Retention behaviour of a bonded reversed phase in a high-performance liquid chromatographic assay of serum theophylline

A simple, rapid assay for serum theophylline is described. Optimisation of the high-performance liquid chromatographic system has allowed the analysis of minimun sample volume (50 micronl) in the shortest time (8 min) with good between-batch precision (C.V. = 8.3%)and accuracy (recovery ca. 100%). The behaviour of theophylline and other xanthine derivatives, theobromine, caffeine and 8-chlorotheophylline, on a bonded octadecyl reversed-phase system suggests that C18 phase effects solute separation by mixed retention mechanisms rather than by pure reversed-phase chromatography. This behaviour is attributed to the participation of underivatised silanol groups in the chromatographic process, and observation corroborated by independent investigation. The mechanism of the silanol participation is difficult to rationalise; however, a normal-phase liquid partition system complies with the experimental data.