Immobilized artificial membrane chromatography: a rapid and accurate HPLC method for predicting bile salt-membrane interactions.
Academic Article
Overview
abstract
To predict bile salt-membrane interactions physiologically, we used an immobilized artificial membrane HPLC column that contains dimyristoyl-phosphatidylcholine molecules covalently linked to silica microspheres. Using a 90% aqueous (10% acetonitrile) mobile phase, 22 species of bile salts and 4 species of fusidates were eluted. Glycine conjugates displayed higher affinity for the column at pH 5.5, eluting later than their taurine-conjugated congeners, but this order was reversed at pH 6.5 and 7.4 as glycine conjugates became fully ionized. Capacity factors decreased logarithmically as functions of increasing temperature, permitting determinations of interaction enthalpies, which ranged from -2.86 to -7.67 kcal/mol. A standard curve was developed from which the enthalpy for an uncommon bile salt could be inferred from its capacity factor at room temperature. Bile salt interaction enthalpies were substantially better correlated than hydrophobic indices by octadecylsilane-HPLC (D. M. Heuman, J. Lipid Res. 1989. 30: 719-730) with equilibrium binding to small unilamellar vesicles and literature values reflecting bile salt-membrane interactions (e.g., biliary phosphatidylcholine secretion), but not with bile salt functions that do not require phospholipid (e.g., micellar cholesterol solubility). This new application should prove valuable for evaluating membrane-active physical-chemical properties as well as therapeutic potential of novel bile salts, particularly when they are available in quantities too small for study by conventional techniques.
