COMPLETE MAPPING OF CRYSTALLIZATION PATHWAYS DURING CHOLESTEROL PRECIPITATION FROM MODEL BILE - INFLUENCE OF PHYSICAL-CHEMICAL VARIABLES OFPATHOPHYSIOLOGIC RELEVANCE AND IDENTIFICATION OF A STABLE LIQUID-CRYSTALLINE STATE IN COLD, DILUTE AND HYDROPHILIC BILE SALT-CONTAINING SYSTEMS

Using complementary physical-chemical techniques we defined five diffe rent crystallization pathways as functions of time (30 days) and incre asing lecithin (egg yolk) content in pathophysiologically relevant mod el biles supersaturated (cholesterol saturation indices, 1.2-2.7) by d ilution of approximate to 29 g/dl bile salt-lecithin-cholesterol micel lar solutions. As evidenced by quasi-elastic light-scattering spectros copy, supersaturation was heralded by the appearance of unilamellar ve sicles. With the lowest lecithin contents, arc-like crystals with habi t and density (d 1.030 g/mL) consistent with anhydrous cholesterol app eared first and evolved via helical and tubular crystals to form plate -like cholesterol monohydrate crystals (d 1.045 g/mL). With higher lec ithin fractions, cholesterol monohydrate crystals appeared earlier tha n are and other transitional crystals. With typical physiological leci thin contents, early liquid crystals (d 1.020 g/mL) were followed by c holesterol monohydrate crystals and subsequent appearances of are and other intermediate crystals. With higher lecithin contents, liquid cry stals were followed by cholesterol monohydrate crystals only, and at t he highest lecithin mole fractions, liquid crystals appeared that did not generate solid crystals. Added calcium increased solid crystal num ber in proportion to its concentration (5-20 mM) but did not influence appearance times, crystallization pathways, or micellar cholesterol s olubilities. Decreases in temperature (37 degrees --> 4 degrees C), to tal lipid concentration (7.3 --> 2.4 g/dL), and bile salt hydrophobici ty (3 alpha,12 alpha --> 3 alpha,7 alpha --> 3 alpha,7 alpha,12 alpha --> 3 alpha,7 beta hydroxylated taurine conjugates) progressively shif ted all crystallization pathways to lower lecithin contents, retarded crystallization, and decreased micellar cholesterol solubilities. The lecithin content of mother biles decreased markedly during crystalliza tion especially where liquid crystals were a coexisting phase at equil ibrium. This systematic study provides a framework for understanding c holesterol crystallization in human and animal biles and for examining factors that influence the kinetics of phase separation.