Procaine incorporation into human erythrocyte membrane--a spin label study

The positioning of procaine among the components of human erythrocyte membranes was investigated by electron paramagnetic resonance (EPR) using several spin labelled molecules: fatty acids 5-, 12- and 16-doxyl-stearic acids (5DSA, 12DSA, 16DSA), procaine (SLP) and a quaternary ammonium salt (CAT4). It was proved that while procaine significantly modified EPR parameters of both CAT4 and 5DSA, it had no effect at the level of the twelfth or sixteenth carbon atom of the membrane lipid chains. Simultaneous incorporation of 5DSA and SLP and SLP in erythrocyte membranes revealed a shorter than 10 A distance between paramagnetic centers of 5DSA and SLP. It is inferred that procaine interacts with the membrane not only at lipid water interface, but also (at least) up to the fifth carbon atom of the hydrophobic tail of phopolipids. Procaine inclusion does not change the transition point (40 degrees C) at which a break appears in the temperature dependence of the erythrocyte volume, hence no strong interaction between the procaine and the cytoskeleton or the integral proteins responsible for this phase transition occurs. In the temperature domain of 25-42 degrees C, procaine seems to be freely diffusible within the membrane (Debye model can be applied).