There are some shortcomings of animal experiments applied in chemical toxicity testing, such as long period, large cost, species differences and dose differences, which limit the use of animal experiments' results in predicting human toxicity.Accordingly, we established a toxicity testing alternative screening system in line with the toxicity endpoints which required in chemical safety evaluation and risk assessment (genotoxicity, carcinogenicity, reproductive toxicity, acute toxicity and general toxicity) based on 3R principles (replacement, reduction, refinement) for animal experiments.This system covers most of the endpoints of toxicity assessment, and molecular biology technology was also applied to integrate the toxicity test, as well as some operation was optimized in order to shorten the experimental period, reduce experimental costs, improve animal welfare.Furthermore, the results from the screening system have higher clinical relevance because it is based on the toxicity mechanisms.

Objective To establish a rapid and sensitive liquid chromatography-tandem mass spectrometry(LC-MS/MS)analysis method for determination of unsymmetrical dimethylhydrazine(UDMH)contents in rat plasma and investigate the toxicokinetic characteristics of UDMH in rats.Methods Twenty-two SD rats were divided into the intravenous injection of 10 mg/kg dose group(4 females)and intragastric administration groups(low,medium and high dose,with 6 rats in each group,half males and half females).The rats were given 10mg/kg by intravenous administration and 10 mg/kg,30 mg/kg,and 90 mg/kg single dose of UDMH by gavage.Blood samples were collected from the orbital venous plexus at 0 hour before administration and at different time points after administration.The plasma samples were extracted with protein precipitation and derivatization before being analyzed using the LC-MS/MS method.Separation was carried out on a ZORBAX column(4.6mm×75mm,3.5 μm),with a mobile phase composed of 0.3％acetonitrile/formic acid at a flow rate of 1 mL/min.Propranolol was used as the internal standard.An electrospray ionization(Turbo Ionspray)source was applied and the mass spectrometer was operated in a positive MRM mode.Quantitative analysis showed that the ionization source unsymmetrical dimethylhydrazine and propranolol was at m/z:192.0→148.1,m/z:260.2→116.1,respectively.The toxicokinetic parameters were analyzed with the DAS 2.1 software.Results Quantification of UDMH exhibited a good linearity within the concentration range of 50-50000 ng/mL,with a linear correlation coefficient greater than 0.9900 and a lower limit of quantification of 50 ng/mL.The average recovery rate of UDMH was 98.1％,compared with 100.5％for the internal standard propranolol hydrochloride.The inter batch precision of standard curve samples ranged from 0.7％to 6.3％,and the relative error was between-7.1％and 6.2％.The inter batch and intra batch precision of quality control samples ranged from1.8％to 19.8％,and the relative error from-9.8％to 0.2％.The main pharmacokinetic parameters of UDMH in rats exposed to 10 mg/kg,30 mg/kg,and 90 mg/kg gavage were UC(0-t):(7624.99±2569.31),(34284.04±6657.15),(84720.88±22354.80)μg/L·h,t1/2:(0.07±0.15),(2.24±1.45),(3.04±0.90)h,Tmax:(0.75±0.27),(0.51±0.29),(0.29±0.10)h,Cmax:(4454.14±1329.45),(19442.45±9121.07),(32334.35±9882.41)μg/L,F:(77.34±26.06)％,(115.92±22.51)％,(95.48±25.19)％.Conclusion The LC-MS/MS method is highly accurate and specific,and is suitable for the toxicokinetic study of UDMH in rats.Single gavage administration of UDMH results in absorption and elimination saturation at a high dose.This study provides data for toxicological studies related to UDMH.