An algae-bacterial symbiotic system and its application in accelerating iron biomineralization in acidic mine wastewater

Abstract

This application discloses an algae-bacterial symbiotic system and its application in accelerating iron biomineralization in acidic mine wastewater, belonging to the field of acidic mine wastewater treatment technology. By simultaneously inoculating *Parachlorella sp. AH02* and *Acidithiobacillus ferrooxidans* LX5, supplemented with a small amount of nutrients, light, and oxygenation, an algae-bacterial symbiotic reaction system is formed. Based on the dual effects of electron shuttle in algal-derived organic matter and seed crystal template regulation by algal cells, high-concentration dissolved Fe(II) in acidic mine water is rapidly oxidized to Fe(III), and Fe(III) is rapidly hydrolyzed to ferric hydroxide minerals, thereby significantly improving the biomineralization efficiency of Fe and contributing to the green and efficient treatment of acidic mine wastewater.

Description

Technical Field

[0001] This application belongs to the field of acidic mine wastewater treatment technology, specifically relating to an algae-bacteria symbiotic system and its application in accelerating iron biomineralization in acidic mine wastewater. Background Technology

[0002] Acid mine wastewater (AMD) originates from the chemical and biological oxidation of sulfide minerals such as pyrite (FeS2) exposed to air during metal mining operations, including waste ore and tailings piles. AMD is reddish-brown in appearance, has a pH of 2.0–4.0, and contains high levels of dissolved iron (200–2000 mg / L) and heavy metals such as copper, manganese, and arsenic (10–100 mg / L). Effective treatment of AMD is a global environmental issue.

[0003] Biomineralization based on *Acidithiobacillus ferrooxidans* (Af) is considered a promising green and low-carbon treatment technology for AMD (Ammonium ferrooxidase). It primarily involves Af biologically oxidizing Fe(II) and Fe(III) through hydrolysis and precipitation, converting the high concentration of soluble Fe in AMD into ferric hydroxide minerals (such as Fe8O8(OH)6SO4) for removal. Subsequently, only a small amount of lime neutralization is needed to achieve AMD discharge compliance (adjusting pH and removing other toxic metals). Compared to traditional AMD treatment technologies (e.g., direct lime neutralization and precipitation, electrodialysis, sulfate reduction), the Af biomineralization method offers advantages such as a wide range of applicable water qualities, low treatment costs, and minimal production of toxic metal sludge.

[0004] However, based on existing research and practice, the biomineralization induced by *Acidithiobacillus ferrooxidans* (Af) generally suffers from bottlenecks in the actual application of AMD treatment, such as slow Fe(II) oxidation rate and low Fe(III) hydrolysis ore formation efficiency. This is because: on the one hand, Af is an inorganic chemoautotrophic microorganism with a very slow growth and reproduction rate, resulting in the complete oxidation of Fe(II) in AMD (Fe... 2+ →Fe 3+ On the other hand, under acidic conditions at room temperature and pressure, the Fe(III) hydrolysis mineralization reaction (Fe...) is often time-consuming; 3+ Gibbs free energy (Δ →Fe8O8(OH)6SO4) r G ° 298 The concentration of Fe in AMD is relatively high, especially during the nucleation stage, which often results in about 50% of the dissolved Fe failing to mineralize, affecting the efficiency of lime neutralization secondary treatment and the final effluent water quality.

[0005] Therefore, there is an urgent need to develop a new method to accelerate the efficient biomineralization of dissolved iron in acidic mine wastewater. Summary of the Invention

[0006] 1. The problem to be solved

[0007] This application addresses the problems of slow Fe(II) oxidation rate and low Fe(III) hydrolysis mineralization efficiency in existing Acidithiobacillus ferrooxidans (Af) biomineralization treatment technologies for AMD. It provides an algae-bacterial symbiotic system and its application in accelerating iron biomineralization, particularly in accelerating iron biomineralization in acidic mine wastewater. This method utilizes an algae-bacterial symbiotic system formed by Acidithiobacillus ferrooxidans (Af) and acidophilic algae, especially Parachlorella sp. AH02 used in this application, to treat AMD, accelerating iron biomineralization, improving the Fe(II) oxidation rate and Fe(III) hydrolysis mineralization efficiency, and achieving the green and efficient conversion of high-concentration soluble iron in AMD into ferric hydroxide sulfate.

