By Anil. K. Patni, Tausif Monif, A. H. Khuroo, Ashok. K. Tiwary
A simple, sensitive and reproducible high performance liquid chromatography tandem mass
spectrometric method was developed and validated for the determination of itraconazole and
hydroxy itraconazole in human plasma using itraconazole-d9 and hydroxy itraconazole-d8,
respectively, as internal standards. The precursor to product ion transitions of m/z 705.4
?392.5, m/z 716.5 ? 402.6, m/z 721.3 ? 408.4 and m/z 729.4? 416.5 were used to detect and
quantify itraconazole, hydroxy itraconazole, itraconazole-d9 and hydroxy itraconazole-d8
respectively. Sample preparation was carried out by solid phase extraction using hydrophilic-
lypophilic balance cartridges and the processed sample was chromatographed on Chromolith
speed rod RP-18e (50 × 4.6 mm) column using 10 mM ammonium formate buffer (pH 4.0):
methanol:: 20:80 v/v as the mobile phase. The weighed (1/x
) calibration curves were linear
over the range of 0.301 to 151.655 ng/ml and 0.946 to 224.908 ng/ml for itraconazole and
hydroxy itraconazole respectively. The RSD of intra-day and inter-day assay was =15%.
Extraction procedure yielded a recovery of 52.07%, 53.73%, 49.68% and 50.16% for
itraconazole, hydroxy itraconazole, itraconazole-d9 and hydroxy itraconazole-d8 respectively.
The validated method was successfully employed for establishing pharmacokinetic parameters of
two formulations of itraconazole 100 mg capsule in 14 healthy male Indian volunteers.
Key Words : itraconazole, hydroxy itraconazole, liquid chromatography tandem mass
spectrometry, human plasma, pharmacokinetic study
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