Determination of irinotecan and its metabolite SN-38 in rabbit plasma and tumors using a validated method of tandem mass spectrometry coupled with liquid chromatography

New tandem mass spectrometric method coupled with liquid chromatography (LC–MS/MS) has been developed to determine the total concentration of camptothecin derivatives (irinotecan and SN-38) regardless of inter-conversion phenomenon between carboxylate and lactone forms. At first, all sample solutions were acidified for 1 h in order to completely convert CPT derivatives into their lactone forms and then CPT derivatives were extracted with organic solution containing diethyl ether and ethyl acetate (2:1, v/v) just after alkalization in the range pH 8.0–8.5 in acid-treated solutions. Analytes were separated on a reverse phase C18 column (150 × 2.1 mm) and eluted isocratically with a mobile phase which consisted of acetonitrile-methanol-buffer (0.1% formic acid, 5 mM ammonium formate) (3:4:3, v/v). CPT derivatives were monitored by tandem mass spectrometry in electrospay-positive ionization and multiple reaction mode programmed to the following transitions (m/z): ‘587.6 → 167.2’ of CPT-11, ‘393.6 → 349.3’ of SN-38 and ‘349.4 → 305.2’ of CPT. The method was validated to have the proper linearity (r2 > 0.99) over the range of 5–1000 ng/ml of CPT-11 and 1–250 ng/ml of SN-38 with good accuracy (89.8–114.3%) and precision (less than 10%). In all stability tests, concentration of CPT-11 and SN-38 had been left in the acceptable range of 88.8–110.7% when sample solutions were acidified before determination of CPT derivatives. Newly developed LC–MS/MS method was suitable for the determination of CPT derivatives of both rabbit plasma and tumor tissues in the pharmacokinetic study.