Simultaneous determination of citalopram, fluoxetine, paroxetine and their metabolites in plasma and whole blood by high-performance liquid chromatography with ultraviolet and fluorescence detection

• Published in: Journal of chromatography. B, Biomedical sciences and applications Vol. 734; no. 2; pp. 229 - 246
• Main Authors: Kristoffersen, L, Bugge, A, Lundanes, E, Slørdal, L
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 12.11.1999; Elsevier Science
• Subjects: Acetonitriles; Biological and medical sciences; Chromatography, High Pressure Liquid - methods; Citalopram; Citalopram - blood; Fluoxetine; Fluoxetine - blood; Humans; Hydrogen-Ion Concentration; Medical sciences; Neuropharmacology; Paroxetine; Paroxetine - blood; Pharmacology. Drug treatments; Psychoanaleptics: cns stimulant, antidepressant agent, nootropic agent, mood stabilizer..., (alzheimer disease); Psychology. Psychoanalysis. Psychiatry; Psychopharmacology; Quality Control; Sensitivity and Specificity; Serotonin Uptake Inhibitors - blood; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet; Paroxetine; Fluoxetine; Citalopram; Human; Biological fluid; Fluorescence detector; Metabolite; HPLC chromatography; Blood plasma; Whole blood; Ultraviolet detector; Analytical method; Antidepressant agent; Quantitative analysis
• ISSN: 0378-4347; 1387-2273
• DOI: 10.1016/S0378-4347(99)00352-7

A method for the simultaneous determination of the three selective serotonin reuptake inhibitors (SSRIs) citalopram, fluoxetine, paroxetine and their metabolites in whole blood and plasma was developed. Sample clean-up and separation were achieved using a solid-phase extraction method with C 8 non-endcapped columns followed by reversed-phase high-performance liquid chromatography with fluorescence and ultraviolet detection. The robustness of the solid-phase extraction method was tested for citalopram, fluoxetine, paroxetine, Cl-citalopram and the internal standard, protriptyline, using a fractional factorial design with nine factors at two levels. The fractional factorial design showed two significant effects for paroxetine in whole blood. The robustness testing for citalopram, fluoxetine, Cl-citalopram and the internal standard revealed no significant main effects in whole blood and plasma. The optimization and the robustness of the high-performance liquid chromatographic separation were investigated with regard to pH and relative amount of acetonitrile in the mobile phase by a central composite design circumscribed. No alteration in the elution order and no significant change in resolution for a deviation of ±1% acetonitrile and ±0.3 pH units from the specified conditions were observed. The method was validated for the concentration range 0.050–5.0 μmol/l with fluorescence detection and 0.12–5.0 μmol/l with ultraviolet detection. The limits of quantitation were 0.025 μmol/l for citalopram and paroxetine, 0.050 μmol/l for desmethyl citalopram, di-desmethyl citalopram and citalopram- N-oxide, 0.12 μmol/l for the paroxetine metabolites by fluorescence detection, and 0.10 μmol/l for fluoxetine and norfluoxetine by ultraviolet detection. Relative standard deviations for the within-day and between-day precision were in the ranges 1.4–10.6% and 3.1–20.3%, respectively. Recoveries were in the 63–114% range for citalopram, fluoxetine and paroxetine, and in the 38–95% range for the metabolites. The method has been used for the analysis of whole blood and plasma samples from SSRI-exposed patients and forensic cases.