Determination of the antidepressant fluoxetine in human plasma by LC with UV detection

A selective and sensitive isocratic high-performance liquid chromatographic (HPLC) method was developed for the quantitative analysis of low concentrations of fluoxetine (FLX) in human plasma, with ultra-violet detection at 226 nm. A reversed-phase column, LiChrospher 60 RP-Select B (125 x 3 mm i.d., 5 microm) (Merck), was used to resolve FLX and diazepam (DZP) (internal standard) from endogenous matrix interferences. FLX was isolated from plasma by liquid-liquid extraction. Two identical HPLC systems were used, both validated under the same study conditions. Each chromatographic separation was completed in 30 min and the results showed a mean relative recovery of 101 and 99.3% and an overall precision (RSD%) of 4.78 and 6.09 for each HPLC system. The standard curve was linear for FLX concentrations over the range of 5.00-50.0 ng ml(-1) (R = 0.997 and 0.998). The limit of quantitation of FLX was 5 ng ml(-1) for both HPLC systems. The method described was applied to the analysis of plasma samples obtained from healthy subjects treated with one single oral dose of 40 mg of fluoxetine.