A method for preparing micron-porous calcite based on microbial mineralization and its application

Abstract

This invention discloses a method for preparing micron-porous calcite based on microbial mineralization and its application, relating to the field of calcite preparation technology. It includes: isolating and purifying a batch of bacteria from a natural environment where carbonate minerals are precipitating; screening strains with a calcium ion sedimentation rate greater than 95% and a magnesium ion sedimentation rate greater than 50%; then selecting one strain to prepare a seed culture; preparing a mineral ion culture medium containing calcium, magnesium, and lithium ions; inoculating the seed culture medium into the mineral ion culture medium and culturing for a period of time; removing all the precipitate from the bottom of the cultured mineral ion culture medium, rinsing and drying it to obtain the final product. This invention uses bacteria to biomineralize and produce porous calcite in an aquatic environment containing calcium, magnesium, and lithium ions. The product has loose, porous particles with a large specific surface area, is environmentally friendly and low-cost, and the preparation process does not emit waste gas or dust, avoiding the ecological damage caused by mining in the natural environment.

Description

Technical Field

[0001] This invention relates to the field of calcite preparation technology, specifically to a method for preparing micron-porous calcite based on microbial mineralization and its application. Background Technology

[0002] Calcium carbonate is an inexpensive, readily available, and widely used inorganic material. With the increasing maturity of science and technology, calcium carbonate products are gradually developing towards diversified crystal forms, ultra-fine particle sizes, and surface modification. Calcite (CaCO3) is an important mineral material, widely used for the adsorption of various inorganic pollutants (cadmium, lead, copper, chromium, zinc, cobalt, etc.) and organic pollutants (oils, dyes, and benzene, etc.) in soil and water due to its significant surface adsorption properties. Currently, the main source of industrial calcite is the mining of natural limestone and marble; however, this inevitably causes irreversible damage to the natural geographical environment, and the mining cost is relatively high. On the other hand, the adsorption capacity of naturally mined calcite is much lower than that of porous calcite, making porous calcite a potentially important industrial material.

[0003] In recent years, many scholars have conducted extensive research on the preparation of porous calcium carbonate. Chen Yanmeng, Mo Huiling, and others studied the carbonation method for preparing porous calcium carbonate, which consists of calcination, digestion, carbonation, filtration, and drying. It mainly involves calcining limestone to obtain CaO and CO2, then reacting the Ca(OH)2 emulsion generated by hydrating CaO with CO2 gas to obtain porous calcium carbonate. CN110589865A discloses a method for preparing loose porous calcite using steel slag. This method is based on obtaining a CaCl2-NH4Cl-NH3-H2O leachate from steel slag, adding sucrose, introducing CO2 into the leachate while simultaneously performing ultrasonic treatment, and then obtaining porous calcite through vacuum filtration, washing, and drying. Currently, the carbonation method is mainly used in China. The porous calcium carbonate produced by this method has advantages such as a large specific surface area, low oil absorption value, and stable product performance. Although the above methods yield porous calcium carbonate, they also have drawbacks in terms of preparation routes, such as high energy consumption, high wastewater discharge, uneven pore size distribution, and irregular structural morphology. Therefore, it is essential to conduct research on the preparation of porous calcium carbonate using other process routes.

[0004] Biomineralization involves the mineralization of metal ions into biominerals through the physiological and biochemical metabolic activities of organisms. This method has been used to precipitate and remove metal ions from contaminated water and soil. Due to the rapid reproduction and efficient biochemical metabolism of bacteria, different types of bacteria have been used to treat water pollution and repair concrete. Currently, industrial production of calcium carbonate mainly involves direct mining from the natural environment, which not only damages the ecological environment but also inevitably generates a lot of wastewater, waste gas, and waste materials. Furthermore, naturally mined calcium carbonate has a dense structure and poor adsorption capacity for wastewater. Microbial mineralization is a novel, cleaner production method; this more economical and environmentally friendly method for preparing microporous calcite warrants further exploration. Summary of the Invention

[0005] One of the objectives of this invention is to provide a method for preparing micron-porous calcite based on microbial mineralization, which solves the technical problems of high energy consumption and high wastewater and waste gas emissions in the prior art of porous calcium carbonate. The micron-porous calcite prepared by microbial mineralization in this invention has high porosity and uniform pore size, and its morphology and structure are easy to control. The preparation process is also more environmentally friendly.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: a method for preparing micron-porous calcite based on microbial mineralization, comprising the following steps in sequence: Step 1, separating and purifying a batch of bacteria from the natural environment where carbonate minerals are precipitating, selecting strains with a calcium ion precipitation rate greater than 95% and a magnesium ion precipitation rate greater than 50%, and then selecting one of the strains to prepare a seed solution.

