Evidence suggests that oxidative modification of low density lipoprotein (LDL) occurs in vivo, increasing the atherogenecity of the particle. A total of 13 subjects (age range 46-78 years) with an LDL cholesterol concentration >3.36 mmol/l consumed each of four diets for 32-day periods. The diets contained 30% energy as fat of which 2/3 was either corn oil or beef tallow with and without 115 mg/4.2 MJ of supplemental cholesterol in the form of cooked egg yolk. The susceptibility of LDL to oxidation was assessed during a challenge with hemin and hydrogen peroxide, and results are expressed as lag time to oxidation in minutes. Addition of moderate amounts of cholesterol to either the corn oil or beef tallow enriched diet resulted in increased susceptibility of LDL to oxidation (decreased lag time): 69+/-22 min versus 96+/-24 min in the corn oil diet with versus without supplemental cholesterol, respectively, P = 0.006; 82+/-20 min versus 96+/-26 min in the beef tallow diet with versus without supplemental cholesterol, respectively, P = 0.025. A stepwise equation indicated that as plasma oleic acid concentrations increased and/or linoleic acid concentrations decreased, lag time increased (decreased susceptibility to oxidation), whereas as dietary cholesterol concentrations increased, lag time decreased (increased susceptibility to oxidation). In conclusion, these data suggest that addition of a moderate amount of dietary cholesterol to a reduced fat diet rich in polyunsaturated or saturated fatty acids increased the in vitro susceptibility of LDL to oxidation.