[0008] 2. Technical Solution

[0009] To solve the above problems, the technical solution adopted in this application is as follows:

[0010] This application provides an algae-bacterial symbiotic system, which includes Acidithiobacillus ferrooxidans and acidophilic algae. The algae-bacterial symbiotic system is formed by simultaneously inoculating the two into a culture medium or wastewater system. In the symbiotic system, the algae and bacteria achieve co-existence and reproduction through the exchange of nutrients and metabolites and mutual influence.

[0011] Furthermore, the aforementioned acidithiobacillus ferrooxidans LX5 is deposited at the China General Microbiological Culture Collection Center (CGMCC) on March 13, 2002, with accession number CGMCCNO.0727. This microorganism was previously screened and deposited by the inventors, as detailed in Chinese invention patent publication number CN1375553A. LX5 was initially classified as Thiobacillus ferrooxidans, but with the development of microbial taxonomy and a deeper understanding of this type of bacteria, it was reclassified into the genus Acidithiobacillus and named Acidithiobacillus ferrooxidans.

[0012] Furthermore, the aforementioned acidophilic algae is Parachlorella sp. AH02, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) on June 5, 2023, with accession number CGMCC NO.40698. This microorganism was previously screened and deposited by the inventors in acidic mine wastewater, as detailed in Chinese Invention Patent Publication No. CN118084206A.

[0013] This application also provides the application of the aforementioned algae-bacteria symbiotic system in accelerating iron biomineralization in acidic mine wastewater.

[0014] Furthermore, the above applications include:

[0015] S1, Acidophilic ferrooxidizobacterium and acidophilic algae are simultaneously inoculated into acidic mine wastewater; the two form an algae-bacterial symbiotic system;

[0016] S2, Add nutrients; the nutrients are used for the growth and proliferation of *Acidithiobacillus ferrooxidans* and / or acidophilic algae;

[0017] S3 involves placing acidic mine wastewater under light conditions for iron biomineralization. After settling, ferric hydroxide minerals and treated wastewater can be obtained. During the oxidation of Fe(II) in acidic mine wastewater by *Acidithiobacillus ferrooxidans*, organic matter with high electron transfer capacity (such as proteins, phospholipids, and unsaturated hydrocarbons) carried or secreted by acidophilic algae acts as an electron shuttle, accelerating electron transfer between Fe(II) and *Acidithiobacillus ferrooxidans*, and accelerating the generation of reactive oxygen species (H2O2 and ·OH, etc.) for the oxidation of Fe(II). During the hydrolysis and precipitation of Fe(III), algal cells play a role in seed crystal template regulation, accelerating the heterogeneous nucleation and crystal growth of Fe(III) minerals. Under the dual action, Fe(II) in AMD can be oxidized to Fe(III) more quickly, and Fe(III) can also be hydrolyzed into minerals more quickly, thus significantly improving the efficiency of iron biomineralization in AMD.

[0018] Furthermore, in the above applications, the inoculation density of *Thiobacillus acidophilus* is 1 × 10⁻⁶. 7 ~1×10 9 per mL.

[0019] Furthermore, in the above applications, the inoculation density of acidophilic algae is 1×10⁻⁶. 6 ~1×10 8 per mL.

[0020] Furthermore, in the above applications, the nutrient agents include any one or more of the following: NaNO3, K2HPO4, MgSO4·7H2O, CaCl2·2H2O, NaCO3, propionic acid, and citric acid.

