Transfer of long-chain fluorescent fatty acids between small and large unilamellar vesicles

Transfer of 12-(9-anthroyloxy)stearic acid (12AS) was measured between small unilamellar vesicles (SUV) and between large unilamellar vesicles (LUV), over a temperature range of 5-50 degrees C. The results of this study clearly establish the biexponential nature of the time dependence of the transfer in a variety of vesicle types and confirm our previous results using egg phosphatidylcholine (EPC) SUV at 25 degrees C (Storch & Kleinfeld, 1986). In our previous study we developed a kinetic model of the transfer process and concluded that the observed time dependence of the transfer of long-chain 12-(9-anthroyloxy) fatty acids (AOFA) was due to transbilayer flip-flop that was much slower than the rate at which the fatty acids (FA) move from the vesicle and into the surrounding aqueous phase (the off step). In the present study, experimental and theoretical advances have allowed us to examine, in detail, predictions of the kinetic model that critically depend upon the slow rate of flip-flop. The current results verify these predictions and demonstrate that slow AOFA flip-flop is rate limiting in at least three different vesicle systems and at all temperatures studied. Moreover, both flip-flop and the off rate constants were almost an order of magnitude smaller in EPC-LUV than in EPC-SUV. Flip-flop was found to be asymmetric (the rate constant for transfer from the inner to outer hemileaflet of the bilayer is approximately twice that from the outer to inner hemileaflet) in SUV but virtually symmetric in LUV. The temperature dependence of transfer was used to determine the thermodynamic activation potentials for the flip-flop and off rate constants.(ABSTRACT TRUNCATED AT 250 WORDS)