[0007] Step 2: Prepare a mineral ion culture medium, which includes: 5 g / L beef extract, 10 g / L tryptone, 10 g / L sodium chloride, 0.02 mol / L calcium ions, 0.02 mol / L magnesium ions, and 0-0.05 mol / L lithium ions.

[0008] Step 3: Add NaHCO3 solution and Na2CO3 solution to the mineral ion culture medium to make the NaHCO3 concentration 0.021 mol / L and the Na2CO3 concentration 0.01 mol / L, and adjust the pH to 7.0 with hydrochloric acid solution.

[0009] Step 4: Add the seed culture from Step 1 to the mineral ion culture medium obtained in Step 3 and culture for a period of time.

[0010] Step 5: Remove all the precipitate from the bottom of the culture medium containing mineral ions, rinse and dry it to obtain the final product.

[0011] In the above-mentioned method for preparing micron-porous calcite based on microbial mineralization, the selected strain in step one has been deposited at the China General Microbiological Culture Collection Center, with accession number CGMCC No. 34171, and the strain is classified as Bacillus licheniformis.

[0012] In the above-mentioned method for preparing micron-porous calcite based on microbial mineralization, the first step of the seed liquid preparation process is as follows: firstly, the purified bacterial water sample is coated onto a solid culture medium; then, the colonies in the solid culture medium are inoculated into a liquid culture medium to obtain the seed liquid.

[0013] The above-mentioned method for preparing microporous calcite based on microbial mineralization uses a solid culture medium with the following formula: 5 g / L beef extract, 10 g / L tryptone, 10 g / L sodium chloride, 20 g / L agar, and pH adjusted to 7.0 with NaOH solution.

[0014] The above-mentioned method for preparing microporous calcite based on microbial mineralization uses a liquid culture medium with the following formula: 5 g / L beef extract, 10 g / L tryptone, 10 g / L sodium chloride, and pH adjusted to 7.0 with NaOH solution.

[0015] The above-mentioned method for preparing microporous calcite based on microbial mineralization uses calcium ions, magnesium ions, and lithium ions, all of which are derived from their chlorides. By adjusting the concentration of lithium ions, microporous calcite with different porosities is obtained, and the particle size distribution of the microporous calcite is 5~30μm.

[0016] The above-mentioned method for preparing microporous calcite based on microbial mineralization uses anhydrous calcium chloride as the chloride containing calcium ions, magnesium chloride hexahydrate as the chloride containing magnesium ions, and anhydrous lithium chloride as the chloride containing lithium ions; the concentration of the anhydrous lithium chloride is 0.005 mol / L.

[0017] In the above-mentioned method for preparing microporous calcite based on microbial mineralization, in step four, the inoculation volume of the seed liquid is 1% of the volume of the mineral ion culture medium, and it is cultured for 4-5 days in a constant temperature shaker with the temperature set at 36℃ and the rotation speed at 100rpm.

[0018] In the above-mentioned method for preparing microporous calcite based on microbial mineralization, step five involves rinsing with distilled water and anhydrous ethanol in turn, for a total of four rinses.

[0019] Another object of the present invention is to provide an application of microporous calcite based on microbial mineralization, wherein the microporous calcite is used to adsorb heavy metal ions in wastewater.

[0020] The preparation mechanism of micron-porous calcite based on microbial mineralization in this invention is mainly as follows: the growth and metabolic activities of microorganisms can change various physicochemical parameters of their hydrochemical environment. During the growth process, bacteria release ammonia (NH3) and carbonic anhydrase (CA), which together increase the pH of the surrounding water environment. At the same time, ammonia and carbonic anhydrase also increase HCO3. - CO3 2- and OH - Ion concentration promotes the precipitation of carbonate minerals. The roles of NH3 and CA can be illustrated by the following chemical reaction.

[0021] .

