1. The uptake of 5-methyltetrahydrofolate (MTHF) and folic acid (pteroylmonoglutamate) by enterocytes and brush-border-membrane vesicles prepared from the small intestine of the 6-d-old male goat was determined using a rapid-filtration assay. 2. Both MTHF and folic acid were taken up by membrane vesicles at 25 degrees and enterocytes at 37 degrees by a pH-dependent mechanism with maximum uptake when the pH of the incubation medium was 5.0. 3. Experiments in which the osmotic pressure of the medium was raised in successive increments with the non-absorbable sugar cellobiose indicated that transport rather than membrane binding was the main component of uptake. 4. Experiments at pH 5.0 showed that uptake of MTHF and folic acid was saturable and that the characteristics of folate transport were similar in both tissue preparations: (a) transport rates of both MTHF and folic acid were constant during the first 1-2 min for a given folate concentration, then declined to reach a steady-state in 10-30 min; (b) initial velocities of transport of MTHF and folic acid increased in proportion to their concentrations up to 7-10 microM, but the rate of increase slowed thereafter until saturation was reached at 20-25 microM (Km for brush-border-membrane vesicles 40.8 and 62.9 microM, Km for enterocytes 50.9 and 55.2 microM for MTHF and folic acid respectively). Values of Vmax for membrane vesicles (pmol/mg protein per min) were 46.5 MTHF or 40.3 folic acid; enterocytes Vmax (pmol/10(7) cells per min) 15.9 MTHF or 30.6 folic acid; (c) uptake of MTHF and folic acid by brush-border-membrane vesicles and enterocytes measured under steady-state conditions approached saturation at 50 microM for each analogue (Km for membrane vesicles 58.1 and 55.2 microM, Km for enterocytes 43.1 and 49.4 microM for MTHF and folic acid respectively; Vmax for membrane vesicles 49.7 and 77.8 pmol/mg protein, for enterocytes 52.8 and 54.7 pmol/10(7) cells for MTHF and folic acid respectively). 5. It was concluded that transport of MTHF and folic acid was by a similar pH-dependent mechanism both in brush-border-membrane vesicles and in intact enterocytes. At pH 5.0, transport involved a saturable carrier-mediated process located in the brush-border membrane.