[0021] Further, the above-mentioned nutrient agents include (g / L): NaNO3 0.1-1.5, K2HPO4 0.01-0.04, MgSO4·7H2O 0.01-0.075, CaCl2·2H2O 0.01-0.036, NaCO3 0.01-0.02, propionic acid 0.01-0.2, and citric acid 0.001-0.01, any one or more of these.

[0022] Furthermore, in the above applications, the lighting conditions include: light intensity of 0.2–2 mW / cm². 2 .

[0023] Furthermore, in the above applications, the iron biogenic mineralization conditions include: 6-8 hours at 15-35℃.

[0024] This application also provides a method for accelerating iron biomineralization in acidic mining wastewater. The method involves forming an algae-bacterial symbiotic system in the acidic mining wastewater, which includes *Thiobacillus ferrooxidans* and acidophilic algae. The algae-bacterial symbiotic system is formed by simultaneously inoculating the two into a culture medium or wastewater system.

[0025] Furthermore, the aforementioned acidithiobacillus ferrooxidans LX5 is deposited at the China General Microbiological Culture Collection Center (CGMCC) on March 13, 2002, with accession number CGMCC NO. 0727.

[0026] Furthermore, the aforementioned acidophilic algae is Parachlorella sp. AH02, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) on June 5, 2023, with accession number CGMCC NO.40698.

[0027] Furthermore, the above-mentioned method for accelerating iron biogenic mineralization in acidic mine wastewater specifically includes the following steps:

[0028] M1 involves simultaneously inoculating acidophilic ferrooxidase and acidophilic algae into acidic mine wastewater; the two form an algae-bacterial symbiotic system.

[0029] M2, nutrient solution added; the nutrient solution is used for the growth and proliferation of *Acidithiobacillus ferrooxidans* and / or acidophilic algae;

[0030] M3 involves placing acidic mine wastewater under light conditions to induce iron biomineralization. After settling, ferric hydroxide minerals and the treated wastewater can be obtained.

[0031] Furthermore, the inoculation density of the aforementioned *Acidithiobacillus ferrooxidans* was 1 × 10⁻⁶. 7 ~1×10 9 per mL.

[0032] Furthermore, the inoculation density of the aforementioned acidophilic algae was 1×10⁻⁶. 6 ~1×10 8 per mL.

[0033] Furthermore, the above-mentioned nutrients include any one or more of the following: NaNO3, K2HPO4, MgSO4·7H2O, CaCl2·2H2O, NaCO3, propionic acid, and citric acid.

[0034] Further, the above-mentioned nutrient agents include (g / L): NaNO3 0.1-1.5, K2HPO4 0.01-0.04, MgSO4·7H2O 0.01-0.075, CaCl2·2H2O 0.01-0.036, NaCO3 0.01-0.02, propionic acid 0.01-0.2, and citric acid 0.001-0.01, any one or more of these.

[0035] Furthermore, the aforementioned lighting conditions include: light intensity of 0.2–2 mW / cm². 2 .

[0036] Furthermore, the aforementioned iron biogenic mineralization conditions include: 6–8 hours at 15–35℃.

[0037] This application also provides the application of the above-mentioned algae-bacteria symbiotic system in the treatment of acidic mine wastewater.

[0038] This application also provides the application of the above-mentioned method for accelerating iron biomineralization in acidic mine wastewater in the treatment of acidic mine wastewater.

[0039] This application provides the application of the above-mentioned method for accelerating iron biomineralization in acidic mine wastewater using an algae-bacteria symbiotic system in the treatment of acidic mine wastewater.

[0040] This application also provides a treatment process for acidic mine wastewater, the process comprising:

[0041] N1, simultaneously inoculate acidophilic ferrooxidase bacillus and acidophilic algae into acidic mine wastewater;

[0042] N2, added nutrients;

[0043] N3 involves placing acidic mine wastewater under light conditions to induce iron biomineralization, followed by static sedimentation to obtain treated wastewater.

[0044] N4 is added to the treated wastewater to adjust the pH and obtain the final effluent.

