Isolated mouse islets were used to compare the effects of three saturated (myristate, palmitate and stearate) and three unsaturated (oleate, linoleate and linolenate) long-chain fatty acids on insulin secretion. By varying the concentrations of fatty acid (250-1250 micromol/l) and albumin simultaneously or independently, we also investigated whether the insulinotropic effect is determined by the unbound or total concentration of the fatty acids. Only palmitate and stearate slightly increased basal insulin secretion (3 mmol/l glucose). All tested fatty acids potentiated glucose-induced insulin secretion (10-15 mmol/l), and the following rank order of potency was obtained when they were compared at the same total concentrations: palmitate approximately = stearate > myristate > or = oleate > or = linoleate approximately = linolenate. The effect of a given fatty acid varied with the fatty acid to albumin molar ratio, in a way which indicated that the unbound fraction is the important one for the stimulation of beta cells. When the potentiation of insulin secretion was expressed as a function of the unbound concentrations, the following rank order emerged: palmitate > myristate > stearate approximately = oleate > linoleate approximately = linolenate. In conclusion, the acute and direct effects of long-chain fatty acids on insulin secretion are due to their unbound fraction. They are observed only at fatty acid/albumin ratios higher than those normally occurring in plasma. Saturated fatty acids are stronger insulin secretagogues than unsaturated fatty acids. Unbound palmitate is by far the most potent of the six common long-chain fatty acids.