A Safety Evaluation And A Concomitant Toxicokinetics Study Of Tibetan Medicine: 25-ingredient Coral Pills

Objective:Establish the safety evaluation:acute toxic test and long-term toxic test of classic Tibetan Medicine:25-ingredient coral pills,to form specific safety evaluation mode:Discover the metabolic dynamics and effect of Copper,Lead,Mercury and Aconitum alkaloids in 25-ingredient coral pills,observe and analysis heavy matal and toxic ingredients'concentration in blood, which metabolic dynamics in tissues.Provide the scientific basis for clinical medicine reasonable and safety.Methods:1.Acute toxic test:The mice were orally administed with 25-ingredient coral pills once with the maximum concentration(0.464g/ml) and the dose volume(0.4ml/10g),continuous observe the toxicity of mice and weigh every day timing within 14 days.2. long-term toxic test:According to acute toxic test results to develop high,medium and low dose groups (0.333,0.667,1.333g (native medicine) /kg).Selected rodents mice and non-rodents beagle intragastrically 90 days.observe the animals'toxicity resulting and severity due to continuous medication,the development and recovery after stopping administrate.3.Concomitant toxicokinetics study and tissue distribution:Take biological samples from Rats and Beagle dogs' long-term toxic test. Detect the concentration of Aconitum Alkaloids with LC-MS-MS in blood and tissues Use DAS2.1.1 to calculate toxicokinetic parameters.4. In vivo distribution of Copper Lead Mercury:Take organs from Rats and Beagle dogs' long-term toxic test to be digested by Microwave Digestion System,to be detected the concentration of Copper,Lead,Mercury by Atomic absorption spectrometer.Results:1.Acute toxic test:The mice fixed static after just being feded,ptosis.breath deeply and fast,hair shaft and other symptoms,which recovered gradually after 4 hours.Continuous observation for 14 days,the appearance,behavior,mental status,urine,color,hair,skin,respiratory,nose,eyes.oral secretiona were no obvious abnomalities.Body weight of mice showed normal growth in the state before and after experimental.It's no significant gross anatomic pathology end of the trial.2. long-term toxic test:It's no changes significantly on each dose group of rats and Beagles general signs,appearance.behavior,feces.food intake,body weight.blood cytology,blood biochemistry,organ weight and histopathlolgical caused by 25 ingredient coral pill.The rats withdrawal 2weeks and 4weeks,Beagles withdrawal 35 days,the indicators also no change significantly caused by 25 ingredient coral pill.3.Concomitant toxicokinetics study and tissue distribution: Endogenous impuritise don't interfere the determination of Aconitum alkaloids and no matrix effects; the linear range were 0.1-200 ng/ml (r=0.997),the extraction recovery were more than 60%,the limit of detection were 0.1 ng/ml,precision and accuracy both meet the determination of biological samples.4. In vivo distribution of Copper Lead Mercury:The long-term toxic test in rat for 3 months of Ershiwuwei coral pills,the mercury levels in kidney,liver of high dose group were significantly higher than control group's(p<0.01,p<0.05),the long-term toxic test in Beagles for 3 months of Ershiwuwei coral pills,mercury levels in kideny,liver,brain of high dose group were significantly higher than control group's(p<0.01, p<0.01, p<0.05),lead level in brain of high dose group was significantly higher than control group's(p< 0.01),withdrawal for recovery 35 days, the mercury levels in brain of high dose group were higher than control group's(p<0.05).Conclusion:1.Safety Evaluation:Long-term toxic rest in rats and Beagles were no obvious toxic effects of Ershiwuwei coral pills.Long-term use of high dose can cause increased secretion of salivary glands and salivation that pointed a certain effect on the autonomic nervous system.2. Concomitant toxicokinetics study and tissue distribution:To determinate the content of alkaloids in rats and Beagles'serum and tissues accurately and sensitively with LC/MS/MS.The toxicokinetic parameters and tissue distribution data from this test can offer scientific basis to take tibetan medicine: 25-ingredient Coral Pills safety and legitimately.3.In vivo distribution of Copper Lead Mercury:The long-term toxic test in rat for 3 months of Ershiwuwei coral pills,mercury were accumulation in kidney and liver of high dose group.Lead were accumulation in heart of high dose group.The long-term toxic test in Beagles for 3 months of Ershiwuwei coral pills,mercury were accumulation in kidney,liver and brain of high dose group.lead was accumulation in brain of high dose group. withdrawal for recovery 35 days,mercury was still accumulation in brain of high dose group.