Carbonic anhydrase excreted by bacteria induces the formation of carbonate minerals

Study on the mechanism of carbonate mineralization in the presence of microorganisms is of great significance for the global carbon cycle, soil formation and evolution and other related issues. In order to clarify the role of extracellular carbonic anhydrase (CA) excreted by bacteria in the process of carbonate mineralization, a series of chemical experiments were carried out in the CA-calcium acetate-tryptone (without carbonate) system for 120 hours. The extracellular CA from MF-2 strains was extracted through precipitation of protein by ammonium sulfate and dissolution of protein by Tris-H2SO4 buffer solution. Synchronously, a series of control experiments without CA were performed. After the experiments, the solid and the liquid phases were separated using the centrifugation method. The mineral species, its chemical composition and morphology were characterized by X-ray diffractometer (XRD) and scanning electron microscope accompanied by energy-dispersive spectrometry (SEM/EDS). The concentration of calcium ion in the solution was determined by the inductively coupled plasma optical emission spectrometer (ICP-OES). The results show that (1) the precipitate with visible amount formed in the system with extracellular CA. With the prolongation of the reaction time, the precipitate weight increased gradually, while the Ca2+ concentration in the solution decreased. The treatment results with 30% hydrogen peroxide show that carbonate mineral was the dominant component in the precipitate. The precipitate formed in the control experiments was significantly less than that in the CA experiments, which was mainly composed of organic substances. This adequately demonstrates that the extracellular CA excreted by MF-2 strain could significantly promote precipitation of carbonate minerals. (2) No crystalline formed in the initial stage of experiments (before 48th h) with CA, and calcite was dominant mineral in the precipitate formed in the middle and late stages. This was obviously different from the incubation experiments with MF-2 strain. Only vaterite formed in the latter case. (3) The mineral morphologies formed by the action of extracellular CA were rhombohedral, spherical and hemi-spherical, in which then rhombohedral mineral dominated. This was also different from the incubation experiments with MF-2 strain (spherical and bowl-shaped in the latter case). The statistical results based on several SEM photographs show that the proportion of rhombohedral-shaped minerals decreased gradually with time, from nearly 100% at 48th h to 84% at 120th h, i.e. spherical and hemi-spherical minerals gradually increased. (4) CA accelerated hydration reaction of carbon dioxide and promoted formation of carbonate minerals. The CO2 participated in the reaction might largely come from air.