Fluoxetine Bioequivalence Study: Quantification of Fluoxetine and Norfluoxetine by Liquid Chromatography Coupled to Mass Spectrometry

Abstract

In this study, the authors assessed the bioequivalence of two fluoxetine tablet formulations in 24 healthy volunteers of both sexes who received a single 20 mg dose of each fluoxetine formulation, and a new sensitive method for the quantification of fluoxetine and norfluoxetine in human plasma was developed. The study was conducted using an open, randomized, two-period crossover design with a 4-week washout interval. Plasma samples were obtained over a 672-hour period. Plasma fluoxetine and norfluoxetine concentrations were analyzed by combined liquid chromatography coupled to mass spectrometry (LC-MS) with positive ion electrospray ionization using selected ion recording (SIR). Kolmogorov-Smirnov's test, histograms, probit plots, and the correlation between norfluoxetine AUC(0-infinity) and fluoxetine AUC(0-infinity) were used to analyze the population distribution. The limit of quantification was 0.15 ng.ml-1 and 0.50 ng.ml-1 for both fluoxetine and norfluoxetine, respectively. Within- and between-run imprecision was less than 13% and 17%, respectively. The pharmacokinetic parameters obtained for fluoxetine and norfluoxetine after the administration of each formulation included AUC(0-672 h), AUC(0-infinity), Cmax, Cmax/AUC(0-672 h), tmax, t1/2, and Ke. The AUC values for fluoxetine were not consistent with a normal distribution, reflecting the existence of two different populations (poor and extensive metabolizers). The mean pharmacokinetic parameters for extensive fluoxetine metabolizers were 27.0 ng ml-1 for Cmax, 2064.0 ng h ml-1 for AUC(0-infinity), and 85.4 h t1/2. The mean pharmacokinetic parameters for norfluoxetine (in extensive metabolizers only) were 2532.0 ng h ml-1 for AUC(0-infinity) and 8.4 ng ml-1 for Cmax. For fluoxetine bioequivalence, the 90% CI of the individual ratio geometric mean for Psiquial/Prozac (including both extensive and poor metabolizers) was 101.6% to 121.1% for AUC(0-672 h) and 86.1% to 102.6% for Cmax. For norfluoxetine, the 90% CI of the individual ratio geometric mean for Psiquial/Prozac (including both extensive and poor metabolizers) was 90.3% to 108.3% for AUC(0-672 h) and 84.5% to 106.3% for Cmax. The new method developed (LC-MS) presented high sensitivity, specificity, and short chromatographic run for the quantification of both fluoxetine and norfluoxetine in human plasma. Since both 90% CI for AUC and Cmax geometric mean ratios were included in the 80% to 125% interval proposed by the U.S. Food and Drug Administration, Psiquial was considered bioequivalent to Prozac according to both the rate and extent of absorption. The finding that there were no significant differences in the bioequivalence assessed by either fluoxetine or norfluoxetine pharmacokinetic parameters indicates that future bioequivalence trials may be performed by quantifying fluoxetine only.

Bioequivalence testing of a new tablet formulation of generic fluoxetine

“…The overall pharmacokinetic profiles offluoxetine and norfluoxetine in the present study were very close, and in agreement with the data previously published in the relevant literature [17,18,23]. Additionally, the ranges of individual maximal concentrations [15.0--53.5 ng/ml (f1uoxetine), 14.5-73.7 ng/ml (Prozac)] obtained in the study confirmed the previous finding that the absorption rate of f1uoxetine is highly variable [7].…”

“…This finding strongly supports the conclusion of Moraes and co-workers [18]that quantifying only the parent drug might satisfy future bioequivalence trials of generic f1uoxetine preparations.…”

“…According to the literature, its sensitivity was comparable to gas chromatographic-mass spectrometric assay [17] but, as expected, it was less sensitive than liquid chromatography coupled to mass spectrometry [18] or the liquid chromatographic-tandem mass spectrometric technique [19,20]. Twenty-four subjects were chosen since this number arbitrarily represents a reliable sample to assess bioequivalence [21,22].…”

Hollow fiber-based liquid phase microextraction combined with high-performance liquid chromatography for extraction and determination of some antidepressant drugs in biological fluids

“…The analytical techniques employed in the recent years for the estimation of sertraline in human plasma include HPLC, GC with popular detectors or in combination with tandem mass spectrometry (i.e. LC-MS/MS and GC-MS/MS) [12][13][14][15][16][17][18][19][20][21][22][23].…”

Indirect enantioseparation of fluoxetine in mouse serum by derivatization with 1R‐(–)‐menthyl chloroformate followed by ultra high performance liquid chromatography and quadrupole time‐of‐flight mass spectrometry

“…Thus, the rate of fluoxetine elimination is fairly slow, with a half-life of 47-72 h [2]. Several analytical methods have been reported for determining the levels of only fluoxetine [3,4] or fluoxetine and its active metabolite, norfluoxetine, in biological fluids for therapeutic drug monitoring [5][6][7] and bioavailability studies [8].…”