Straightforward solid-phase extraction method for the determination of verapamil and its metabolite in plasma in a 96-well extraction plate.

ABSTRACT

Straightforward solid-phase extraction (SPE) methods were developed for the determination of verapamil and its metabolite in a plasma matrix. The spiked plasma sample was pretreated with 2% phosphoric acid followed by two different SPE methods using a Waters Oasis HLB 96-well extraction plate. Recoveries greater than 90% were obtained using both a generic and a selective SPE methods. The generic method is a good starting protocol, and it is applicable to a wide range of compounds. This generic method consists of using 5% methanol as the wash solvent, and 100% methanol for the elution. The limitation of the non-specific method is that it does not remove all plasma constituents that interfere with the quantitation of the metabolite, norverapamil. A second, more specific method was developed using the same Oasis HLB sorbent which removes more plasma interferences and provides cleaner extracts for the HPLC-UV analysis. This selective method uses both the methanol concentration and the pH advantageously to preferentially isolate the analytes of interest from a complex sample matrix. Recoveries of greater than 90% with R.S.D.s less than 3.8% were obtained with this selective method.