# Effect of Temperature, pH, and Reaction Duration on Microbially Induced Calcite Precipitation
###### Abstract  
In this study, the amount of calcite precipitate resulting from microbially induced calcite precipitation (MICP) was estimated in order to determine the optimal conditions for precipitation. Two microbial species (_Staphylococcus saprophyticus_ and _Sporascrina pasteurii_) were tested by varying certain parameters such as (1) initial potential of hydrogen (pH) of urea-CaCl\({}_{2}\) medium, (2) temperature during precipitation, and (3) the reaction duration. The pH values used for testing were 6, 7, 8, 9, and 10, the temperatures were 20, 30, 40, and 50 \({}^{\circ}\)C, and the reaction durations were 2, 3, and 4 days. Maximum calcite precipitation was observed at a pH of 7 and temperature of 30 \({}^{\circ}\)C. Most of the precipitation occurred within a reaction duration of 3 days. Under similar conditions, the amount of calcite precipitated by _S. saprophyticus_ was estimated to be five times more than that by _S. pasteurii_. Both the species were sensitive to temperature; however, _S. saprophyticus_ was less sensitive to pH and required a shorter reaction duration than _S. pasteurii_. MICP; optimal condition; Staphylococcus saprophyticus; Sporosarcoma pasteurii 2018 8 1272 2018  
at its peak at this temperature. It was reported that the microbial activity and growth was slightly higher at 30 \({}^{\circ}\)C than at 20 \({}^{\circ}\)C [21]. Likewise, an increased calcite production was reported when the temperature was increased from 20 to 50 \({}^{\circ}\)C, while there was no precipitation at a temperature of 60 \({}^{\circ}\)C or higher [22; 23]. Some researchers indicate that urease activity does not have a strong correlation with the engineering properties of MICP treated soil. For instance, the strength of MICP treated coarse sand was found to be higher when soil improvement was performed at a relatively lower temperature of 20 \({}^{\circ}\)C [15]. Interestingly, even with the same amount of calcium carbonate precipitated in soil, the strength of MICP treated soil would differ. [PERSON] et al. (2016) measured the unconfined compressive strength of the soil at temperatures of 4, 25, and 50 \({}^{\circ}\)C, and reported that soil improvement was most effective at 25 \({}^{\circ}\)C for the same calcium carbonate content in the treated soil [12]. Moreover, more calcite precipitation does not necessarily result in higher strength if the precipitation occurs at a different temperature [24]. This is likely due to the crystal size resulting from a change in CaCO\({}_{3}\) solubility, which varies with temperature and pH [25]. The CaCO\({}_{3}\) solubility decreases with an increase in temperature and pH, thus affecting calcite precipitation. Another factor that highly influences MICP is the pH, with an alkaline environment known to be favorable to the process [24; 26]. Through hydrolysis, urea (CO(NH\({}_{2}\))\({}_{2}\)) is hydrolyzed by the urea enzyme into ammonium (NH\({}_{4}\)\({}^{+}\)) and carbonate ions (CO\({}_{3}\)\({}^{2-}\)). This CO\({}_{3}\)\({}^{2-}\) is combined with calcium ions (Ca\({}^{2+}\)) from the supplied medium to form calcium carbonate (CaCO\({}_{3}\)). \[\text{CO(NH${}_{2}$)}_{2}+2\text{H${}_{2}$O}\to 2\text{NH${}_{4}$}^{+}+ \text{CO${}_{3}$}^{2-}\]  
The urea enzyme is optimally active at a specific range of pH; the optimum pH for active enzyme ranges between 7 and 8.0 [9; 27; 28; 29]. Its activity peaked at a pH close to 8.0 and gradually decreased at a pH of 8 or higher. It was reported that the optimum pH for the action of most of the microbial ureases is around 7 [30], while that for _Sporosarcina pasteurii_, which is the most common microbe for MICP, is 8 [27; 31]; and the precipitation amount increased with pH and converged to a certain value at a pH range of 8.7 to 9.5 [27; 32; 33; 34]. Other microbes such as _Bacillus megaterium_ and _Bacillus sphaericus_ have an optimum pH value of 7 and 8, respectively [29; 35].