Determination of Midodrine and its Active Metabolite Desglymidodrine by LC-MS/MS and its Application in Human Pharmacokinetic Studies

Abstract

Background: Midodrine (MD) is a prodrug which is converted into Desglymidodrine (DMD) after oral administration. </P><P> Objective: The aim of the present study is to develop and validate a precise, accurate liquid chromatography- tandem mass spectroscopy (LC-MS/MS) method for the separation and detection of Midodrine and Desglymidodrine. Methods: The quantification of prodrug Midodrine (MD) and its active metabolite Desglymidodrine (DMD) in human plasma was performed using simple and economical protein precipitation method. Caffeine was used as an internal standard (IS). LC separation was carried out using Jones C18 column (4.6mm x 150mm, 3µm). Isocratic elution was performed using 10mM ammonium formate (pH 4.0 adjusted with formic acid): methanol 30:70, v/v as a mobile phase, at a constant of flow of 0.5 ml/min. Results: Mass spectrometric detection was carried out at positive electrospray ionization with proton adducts at m/z 255.0˃237.1, 198.1˃180.2 and 195.0˃138.1 for Midodrine, Desglymidodrine and caffeine (IS) respectively, in MRM mode. The method was validated over a linear concentration range of 0.3-110 ng/ml (r2 =0.996 and r2 =0.9988) for both Midodrine and Desglymidodrine. The recovery of Midodrine and Desglymidodrine were found to be 99 ± 0.12%. The precision (intra-day and inter-day) and accuracy studies fulfilled the acceptance criteria. Conclusion: The method shows to be stable for the studied stability parameters and was successfully applied for clinical pharmacokinetic studies.