Adsorption Of Cadmium And Copper Ions By Immobilized Recombinant Metallothioneins

Heavy metals pollution has a serious impact on human health.The accumulation of heavy metal ions in the food chain leads to the excessive content of heavy metals in food.The pollution of heavy metal ions also occurred in the process of food production,storage,and transportation.Metal pollution has become one of the main reasons affecting the quality and safety of Chinese food.Therefore,effective methods for heavy metal detection and removal are important for food safety.Different metallothioneins effectively chelate different heavy metal ions.Tetrahymena thermophila has five different metallothioneins which can effectively chelate Cu²⁺ and Cd²⁺.The immobilization technology has lower cost,good adsorption performance and efficiency,and does not harm the environment in the process of heavy metal treatment.In the study,sodium alginate-polyvinyl alcoholsilicon dioxide was used as carrier material to prepare immobilized spheres encapsulated with recombinant metallothionein,and their adsorption properties for heavy metal ions were investigated.The main results are as follows:1.The concentration of sodium alginate was 2%,barium chloride was 3%,and the dosage of silica was 5 mg/ml.Using single factor test and orthogonal test,the conditions for preparing immobilized spheres for adsorbing cadmium ions were as follows: polyvinyl alcohol 1%,boric acid 1%,20 mg/m L recombinant metallothionein MTT1 Escherichia coli,cadmium ion 7.53 mg;the conditions for preparing immobilized spheres for adsorbing copper ions were: polyvinyl alcohol 2%,boric acid 1%.20 mg/m L recombinant MTT2 Escherichia coli with copper ion adsorption capacity of 7.44 mg.2.Immobilized spheres were prepared by recombinant metallothionein MTT1,MTT2,MTT5,TM1,TM2 and GST tags respectively,and corresponding immobilized crushed spheres were prepared at the same time,and Cd²⁺,Cu²⁺ adsorption experiments were carried out.The results showed that the immobilized recombinant metallothionein MTT1 had better cadmium ion adsorption performance and the immobilized recombinant metallothionein MTT2 had better copper ion adsorption performance.The adsorptive capacity of immobilized crushing bacteria for heavy metal ions was significantly higher than that of immobilized whole bacteria.The adsorptive capacity of immobilized recombinant metallothionein MTT1 crushing bacteria was 7.45 mg for cadmium ions,and that of immobilized recombinant metallothionein MTT2 crushing bacteria was 7.15 mg for copper ions.3.Scanning electron microscopy?SEM?results show that there are a lot of folds on the surface of immobilized crushed bacteria,the internal structure is relatively loose,there are a lot of pore,and the spheres have a large specific surface area.This structure is beneficial to the adsorption of heavy metal ions by immobilized spheres.The diameter,density and mechanical strength of immobilized spheres were characterized.The shape of immobilized spheres was stable and the mechanical strength was high.4.From the aspects of p H value,time and initial concentration of heavy metal ions,the optimum parameters of adsorption of Cd²⁺,Cu²⁺ by immobilized spheres were established.Gradients were set up and adsorption experiments were carried out.The results showed that the adsorption performance of immobilized spheres was better when the p H value was about 4.The cadmium ion adsorption capacity of immobilized recombinant metallothionein MTT1 and immobilized recombinant metallothionein MTT2 was 7.86 mg and the copper ion adsorption capacity of immobilized recombinant metallothionein MTT2 was 7.45 mg respectively.When the adsorption time was 4 h and the initial concentration of heavy metal ions was 200 g/m L,the adsorption of Cd²⁺,Cu²⁺ on immobilized spheres reached a dynamic equilibrium state.The adsorption capacity of Cd²⁺ and Cu²⁺ on immobilized MTT1 crushing bacteria reached 7.95 mg,and the adsorption capacity of copper ion on immobilized MTT2 crushing bacteria reached 7.65 mg.In the experiment of reusability of immobilized spheres,immobilized spheres showed certain recyclability.5.By fitting the kinetic equation,it is found that the R2 of the quasisecond-order kinetics of Cd²⁺ and Cu²⁺ adsorption are 0.9963 and 0.9970,respectively.The adsorption process is more in line with the description of the quasi-second-order kinetics,which indicates that chemical adsorption is the key step to control the reaction rate in the adsorption process.The results of fitting isotherm model show that R2 of Langmuir model are 0.9692 and 0.9395,respectively.Langmuir isotherm model can describe the adsorption performance of immobilized spheres for Cd²⁺,Cu²⁺.The adsorption of immobilized spheres is on the monolayer.In conclusion,the immobilized recombinant metallothionein spheres were successfully prepared.The immobilized spheres have stable forming capacity,high mechanical strength,larger specific surface area,excellent adsorption property of cadmium and copper ions,and reusability.