Elderly patients with alcoholism often require surgery and receive nitrous oxide (N2O) as a component of their anesthetic. Since aging, ethanol, and N2O may all perturb folate and/or vitamin B12 metabolism, we examined the combined influence of these parameters on vitamin B12/folate status in a rodent model. Aged male Fischer 344 rats (24 months old) were given a liquid ethanol diet (35% of calories as ethanol) and control rats were pair-fed a liquid diet with carbohydrate substituting for the caloric content of ethanol. After receiving liquid diets for 7 weeks, rats were exposed to 60% N2O/40% 0(2) for 6 h. Urinary excretion of formic acid, formiminoglutamic acid (FIGLU), and methylmalonic acid (MMA) were used as indirect markers of folate/vitamin B12 status. In both the aged ethanol-fed and control groups, excretion of formic acid and FIGLU markedly increased the first day after N2O exposure and returned towards background values by the second day. No changes occurred in MMA excretion. Exposure to N2O decreased methionine synthase activities in liver, kidney and brain, and recovery of methionine synthase activities occurred over a period of 4 days in both the aged ethanol-fed and control groups. Ethanol treatment for 7 weeks combined with acute exposure to N2O did not deplete the aged rats of folate or vitamin B12 in blood, liver, kidney or brain. Thus, in this animal model, aging, chronic ethanol administration, and acute N2O exposure did not act synergistically to produce prolonged and severe disturbances in folate and vitamin B12 metabolism.