[0022] .

[0023] .

[0024] .

[0025] In the presence of bacteria Ions can be easily precipitated based on the following chemical reactions.

[0026] .

[0027] In addition, extracellular polymeric substances (EPS) serving as nucleation sites also play a crucial role in the biomineralization of calcium carbonate. EPS provides nucleation sites for minerals because they contain abundant amino acids, polysaccharides, and other substances, and the free hydroxyl ions have a large negative charge in an alkaline environment, which is beneficial for calcium ion absorption. Added magnesium ions react with CO32-... 2- The formation of soluble complexes increases local supersaturation and accelerates nucleation. Furthermore, magnesium ion adsorption partially blocks crystal growth sites, resulting in finer micron-sized particles.

[0028] Compared with the prior art, the present invention brings the following beneficial technical effects: (1) The porous calcite produced by microbial action has a particle size distribution of approximately 5~30μm. The surface and interior of these calcite particles are covered with micron-sized pores, and micron-sized calcite with different porosities can be prepared by changing the concentration of lithium ions. Compared with directly mined calcite, this micron-sized calcite has a larger specific surface area and macropores, which is beneficial for adsorbing heavy metal ions and other pollutants in wastewater.

[0029] (2) Compared with chemical precipitation of calcium carbonate, biomineralization precipitation of microporous calcite mainly relies on bacterial growth to increase pH value, produce ammonia, and change HCO3. - CO3 2- and OH -The low ion concentration allows for mineral precipitation under conditions of low metal ion concentration, reducing the consumption of chemicals and energy. Using bacteria to biomineralize microporous calcite from aquatic environments containing calcium, magnesium, and lithium ions simplifies the preparation process and contributes to sustainable environmental remediation strategies. Attached Figure Description

[0030] Figure 1 X-ray diffraction (XRD) characterization of micron-porous calcite.

[0031] Figure 2 The graph shows the relationship between lithium ion concentration and calcite surface porosity. The calcite surface porosity was calculated using ImageJ software.

[0032] Figure 3 This is a graph showing the changes in bacterial concentration (od) and pH value in liquid culture medium.

[0033] Figure 4 The calcite obtained from mineralization and the natural calcite in relation to Cu 2+ The pattern of removal rate change over time.

[0034] Figure 5 To adsorb Cu 2+ After the experiment, micron-pore calcite samples were taken for energy dispersive spectroscopy (EDS) scanning electron microscopy.

[0035] Figure 6 The image shows the energy-dispersive X-ray spectrum (EDS) at point 1.

[0036] Figure 7 This is a scanning electron microscope image of microporous calcite produced by mineralization at a lithium ion concentration of 0 mol / L.

[0037] Figure 8 This is a scanning electron microscope image of microporous calcite produced by mineralization at a lithium ion concentration of 0.005 mol / L.

[0038] Figure 9 This is a scanning electron microscope image of microporous calcite produced by mineralization at a lithium ion concentration of 0.01 mol / L.

[0039] Figure 10 This is a scanning electron microscope image of microporous calcite produced by mineralization at a lithium ion concentration of 0.02 mol / L.

[0040] Figure 11 This is a scanning electron microscope image of microporous calcite produced by mineralization at a lithium ion concentration of 0.03 mol / L.

[0041] Figure 12 This is a scanning electron microscope image of microporous calcite produced by mineralization at a lithium ion concentration of 0.04 mol / L.

[0042] Figure 13 This is a scanning electron microscope image of microporous calcite produced by mineralization at a lithium ion concentration of 0.05 mol / L.

[0043] Figure 14 Scanning electron microscope image of calcite prepared without seed culture. Detailed Implementation

[0044] The technical solutions in the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings.

[0045] In the description of this application, unless otherwise stated, " / " means "or". For example, A / B can mean A or B. The "and / or" in this document is merely a description of the relationship between related objects, indicating that three relationships can exist. For example, A and / or B can represent three cases: A exists alone, A and B exist simultaneously, and B exists alone.

[0046] The technical solution of this application will be further described in detail below with reference to the accompanying drawings.

[0047] The magnesium ions, calcium ions, and lithium ions mentioned in this invention all originate from their chlorides. For example, magnesium ions are derived from magnesium chloride hexahydrate, calcium ions from anhydrous calcium chloride, and lithium ions from anhydrous lithium chloride, all of which can be purchased through commercial channels.

