1. Everted sacs of rat jejunum were used to study the transport of 5-methyltetrahydrofolic acid at various concentrations and at different pHs. 2. The transport of 5-methyltetrahydrofolic acid appeared to be linear with increasing incubation time at a 5-methyltetrahydrofolic acid concentration of 10(-5) M in the incubating medium. Tissue uptake was much higher than serosal uptake. 3. At 'zero' concentration gradient of 5-methyltetrahydrofolic acid, the transport from mucosal to serosal sides was negligible but tissue uptake was appreciable. 4. Reducing the incubation temperature from 37 to 27 degrees C gave a Q10 value of 1-8. 5. The characteristics of 5-methyltetrahydrofolic acid uptake suggested a non-saturable transport mechanism. 6. Studies were also carried out with isolated mucosal epithelial cells at different incubation times and at different pH values. 7. The over-all results suggest that 5-methyltetrahydrofolic acid transport is most likely a passive diffusion of its zwitterion and also a solvent drag with water flow. 8. The 5-methyltetrahydrofolic acid may be converted to its zwitterion in an acid microclimate at the surface of the intestinal absorbing cell.