| BackgroundHydroxyurea is antitumor drug of DNA nucleotide inhibitors, Once was applied for blood disease malignancy, sickle cell anemia and myeloproliferative syndrome.But it is so serious aderse effects that clinical application of it is limited. (1-(4-bromobenzyl)-1-(4-bromobenzyloxy)urea,HY-D11) is one of the most promising series of derivatives which were come from HU. In vivo and vitro tests on animals it presented strong antitumor effect, and low toxicity.Pharmacokinetic studies is very important in new drug development,and investigated the process of HY-D11 in vivo, and the metabolism is important significance for HY-D11 as an new anti-tumor drug.ObjectivesThe aim of this study is to investigate the pharmacokinetic of 1-(4-bromobenzyl)-1-(4-bromobenzyloxy)urea profiles in rats and metabolism of 1-(4-bromobenzyl)-1-(4-bromobenzyloxy)urea in rat liver microsome incubation system,and provide theoretical and experimental basis of research and development for the HY-D11 as new antineoplastic drug.Methods①Seting up the HPLC analyse method of HY-D11 to detect the concentration of HY-D11 in rat plasma and live microsomal.②Investigate the effect of incubation time,substrate concentration and microsomal protein concentration on the enzyme kinetics of the metabolism of HY-D11 in rat live microsome incubation solution.Various selective CYP inhibitors (CYP1A inhibitor:α-naphthoflavone,CYP2B inhibitor:sertraline,CYP2C inhibitor: quercetin,CYP2D inhibitor:yohimbine, CYP2E inhibitor:disulfiram and CYP3A inhibitor:troleandomycin) were used to investigate their inhibitory effects on the metabolism of HY-D11. Oral administration of HY-D11 to six SD rats with 66.7mg/kg, collecting plasma samples before and after administration of HY-D11 in different time. Detecting the concentration of HY-D11 in plasma with RT-HPLC.Acquained the pharmacokinetic parameters by using the DAS2.0 software.③Oral administration of HY-D11 to six SD rats with 66.7mg/kg, collecting plasma samples before and after administration of HY-D11 in different time. Detecting the concentration of HY-D11 in plasma with RT-HPLC.Acquained the pharmacokinetic parameters by using the DAS2.1 software.Results①The HPLC detection method of HY-D11:mobile phase:cetonitrile-10mM ammonium acetate(65:35,V:V); Velocity:1.0 ml/min; detected wavelength:225nm; sensitivity:0.004Aufs; Column temperature:room temperature.②The metabolism of HY-D11 in live microsome incubation solution was nearly linear time dependence, When put HY-D11 in live microsome incubation solution in 37℃from 0 to 60 min. When incubation time is more than 60min, HY-D11 metabolic rates was decreased, so choose 60 min as experimental incubation time. The elimination of HY-D11 was increased with the increasing protein concentration. The elimination of HY-D11 was linear increase when the protein concentration was 0.25 to 1.5 mg/ml,so we choosed the 1.0 mg/mL liver microsomal protein.The metabolism of HY-D11 in live microsome incubation solution showed the characteristic of enzyme kinetics:In the concentration range of 0-16μM, the disappeared rate of Ost in live microsome incubation solution all linearly increase, when the concentration above 16μM, the metabolism of HY-D11 showed saturation. When the live microsome incubation solution maintained sealed in air at 37℃in a shaking bath for 0-90 min, the disappeared rate of Ost all linearly increased.③a-naphthoflavone(an inhibitor of CYP1A),sertraline (an inhibitor of CYP2B),quercetin (an inhibitor of CYP2C),yohimbine (an inhibitor of CYP2D),disulfiram (an inhibitor of CYP2E) did not have effect on the HY-D11 metabolism in live microsome incubation solution(P>0.05).troleandomycin,an inhibitor of CYP3A,reduced HY-D11 metabolism in live microsome incubation solution. (P<0.05).④The pharmacokinetic profiles of HY-D11 is described by two compartment model legitimately after oral administration of HY-D11 (66.7 mg/kg) in rats. Tmax is 3.67±0.52 h, Cmax is 1610.35±130.28 ng-mL-1, Vz/F is 9540.42±1530.62 mL-kg-1.tl/2z is 3.66±0.25 h,MRT(0-∞) is 7.05±0.29 h.ConclusionsThe HPLC detection method of HY-D11 we have established are Sensitivity and reliability.The linear range of HY-D11 in rat plasma was 50 to 2400 ng·mL-1. The linear range of HY-D11 in rat live microsome incubation was 0.25 to 32μM. The recovery of HY-D11 in low.medium and high concentration were between 85%and 115%, RSD all were less than 15%. It could meet nalysis requirements.The metabolism of HY-D11 in live microsome incubation solution showed the characteristic of enzyme kinetics,a-naphthoflavone (an inhibitor of CYP1A),sertraline (an inhibitor of CYP2B),quercetin (an inhibitor of CYP2C),yohimbine (an inhibitor of CYP2D),disulfiram (an inhibitor of CYP2E) did not have effect on HY-D11 metabolism in live microsome incubation solution(P>0.05),It Shows that CYP2B, CYP2C, CYP1A, CYP2D, CYP2E don't participate in HY-D11 metabolism in rat liver. Troleandomycin, an inhibitor of CYP3A, reduced HY-D11 metabolism in live microsome incubation solution. (P<0.05).It shows that the metabolism of HY-D11 in rats may be mediated by CYP3A.The pharmacokinetic profiles of HY-D11 were described by two compartment model legitimately after oral administration of HY-D1166.7mg/kg in rats.main dynamics parameters:Tmax is 3.67±0.52 h, it means the absorbtion of HY-D11 is very fast; Cmax is 1610.35±130.28 ng-mL-1,it was much larger the concentration of Lowest effectie antibacterial concentrations in vitro; Vz/F is 9540.42±1530.62 mL-kg-1,it means that the distribution of HY-D11 is very aboad;1/2z and MRT(O-∞)) are is 3.66±0.25 h,7.05±0.29 h,respectively,it demonstrated that the elimination of HY-D11 is fast. |
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