Analysis Of EPS Binding Characteristics And Toxicity Effects Of Different Concentrations Of Lead Ions And Klebsiella Trevisan

In recent years,with the rapid development of human activities and agriculture and industry,industrial wastewater containing a large amount of heavy metal ions has been discharged into rivers,lakes,rivers and marine waters,causing serious pollution to soil and waters.With the widespread application of lead in various industries,the accumulation and transfer of lead ions in soil and water bodies will not only seriously affect the microbial activity in the soil,but also the yield and quality of crops.It affects the growth and development of aquatic animals and aquatic plants,and can enter the food chain and endanger human health.Nitrogen-fixing bacteria are good at taking nitrogen from the air.They can convert nitrogen that cannot be absorbed by plants in the air into nitrogen fertilizer,which is continuously available for plants to enjoy.Utilizing nitrogen-fixing microorganisms can not only increase crop yield,reduce fertilizer use and production costs,but also reduce soil and water pollution,maintain ecological balance,and promote sustainable agricultural development.And because of the heavy use of pesticides and fertilizers,as well as the industrial"three wastes"and other heavy metals that have entered the soil,the lead seriously exceeds the standard,soil heavy metals cannot be decomposed by soil microorganisms,and tritium is easy to accumulate,migrate and transform,which seriously affects soil quality.A kind of microorganism,its growth,enzyme activity,cell membrane permeability,etc.will also be affected by the heavy metal lead,thereby reducing the nitrogen-fixing ability of nitrogen-fixing microorganisms.In this paper,the nitrogen-fixing functional bacteria-Klebsiella trevisan was used as the research object,and the heavy metal pb²⁺was used as the target pollutant.The binding mechanism and toxicity of lead to nitrogen-fixing bacteria.two-dimensional Fourier transform infrared correlation spectroscopy(2D-FTIR-COS)and other methods,the mechanism of the combined action of EPS and Pb was analyzed;And by measuring the growth trend,morphological changes,antioxidant enzyme activities(SOD,MDA),growth inhibition rate,cell membrane permeability of nitrogen-fixing functional bacteria under different concentrations of lead ion(Pb²⁺)stress,the lead ion pairs of different concentrations were analyzed toxic effects of nitrogen-fixing functional bacteria.The paper mainly obtained the following research conclusions:(1)In order to study the interaction mechanism between EPS and lead ions,three-dimensional fluorescence,synchronous fluorescence fluorescence,two-dimensional Fourier transform infrared correlation spectrum,and two-dimensional fluorescence spectrum were used for analysis.The three-dimensional fluorescence spectrum results showed that four substances exist in EPS.They are fragrant proteins,tryptophan protein substances,tryptophan and tyrosine compounds.The results of synchronous fluorescence spectroscopy show that proteinoids are the main constituents of EPS.When the concentration of lead ions gradually increases,the fluorescence intensity of EPS decreases.In order to study the binding mechanism of EPS and heavy metal lead ions,Stern-Volmer was performed on synchronous fluorescence data.Linear fit model,quenching rate constants K for tryptophan and proteinoids.They are 5.45×10¹¹ L/mol/s and 8.51×10¹¹ L/mol/s,which are both higher than the maximum scattering collision quenching constant.The results show that the quenching process is mainly static quenching.By applying the modified Stern-Volmer equation and fitting the experimental values,K_A was calculated to be 0.76×10⁴ L/mol and 0.69×10⁴L/mol,which indicates that Pb²⁺and EPS have a strong interaction.In addition,the binding sites were fitted and the static fluorescence quenching process was analyzed.The maximum log K of tryptophan and proteinoids was 7.36 L/mol and 3.43 L/mol,respectively.Tryptophan There are one type of binding site for class-like substances,and two types of binding sites for protein-like substances.After two-dimensional fluorescence spectrum analysis,the synchronous spectrum results showed that the fluorescence intensity of the aromatic protein(280 nm)was greater than that of the tryptophan protein(225 nm),and the directions of the two peaks were consistent.Asynchronous spectrum shows that with the continuous addition of Pb²⁺concentration,the order of binding of aromatic proteins and amino acids is 225 nm<280 nm.And two-dimensional Fourier transform infrared correlation spectroscopy can prove that the combination of heavy metal lead ion lead and EPS has an effect on the C=O,CH groups of proteins in the EPS and the-OH,-COOH and CO groups in the polysaccharide And the binding order is-COOH>aromatic amides C=O>C=O>CH>CO in the carboxyl group.(2)When nitrogen-fixing functional bacteria are under heavy metal Pb²⁺stress,the lower the OD value measured as the concentration of Pb²⁺increases,it can be judged that Pb²⁺mainly inhibits the division and reproduction of the nitrogen-fixing functional bacteria during the logarithmic growth phase,which significantly reduces the number of live bacteria.And its SOD and MDA enzyme activities were affected,SOD enzyme activity was reduced,and when the lead ion concentration was 200 mg/L,the SOD activity was reduced by 76.58%compared with the control group.It was proved that the greater the concentration of lead ions,the greater the free radicals generated,which inhibited the activity of SOD enzymes and reduced the activity-like scavenging ability;the MDA content increased,which indicates that under the stress of lead ions,nitrogen-fixing functional bacteria produced more propionic acids.Dialdehyde(MDA),which causes lipid peroxidation of cells,produces oxidative damage,and damages nitrogen-fixing bacterial cells to a greater extent.With the increase of Pb²⁺concentration,the inhibition rate of nitrogen-fixing functional bacteria cells continued to increase.When the concentration of Pb²⁺increased to 200 mg/L,the inhibition rate reached 35.12%.Increased proportion of dead cells.It also proves that lead ions have a strong toxic effect on nitrogen-fixing bacteria and can inhibit cell survival.