| Objective:The metabolic kinetics study on 5-bromo-2-fluorobenzonitrile was conducted with integral animal experiments and in vitro metabolism methods, to find out main metabolic kinetic parameters and characteristics of 5-bromo-2-fluorobenzonitrile in the body and compare the differences in metabolism between rats and human, and for the purpose of providing data for poison effect research and extrapolating poison effect of 5-bromo-2-fluorobenzonitrile from animals to human being. Methods:A specific, sensitive gas chromatography(GC) analytical method was developed for 5-bromo-2-fluorobenzonitrile. 5-bromo-2-fluorobenzonitrile were administrated to 3 groups of rats, 4 animals in every group, respectively in single doses of 20 mg/kg by caudal vein injection, 20 mg/kg by gavage and 500 mg/kg by gavage. Whole blood was collected continually from orbital venous plexus after the administration. The concentrations of 5-bromo-2-fluorobenzonitrile in plasma were analyzed with the software DAS 3.0, and then provided kinetic parameters. 5-bromo-2-fluorobenzonitrile were administrated to another 8 rats continually for 7 times with interval of 24 hours. Blood was collected from 4 rats before each administration for analyzing the trough concentrations, while blood was collected continually from another 4 rats after the 7th administration. Accumulation index was figured out respectively by trough concentrations, Cmax and AUC. Equilibrium dialysis method was used to analyzed the protein binding ratio of 5-bromo-2-fluorobenzonitrile in rats and human plasma respectively at the concentration levels of 500, 5000 and 50000 ng/ml, 3 repeats in each level. Metabolic incubation systems was established respectively with SD rat and human liver microsomes in vitro. 110 ?l solution was withdrawn from the incubation systems respectively 0, 10, 30, 60 and 90 min after incubating, and then stoped the metabolic reaction. The residual amounts of 5-bromo-2-fluorobenzonitrile were analyzed and metabolic half-life of 5-bromo-2-fluorobenzonitrile incubating with liver microsomes in vitro was figured out. Results:Based on administration by intravenous injection, the statistical moment parameter Vz of 5-bromo-2-fluorobenzonitrile in SD rats is 2.04±0.67 L/kg, which is far larger than total body water content. t1/2z is 4.17±1.92 h, and CLz is 0.36±0.10 L/h/kg, which means a general elimination rate. Model fitting results is fitted well to two-compartment open model. In compartmental model, t1/2? is 0.135±0.065 h, which leads to a fast distribution, apparent volume of distribution(v1) is 1.862±0.559 L/kg, elimination half life(t1/2?) is 10.413 ± 2.188 hours, and clearance rate(cl) is 0.277 ± 0.06 l/h/kg.Based on administration by gavage in single dose of 20 and 500 mg/kg(dose-ratio is 1:25), the basic toxicokinetics features Tmax of 5-bromo-2-fluorobenzonitrile in SD rats are 5.58±2.06 and 9.50±19.2 h respectively, MRT0-? are 8.63±1.39 and 25.37±4.62 h respectively, and t1/2z are 4.68±0.69 and 13.60±12.08 h respectively, all of which have a tendency to extend with the increase of dose. AUC0-? are 30248.3±4817.4 and 1148043.8±276793.5 ng·h·m L-1 respectively, with a specific value of 1.0 : 38.0, which also showes a nonlinear proportional increase with the increase of dose. The absorption fraction is 47.3±7.5% in 20 mg/kg group while 71.8±17.3% in 500 mg/kg group.Based on administration by gavage for 7 times with interval of 24 hours, Tmax of the first-administrated group is prolonged with the increase of dose comparing with the lastadministrated group, Cmax is higher, t1/2z and MRT0-? are prolonged lightly, while AUC under interval period are significantly higher. Trough concentration remained unchanged after more times of administration. Accumulation index figured out by trough concentrations is 0.21, while 1.24 by Cmax and 1.42 by AUC.Protein binding ratio of 5-bromo-2-fluorobenzonitrile at the concentration levels of 500, 5000 and 50000 ng/ml are respectively 83.5%, 88.8% and 88.6% in rats plasma, and 85.2%, 89.0% and 91.1% in human plasma. Protein binding ratio of both kinds of plasma have no significant statistical differences at the low and medium concentration(P=0.078, P=0.234), while it shows significant statistical differences at the high concentration(P=0.001), but with few absolute difference.Metabolic half-life of 5-bromo-2-fluorobenzonitrile incubating with rats and human liver microsomes and control solution in vitro are respectively 58.6, 59.2 and 65.0 min, which shows no significant differences. Conclusions:The metabolism of 5-bromo-2-fluorobenzonitrile after administration by intravenous injection is fitted well to two-compartment open model with t1/2z of 4.17±1.92 h. 5-bromo-2-fluorobenzonitrile distributes less in extracellular fluid and possible enriches in the body. 5-bromo-2-fluorobenzonitrile shows high absorption ratio and nonlinear kinetics feature after administration to rats by gavage in the dose range from 20 to 500 mg/kg, that is, with increase of dose, absorption is delayed, elimination is saturated, and the whole exposed quantity showes a nonlinear proportional increase. Repeated administrations of 5-bromo-2-fluorobenzonitrile by gavage lead to absorption delay and slightly less elimination,but shows no obvious accumulation.The potein binding ratio and metabolism in liver microsomes of 5-bromo-2-fluorobenzonitrile in SD rats is accordant to those in human, and both with high potein binding ratio(more than 80%). 5-bromo-2-fluorobenzonitrile will decompose in body fluid at the temperature of 37?, and will not be cleared mainly in neither rats nor human liver microsomes. |
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