Study Of The Residues And Toxic Effect Of Dbdmh In Gift Tilapia

In aquaculture,1,3-dibromo-5,5-dimethylhydantoin DBDMH is a new type of sterilization agent,rendering positive effects in treating aquatic animal diseases that is being widely used and currently considered to be the most promising disinfectant.Although DBDMH is being used to kill harmful bacteria in water it is forming aquatic organism residues,which may be causing differences in the degree of toxicity to aquatic organism breeding mechanisms.In view of the above,for this study,GIFT tilapia were used as primary subjects at setting different DBDMH concentration gradients to acquire an in-depth knowledge of the sub-acute toxicity test results.In the experiments,bromine ion degradation and its residues in the GIFT tilapia muscle were detected by ion chromatography method,while the antioxidative enzymes activities in the hepatopancreas were detected and observed in the liver and pancreas tissue sections.In addition,the DNA damage of hepatopancreas was detected by Single cell gel electrophoresis.This research was done in order to investigate the toxic effects of DBDMH on GIFT tilapia and the results were as follows:(1)Residues and degradation of DBDMH in the GIFT TilapiaThis dissertation is inclined to develop an ion chromatography method to identify and determine the bromine ions in the aquatic products.It also intends to use the said method to find the residues and degradation of bromine ions in GIFT Nile Tilapia to be treated under the conditions of 1,3-dibromo-5,5-dimethylhydantoin(DBDMH).For the said purpose,we sampled and made the analysis of the aquatic products after a series of pretreatments like homogenizing and centrifuging the fish muscles.In so doing,it was possible to separate bromine ions from the anion exchange column Cs12A and detected the bromine ions with the conductivity detector through the Dionex ICS-3000 Ion Chromatograph.The linear range of the calibration curve for bromine ions was also discovered between 10 and 200 mg/kg with a correlation coefficient of 0.9963.At the different addictive levels,we confirmed that the recovery rating range could reach as high as 82.58 to 86.14%,with the relative standard deviations from 4.61 to 6.54%.Furthermore,we also confirmed that the method could be successfully applied to the quantitative determination of bromine ions in the aquatic products.Nevertheless,we disclosed the limitation range of the method in detecting the bromine ions in fish muscles was under 0.2μg/mL.In addition,in comparison with other detection methods i.e.optical analysis and electrochemical analysis,the developed ion chromatography has proven much easier and higher in operation with fine goal availability.To be accurate,it could be said that we gained from the detection of the bromine ions in the muscles of GIFT Nile tilapia under DBDMH disinfection treatment as shown in the concentrations of 0 to 0.3 mg/L DBDMH we gained almost free bromine ions in the muscles,although the concentration content of 1.5mg/L DBDMH may still result in some bromine ions as a time-effect.Moreover,it remained likely that on the 16-th day,some bromine ions could still be found with the maximum contents of 21.6 mg/kg in the muscle,which complied with the regulations of international food code.Although the results we obtained indicated that the bromine ions tend to increase with time,its concentration content in the muscles was proven to decrease 10 days later from the recovery moment.In spite of this,we found the remaining bromine ions in the fish muscles was under 0.3 to 0.4 mg/L DBDMH a treatment proven to be harmless to human beings in accordance with this dissertation.(2)Effects of DBDMH on antioxidant defense system of GIFT tilapiaFish were exposed to sub-lethal concentrations of 0.06,0.3 and 1.5 mg/L for 16 days and then transferred to DBDMH-free water for 10 days.Hepatic antioxidant factors,including superoxide dismutase(SOD),catalase(CAT),total-glutathione(T-GSH),oxidized glutathione(GSSG),glutathione-S-transferase(GST),glutathione reductase(GR),glutathione peroxidase(GPx),were measured at 1,2,4,8 and 16 days on starting the experiment and at 10 days after transferring to DBDMH-free water.The results showed significant increments(P<0.05)then decrements in SOD,CAT,GPx activities and GSH/GSSG ratios,decrement then increment in GR,GST activities accompanied by decrements in T-GSH levels following DBDMH exposure in GIFT tilapia in 0.06,0.3,1.5 mg/L over the 16-day exposure period,suggesting the presence of oxidative stress.Recovery data showed that the effects produced by 0.06-1.5 mg/L DBDMH were irreversible.However,the amount of DBDMH applications in actual production have little damage for the degradation.The study results suggest that antioxidant responses could be used as potential biomarkers for monitoring residual DBDMH in aquatic environments.(3)Study on slices observed of DBDMH in hepatopancreas of tilapia We use H-E staining method to explore DBDMH on chronic toxicity test of liver pancreas samples for observation of paraffin slice.The results showed that:with the time going and the increase of the concentration of test,the hepatopancreatic damage gradually aggregate.The damage showed liver cells swelling,liver cell hydropic degeneration,hepatic steatosis,liver blood sinus congestion,central hepatic venous congestion and other symptoms.Recovery data showed that the liver tissue damage produced by 0.06～1.5 mg/L DBDMH were reversible.(4)Effects of DBDMH on DNA damage in hepatopancreas of tilapiaOn the 4th,8th and 16th days under the DBDMH exposed,the DNA damage to the liver with Single cell gel electrophoresis was determined,using DNA damage index,such as the tail DNA content(TDNA%),tail length(TL),tail moment(TM)and Olive tail moments(OTM).The results showed the extent of DNA damage variation aggravated with the increment of concentration and time,especially,in the 16th day under the DBDMH exposure,the DNA damage was the most severe.In the short-term exposure of 4 days,compared with the control group,the extent of DNA damage variation had no significant differences(P<0.05).During the 8 days exposure,the extent of DNA damage tended to be gentle in the low-dosage group,however,in the high-dose group,the extent of DNA damage variation increased significantly(P<0.05).DNA damage became more severe through time,revealing that DBDMH with high concentrations had a time effect relationship with DNA damage.