Bioremediation Of Cd²⁺/Pb²⁺ And Calcium Carbonate Precipitation Induced By Urease Producing Bacteria

Due to the rapid development of industry,the persistent accumulation and non biodegradability of heavy metals in industrial activities pose a serious threat to the environment.In many methods of treatment of heavy metals,microbially induced carbonate precipitation?MICP?as an emerging and a quite promising bioremediation technology has begun to apply to the removal of heavy metals.its main principle is that in the induction of calcium ions,urease producing microorganisms depositing calcium ions and heavy metal ions in the form of carbonate minerals in the environment through the biological catalysis of urease so that it can realize the the fixation of calcium ions and heavy metal ions.The research include the followings:the isolation of urease-producing-bacteria;the effects of temperature,pH,urea concentration,Ca²⁺concentration on urease-producing-bacteria's growth and the activity of urease;the effect of calcium source types on MICP's product crystal;Based on the best physiological and biochemical conditions and calcium sources,the removal effect of Cd²⁺and Pb²⁺by urease producing bacteria in the process of MICP was analyzed,and the mechanism of heavy metal removal was explored preliminarily.The main results are as follows:?1?Three urease-producing-bacteria were isolated from the activated sludge which acclimated by urea.They were named UPB1,UPB2 and UPB3 which were identified as Bacillus sp,Bacillus cereus,Micrococcus by sequencing 16S rRNA gene and the strain of Bacillus sp?UPB1?had the ability of inducing calcium precipitation obviously.The urease activity could reach 11.33 mmol·L^-1·min^-1.?2?The effects of temperature,pH,urea concentration and Ca²⁺concentration on the growth and urease activity of UPB1 bacteria were investigated by single factor experiment,the optimum conditions for the MICP medium of UPB1 bacteria were determined:the optimum temperature for the growth and producting urease of UPB1was 30?,the optimum p H was about 9.0,the favorable urea concentration was 30 g·L^-1 for producing urease and the favorable Ca²⁺concentration was 0.25 mol·L^-11 for the highest efficiency of MICP.?3?Based on the best medium for growth and the highest urease activity of UPB1bacteria,the crystal characteristic of CaCO₃ induced by UPB1 bacteria through 4different calcium sources of calcium chloride,calcium acetate,calcium lactate and calcium gluconate had been explored by the field emission electron microscopy.The result showed that:the UPB1 induced Ca Cl₂ to form the most thermodynamically stable rhombohedral Ca CO₃?calcite?,induced Ca?CH3COO?₂ to form metastable CaCO₃which made of vaterite,induced calcium lactate and calcium gluconate to form a very complex crystal CaCO₃ which the crystals and crystals were embedded into a dense whole.Among of them,the Ca CO₃ crystals formed by using calcium lactate as the calcium source had no regular shape,while the CaCO₃ crystals formed by using calcium gluconate as a calcium source were combined into a regular sphere.?4?Based on the best calcium source CaCl₂,the dynamic process of CaCO₃deposition induced by UPB1 bacteria was observed by metallurgical microscope,the result showed that:the CaCO₃ crystals induced by UPB1 bacteria taking the mycelium cells as the core.The CaCO₃ crystal adhered on the cells,which led to the formation of a large number of pores,and with the advancement of the MICP process,the number of pores were increasing and the diameter of pore decreased gradually.The shape of Ca CO₃crystal was shown as the small particles at first,and then gradually collected into thin slices,the rhombohedral type Ca CO₃ crystal was formed at last.The UPB1 had provided effective nucleation sites for CaCO₃ crystal during the process of MICP.?5?Based on the principle of MICP,the differences on the removal of heavy metals Cd²⁺,Pb²⁺by UPB1 bacteria was analyzed in the medium which containing Ca²⁺and without it at first;and then the micro-analysis of MICP products was carried out by SEM and EDS,the mechanism of removal of heavy metals by UPB1 bacteria was discussed;the acid resistance analysis of MICP products by a series of concentration gradient hydrochloric acid was explored at last.The result showed that:in the medium containing Ca²⁺,the removal rates of Cd²⁺and Pb²⁺by UPB1 were 95.94%and 99.56%respectively,which were higher than those of medium without Ca²⁺,which the removal rates of Cd²⁺and Pb²⁺were 87.06%and 96.44%.But the MICP coprecipitation products Ca_x Cd_y?CO₃?_z?Ca_x Pb_y?CO₃?_z were less aciduric than the products of Cd CO₃?Pb CO_3.In the process of mineralization of heavy metals,the urease plays a key role that the short distance non covalent bond?the hydrogen bond,ionic bond and hydrophobic bond?was formed by the combination of enzyme activity center and substrate molecule,which changed or polarized the form of the bottom price key.The heavy metal ions were then combined with the substrates that have been polarized,making the heavy metal to be fixed.Secondly,because the ionic radius of Cd²⁺and Pb²⁺is close to Ca²⁺,the replacement reaction occurs in the process of UPB1 bacteria induced calcite deposition,so that the Cd²⁺and Pb²⁺entered into calcite to form coprecipitation.