Objective: Prickly Pear (PP) fruit is proposed to have anti-atherosclerotic and anti-hyperglycemic effects. The aim of this study was to examine the effects of a single consumption of PP juice on modifiable blood and physiological markers of cardiovascular disease risk in healthy men using a postprandial hyperlipidemia model. Methods: This was a double-blind, randomized, placebo-controlled, crossover trial with 17 healthy men (body mass index 22.6 ± 2.04 kg/m2; 29.5 ± 7.19 y of age). Participants consumed PP juice (250 mL; 45 mg betalain content; reduced fiber) or a simple placebo drink (water-based), with a high-fat muffin (50 g fat) to determine potential effects on physiologic and biological responses, for up to 3 h post-consumption (hourly, 2 sessions, 7-d washout period). Blood pressure, heart rate variability (HRV), total cholesterol (TC), triacylglycerides (TGs), low-density and high-density lipoprotein cholesterol (LDL-C and HDL-C, respectively), and glucose were measured. Results: Key findings included a lower HRV measure after PP consumption (main effect for group, P ≤0.001–0.020) but no differences for TC, TG, LDL-C, or HDL-C. Conclusion: Consumption of PP (with high-fat muffin), did not alter traditional cardiovascular disease risk responses but rather markers of HRV, beyond an expected increase in glucose attributed to the carbohydrate content of the trials foods. Additionally, macronutrient content is important when understanding HRV responses to meals.