[0045] Furthermore, the aforementioned Acidithiobacillus ferrooxidans (Af) is Acidithiobacillus ferrooxidans LX5, deposited at the China General Microbiological Culture Collection Center (CGMCC) on March 13, 2002, with accession number CGMCC NO.0727.

[0046] Furthermore, the aforementioned acidophilic algae is Parachlorella sp. AH02, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) on June 5, 2023, with accession number CGMCC NO.40698.

[0047] Furthermore, the inoculation density of the aforementioned *Acidithiobacillus ferrooxidans* was 1 × 10⁻⁶. 7 ~1×10 9 .

[0048] Furthermore, the inoculation density of the aforementioned acidophilic algae was 1×10⁻⁶. 6 ~1×10 8 per mL.

[0049] Furthermore, the above-mentioned nutrients include any one or more of the following: NaNO3, K2HPO4, MgSO4·7H2O, CaCl2·2H2O, NaCO3, propionic acid, and citric acid.

[0050] Further, the above-mentioned nutrient agents include (g / L): NaNO3 0.1-1.5, K2HPO4 0.01-0.04, MgSO4·7H2O 0.01-0.075, CaCl2·2H2O 0.01-0.036, NaCO3 0.01-0.02, propionic acid 0.01-0.2, and citric acid 0.001-0.01, any one or more of these.

[0051] Furthermore, the aforementioned lighting conditions include: light intensity of 0.2–2 mW / cm². 2 .

[0052] Furthermore, the aforementioned lighting conditions include: a light intensity of 1.5 mW / cm². 2 .

[0053] Furthermore, the aforementioned lighting conditions include: a light intensity of 2 mW / cm². 2 .

[0054] Furthermore, the aforementioned iron biogenic mineralization conditions include: 6–8 hours at 15–35℃.

[0055] Furthermore, the aforementioned alkaline substances include lime.

[0056] Furthermore, the pH is adjusted to 7.0–7.5.

[0057] 3. Beneficial effects

[0058] Compared with the prior art, the advantages of this application are as follows:

[0059] (1) This application provides an algae-bacterial symbiotic system and its application in accelerating iron biomineralization in acidic mine wastewater. By simultaneously inoculating Acidithiobacillus ferrooxidans (Af) and Parachlorella sp. AH02, an algae-bacterial symbiotic system is formed. In acidic mine wastewater, the algae-bacterial symbiotic system, on the one hand, accelerates the electron transfer between Fe(II) and Af through organic matter (proteins, phospholipids, unsaturated hydrocarbons, etc.) with high electron transfer capacity carried or secreted by the acidophilic algae as an electron shuttle, and accelerates the generation of reactive oxygen species (H2O2 and ·OH, etc.) for the oxidation of Fe(II); on the other hand, the algal cells of the acidophilic algae can serve as seed crystal templates to accelerate the heterogeneous nucleation and crystal growth of Fe(III) minerals. Under the combined effect of the above two aspects, Fe(II) in AMD is rapidly oxidized to Fe(III) and Fe(III) is rapidly hydrolyzed to form minerals, thereby greatly improving the efficiency of dissolved iron biomineralization in AMD. Compared with the existing AMD-based iron bioore-forming treatment based on Acidobacterium ferrooxidans, this method effectively solves the problems of slow Fe(II) oxidation rate and low Fe(III) hydrolysis ore-forming efficiency, and significantly improves wastewater treatment efficiency.

[0060] (2) The algae-bacterial symbiotic system provided in this application and its application in accelerating iron biomineralization in acidic mine wastewater, wherein the acid-loving algae is Parachlorella sp. AH02 and the acid-loving ferrooxidans is Acidithiobacillus ferrooxidans LX5, both of which are isolated from acidic mine wastewater and are indigenous microorganisms with strong environmental adaptability; the nutrients required for their growth are widely available and inexpensive.