[0048] The main technical concept of this invention is to prepare microporous calcite by using bacteria to biomineralize an aquatic environment containing calcium, magnesium, and lithium ions. This is a new method that is different from the existing physicochemical methods.

[0049] This invention utilizes bacteria screened from the Lan Hai Xin Yuan mining well in Dongying Economic and Technological Development Zone, Dongying City. The bacteria have been deposited at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, on April 11, 2025, with accession number CGMCC No. 34171, and classified as *Bacillus licheniformis*. This strain was used for biomineralization to obtain microporous calcite. Its biomineralization mechanism and mineral characteristics were explored, and its effectiveness was compared with that of natural calcite in adsorbing heavy metal ions.

[0050] First, a preliminary assessment of the bacteria and their mineralization capacity is conducted. The following sections provide a detailed explanation of bacterial isolation and purification, evaluation of bacterial mineralization capacity, bacterial preservation, growth of Bacillus licheniformis, changes in aquatic pH, and ammonia detection.

[0051] (1) Bacterial isolation and purification: The liquid culture medium consists of the following components: 5 g / L beef extract, 10 g / L tryptone, 10 g / L sodium chloride, and pH 7.0; the solid culture medium is the same as above, with an additional 20 g / L of agar powder added.

[0052] Take 100 mL of well water sample from the LanHaiXinYuan well and add it to 500 mL of sterile liquid culture medium. Seal the container and incubate in a constant temperature shaker until the liquid becomes turbid. In a laminar flow hood, spread 20 μL of the turbid liquid culture medium onto the surface of sterile solid culture medium, seal the container, and incubate at 37°C until colonies are visible on the plate. Pick a single colony and inoculate it into the liquid culture medium until the liquid becomes turbid, then inoculate it into the solid culture medium. Repeat the above process until the solid culture medium contains colonies with a uniform morphology.

[0053] (2) Evaluation of bacteria with mineralization ability: Each isolated and purified bacterium was subjected to a microbial mineralization experiment in a mineral ion medium containing calcium ions (0.01 mol / L) and magnesium ions (0.01 mol / L) (beef extract 5 g / L, tryptone 10 g / L, sodium chloride 10 g / L, pH=7.0). The strains with a calcium ion precipitation rate greater than 95% and a magnesium ion precipitation rate greater than 50% can be used in this method.

[0054] (3) Preservation of bacteria: The strain has been preserved and classified as Bacillus licheniformis, with accession number CGMCC No.34171.

[0055] (4) Growth, pH changes and ammonia detection of Bacillus licheniformis: In order to investigate bacterial growth and pH changes, the seed culture was inoculated into liquid culture medium (initial pH value was 7.0) at a volume of 1% of the liquid culture medium and placed in a constant temperature incubator (37℃, 100rpm). The cell concentration was measured at 600 nm wavelength using a spectrophotometer at different time points.

[0056] Figure 3 The graph shows the relationship between bacterial concentration (od) and pH changes in the inoculated liquid culture medium. Bacterial growth can be divided into three phases: stasis phase (0-5 h), logarithmic growth phase (5-20 h), and stationary phase (20-130 h). The pH value rapidly increased from 7.0 to 7.83, then continued to rise to 9.03, finally stabilizing at around 9. This indicates that the presence of bacteria increased the pH value in the aquatic environment, which promoted mineral precipitation.

[0057] To investigate the cause of the pH increase, the ammonia-producing ability of *Bacillus licheniformis* was tested. The composition of 100 mL of liquid culture medium used for the ammonia test was as follows: 0.5 g tryptone, 0.05 g K₂HPO₄, and 0.05 g MgSO₄, with the pH adjusted to 7.0-7.2. Nessler's reagent was prepared by adding 16.0 g NaOH, 7.0 g KI, and 10.0 g HgI₂ to 100 mL of distilled water. Simultaneously, a seed culture was inoculated into 10 mL of liquid culture medium (1% of the liquid culture medium volume), designated as the experimental group; another group was inoculated with the same volume of sterile distilled water, designated as the control group. After 24 hours of incubation, 3-5 drops of Nessler's reagent were added to both the experimental and control groups, and the color change was observed. The experimental group turned a deep yellow, indicating ammonia production, while the control group remained pale yellow, indicating the absence of ammonia. The presence of ammonia is a significant reason for the pH increase in the culture medium.

