In a recent study of older participants (age >/=60 years) in the 1999-2002 National Health and Nutrition Examination Survey (NHANES), we showed that a combination of high serum folate and low vitamin B(12) status was associated with higher prevalence of cognitive impairment and anemia than other combinations of vitamin B(12) and folate status. In the present study, we sought to determine the joint influence of serum folate and vitamin B(12) concentrations on two functional indicators of vitamin B(12) status, total homocysteine (tHcy) and methylmalonic acid (MMA), among adult participants in phase 2 of the NHANES III (1991-1994) and the NHANES 1999-2002. Exclusion of subjects who were <20 years old, were pregnant, had evidence of kidney or liver dysfunction, or reported a history of alcohol abuse or recent anemia therapy left 4,940 NHANES III participants and 5,473 NHANES 1999-2002 participants for the study. Multivariate analyses controlled for demographic factors, smoking, alcohol use, body mass index, self-reported diabetes diagnosis, and serum concentrations of creatinine and alanine aminotransferase revealed significant interactions between serum folate and serum vitamin B(12) in relation to circulating concentrations of both metabolites. In subjects with serum vitamin B(12) >148 pmol/liter (L), concentrations of both metabolites decreased significantly as serum folate increased. In subjects with lower serum vitamin B(12), however, metabolite concentrations increased as serum folate increased starting at approximately 20 nmol/L. These results suggest a worsening of vitamin B(12)'s enzymatic functions as folate status increases in people who are vitamin B(12)-deficient.