[0061] (3) The algae-bacterial symbiosis system provided in this application and its application in accelerating iron biomineralization in acidic mine wastewater are used to treat acidic mine wastewater. The entire algae-bacterial symbiosis system has a simple treatment process, is easy to operate, is green, efficient, economical and practical. Attached Figure Description

[0062] Figure 1This is a process flow diagram of algae-bacteria symbiotic biological mineralization treatment of acidic mine wastewater.

[0063] Figure 2 The dissolved Fe(II) concentration (a), Fe mineral yield (b), microstructure (c), and XRD identification (d) during the treatment of acidic mine wastewater by algae-bacteria symbiotic biomineralization are analyzed.

[0064] Figure 3 This refers to the changes in the appearance of acidic mine wastewater before and after treatment by algae-bacteria symbiotic biological mineralization. Detailed Implementation

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

[0066] It should be noted that terms such as "upper", "lower", "left", "right", and "middle" used in this specification are only for clarity of description and are not intended to limit the scope of implementation. Changes or adjustments to their relative relationships, without substantially altering the technical content, should also be considered as within the scope of this application.

[0067] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains; the term “and / or” as used herein includes any and all combinations of one or more of the associated listed items.

[0068] Unless otherwise specified in the examples, the procedures should be performed under standard conditions or conditions recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all commercially available products.

[0069] As used herein, the term “about” is used to provide for the flexibility and imprecision associated with a given term, measure, or value. Those skilled in the art can readily determine the degree of flexibility for a particular variable.

[0070] As used herein, the term “at least one of…” is intended to be synonymous with “one or more of…”. For example, “at least one of A, B, and C” explicitly includes only A, only B, only C, and combinations thereof.

[0071] Concentration, amount, and other numerical data may be presented in range format herein. It should be understood that such range format is used solely for convenience and brevity and should be flexibly interpreted to include not only the values ​​explicitly stated as the limits of the range, but also all individual values ​​or subranges encompassed within the range, as if each value and subrange were explicitly stated. For example, a range of values ​​from about 1 to about 4.5 should be interpreted to include not only the explicitly stated limits of 1 to 4.5, but also individual numbers (such as 2, 3, 4) and subranges (such as 1 to 3, 2 to 4, etc.). The same principle applies to ranges that describe only a single value, such as “less than about 4.5,” which should be interpreted to include all the aforementioned values ​​and ranges. Furthermore, this interpretation should apply regardless of the breadth of the range or characteristic described.

[0072] As used in this article, "algae" refers to a group of thallus plants that lack differentiation into roots, stems, and leaves, have no vascular tissue, whose zygotes do not develop into embryos, whose reproductive organs are single-celled, contain photosynthetic pigments, and are photosynthetically autotrophic, producing oxygen. They include eukaryotic algae and prokaryotic algae. The vast majority of species live in water.

[0073] In this application, "synchronous inoculation" refers to the process of inoculating two or more different microorganisms into the same culture system or environment at the same time point or within a very close time interval.

[0074] Example 1

[0075] This embodiment provides an algae-bacteria symbiotic system and its application in accelerating iron biomineralization in acidic mine wastewater.

[0076] In this embodiment, the algae-bacterial symbiotic system includes Acidophilus ferrooxidans and acidophilic algae. The algae-bacterial symbiotic system is formed by simultaneously inoculating the two into a culture medium or wastewater system. In the symbiotic system, algae and bacteria achieve co-existence and reproduction through the exchange of nutrients and metabolites and mutual influence.

[0077] In this embodiment, the acidithiobacillus ferrooxidans (Af) is Acidithiobacillus ferrooxidans LX5, which is deposited at the China General Microbiological Culture Collection Center on March 13, 2002, with accession number CGMCC NO.0727.

[0078] In this embodiment, the acidophilic algae is Parachlorella sp. AH02, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) on June 5, 2023, with accession number CGMCCNO.40698, and the deposit address is Institute of Microbiology, Chinese Academy of Sciences, No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0079] In this embodiment, the algae-bacteria symbiotic system is used to accelerate the biomineralization of iron in acidic mining wastewater, including:

[0080] Acidophilic ferrooxidase and acidophilic algae were simultaneously inoculated into acidic mine wastewater;

[0081] Add nutrients;

[0082] Iron biomineralization is achieved by exposing acidic mine wastewater to light conditions.