[0058] The following is a detailed description of a method for preparing micron-porous calcite based on microbial mineralization according to the present invention. In this invention, in order to improve the purity of bacteria inoculated into the mineral ion culture medium and improve the mineralization efficiency, the purified bacterial aqueous sample is first inoculated into a solid culture medium, then the colonies in the solid culture medium are inoculated into a liquid culture medium to obtain a seed solution, and finally the seed solution is inoculated into the mineral ion culture medium.

[0059] Specifically, the steps are as follows: Step 1: Take a purified bacterial sample and spread it onto the surface of a solid culture medium. The solid culture medium formula is: 5 g / L beef extract, 10 g / L tryptone, 10 g / L sodium chloride, 20 g / L agar, and adjust the pH to 7.0 with NaOH solution. Sterilize the solid culture medium in an autoclave at 121°C for 20 minutes. In a clean bench, dilute the bacterial sample with sterile distilled water. 0 times, 10 -1 times, 10 -2 times, 10 -3 times, 10 -4 times, 10 -5 The mixture was applied to the surface of the condensed solid culture medium, sealed, and then placed in a static incubator (temperature close to 37°C) for incubation.

[0060] Step 2: Inoculate the bacteria from the solid culture medium into the liquid culture medium; wait for colonies to grow on the solid culture medium (which takes 2-4 days), then prepare the liquid culture medium in an Erlenmeyer flask. The liquid culture medium formula is: 5 g / L beef extract, 10 g / L tryptone, 10 g / L sodium chloride, and adjust the pH to 7.0 with NaOH solution; sterilize the liquid culture medium in an autoclave at 121℃ for 20 minutes; in a laminar flow hood, use a pipette with a sterile tip to pick up a small amount of colonies from the solid culture medium and inject it into the liquid culture medium that has cooled to room temperature. Seal the flask and incubate it in a shaker for 2-3 days to obtain the seed culture; set the shaker temperature to 36℃ and the shaking frequency to 100 rpm.

[0061] Step 3: Preservation of bacterial strains; When the cell concentration od value of the seed culture cultured on the shaker is greater than or equal to 1 when measured by spectrophotometer at a wavelength of 600 nm, prepare a 30% glycerol solution with glycerol and distilled water and sterilize it in an autoclave. Mix the sterilized glycerol solution with the seed culture at a volume ratio of 1:1 in a sterile centrifuge tube, seal it with sterile sealing film, and store it in the freezer (temperature below -20℃). The preserved bacterial strains can be directly used for subsequent experiments.

[0062] Step 4: Inoculate the seed culture into the mineral ion medium. Prepare the mineral ion medium with the following formula: 5 g / L beef extract, 10 g / L tryptone, 10 g / L sodium chloride, 0.02 mol / L magnesium ion concentration, 0.02 mol / L calcium ion concentration, and lithium ion concentration controlled at 0~0.05 mol / L. Multiple lithium ion concentration gradients can be set to adjust the calcite porosity. Then, sterilize the mineral ion medium in an autoclave. In a laminar flow hood, prepare 0.8 mol / L NaHCO3 solution and 0.8 mol / L Na2CO3 solution using sterile distilled water. Filter the NaHCO3 and Na2CO3 solutions separately into sterile conical flasks using a sterile needle filter. Add 4 ml of filtered NaHCO3 solution and 2 ml of filtered Na2CO3 solution to every 150 ml of mineral ion medium, and adjust the pH to 7.0 with hydrochloric acid. In the laminar flow hood, inoculate the seed culture with a seed culture volume of 1% of the medium volume.

[0063] Step 5: Seal the inoculated mineral ion culture medium and place it in a constant temperature shaker for 5 days.

[0064] Step 6: Take out the mineral ion culture medium that has been cultured in the shaker, remove all the precipitate at the bottom of the medium, rinse it three times in turn with distilled water and anhydrous ethanol, and then dry it in a drying oven at a constant temperature of 43°C to obtain the product microporous calcite.

[0065] The present invention will be further described below with reference to specific embodiments.

