Partitioning of Amphiphiles between Coexisting Ordered and Disordered Phases in Two-Phase Lipid Bilayer Membranes

Abstract

The partition coefficients ( K P) of a series of single-chain and double-chain fluorescent amphiphiles, between solid ordered (P β′ and L β ) and liquid disordered (L α of the type l d) lipid phases coexisting in the same lipid bilayer, was studied using steady-state fluorescence emission anisotropy. The single-chain amphiphiles were N-(7-nitrobenzoxa-2, 3-diazol-4-yl)-alkylamines, and the double-chain amphiphiles were N-(7-nitrobenzoxa-2, 3-diazol-4-yl)-phosphatidylethanolamines with chain lengths of 12–18 carbon atoms. Saturated 18-carbon alkyl/acyl chain compounds were also compared with Δ 9- cis unsaturated chains of the same chain length. The fluorescence anisotropy of the probes was examined in lipid bilayers (multilamellar vesicles) prepared from an equimolar mixture of dilauroylphosphatidylcholine and distearoylphosphatidylcholine and studied as a function of temperature through the entire temperature range of coexistence of ordered gel phases and a disordered fluid phase in this system. The unsaturated chain amphiphiles partitioned exclusively into the fluid phase whenever this phase was present, as did the saturated chain amphiphiles with the shortest chains (C 12:0), while K P ranges between 1 and 2, in favor of the L β solid phase, for the amphiphiles with long saturated (C 18:0) alkyl/acyl chains, with intermediate behavior for the intermediate chain lengths. All probes appeared to be totally excluded from P β′ solid (gel) phases. The technique was also used to determine partitioning of some of the probes between coexisting liquid ordered (cholesterol-containing) ( l o) and liquid disordered ( l d) L α phases. In this case the ratio of signal amplitude to noise allowed us to obtain a qualitative, but not quantitative, measure of the phase partitioning of the probes. We conclude that the partitioning behavior of the probes examined between coexisting l o and l d phases is qualitatively similar to that observed between solid ordered and liquid disordered phases.