[0083] In this embodiment, an algae-bacterial symbiosis system is used to treat acidic mine wastewater. Through algae-bacterial symbiosis, the biomineralization of iron in the acidic mine wastewater is accelerated, thereby removing iron from the wastewater. The process is as follows: Figure 1 As shown, it includes:

[0084] Acidophilic ferrooxidase and acidophilic algae were simultaneously inoculated into acidic mine wastewater;

[0085] Add nutrients;

[0086] Acidic mine wastewater is subjected to light conditions to induce iron biomineralization, and after settling, the treated wastewater is obtained.

[0087] Add an alkaline substance (lime) to the treated wastewater to adjust the pH to 7.0-7.5 to obtain the final effluent.

[0088] In this embodiment, the water sample was mine water from an iron mine in Ma'anshan, Anhui Province. The pH value was 2.7, the dissolved Fe(II) content was 615 mg / L, and the contents of heavy metal ions Mn, Cu, Zn and Ni were 393 mg / L, 37 mg / L, 36 mg / L and 3 mg / L, respectively.

[0089] In this embodiment, an algae-bacterial symbiosis system is used to treat acidic mine wastewater. Through algae-bacterial symbiosis, the system accelerates the biomineralization of iron in the acidic mine wastewater and removes iron from it. Specifically, this includes:

[0090] Inoculation: Add algae to the water sample at a density of 1×10⁻⁶. 7 Parachlorella sp. AH02 cells / mL and a bacterial density of 5 × 10⁻⁶ cells / mL 7Acidithiobacillus ferrooxidans LX5 (units / mL);

[0091] Add nutrient: 0.5g / L NaNO3;

[0092] Biological mineralization treatment: 1.5 mW / cm 2 The system was treated under light and at 18°C ​​for 8 hours. Electrochemical workstation measurements showed that the electron acceptor and electron supply capacity of this treatment system were 0.9 mmol e. - / g C(-0.45V) and 0.7mmol e - / g C(0.61V); Liquid chromatography analysis showed that the cumulative concentration of hydroxyl radicals (·OH) reached 25 μM; analysis of the treated water sample revealed that the solution pH was 2.1 and the Fe(II) content decreased to 22 mg / L. Figure 2 (a) At the same time, a large amount of reddish-brown precipitate was produced. Figure 2 (b); SEM and XRD analysis showed that the precipitate had a burr-like appearance. Figure 2 c) The chemical composition is ferric hydroxide mineral ( Figure 2 (d)

[0093] Solid-liquid separation: After biological mineralization treatment, the water sample is allowed to settle. Lime is added to the supernatant to adjust the pH to 7.5, yielding the final effluent. Figure 3 According to ICP testing, the concentration of heavy metals in the effluent meets the discharge standards.

[0094] Example 2

[0095] This embodiment provides an algae-bacteria symbiotic system and its application in accelerating iron biomineralization in acidic mine wastewater.

[0096] In this embodiment, the water sample is acidic water from a natural mining lake in Liuhe District, Nanjing, with a pH of 2.5, a dissolved Fe(II) content of 1080 mg / L, and heavy metal ions Mn and Zn contents of 45 mg / L and 37 mg / L, respectively.

[0097] In this embodiment, the acidithiobacillus ferrooxidans (Af) is Acidithiobacillus ferrooxidans LX5, which is deposited at the China General Microbiological Culture Collection Center on March 13, 2002, with accession number CGMCC NO.0727.

