Abstract
The effect of temperature on the retention and multiple separation of hydrocortisone, testosterone, 17α-methyltestosterone, prednisone, cortisone and 17α-hydroxyprogesterone in reversed-phase liquid chromatography has been studied. Capacity factors ( k′) of the steroids were measured using a mobile phase modified with different concentrations of β-cyclodextrin (from 0–16 mM), a fixed solvent composition (acetonitrile—water) and a wide range of column temperatures (from 5–80°C). The plots of capacity factors vs. reciprocal of absolute temperature are nonlinear in every case when mobile phase modified with β-cyclodextrin was used. Particularly strong nonlinearity was observed at lower temperature and at higher β-cyclodextrin concentration. The complex chromatograms were evaluated using optimization parameters such as capacity factor of the last-eluted peak ( k′ max), the smallest resolution between adjacent peaks ( R s; min) and relative resolution product ( r). The results presented describe precisely the role of temperature in high performance liquid chromatography systems in which mobile phases modified with cyclodextrin were used.
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