[0066] Example 1: A method for preparing micron-porous calcite based on microbial mineralization according to the present invention includes the following steps: Step 1: Select a water sample of Bacillus licheniformis and dilute it with sterile distilled water at a concentration of 10 g / L in a clean bench. 0 times, 10 -1 times, 10 -2 times, 10 -3 times, 10 -4 times, 10 -5 Apply six gradients of the mixture to the surface of the sterilized and condensed solid culture medium, seal the contents, and incubate in a static incubator for two days.

[0067] Step 2: Prepare liquid culture medium in an Erlenmeyer flask and sterilize it in an autoclave. In a laminar flow hood, use a pipette with a sterile tip to pick up a small amount of colonies from the solid culture medium and inject it into the liquid culture medium cooled to room temperature. Seal the flask and incubate it in a shaker for 3 days to obtain the seed culture.

[0068] Step 3: Prepare the mineral ion culture medium, setting the calcium ion concentration to 0.02 mol / L, the magnesium ion concentration to 0.02 mol / L, and the lithium ion concentration to 0 mol / L. Sterilize the mineral ion culture medium in an autoclave. In a laminar flow hood, prepare 0.8 mol / L NaHCO3 solution and 0.8 mol / L Na2CO3 solution using sterile distilled water. Filter the NaHCO3 solution and Na2CO3 solution separately into sterile conical flasks using a syringe filter. Add 4 ml of filtered NaHCO3 solution and 2 ml of Na2CO3 solution to every 150 ml of mineral ion culture medium, and adjust the pH to 7.0 with hydrochloric acid. In the laminar flow hood, inoculate each mineral ion culture medium with seed culture at 1% of the culture medium volume.

[0069] Step 4: After sealing the inoculated mineral ion culture medium, place it in a shaker for 5 days.

[0070] Step 5: Take out the mineral ion culture medium that has been cultured in the shaker, tilt the mineral ion culture medium at 30°, use a pipette with the tip removed, and remove all the precipitate at the bottom of the mineral ion culture medium into a centrifuge tube. Rinse the centrifuge tube three times in turn with distilled water and anhydrous ethanol, and then dry it in a drying oven at a constant temperature of 43°C to obtain the microporous calcite product.

[0071] The electron microscopy image of the microporous calcite prepared in this embodiment is as follows. Figure 7 As shown.

[0072] Example 2: The difference from Example 1 is that the lithium ion concentration in the mineral ion culture medium in step three is 0.005 mol / L; the electron micrograph of the microporous calcite prepared in this example is shown below. Figure 8 As shown.

[0073] Example 3: The difference from Example 1 is that the lithium ion concentration in the mineral ion culture medium in step three is 0.01 mol / L; the electron micrograph of the microporous calcite prepared in this example is shown below. Figure 9 As shown.

[0074] Example 4: The difference from Example 1 is that the lithium ion concentration in the mineral ion culture medium in step three is 0.02 mol / L; the electron micrograph of the microporous calcite prepared in this example is shown below. Figure 10 As shown.

[0075] Example 5: The difference from Example 1 is that the lithium ion concentration in the mineral ion culture medium in step three is 0.03 mol / L; the electron micrograph of the microporous calcite prepared in this example is shown below. Figure 11 As shown.

[0076] Example 6: The difference from Example 1 is that the lithium ion concentration in the mineral ion culture medium in step three is 0.04 mol / L; the electron micrograph of the microporous calcite prepared in this example is shown below. Figure 12 As shown.

[0077] Example 7: The difference from Example 1 is that the lithium ion concentration in the mineral ion culture medium in step three is 0.05 mol / L; the electron micrograph of the microporous calcite prepared in this example is shown below. Figure 13 As shown.

[0078] Comparative Example 1: Unlike Example 1, no seed culture was inoculated in the mineral ion culture medium. The electron micrograph of the calcite prepared in this comparative example is shown below. Figure 14 As shown.

[0079] XRD characterization of micron-porous calcite as follows Figure 1 The measurement range (2θ) is 20°–60°. According to the phase search software MDI Jade 6, the peak positions (2θ) in the figure are 23.117°, 29.499°, 36.055°, 39.516°, 43.268°, 47.702°, and 48.668°, which are characteristic diffraction peaks of Magnesium calcite,syn (magnesium-containing calcite). The magnesium content is relatively low, and syn indicates that the mineral is a laboratory-synthesized mineral rather than a natural mineral.

