Effects Of Heavy Metal Cd And Cu On Toxicity Of Tegillarca Granosa

In this paper, Tegillarca granosa was chosen as research object. In order to understand the effect of heavy metal to its digestive enzymes activity and antioxidant enzymes activity, different concentration gradients of Cd2+,Cu2+,(Cd2++Cu2+) and enzymology analysis method were utilizing. DNA damage detection methods were established at the same time. Further study on the influence of heavy metal on DNA chain rupture and DNA methylation were also carried out, which help to investigate the toxic mechanisms of heavy metal and determine the sensitive and reliable monitoring index. The experiment results indicate that:1. Analysis of digestive enzyme activityAmylase enzyme:For low concentration treatment, enzyme activity increased (which meant being induced) at the early period, and then decreased (which meant being inhibited) at the later period. While for high concentration treatment, enzyme activity stayed low level throughout the experiment. For each treatment, the toxicity of high concentration was greater than that of the low concentration. For different heavy metal ions and their combinations, the toxicity of the Cu2+group was the highest, followed by the (Cd2++Cu2+) group, and the Cd2+group was the lowest.Lipase enzyme:Enzyme activity increased at the early experimental period, and then decreased moderately at the later period. For each treatment, the toxicity of high concentration was greater than that of the low concentration. For different heavy metal ions and their combinations, the toxicity of the (Cd2++Cu2+) group was the highest, followed by the Cu2+group while the Cd2+group was lowest.Pepsin:Enzyme activity increased at the early experimental period, then experiencing a repeated rises and falls. The later period was a more complicated case. For each treatment, the toxicity of high concentration was greater than that of the low concentration. For different heavy metal ions and their combinations, the toxicity of the (Cd2++Cu2+) group was the highest, followed by the Cu2+group, and the Cd2+group was lowest.2. Analysis of antioxidant enzyme activityThe alteration of SOD activity was associated with CAT in Tegillarca granosa visceral mass. Both of them showed the trend of first increase and then decrease, exhibiting obvious time-dose effect which can be selected as heavy metal ion pollution detection index. For SOD, the toxicity of the Cu2+group was the highest, followed by the (Cd2++Cu2+) group while the Cd2+group was the lowest. For CAT, the toxicity of the (Cd2++Cu2+) group was higher than the Cu2+group and Cd2+group.3. Establishment and improvement of single cell gel electrophoresis methodThe best condition:single frosted glass slide and double system glue method were chosen. The concentration of both normal melting point agarose and low concentration agarose were0.7%, while colloid cell density was2.1×105cells/mL and the unwinding time was30min. The dying condition was with20ul20ug/mL EB and gradient ethanol elution and washing step was added. The results show that this method was of high accuracy and reliability.4. Study of the damage to blood cell DNAThe blood cell DNA was seriously damaged in each treatment and the damage degree rose with the increment of heavy metal ion concentration. The Potential genetic toxicity of Cu2+was higher than that of Cd2+. The toxicity of low concentration (Cd2++Cu2+) group was inferior than that of Cd2+group and Cu2+group, which showed an antagonistic effect. While the toxicity of high concentration (Cd2++Cu2+) group was higher than separated group, showing a collaborative effect. Test also revealed that TM and OTM as indicators responded sensitively.5. Establishing the method of determining total DNA methylation levelTotal DNA methylation level was detected by HPLC, in which6%methanol-5mmol/L ealkylsulfonate-0.2%triethylamine was selected as mobile phase at the pH of4. The fourth pretreatment method was applied and a standard equation was figured out, showing a high correlation coefficient, precision and recovery rate. Thus such method met the detection requirements.6. Influence on the total DNA methylation levelDNA methylation level increased abnormally under each treatment group, varying with time and concentration. The influence on gill was greater than that on foot. For the effect in the same tissue, the toxicity of high concentration was greater than that of the low concentration, and the toxicity of the Cu2+group was higher than the Cd2+group. Cd2+and Cu2+in (Cd2++Cu2+) group showed an antagonistic effect under low concentration and a collaborative effect under high concentration.