Surfactant partitioning into lipid vesicles was studied using isothermal titration calorimetry (ITC), comparing the behavior of four surfactants with current or potential application in contraception and the prevention of sexually transmitted diseases: nonoxynol-9 (N-9), the amphoteric mixture known as C31G, benzalkonium chloride (BZK), and sodium dodecyl sulfate (SDS). Membranes varied in composition from a single-component system, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, to a complex lipid mixture that models the sperm plasma membrane. The partitioning of N-9 into the membranes was found to be endothermic in contrast to the other surfactants studied. For all. four surfactants, the partition coefficient decreased as the membrane cholesterol content increased. Surfactant translocation across the membrane leaflets was also determined, with SDS being the only surfactant of the four not to exhibit "flip-flop" on experimental time scales. The results of these studies shed light on the process of surfactant-induced membrane permeabilization.