Determination of fluoxetine and norfluoxetine in human plasma by high-pressure liquid chromatography with fluorescence detection

Fluoxetine is an atypical antidepressant drug, which selectively inhibits the neuronal reuptake of serotonin, and is widely used in the treatment of depressive disorders. The aim of this research is the development of an HPLC method with fluorescence detection for the monitoring of fluoxetine plasma levels. The determination requires no more than 250 μl of plasma, which undergo solid phase extraction (SPE), then are injected in the HPLC. For the analytical separation a reversed phase C8 column (150×4.6 mm I.D.) was used, while the mobile phase was a mixture of acetonitrile and water containing perchloric acid and tetramethylammonium perchlorate (flow rate: 1 ml min−1). The very low levels of analytes in plasma required the employment of a fluorescence detector (λexc=230 nm, λem=290 nm), which also granted a good selectivity. Fluoxetine is revealed as a single peak at a retention time of 9.7 min, while norfluoxetine, the main metabolite of fluoxetine, is revealed at a retention time of 8.1 min. Linearity was obtained over the concentration range 8–200 ng ml−1 for both substances. The method seems suitable, in accuracy and precision, for the determination of fluoxetine plasma levels of patients; furthermore, it is rapid and sensitive.