Quantitation of Irinotecan and its two major metabolites using a liquid chromatography–electrospray ionization tandem mass spectrometric

A sensitive and selective liquid chromatographic–tandem mass spectrometric (LC–MS/MS) method for the determination of Irinotecan (CPT-11) and its metabolites in human plasma has been developed. Samples were prepared after protein precipitation and analyzed on a C18 column interfaced with a Q-Trap tandem mass spectrometer. Positive electrospray ionization was employed as the ionization source. The mobile phase consisted of acetonitrile–water (0.05% formic acid), using gradient procedure. The analytes and internal standard camptothecin were both detected by use of multiple reaction monitoring mode. The method was linear in the concentration range of 10.0–2000.0 ng/ml for CPT-11 and 0.5–200.0 ng/ml for 7-ethyl-10-hydroxycamptothecin (SN-38), respectively. The lower limit of quantification (LLOQ) was 10 ng/ml for CPT-11 and 0.5 ng/ml for SN-38. The intra- and inter-day relative standard deviation across three validation runs over the entire concentration range was less than 10.6%. The accuracy determined at three concentrations was within ±11.4% in terms of relative error. Due to the unavailability of standard for 7-ethyl-10-O-glucuronyl-camptothecin (SN-38G) and the importance of knowing the concentration of this metabolite, we developed a method for analysis SN-38G by taking advantage of the quantitive conversion of SN-38G to SN-38 using glucuronidase. This enzymatic method of identification and quantitation of gluconated compound can be widely used when the standard for phase II glucuronide metabolites are not available.