We validated whether a shorter methionine loading test is as accurate as the original 6-h test in identifying hyperhomocysteinemic patients and investigated determinants of fasting and post-load homocysteine concentration. Plasma homocysteine was determined in EDTA-blood from women with a history of pre-eclampsia (n=106) after 12 h fasting and 3 and 6 h after an oral methionine load (0.1 g/kg body weight). The 677C>T polymorphism in the methylenetetrahydrofolate reductase (MTHFR) gene, vitamin B6, vitamin B12, folate and creatinine were measured as determinants of homocysteine concentration. Good correlation and agreement between 3-h and 6-h plasma concentration of post-load (r=0.93, Kendall's tau-b=0.85) and delta (post-load minus the fasting value; r=0.90, Kendall's tau-b=0.79) homocysteine was observed and gross misclassification did not occur after division of 3-h and 6-h homocysteine scores into quartiles. Multiple linear regression revealed MTHFR 677 TT (p=0.01), folate (p=0.04) and vitamin B12 (p=0.06) as determinants of fasting homocysteine concentration; only MTHFR 677TT was related to 3-h (p=0.04) and 6-h (p=0.004) post-load homocysteine concentration. The MTHFR 677TT genotype resulted in >30% higher fasting and 3-h and 6-h post-load homocysteine concentrations compared to the wild-type CC genotype. This study shows that the 3-h methionine loading test is as good as the 6-h methionine loading test in identifying hyperhomocysteinemic patients. Furthermore, remethylation parameters (MTHFR 677C>T) strongly affect both fasting and post-load homocysteine.