1) In situ and in vitro absorption experiments of nalidixic acid from the gastrointestinal tracts of rats were studied. 2) Results obtained from both methods were in favor of pH-partition hypothesis. Namely, absorption rate of unionized form of nalidixic acid was faster than that of ionized form of nalidixic acid. 3) Absorption rate of nalidixic acid in situ was faster than that of its drug in vitro by ten times. This might be due to the volume of solution, the length of the intestine and other physiological conditions of rats.