| 20(5)-Camptothecin (CPT) is a natural alkaloid extracted from the leaves and fruits of Camptotheca acuminata, and 9-nitro-20(S)-camptothecin (9-NC), an analog of CPT, has been proved the antitumor activity. The antitumor effect of CPT and its analogs is thought to be based on its inhibition of DNA topoisomerase I. In this study, the pharmacokinetic characters of 9-NC in different animals were systematically investigated with HPLC and LC/MS/MS method.The dissertation is summarized as follows:1. Investigation of absorption of 9-NC in different speciesAn HPLC-UV assay and a more sensitive LC/MS/MS method were developed to determine the concentrations of 9-NC in plasma, and successfully applied to investigate the pharmacokinetics in rats and dogs.The analytes were extracted from plasma samples by liquid-liquid extraction, separated through a Hypersil BDS C18 column, and camptothecin was used as the internal standard. The mobile phase consisting of a mixture of acetonitrile-water-formic acid (35: 65: 2, v: v: v) was applied in the HPLC method, and the UV detector was set at 370 nm. The linear range of concentrations for 9-NC in plasma were 25-1600 ug L-1. The lower limit of quantitation (LLOQ) was 25 ug L-1. By the LC/MS/MS method, the analytes were detected by tandem mass spectrometry with an electrospray ionization interface, and the mobile phase consisted of a mixture of acetonitrile-water-formic acid (80: 20: 2, v: v: v). Selected reaction monitoring (SRM) using the precursor → product ion combination of m/z 435.0 → 394.0, and m/z 390.0 → 349.0 was used to quantify 9-NC and internal standard, respectively. The linear calibration curves were obtained in the concentration range of 0.5-50 ug L-1, and the lower limit of quantitation (LLOQ) was 0.5 u g L-1.After iv administration of 9-NC at the doses of 1.5, 3, and 6 mg kg-1 to rats, the AUC-dose relationship was evaluated to be linear (r > 0.85, P < 0.001). Following ig administration of 9-NC at the doses of 3, 6, and 12 mg kg-1, the linearity between Cmax and doses was good (r = 0.65, P < 0.01), however, the AUC-dose relationshipwas nonlinear. The results of the compartmental analysis indicated that the kinetic process of 9-NC in rats in vivo was best fitted to a two-compartment model. The absolute oral bioavailability of 9-NC in rats was calculated to be 14.6%.Following treatment with 9-NC at 0.5, 1, and 2 mg kg-1 to dogs by iv, the AUC-dose relationship was evaluated to be linear (r > 0.96, P < 0.001). After ig administration of 9-NC at the doses of 1, 2, and 4 mg kg-1 to dogs, AUC- and Cmax-dose relationships were evaluated to be linear (r > 0.76, P < 0.001). The results of the compartmental analysis indicated that the kinetic process of 9-NC in dogs in vivo was best fitted to a two-compartment model. The absolute oral bioavailability of 9-NC in dogs was calculated to be 6.1%.At the plasma concentrations of 9-NC 100, 250, 500, and 1000 ug L-1, theplasma protein binding of 9-NC in rats were 95.9+0.8, 96.2 + 0.4, 97.6+0.4, and 96.0 + 0.5%, respectively.The results above revealed a low absolute bioavailability of 9-NC and a high binding between the drug and the plasma protein.2. Investigation of the distribution of 9-NC in ratsAn HPLC-UV method was developed to measure the concentration of 9-NC in tissues, and the distribution characters of 9-NC in rats in vivo were studied.The analytes were extracted from tissue homogenate samples by liquid-liquid extraction, separated through a Hypersil BDS C18 column, and camptothecin was used as the internal standard. The mobile phase consisted of a mixture of acetonitrile-water-formic acid (30: 70: 2, v: v: v), and the UV detector was set at 370 nm. The linear range of concentrations for 9-NC in heart, lung, spleen, stomach, fat, womb, and ovary was 10-1000 ng g-1, and the lower limit of quantitation (LLOQ) was 10 ng g-1. The linear range of concentrations for 9-NC in brain, kidney, liver, intestine, smooth muscle, skeletal muscle, and tectical was 5-500 ng g-1, and the lower li... |
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