[0098] In this embodiment, the acidophilic algae is Parachlorella sp. AH02, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) on June 5, 2023, with accession number CGMCCNO.40698, and the deposit address is Institute of Microbiology, Chinese Academy of Sciences, No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0099] In this embodiment, an algae-bacterial symbiosis system is used to treat acidic mine wastewater. Through algae-bacterial symbiosis, the system accelerates the biomineralization of iron in the acidic mine wastewater and removes iron from it. Specifically, this includes:

[0100] Inoculation: Add algae to the water sample at a density of 3 × 10⁻⁶. 6 Parachlorella sp. AH02 cells / mL and a bacterial density of 2×10⁻⁶ cells / mL 8 Acidithiobacillus ferrooxidans LX5 (units / mL);

[0101] Added nutrients: 0.01g / L NaCO3, 0.02g / L propionic acid;

[0102] Biological mineralization treatment: 2mW / cm 2 The algal-derived organic matter was treated under light and at 28°C for 24 hours. Electrochemical workstation measurements showed that the electron acceptor and electron donor capacity of the algal-derived organic matter in this treatment system were 0.25 mmol e. - / g C and 0.32mmole - / g C; the cumulative concentration of H2O2 was 37μM as detected by liquid chromatography; analysis of the treated water sample revealed that the soluble Fe(II) content decreased to 45mg / L, and a large amount of reddish-brown precipitate was produced; XRD analysis showed that the precipitate was ferric hydroxide sulfate.

[0103] Solid-liquid separation: After the biological mineralization treatment, the water sample is allowed to settle and lime is added to the supernatant to adjust the pH to 7.0, and the final effluent is obtained.

[0104] Comparative Example 1

[0105] This comparative example provides the application of Acidithiobacillus ferrooxidans LX5 alone in the biological mineralization and iron removal of acidic mine wastewater, without the addition of the acidophilic algae Parachlorella sp. AH02.

[0106] Referring to Example 1,

[0107] The water sample was the acidic mine wastewater from Example 1. The specific steps are as follows:

[0108] Inoculation: Add a bacterial density of 5 × 10⁶ to the water sample. 7 Acidithiobacillus ferrooxidans LX5 (units / mL);

[0109] Add nutrient: 0.5g / L NaNO3;

[0110] Biological mineralization treatment: 1.5 mW / cm 2 The water was treated under light and at 18℃ for 8 hours; no current signal was detected during the entire treatment process; the cumulative concentration of hydroxyl radicals was 10 μM; analysis of the treated water sample revealed that the solution pH was 2.4 and the Fe(II) content decreased to 126 mg / L. Figure 2 In (a), a certain amount of reddish-brown precipitate was produced. Figure 2 (b); SEM and XRD analysis showed that the precipitate had a burr-like appearance. Figure 2 c) The chemical composition is ferric hydroxide mineral ( Figure 2 (d)

[0111] Solid-liquid separation: The water sample obtained after biological mineralization treatment is allowed to settle. Lime is added to the supernatant to adjust the pH to 9.0. The concentration of heavy metals in the effluent meets the requirements for discharge standards.

[0112] Comparative Example 2

[0113] This comparative example provides the application of adding only the acidophilic algae Parachlorella sp. AH02 in the biological mineralization and iron removal of acidic mine wastewater, i.e., without adding Acidithiobacillus ferrooxidans LX5.

[0114] Referring to Example 1,

[0115] The water sample was the acidic mine wastewater from Example 1. The specific steps are as follows:

[0116] Inoculation: Add algae to the water sample at a density of 1×10⁻⁶. 7 Parachlorella sp. AH02 cells / mL;

[0117] Add nutrient: 0.5g / L NaNO3;

[0118] Biological mineralization treatment: 1.5 mW / cm 2 The sample was treated under light and at 18°C ​​for 8 hours; throughout the treatment process, the detected electron acceptor and electron supply capacity were 0.9 mmol e. - / g C and 0.7mmol e - / g C; but no hydroxyl radicals were observed to be generated; the pH of the treated water sample was 2.7, and the Fe(II) content was 612 mg / L ( Figure 2 (a) and no obvious precipitate was observed. Figure 2 (b)

[0119] Solid-liquid separation: The water sample after biological mineralization treatment is allowed to settle, and lime is added to the supernatant to adjust the pH to 12.0. The concentration of heavy metals in the obtained effluent meets the requirements for discharge standards.

