A number of reports suggest that shift workers have an increased risk of coronary heart disease (CHD). One contributing factor may be the consumption of meals at night with consequent altered postprandial responses. This study investigated circulating triacylglycerol (TAG), a possible risk factor for CHD, after meals during a simulated day and night shift. Twenty-five healthy participants (10 women and 15 men) were studied. They were given a pre-meal at 0800 h and a test meal at 1330 h on a simulated day shift and then an identical pre-meal at 2000 h and test meal at 0130 h, respectively, on a simulated night shift with maintained wakefulness. Blood was sampled for 9 h after the test meal for analysis of basal and postprandial plasma TAG levels. ANOVA for repeated measures indicated higher TAG in men compared with women (p < 0.0001) and higher responses at night in both genders (p = 0.027). Incremental area under the curve (IAUC) analysis indicated that men had significantly increased postprandial TAG levels at night compared with the day: (IAUC 0-540 min, mean +/- SEM) 253.29 +/- 28.73 versus 148.33 +/- 17.28 mmol/L x min, respectively, p = 0.025. In women, night and day responses (61.16 +/- 8.93 versus 34.09 +/- 7.87 mmol/L x min, respectively, p = 0.457) were not significantly different. Circulating TAG remained elevated for longer at night in the men compared with the women (p = 0.009). This study demonstrates the existence of gender and time-of-day differences in TAG responses to a meal. These raised TAG levels at night, for a prolonged time in men, may be relevant to the increased risk of CHD in shift workers.