Study On The Mineralization Performance Of Bacillus Sp. Under Low Nitrogen Condition

During the period of service,concrete is subject to cracking on the surface or deep inside the structure,subsequently resulting in the ingress of harmful substances,which may damage the cementitious structure and shorten the service life.There are many kinds of microorganisms in natural environment able to induce mineral deposition.In recent years,the use of mineralization microorganisms to repair concrete cracks has been focused.Considering the high alkalinity inside concrete structure,vegetative cells are difficult to be introduced into the concrete during the preparation stage,so the mineralized microorganisms with sporulation capability are generally used in the self-healing process of concrete cracks.In this case,sporulation and spore germination capacity of the self-healing microorganisms are the key factors to ensure the efficiency of self-healing behavior.On the other hand,in the process of self-healing,microorganisms may produce some by-products such as ammonia that have negative effects on the environment as utilizing nitrogen sources.In order to reduce this environmental risk,it is necessary to screen the mineralized microorganisms with low nitrogen requirement and systematically study the mineralization performance of mineralized bacteria under low nitrogen conditions.In this study,the conditions of sporulation of a mineralized bacterium Bacillus sp.B8,was investigated and optimized.Single factor experiment showed that the optimal inoculation amount was 1×10⁷cells/mL.The optimum carbon source was starch with the optimum concentration of 0.5 g/L.The optimal nitrogen source was yeast extract and the optimal concentration was 2 g/L.Certain concentrations of Mg²⁺(0.24 g/L)and Mn²⁺(4.8 mg/L)can promote spore production.On the basis of single factor experiment,orthogonal experiment was used to optimize the factors,and the results showed that both the inoculation amount and concentration of yeast extract have significant influences on spore yield.After optimization,the inoculation amount,starch,yeast extract,Mg²⁺and Mn²⁺dosage were 1×10⁷ cells/mL,5,5,0.48 and 4.8×10^-3g/L,respectively,and the spore concentration reached up to 1.54×10⁹ spores/mL,about 10 times higher than before optimization.The spore germination experiment showed that the spores need about 4h to complete germination process.The most suitable germinant was inosine with the optimum concentration being 2 g/L.The optimal value of pH was 10.Na⁺ can promote spore germination,whereas Ca²⁺inhibited spore germination.The inhibitory effect of Ca²⁺on germination can be alleviated with the presence of Na⁺,and the optimal Na⁺concentration was 24 g/L.In the process of self-healing,in order to reduce the negative impact of byproducts of microbial nitrogen metabolism on the environment,the mineralization activity of B8 was studied under low nitrogen(0.2 g/L)condition.The results showed that B8 had the advantage of mineralization in the low nitrogen(0.2 g/L)environment,and the optimal nitrogen source was sodium nitrate.The optimum carbon source was sodium formate with the optimum concentration of 18 g/L.The optimum initial spore concentration was 4×10⁹ spores/mL.When the yeast extract concentration was 5 g/L in the process of sporulation,the mineralization ability of spores was the best,and the CaCO₃ production was 27.53 mM.