The association of moderately elevated total homocysteine (tHcy) levels with coronary artery disease is increasingly being recognized. However, the role of genetic influence on plasma tHcy levels is not completely understood. We studied 1,055 individuals with respect to the effect of two silent polymorphisms, the 699C--> T and the 1080C-->T, of the cystathionine beta-synthase (CBS) gene on plasma tHcy levels. Individuals who were heterozygous or homozygous for the T699 allele had lower post-methionine load (PML) tHcy levels when compared to individuals with the C/C genotype. This association was statistically significant (p = 0.005) for the T/T genotype compared to the C/C genotype and became even more significant (p = 0.000002) when individuals carrying the 68-bp insertion (844ins68) and the T1080 allele were excluded from the analysis. With regard to the 1080C-->T polymorphism, the T1080 allele was associated with significantly lower PML tHcy levels only when individuals carrying the 844ins68 and T699 allele were excluded from the study (p = 0.01 for 1080T/T genotype compared to 1080C/C genotype). We speculate that the 699C-->T and 1080C-->T polymorphisms may be in linkage disequilibrium with regulatory elements that upregulate CBS gene transcription.