[0080] Morphological analysis was performed on the products prepared in Examples 1 to 7 and Comparative Example 1. Scanning electron microscopy (SEM) images of the products obtained in the corresponding examples show that the calcite prepared using microorganisms contains many pores. This is because, under the action of bacteria, carbonate and calcium ions accumulate and precipitate around the calcite, and the pores in the calcite are traces of bacterial growth activity. The morphology of the calcite gradually changes with increasing lithium ion concentration, which may indicate that changes in lithium ion concentration affect bacterial growth and metabolism. Figure 14 Minerals produced without bacterial culture medium can be observed to have no pores and are plate-like or spherical in shape.

[0081] Figure 2 The graph shows the change in surface porosity corresponding to lithium ion concentration, plotted using Origin 2021 software. The surface porosity in Table 1 is the average porosity. The graph shows that the surface porosity of micron-sized calcite first increases and then decreases with higher lithium ion concentrations. + The highest porosity was observed at a concentration of 0.005 mol / L. Figures 7 to 13 It can also be observed that as the lithium ion concentration increases, the porosity of calcite first increases and then decreases, but its surface becomes rough with a large number of tiny protrusions, and the morphology of calcite changes significantly.

[0082] Table 1 shows the correlation between lithium ion concentration and calcite surface porosity.

[0083] Lithium ion concentration (mol / L) Surface porosity (%) 0 42.46 0.005 46.01 0.01 39.92 0.02 35.87 0.03 34.48 0.04 32.04 0.05 29.7

[0084] The application of the microporous calcite prepared in Examples 1 to 7 is described in detail below.

[0085] The calcite mineralized according to this invention and natural calcite were added to simulated wastewater to test the adsorption performance of copper in the wastewater. The adsorption equilibrium time, removal rate and adsorption capacity of the two were investigated.

[0086] Specific experimental steps: A certain mass of copper nitrate trihydrate was dissolved in distilled water to prepare a 24 mg / L copper ion solution as a simulated wastewater solution. 50 ml of the simulated wastewater solution was placed in a polyethylene bottle, and 0.2 g of mineralized calcite and natural calcite with different lithium ion concentrations were added. All experiments were conducted in a constant-temperature shaker at 100 rpm and a temperature of 25°C. At different time intervals, 1 mL of the supernatant was extracted and diluted 10 times with distilled water. The solution was then filtered through a 0.22 μm hydrophilic microporous membrane. The filtered solution was analyzed for Cu content using a flame atomic absorption spectrophotometer. 2+ concentration.

[0087] Removal rate = (1).

[0088] Adsorption capacity q e = (2).

[0089] In formula (1): Initial concentration, mg / L; To balance the concentration, mg / L.

[0090] In formula (2): The volume of the solution is in L; Based on the mass of the material, in grams; The adsorption amount is expressed in mg / g.

[0091] Conclusion: From Figure 4 It can be seen that with the increase of time, the effect on Cu 2+ The removal rate gradually increases; in the initial stage of adsorption, mineralized calcite and natural calcite show the best removal rate for Cu. 2+ The adsorption rates of ions were all very fast, with all mineralized calcite showing a rapid adsorption rate for Cu within 15 minutes. 2 + The removal rate increased from 0% to over 60%, while natural calcite only reached 25%, indicating poor adsorption performance. In the first 30 minutes of the reaction, the adsorption efficiency of calcite with different lithium-ion modulators varied slightly; as time progressed, the adsorption efficiency of each mineralized calcite for Cu... 2 + The removal rates were relatively consistent, reaching as high as 85%, with an average adsorption capacity of 5.1 mg / g, while the adsorption capacity of natural calcite was 1.8 mg / g; mineralized calcite showed a similar adsorption capacity for Cu. 2 + The adsorption rate of ions gradually decreased over time, and the reaction reached equilibrium after 30 minutes. Figure 5 To adsorb Cu 2+ Subsequent scanning electron microscope (SEM) images of mineralized calcite were obtained by spot scanning at five locations. Figure 6 The energy dispersive spectroscopy (EDS) spectrum for site 1 shows the elemental composition and distribution of the sample. A distinct peak for Cu (copper) indicates adsorbed copper ions or copper compounds (such as CuO or CuCO3), providing important evidence of copper ion adsorption by calcite. Tables 2 and 3 show the weight and atomic percentages for site 1, confirming that calcite (CaCO3) is the main phase, and the Cu content further confirms Cu... 2+ Adsorbed by calcite.

