Deficient levels of folic acid and vitamin B12 are associated with elevated chromosome damage rate and high concentrations of homocysteine in the blood. We have therefore performed a study to determine the prevalence of folate deficiency, vitamin B12 deficiency and hyperhomocysteinemia in 64 healthy men aged between 50 and 70 years, and evaluate the relationship of these micronutrient levels in the blood with the micronucleus frequency in peripheral blood lymphocytes. We also performed a placebo-controlled, double-blind intervention study to determine whether supplementation of the diet with a daily dose of 0.7 mg (as a supplement in cereal) or 2.0 mg (in a tablet) over a period of 4 months resulted in a significant alteration of folate status, homocysteine status and the micronucleus index. Twenty-three per cent of the men were serum folate deficient (<6.8 nmol/l), 16% were red blood cell folate deficient (<317 nmol/l), 4.7% were vitamin B12 deficient (<150 pmol/l) and 37% has plasma homocysteine levels >10 micromol/l. In total, 56% of the men had one or more abnormal blood values for folate, vitamin B12 or homocysteine. The micronucleus index of these men (n = 34) in cytokinesis-blocked binucleated cells (19.2 +/- 1.1) was significantly elevated (P = 0.02) when compared to the micronucleus index of the rest of the men who had normal levels of folate, vitamin B12 and homocysteine (16.3 +/- 1.3, n = 30). Interestingly, the micronucleus index in men with normal folate and vitamin B12, but homocysteine levels >10 micromol/l (19.4 +/- 1.7, n = 15) was also significantly higher (P = 0.05) when compared to those with normal folate, vitamin B12 and homocysteine. This novel result was also supported by the observation that the micronucleus index and plasma homocysteine were significantly (P = 0.0086) and positively correlated (r2 = 0.172) in those subjects who were not deficient in folate or vitamin B12. The micronucleus index was not significantly correlated with folate indices, but there was a significant (P = 0.013) negative correlation with serum vitamin B12 (r2 = 0.099). Daily supplementation of the diet with 0.7 mg free folic acid in cereal for 2 months followed by 2.0 mg free folic acid via a tablet produced a 4-fold increase in plasma folate, a 2.6-fold increase in red blood cell folate and a 11% reduction in plasma homocysteine; however, these changes were not accompanied by a reduction in the micronucleus index. In conclusion, it is apparent that elevated homocysteine status, in the absence of vitamin deficiency and low, but not deficient, vitamin B12 status are important risk factors for increased chromosome damage in lymphocytes.