Abstract
A sensitive and reproducible liquid chromatography–mass spectrometric (LC-MS) method has been developed and validated for the quantifi cation of doxycycline in human plasma, after liquid-liquid extraction (LLE). Best chromatographic resolution was achieved on a reverse-phase Phenomenex C18 column using the mobile phase acetonitrile–5mM ammonium acetate (80:20) in isocratic elution with a total run time of 3.5 min. The analyte, doxycycline was detected by using an electrospray ionization mass spectrometer in the selected ion monitoring (SIM) mode. Linear plot was obtained in the concentration range of 0.5–5.0 µg/ml (r2 = 0.998). Lower limit of quantifi cation (LLOQ) was found to be 100 ng/ml in 500µl of plasma. Average recovery of the analyte was found to range from 96.08 to 104.60% in plasma at the concentrations of 1.0, 3.0 and 5.0 µg/ml. Intra- and inter-day relative standard deviation was found to be 0.13-0.74 and 0.11-0.34 respectively. The present method was successfully applied in the pharmacokinetic study of doxycycline in human plasma.
Highlights
Doxycycline a member of tetracycline antibiotic group is commonly used to treat chronic prostatitis and syphilis
Limited analytical techniques were available in the assay of doxycycline such as HPLC-DAD as well as liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS)
Application of the assay: The developed liquid chromatography–mass spectrometric (LC-MS) assay method was used in the pharmacokinetic study after oral (100.0 mg) administration of doxycycline to human volunteers
Summary
Doxycycline a member of tetracycline antibiotic group is commonly used to treat chronic prostatitis and syphilis. Stock solution of doxycycline was prepared by dissolving the accurately weighed reference compound in water and acetonitrile (1:1) to give a final concentration of 1 mg/ml, stored at 4oC until it is used. The solution was serially diluted with water and mixed with blank human plasma to achieve standard working solutions at concentration of 0.5, 1.0, 2.0, 3.0, 4.0, and 5.0 g/ml for doxycycline, respectively.
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