[0092] Table 2: Quantitative Analysis of Elemental Composition (by Weight Percentage)

[0093] C O Mg Ca Cu 8.35 55.50 2.30 30.62 2.36

[0094] Table 3: Atomic percentage of elemental composition in quantitative analysis

[0095] C O Mg Ca Cu 13.66 68.16 1.86 15.01 0.73

[0096] It should be noted that those skilled in the art should recognize that the above embodiments are only used to illustrate this application and are not intended to limit this application. Any appropriate changes and variations made to the above embodiments within the essential spirit and scope of this application fall within the scope of protection claimed by this application.

Claims

1. Use of a microporous calcite based on microbial mineralization for the adsorption of heavy metal ions in wastewater, characterized in that, The preparation method of the microporous calcite comprises the following steps in sequence: Step one, the strain which has been preserved in China General Microbiological Culture Collection Center, preservation number is: CGMCC No. 34171, classification and naming is: Bacillus licheniformis (Bacillus licheniformis) Bacillus licheniformis ) is prepared into seed liquid; In the mineral ion culture medium, 5 g / L of beef extract, 10 g / L of trypsin peptone, 10 g / L of sodium chloride, 0.02 mol / L of calcium ions, 0.02 mol / L of magnesium ions and 0.005 mol / L of lithium ions are included. In the mineral ion culture medium, 5 g / L of beef extract, 10 g / L of trypsin peptone, 10 g / L of sodium chloride, 0.02 mol / L of calcium ions, 0.02 mol / L of magnesium ions and 0.005 mol / L of lithium ions are included. In the mineral ion culture medium, 5 g / L of beef extract, 10 g / L of trypsin peptone, 10 g / L of sodium chloride, 0.02 mol / L of calcium ions, 0.02 mol / L of magnesium ions and 0.005 mol / L of lithium ions are included. The seed liquid prepared in step one is inoculated into the mineral ion culture medium obtained in step three and cultured for a period of time. The precipitate at the bottom of the mineral ion culture medium obtained through the culture is taken out, washed and dried, and thus the microporous calcite is obtained. The calcium ions, the magnesium ions and the lithium ions are all from their chlorides, and the microporous calcite with different porosities can be obtained by adjusting the concentration of lithium ions. The calcium ions are from anhydrous calcium chloride, the magnesium ions are from magnesium chloride hexahydrate, and the lithium ions are from anhydrous lithium chloride.

2. Use of a microporous calcite based on microbial mineralization according to claim 1 for the adsorption of heavy metal ions in wastewater, characterized in that: In step four, the inoculation volume of the seed liquid is 1% of the volume of the mineral ion culture medium, and the seed liquid is cultured in a constant-temperature shaker for 4-5 days at a temperature of 36 DEG C and a rotating speed of 100 rpm.

3. Use of a microporous calcite based on microbial mineralization according to claim 2 for the adsorption of heavy metal ions in wastewater, characterized in that, In step one, the preparation of the seed liquid comprises the following steps: first, the purified bacterial water sample is smeared on a solid culture medium; and then the colonies on the surface of the solid culture medium are inoculated into a liquid culture medium to obtain the seed liquid.

4. Use of a microporous calcite based on microbial mineralization according to claim 2 for the adsorption of heavy metal ions in wastewater, characterized in that, The formula of the solid culture medium is as follows: 5 g / L of beef extract, 10 g / L of trypsin peptone, 10 g / L of sodium chloride and 20 g / L of agar, and the pH is adjusted to 7.0 by using a sodium hydroxide solution.

5. Use of a microporous calcite based on microbial mineralization according to claim 1 for the adsorption of heavy metal ions in wastewater, characterized in that: The formula of the liquid culture medium is as follows: 5 g / L of beef extract, 10 g / L of trypsin peptone and 10 g / L of sodium chloride, and the pH is adjusted to 7.0 by using a sodium hydroxide solution. In step five, the precipitate is washed with distilled water and anhydrous ethanol in turn for 4 times.

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