The use of methyl-β-cyclodextrin to solubilize cholesterol prior to coating onto a C18 stationary phase

The use of methyl-β-cyclodextrin (MBCD) as a mobile phase additive in reversed-phase liquid chromatography is explored, with the primary goal of using MBCD to solubilize cholesterol in reversed-phase mobile phases for cholesterol-coating of C18 stationary phases. MBCD is shown to increase the solubility of cholesterol in typical reversed-phase mobile phases, especially when the stoichiometric ratio of MBCD to cholesterol exceeds 2:1. Additional equivalents of MBCD further increase solubility, or allow for weaker solvents to be used. The use of weaker solvents allows for larger coating levels of cholesterol onto a C18 stationary phase than are possible without the use of MBCD. Stationary phases coated with cholesterol using MBCD as a co-additive have different selectivity than uncoated phases, especially with regards to phenyl and shape selectivity. Further, the use of MBCD as a mobile phase additive for the elution of cholesterol is examined. It is seen via van’t Hoff analysis that the reduction in retention of cholesterol when MBCD is added to the mobile phase is enthalpically driven.