A metabolic study (84-d) was conducted to investigate the folate status response of pregnant subjects (n = 12) during their second trimester and nonpregnant controls (n = 12) to folate intakes approximating the current (400 microg/d) and former (800 microg/d) recommended dietary allowance (RDA). The overall goal of the study was to provide metabolic data to assist in the interpretation of the current RDA for folate. Subjects were fed a controlled diet containing 120 +/- 15 microg/d (mean +/- SD) folate and either 330 or 730 microg/d synthetic folic acid. Outcome variables between and within supplementation groups were compared at steady state. Serum folate was higher (P </= 0.05) in pregnant women consuming 850 compared with 450 microg/d (44.6 +/- 13.4, 26.3 +/- 11.3 nmol/L, respectively, mean +/- SD). No differences (P > 0.05) were detected in serum folate between pregnant and nonpregnant women within the same supplementation group. Urinary 5-methyl-tetrahydrofolate excretion was greater (P </= 0.05) in pregnant women consuming 850 compared with 450 microg/d (198.0 +/- 100.4, 9.5 +/- 3.2 nmol/d, respectively). No differences (P > 0.05) in 5-methyl-tetrahydrofolate excretion were detected between pregnant and nonpregnant women within supplementation groups. Differences (P </= 0.05) were not detected in red cell folate between pregnant women consuming either 450 or 850 microg/d (1452.5 +/- 251.8, 1733.5 +/- 208.5 nmol/L, respectively) or between pregnant and nonpregnant women consuming 450 microg/d. Our data suggest that 450 microg/d (dietary folate + synthetic folic acid) is sufficient to maintain folate status in pregnant women. This level of intake equates to approximately 600 microg/d dietary equivalents, assuming 50 and 75% availability of dietary folate and synthetic folic acid consumed with meals, respectively.