The mechanism of efflux of melphalan from L5178Y lymphoblasts in vitro was investigated. Evidence was obtained that drug efflux occurs by a process different from the influx mechanism. A time course of efflux followed a first-order process for at least 5 min, with a rate constant K of 0.13 +/- 0.05 (S.D.) min-1, and approximately 80% of drug taken up by the cells was exchangeable. From a kinetic analysis of melphalan efflux, it was not possible to determine if the mechanism of drug efflux was simple diffusion or a technically nonsaturable carrier-mediated process. The Q10 for drug efflux varied from 1.2 to 1.8, values intermediate between those expected for simple diffusion and a carrier-mediated process. Furthermore, drug efflux was sodium independent and was not inhibited by the presence of amino acids on the same side of the membrane, findings which support the concept that efflux was by simple diffusion. The presence of a wide variety of amino acids in the extracellular medium was found to stimulate melphalan efflux. An evaluation of the structure-activity relationship disclosed that amino acids containing hydroxyl, acidic, or amide side chains were most active; however, a distinct pattern between amino acid structure and stimulation of efflux was not established. The stimulation process was concentration dependent and was not restricted to either L5178Y cells or to melphalan as transport substrate.