil and isoflavones (FO + ISO). Serum collected at baseline and at 2, 4, and 6 h postprandially was analyzed for fatty acids, isoflavones, TG, and oxidative stress biomarkers (lipid hydroperoxides, oxidized-LDL, total antioxidant status). FO significantly increased serum (n-3) PUFA and ISO increased serum isoflavones. The study meal significantly increased serum total fatty acids and TG without affecting oxidative stress biomarkers. Serum TG and oxidative stress biomarkers did not differ between treatments. The FO and ISO were bioavailable but did not attenuate the postprandial rise in serum TG. Neither the study meal nor the FO or ISO induced significant changes in oxidative stress biomarkers. The current study adds to a limited literature on the acute effects of FO and ISO interventions on postprandial biomarkers of CVD risk.