The molecular characteristics of the neuropeptide substance P (SP), its agonist [Sar9,Met-(O2)11]SP, and three of its antagonists [D-Arg1,D-Pro2,D-Trp7,9,Leu11]SP, [D-Arg1,D-Trp7,9,Leu11]SP, and [D-Pro2,D-Trp7,9]SP were investigated at the air/water interface and when bound to lipid monolayers and bilayers. Measurement of the Gibbs adsorption isotherm showed that the surface areas of SP and its agonist (240 +/- 5 A2 at biologically relevant concentrations) were distinctly larger than those of the antagonists (138 +/- 5 A2) [Seelig, A. (1990) Biochim. Biophys. Acta 1030, 111-118]. The surface activity of the peptides increased in the order [Sar9,Met(O2)11]SP less than SP less than [D-Pro2,D-Trp7,9]SP less than [D-Arg1,D-Trp7,9,Leu11]SP = [D-Arg1,D- Pro2,D-Trp7,9,Leu11]SP and correlated with the respective binding affinities to lipid membranes. The agonist did not insert into neutral and negatively charged bilayers or into densely packed lipid monolayers (at surface pressures greater than 31 mN/m). In contrast, the three antagonists gave rise to a strong binding both to neutral and to charged lipid monolayers and bilayers. The degree of binding was evaluated from the area increase of lipid monolayers upon peptide insertion, and the binding isotherms were analyzed in terms of the Gouy-Chapman theory. At the monolayer-bilayer equivalence pressure of approximately 32 mN/m, the binding can be described by a surface partition equilibrium with binding constants of (4.5 +/- 0.1) x 10(3) M-1 for [D-Pro2,D-Trp7,9]SP and (1.3 +/- 0.1) x 10(4) M-1 for both [D-Arg1,D-Trp7,9,Leu11]SP and [D-Arg1,D-Pro2,D-Trp7,9,Leu11]SP for pure palmitoyloleoylphosphatidylcholine (POPC) membranes.(ABSTRACT TRUNCATED AT 250 WORDS)