Complementing urea hydrolysis and nitrate reduction for improved microbially induced calcium carbonate precipitation

Microbial-induced CaCO₃ precipitation has been widely applied in bacterial-based self-healing concrete. However, the limited biogenetic CaCO₃ production by bacteria after they were introduced into the incompatible concrete matrix is a major challenge of this technology. In the present study, the potential of combining two metabolic pathways, urea hydrolysis and nitrate reduction, simultaneously in one bacteria strain for improving the bacterial CaCO₃ yield has been investigated. One bacterial strain, Ralstonia eutropha H16, which has the highest Ca²⁺ tolerance and is capable of performing both urea hydrolysis and nitrate reduction in combined media was selected among three bacterial candidates based on the enzymatic examinations. Results showed that H16 does not need oxygen for urea hydrolysis and urease activity was determined primarily by cell concentration. However, the additional urea in the combined medium slowed down the nitrate reduction rate to 7 days until full NO₃ ⁻ decomposition. Moreover, the nitrate reduction of H16 was significantly restricted by an increased Ca²⁺ ion concentration in the media. Nevertheless, the overall CaCO₃ precipitation yield can be improved by 20 to 30% after optimization through the combination of two metabolic pathways. The highest total CaCO₃ precipitation yield achieved in an orthogonal experiment was 14 g/L. It can be concluded that Ralstonia eutropha H16 is a suitable bacterium for simultaneous activation of urea hydrolysis and nitrate reduction for improving the CaCO₃ precipitation and it can be studied later, on activation of multiple metabolic pathways in bacteria-based self-healing concrete.