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Enantiomeric separation and quantification of fluoxetine (Prozac®) in human plasma by liquid chromatography/tandem mass spectrometry using liquid‐liquid extraction in 96‐well plate format
Author(s) -
Shen Zhongzhou,
Wang Sui,
Bakhtiar Ray
Publication year - 2002
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.580
Subject(s) - chemistry , chromatography , atmospheric pressure chemical ionization , mass spectrometry , selected reaction monitoring , elution , tandem mass spectrometry , liquid chromatography–mass spectrometry , triple quadrupole mass spectrometer , ammonium acetate , high performance liquid chromatography , analytical chemistry (journal) , chemical ionization , ionization , organic chemistry , ion
Fluoxetine (Prozac®) is currently one of the widely prescribed selective serotonin reuptake inhibitors (SSRIs) for the treatment of depression. A high‐throughput sample preparation procedure using liquid‐liquid extraction (LLE) in a 96‐well plate format in conjunction with liquid chromatography/tandem mass spectrometry (LC/MS/MS) was developed and validated for quantification of fluoxetine enantiomers in human plasma. After addition of internal standard and ammonium hydroxide, samples were extracted with ethyl acetate. The organic extract was evaporated to dryness and reconstituted in methanol. Where possible, sample transfer and LLE steps were automated using a Tomtec Quadra 96 workstation. Adequate separation of fluoxetine enantiomeric pairs (resolution of 1.17) was achieved on a vancomycin column eluted with methanol containing 0.075% (by weight) ammonium trifluoroacetate. A triple quadrupole mass spectrometer, operated in the multiple reaction monitoring mode at m/z 310 → 44 for fluoxetine enantiomeric pairs and m/z 287 → 241 for oxazepam (internal standard), was used. Analysis was performed in the positive ion mode using atmospheric pressure chemical ionization (APCI). The standard curve range was 2.0–1000 ng/mL for each fluoxetine enantiomer. The intra‐ and inter‐day precision and accuracy of the quality control (QC) samples were <12.5% (CV) and <13.6% (CV), respectively, for each fluoxetine enantiomer; the correlation coefficient was >0.990. Method ruggedness was demonstrated by the reproducible performance of the assay during a 3‐day validation period. Copyright © 2002 John Wiley & Sons, Ltd.