SummaryThe low temperature effect has been investigated in reversed phase micro high performance liquid chromatography with variousaqueous mobile phasesystems by anapproach basedon enthalpy-entropy compensation. The compensation temperatures, T,, were determined for these systems, and the results show that the retention mechanism in a lower temperature column is similar to that in a normal temperature column in the reversed phase mode at mobile phase water contents above 9.8%. At lower water contents, the separation mechanism is different from that in the reversed phase mode, but otherwise very similar to that in the normal phase mode.
The free fractions of five drugs were determined in plasma treated with EDTA and in serum. The free fractions of phenytoin, sodium valproate, and phenobarbital in serum were significantly higher than in plasma (P less than 0.05). In contrast, the free fractions of carbamazepine and theophylline were significantly lower in serum than in plasma. Although we observed minor differences in the protein patterns obtained with two-dimensional electrophoresis, these did not have an important influence on protein binding. No significant differences were observed between plasma and serum in the physiological data, except for pH. Binding of these drugs by protein was shown to be pH-dependent. We conclude that serum may be better than plasma for determination of the free fraction, because the free fraction in plasma is affected by anticoagulants such as EDTA salts and heparin.
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