R 56865 and flunarizine displace Ca2+ from phosphatidylserine monolayers in a stoichiometric manner

The monovalent cationic drugs, R 56865 and flunarizine added to the subphase, displaced Ca2+ from phosphatidylserine (PS) monolayers at pH 5 with EC50 values of 1.2 X 10(-6) and 1.3 X 10(-6) mol/l. The EC50 of flunarizine for the increase in surface pressure due to incorporation of the drug into the PS film was 1.8 X 10(-6) mol/l. The maximal binding of both drugs to PS monolayers was estimated from experiments in which the drug was added to the spreading solution for the monolayer. The maximal molar binding ratio of R 56865 was identical to twice the maximal Ca2+ binding (2 X 0.13 Ca/PS) to the monolayer. The uptake of [14C]R 56865 into PS monolayers could be measured directly and was well described by a Langmuir isotherm in which the EC50 for 45Ca displacement was taken as the concentration for half-maximal binding, and the molar ratio for twice the Ca2+ binding was taken as maximal molar binding ratio. These results show that two molecules of the univalently charged R 56865 displace one molecule of Ca2+. Maximal binding of the cationic drug flunarizine to PS monolayers showed a similar stoichiometry. 45Ca displacement data may thus be useful for predicting monolayer and perhaps also membrane concentrations of the charged species of cationic drugs.