Received for publication March 3, 2004.
Revised April 13, 2004.
Accepted for publication April 14, 2004.

Dipole Potential and Headgroup Spacing Are Determinants for the Membrane Partitioning of Pregnanolone

Abstract

ABSTRACT The membrane interactions of pregnanolone, an intravenous general anesthetic steroid were characterized using fluorescence spectroscopy and monolayer technique. Di-8-ANEPPS, a membrane dipole potential () sensitive probe, revealed pregnanolone to decrease similarly as previously reported for other anesthetics. The decrement in was approx. 16 mV and 10 mV in dipalmitoylphosphatidylcholine (DPPC) and DPPC:cholesterol (90:10, mol:mol) vesicles, respectively. Diphenylhexatriene (DPH) anisotropy indicated pregnanolone to have a negligible effect on the acyl chain order. In contrast, substantial changes were observed for the fluorescent dye Prodan, thus suggesting pregnanolone to reside in the interfacial region of lipid bilayers. Langmuir balance studies indicated increased association of pregnanolone to DPPC monolayers containing cholesterol or 6-ketocholestanol at surface pressures >20 mN/m, as well as to monolayers of the unsaturated 1-palmitoyl-2-oleoylphosphatidylcholine (POPC). In the same surface pressure range the addition of phloretin, which decreases , reduced the penetration of pregnanolone into the monolayers. These results suggest membrane partitioning of pregnanolone to be influenced by the spacing of the phosphocholine headgroups as well as membrane dipole potential. The latter can be explained in terms of electrostatic dipole-dipole interactions between pregnanolone and the membrane lipids with their associated water molecules. Considering the universal nature of these interactions, they are likely to affect membrane partioning of most, if not all, weakly amphiphilic drugs.

Key words: Fluorescence techniques, Optical spectroscopy (fluorescence, DC, etc.), Gases/general anesthetics