Liquid Chromatography-Tandem Mass Spectrometry Method For The Quantification Of A Potent H-3 Receptor Antagonist Conessine In Serum And Its Application To Pharmacokinetic Studies

Conessine, a steroidal alkaloid obtained from the bark and seeds of the plant species of Apocynaceae family, elicits a histamine antagonistic action, selectively for the H-3 histaminergic receptors. This alkaloid is used mainly for the treatment of dysentery and helminthic disorders. For the quantification of conessine in serum, a liquid chromatography-tandem mass spectrometry method was developed. Chromatographic separation was achieved on a Zorbax SB-CN column (100 x 4.6 mm, 3.5 mu(m)), and a mobile phase consisting of 90% methanol in aqueous ammonium acetate buffer (pH 3.5) with 0.1% (v/v) formic acid at an isocratic flow rate of 0.6 ml/min at 40 degrees C provides efficiency in separation. A volume of 40 mu l was injected each time and the run time for each sample was 5 min. Phenacetin (internal standard) was added to 50 ml of serum sample prior to liquid-liquid extraction using 3% isopropanol in n-hexane. The detection was performed on a 5500 QTRAP mass spectrometer by multiple reaction monitoring mode via electrospray ionization source. The multiple reaction monitoring of conessine and IS was m/z 357.4 to m/z 312.1 and m/z 180.1 to m/z 138.1, respectively. The method that showed selectivity and linearity in the range of 1-200 ng/ml was validated in terms of sensitivity, accuracy, precision and stability. The detection and quantitation limits were recognized at 0.1 and 1 ng/ml, respectively. The intra-and inter-day precision and accuracy fulfils the acceptance criteria. Applying the method to the pharmacokinetic studies in rats, conessine showed a peak serum concentration at 2 h post oral dose with a good bioavailability of 71.28 +/- 4.65%.