Copper And Cadmium Ion Of Calyx Flower Arm End Rotifers (biggest Alyciflorus) Toxicity And The Effect Of Sod And Cat Enzyme Activity

Aquatic ecosystem is extreme vulnerable to heavy metals, and aquatic organisms have been commonly used to evaluate the consequence of metal pollutions by scientists of aquatic ecology and environmental science.Rotifera are microscopic aquatic animals living in freshwater environments and are critical to ecotoxicologic studies due to their simple availability, rapid proliferation, short living period, and high sensitivity to pollutant. Brachionus calyciflorus (BC) are a species of plankton and are able to change their population dynamics, antioxidant defense system, and expression of heat shock proteins (HSP) upon the stimulation by heavy metal environment.This study sought to determine the impact of Cu2+and Cd2+, two classic metal pollutant, on BC survivability and rest egg formation. We also evaluated the influence of these ions on the enzymatic activity of SOD and CAT within BC. And finally, we determined the impact of heavy metals on the expression of Hsp70in BC. The results are as following:1.Acute toxicity assays indicated that the lethal concentration50(LC50) is58.53,24.46, and14.87μg L-1when BC are exposed by Cu2+, the95%confidence limit were36.49-93.89,21.62-27.69and13.95-15.86μg L-1; and1.48,0.53, and0.31mg L-1when exposed by Cd2+for24h,48h, and96h, the95%confidence limit werel.09-2.04、0.42-0.67and0.27-0.36mg L-1, respectively. LC50of the two ions are reverse correlated with the exposure time.2.BC exposed with the lethal concentration of Cu2+and Cd2+produced rest eggs from day1and the amount of rest eggs significantly increased (P<0.05) compared with those cultured under the normal condition. The production of rest eggs in both groups increased in the first few days until day3, followed by a decreased production, which paralleled with population dynamics of BC.3.We evaluated the impact of Cu2+and Cd2+on the enzymatic activity of SOD and CAT using time-course and dose-dependent assays.(1) At the same time (24h) of different concentrations of Cu2+environment under the stress of SOD and CAT enzyme activity is increased first and then decreased, significantly (P<0.05), but in the Cd2+under environmental stress, rotifer SOD and CAT enzyme activity change tendency is not obvious (P>0.05). Overall Cu2+or Cd2+of different concentrations of stress, Cu2+is rotifers in30μg L-1, Cd2+mg L-1SOD rotifer in0.8mg L-1and CAT enzyme activity of all peak, and then in60μg L-1and1.6mg L-1Valley, significant difference (P<0.05). Peak above all, valley value low and groups including the control group, there were significant differences (P<0.05).(2) The activity of both SOD and CAT increased along with the exposure time by Cu2+or Cd2+. The peak enzymatic activity was observed after exposed by Cu2+and Cd2+for6h and12h, respectively. After the6h and12h, respectively, the enzymatic activity started to deline for BC exposed in Cu2+and Cd2+. After24h, more than half BC died due to high concentrations of Cu2+or Cd2+, leading to low enzymatic activities.4.Under heavy metal environmental stress,the resting eggs of collection had hatched.In the different concentrations of Cu2+or Cd2+stress, as well as the same concentration and different stress time, SOD and CAT enzyme activity and its consistent performance trend, Cu2+SOD and CAT under environmental stress enzyme activity is increased first and then decreased, significant difference (P<0.05).But under Cd2+environmental stress, rotifer SOD and CAT enzyme activity change tendency is not obvious (P>0.05), but the hatching of resting eggs of rotifers showed the same environment stress SOD and CAT represent the antioxidant defense system and its improved sensitivity compared, enzyme activity increased, especially the peak and valley value of enzyme activity, significant difference (P<0.05)5.Total protein was extracted from BC after exposed by Cu2+or Cd2+. HSP70could not be detected through western blot using either monoclonal or polyclonal antibodies against Hsp70. But we could not rule the possibility that HSP70from BC were failed to be recognized by these antibodies, which are normally used to interact with human-, mice-, and fish-derived HSP70.6.The homologous cloning strategy, in order to rotifer RNA as template, using degenerate primers amplified by RT-PCR, the fragment size, far from reports, to strip is not bright, and diffusion. Can be judged to be not specificity of Brachionus calyciflorus HSP70gene partial sequence. This is the main issue of degenerate primers not, need to try different primers.