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Colloidal phases (self-assemblies) in aqueous dispersions of selected binary bile salt/phospholipid blends were studied utilizing the combined analytical approach of asymmetrical flow field-flow fractionation (AF4) and multi-angle laser light scattering (MALLS) in order to resolve the co-existence of different colloidal assemblies. The binary blends were prepared by freeze-drying from tert-butanol/water co-solvent solutions. The blends contained one of two bile salts (sodium taurocholate (TC) or sodium glycodeoxycholate (GDX)) and a mono- or di-acyl phospholipid (lyso-phosphatidylcholine (L-PC) and phosphatidylcholine (PC), respectively). Bile salt and phospholipid (PL) concentrations and their respective ratios were varied systematically within the physiological range found in human intestinal fluids. Furthermore, the BCS class II drug Celecoxib was incorporated in selected blends to assess its potential impact on colloidal phases. To further investigate the smallest self-assemblies observed in AF4/MALLS analysis, dispersions of TC and GDX, respectively, were prepared and analyzed by dynamic light scattering (DLS).AF4/MALLS analysis revealed that binary bile-salt/phospholipid blends form three distinct particle fractions, when the concentration of bile-salt was sufficiently high (≥3.5 mM). Those fractions were assumed to be very small pure bile-salt dimeric/oligomeric self-assemblies (ϕ ≈ 2–3 nm), mid-sized mixed micelles (ϕ ≈ 10–50 nm) and large liposomes/aggregates (ϕ ≈ 150–280 nm). If present, Celecoxib was found solubilized within the structures, but at the lowest TC concentration triggered the formation of an additional (vesicular) phase.Separation of colloidal phases in artificial intestinal fluids by flow field-flow fractionation.