NMR studies of the bile salt-lecithin mixed micelle

Abstract

Sodium cholate labelled with deuterium at the 7(beta) position has been prepared. The deuterium NMR linewidths were used to calculate the relaxation times (T(,1)'s) of this nucleus. A plot of ln T(,1) vs. the reciprocal temperature gave the activation energy for tumbling of the cholate molecule. Use of the Debye-Stokes-Einstein relationship confirmed that sodium cholate forms tetramers above the critical micellear concentration, but demonstrated that tetramer formation is not complete until the bile salt reaches a concentration of 0.1 M. Once formed, the tetramers do not break up with increasing temperature. The presence of egg yolk lecithin decreased the deuterium T(,1)'s at higher temperature only. This was interpreted on the basis of an equilibrium between tetrameric cholate and cholate bound to lecithin. In the presence of NaCl a similar equilibrium was observed between tetramers of cholate and very large cholate micelles. The ('31)P NMR of lecithin was studied in the presence of sodium cholate and desoxycholate. The sample initially displayed two peaks, corresponding to two species undergoing chemical exchange. The upfield peak had a T(,1) and chemical shift typical of lecithin in a bilayer. This peak disappeared after 2 days, which contradicts the Small model of the bile salt-lecithin mixed micelle. The remaining peak is postulated to correspond to the cylindrical aggregates observed by Small. A model is proposed whereby cholate causes the bilayer vesicles of lecithin to break up. This forms bilayer discs, which exist in equilibrium with the cylinders. At pH 13 cholate was found to rapidly hydrolyze lecithin, suggesting that the bile salt molecules cluster around the lecithin aggregates with their carboxylate tails inward.