Influence of MTHFR genotype on whole blood folate measured by LC/MS/MS, microbiologic and BioRad assay

Red blood cells of individuals who carry two copies of the 5‐methylenetetrahydrofolate reductase polymorphism (MTHFR C677T) contain other reduced folate forms in addition to 5‐methyl‐tetrahydrofolic acid (5CH 3 THF). We analyzed 112 WB lysates (C/C n = 51, C/T n = 44, T/T n = 13) for folate species by LC/MS/MS and for total folate (TFOL) by microbiologic (MA) and BioRad (BR) assay. Subjects with C/C and C/T genotype displayed the same pattern of folate species distribution (mean [SD]): 5CH 3 THF (83% [3%]), 5‐formyltetrahydrofolic acid (5CHOTHF, 14% [2%]), 5,10‐methenyltetrahydrofolic acid (5,10CH=THF, 2% [2%]), and tetrahydrofolic acid (THF, 1% [1%]). Subjects with T/T genotype showed a different pattern: 5CH 3 THF 62% [20%], 5,10CH=THF 19% [15%], 5CHOTHF 15% [2%], and THF 5% [4%]. The sum of folate species (mean [SD], nmol/L) by LC/MS/MS (406 [127]) correlated well with TFOL (nmol/L) measured by MA (444 [144], R 2 = 0.89) and by BR (248 [74], R 2 = 0.80). TFOL by LC/MS/MS was slightly lower than TFOL by MA: 6%, 11%, and 17% for subjects with C/C, C/T, and T/T genotype. TFOL by BR was much lower than TFOL by LC/MS/MS: 41%, 40%, and 27% for subjects with C/C, C/T, and T/T genotype. This is in agreement with spiking recovery findings in WB lysate for the BR: 5CH 3 THF 51%, 5CHOTHF 17%, THF 152%, and 5,10CH=THF 115%. Contrary to previous reports, we cannot confirm lower TFOL concentrations in subjects with T/T genotype.