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Quantization of Didanosine in Human Plasma using HighPerformance Liquid Chromatography–Tandem Mass Spectrometry

Hemasree Sura , Sumadhuri Bonthu , T. E. G. K. Murthy


Present study reports the development and validation of Didanosine in human plasma by LCMS/MS using electron spray ionization technique. Zidovudine was used as an internal standard. The chromatographic separation of analyte and internal standard was achieved by using ACE 5μ, C18 50*4.6mm column as stationary phase and 5mM Ammonium Acetate : methanol (5:95) as mobile phase at a flow-rate of 0.8 ml/min. MS detection was performed at transitions of m/z 235.000/135.100 and 266.000/193.000 in multiple reaction monitoring for didanosine and zidovudine at negative mode. Didanosine was extracted from the plasma by solid phase extraction using Orochem-30mg cartridges. The present method was found to be linear over the concentration range of 10.022 - 3003.561 ng/ml (r2- 0.9983). The limit of quantification of Didanosine in plasma was found to be 10.022ng/ml. The retention times of Didanosine and Zidovudine were found to be 0.77 min and 0.79 min respectively. The analyte was found to be stable under various stability tests such as freeze-thaw, bench top, wet extract, dry extract, auto sampler and interim studies. This simple, rapid and specific validated method was successfully applied for the faster analysis of Didanosine in human plasma in bioavailability and bioequivalence studies.

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