Variations of a recently proposed rat bioassay were examined and applied to the measurement of biologically available folate. The concentration of hepatic folate was found to be inconsistent as a response indicator, although fasting levels of plasma folate accurately reflected available dietary folate. Studies involving the control of coprophagy suggested that fecal folate significantly contributes to the folate status of the rat. Bioassays employing multiple dietary levels of dried orange juice solids or blanched cabbage without control of coprophagy were conducted to determined available folate by slope-ratio methods. The apparent bioavailability of endogenous folate in the dried orange juice and cabbage was 146 and 68%, respectively, relative to folic acid. The presence of dried orange juice solids or cabbage did not inhibit the utilization of folate pentaglutamate added to the diets. These results illustrate the usefulness and several limitations of the rat bioassay for evaluating the bioavailability of folate. However, the appropriateness of the rat as a model of human folate digestion and absorption requires further investigation.