PICOGRAM DETERMINATION OF AN AVERMECTIN ANALOG IN DOG PLASMA BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH FLUORESCENCE DETECTION

A sensitive and automated method for the determination of a new averme ctin analog (MK-324) in dog plasma has been developed and validated. T he drug and internal standard were extracted from plasma via solid-pha se extraction using an automated Gilson ASPEC XL system. The extracts were evaporated to dryness and reconstituted in a mixture of N,N-dieth ylmethylamine and acetonitrile. Automated pre-column derivatization of the reconstituted extracts using trifluoroacetic anhydride was perfor med prior to injection onto the HPLC system. The fluorescent derivativ es of the drug and internal standard were chromatographed using a Zorb ax RX-C18 column and a mobile phase composed of acetonitrile, water an d tetrahydrofuran [6:1:1 (v/v/v)]. The assay was validated in the conc entration range of 100 - 5,000 pg/mL. Its applicability, inter-day per formance and specificity were demonstrated by analyzing plasma samples from six studies with MK-324.