| Objective:1.To study the absorption,distribution and excretion of orally administered nano zinc oxide in rats.2.To study the short term toxic effects of nano zinc oxide through a 28-day repeated oral administration in rats,the distribution of nano zinc oxide in differentorgans and its effect on intestinal immune function were studied.3.To study the subchronic toxicity of nano zinc oxide through a 90-day repeated oral administration in rats,and to compare the difference in toxicity with common size zinc oxide and zinc sulfate.The general toxicity and the toxic effect on male reproductive system were evaluated and compared.Methods:1.Metabolic characteristics of nano zinc oxide in rats:SD rats were randomly divided into control group,and nano zinc oxide groups.There were 4 males and 4 females respectively in the control group.The nano zinc oxide group included four subgroups(N1 to N4,),each subgroup has 4 females and 4males.The control group was orally administered with 0.5% GLU + 3% FBS.The nano zinc oxide group were orally administered with nano zinc oxide suspension at a dose of 350 mg / kg BW.Blood samples were collected from the jugular vein of rats before gavage and after gavage at 1 h,2 h,3 h,4 h,5 h,6 h,10 h,24 h,48 h,72 h,and 168 h.Urine and feces samples of rats were collected after gavage for 0-24 h,24-48 h,48-72 h,and 144-168 h.Animals in one subgroup of nano zinc and normal size zinc oxide were sacrificed at 6 h,1 d,3 d,and 7 d after gavage.and tissues including heart,liver,spleen,lung,kidney,brain,testis(male),and lymph nodes werecollected.The samples of blood,tissues,viscera,urine and feces of the rats were digested and the contents of zinc were determined using Atomic Absorption Spectrophotometer.2.Short-term oral toxicity of nano zinc oxide and its effect on intestinal immunity in rats:Weaning SD rats were randomly divided into 4 groups: the control group and low-,medium-and high-dose nano zinc oxide groups(10 rats/sex/group).The rats were intragastrically administered with nano zinc oxide at doses of 0,87.5,175 and350 mg/kg BW/day for 28 days,respectively.Animals were euthanized for necropsy,and selected organs were weighed and fixed for histological examination.Flow cytometric analysis of lymphocyte subsets of Peyer's patch and the level of secretory immunoglobulin A(SIgA)in intestinal fluid were detected.The concent of zinc in blood,liver,kidney,brain and testis of rats were measured by using Atomic Absorption Spectrometer after digestion.3.Study on 90-day oral toxicity of nano zinc oxide and its effect on reproduction of male rats170 weaning SD rats were randomly divided into 7 groups(10 rats/sex/group):the control group,nano zinc oxide group(87.5,175,and 350 mg/kg BW,respectively),common size zinc oxide group(350 mg/kg BW),and zinc sulfate group(370 and 700mg/kg BW,respectively).Animals were given corresponding test substance by gavage for 90 days.Three satellite groups(5 rats/sex/group)were set up in the experiment including the control group,nano zinc oxide group(350 mg/kg BW),and zinc sulfate group(700 mg/ kg BW),observation of rats in satellite group maintained for 28 days after the last dosing.The changes in body weight and food intake were recorded.At the end of the study,blood samples were collected fore the measurement hematology and clinicalbiochemistry.All animals were euthanized for necrospy,selected organs were weighed and fixed for pathological examination.Sperm analysis was performed for male rats.For animals in satellite groups,hematology and clinical biochemistry detections were conducted in the middle of the experiment(On the 45 th day after gavage).hematology and clinical biochemistry,gross anatomy and sperm analysis were performed after the recovery period.Results:1.Compared with the control group,there was no biological difference in the zinc content in blood,liver,kidney,spleen and other organs of the rats at each time point after single gavage of nano zinc oxide group.In the nano zinc oxide group,87.38% zinc was excreted by feces of male rats after 48 h,86.60% zinc was excreted by feces of female rats after 48 h.2.In the 28-day short term oral study,there were no toxicologically significant changes in body weight,absolute and relative organ weights and zinc content in organs in all groups.Histopathological examination shows an increased incidence of focal epithelial cell exfoliation in gastric mucosa,inflammatory cell infiltration and edema in gastric submucosa,and villous epithelial cell exfoliation in small intestine.Compared with the control group,the number of lesions in the high dose group increased significantly.Lymphocytes phenotyping analysis shows a significant increasement in percentage of T lymphocytes and decreasement in percentage of natural killer(NK)cell in Peyer's patch.The concentration of SIgA had no significant difference between dose groups.3.In the 90-day subchronic study,compared with the control group,the body weight of the male rats in nano zinc oxide high dose group gradually decreased from the 4th week.The body weight of male rats in the zinc sulfate group and the common size zinc oxide group decreased in a larger extent than that of the nano zinc oxidegroup.For rats in satellite groups,the body weight of males in nano zinc oxide high dose group and zinc sulfate high dose group decreased from the third week to the recovery period..No significant difference was found in the body weight of female rats during the 90-day experiment and recovery period.No significant difference were found in the hematological parameters during the middle of the experiment,the 90-day experiment,and recovery period.The serum albumin and total protein of females in nano zinc oxide high dose group and zinc sulfate high dose group decreased significantly during the middle of the experiment.At the end of 90 days,the serum albumin of females in common size zinc oxide group,zinc sulfate high dose and zinc sulfate low dose group decreased,the total protein of females in nano zinc oxide medium and high dose group,common size zinc oxide group and zinc sulfate high dose group decreased,the alkaline phosphatase of females in nano zinc oxide medium and high dose group,common size zinc oxide group and zinc sulfate high dose group increased.The serum albumin and total protein of females decreased in zinc sulfate high dose group,the alkaline phosphatase of females increased in nano zinc oxide high dose group and zinc sulfate high dose group during recovery period.At the end of 90-day study and the recovery period,there were no abnormalities in the gross anatomy and organ weight of rats in zinc oxide groups,the relative weight of liver of male rats in the high dose zinc sulfate group increased in the 90-day studywhile no difference was found after recovery period.There was no significant difference in organ weights in female rats among all groups in 90-day experiment and after recovery period.Compared with the control group,the sperm motility of the male rats in the medium and high dose groups of nano zinc oxide and the low and high dose groups of zinc sulfate decreased significantly in 90-day experiment,the sperm motility of the nano zinc oxide high dose group and zinc sulfate high dose group decreased significantly after the recovery period.Conclusion:1.Nano zinc oxide and common size zinc oxide did not accumulate in the organs after gavages.The excretion of zinc was mainly via feces within 24 hours,the excretion of both zinc oxide through feces within 48 hours is more than 80%.2.After oral ingestion of nano zinc oxide for 28 days,nano zinc oxide did not accumulate in the liver,kidneys and other organs of rats,but it could cause pathological changes such as focal epithelial cell exfoliation in gastric mucosa and edema in gastric submucosa and affects the intestinal immunity of rats.3.Under this experimental conditions,nano zinc oxide,common size zinc oxide and zinc sulfate can have toxic effects on rats,such as weight loss of males,serum albumin and total protein decreasement and alkaline phosphatase increasement of females.The intake of nano zinc oxide and zinc sulfate can cause a decrease in sperm motility.These effects still exist after the recovery period.The contents of zinc in liver and kidney of male and female rats in the high dose groups of nano zinc oxide,common size zinc oxide and zinc sulfate increased,but returned to normal level after recovery period. |
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