[0120] Comparative Example 3

[0121] This comparative example provides a method for accelerating iron biomineralization in acidic mine wastewater through the symbiotic relationship between *Thiobacillus ferrooxidans* and *Algae* without synchronous inoculation, and its application.

[0122] Referring to Example 1, the difference is that Acidithiobacillus ferrooxidans LX5 and Parachlorella sp. AH02 were not inoculated synchronously. The specific steps are as follows:

[0123] Inoculation: Add a bacterial density of 5 × 10⁶ to the water sample. 7 Acidithiobacillus ferrooxidans LX5 (units / mL);

[0124] Add nutrient: 0.5g / L NaNO3;

[0125] Biological mineralization treatment: 1.5 mW / cm 2 The algae were first treated under light and at 18°C ​​for 4 hours, and then added at a density of 1×10⁻⁶. 7 Parachlorella sp. AH02 cells / mL was cultured under the same conditions for another 4 hours; the electron accepting capacity and electron supply capacity of this system were measured to be 0.3 mmol / mL. - / g C and 0.2mmol e - / g C; the cumulative concentration of hydroxyl radicals was about 10μM; the water sample after treatment was analyzed and the solution pH was 2.5, the Fe(II) content decreased to 115mg / L, and a certain amount of reddish-brown precipitate was produced; XRD detection showed that the precipitate was ferric hydroxide mineral.

[0126] Solid-liquid separation: After the biological mineralization treatment, the water sample is allowed to settle. Lime is added to the supernatant to adjust the pH to 9.0. The concentration of heavy metals in the obtained effluent meets the requirements for discharge standards.

Claims

1. The application of an algae-bacterial symbiotic system in accelerating iron biomineralization in acidic mine wastewater, wherein the algae-bacterial symbiotic system comprises *Thiobacillus ferrooxidans* and acidophilic algae. The acidophilic ferrooxidizobacterium is Acidithiobacillus ferrooxidans LX5 is deposited at the China General Microbiological Culture Collection Center (CGMCC) on March 13, 2002, with accession number CGMCC NO. 0727. The acidophilic algae is *Chlorella vulgaris*. Parachlorella sp. AH02 is deposited at the China General Microbiological Culture Collection Center (CGMCC) on June 5, 2023, with accession number CGMCC NO.40698.

2. The application according to claim 1, characterized in that, The applications include: S1, Acidophilic ferrooxidizobacterium and acidophilic algae were simultaneously inoculated into acidic mine wastewater; S2, with added nutrients; S3 involves placing acidic mine wastewater under light conditions to induce iron biomineralization.

3. The application according to claim 2, characterized in that, The inoculation density of the *Acidithiobacillus ferrooxidans* was 1 × 10⁻⁶. 7 ~1×10 9 The inoculation density of the acidophilic algae was 1 × 10⁶ cells / mL. 6 ~1×10 8 per mL.

4. The application according to claim 2 or 3, characterized in that, The nutrient solution comprises (g / L): any one or more of the following: NaNO3 0.1~1.5, K2HPO4 0.01~0.04, MgSO4·7H2O 0.01~0.075, CaCl2·2H2O 0.01~0.036, NaCO3 0.01~0.02, propionic acid 0.01~0.2, and citric acid 0.001~0.

01.

5. The application according to claim 4, characterized in that, The illumination conditions include: illumination intensity of 0.2~2 mW / cm². 2 ; The iron biogenic mineralization conditions include: a reaction at a temperature of 15~35℃ for 6~8 h.

6. A treatment process for acidic mine wastewater, characterized in that, The process includes: N1, *Thiobacillus ferrooxidans* and *Algae ferrooxidans* were simultaneously inoculated into acidic mine wastewater; the *Thiobacillus ferrooxidans* was... Acidithiobacillus ferrooxidans LX5, deposited at the China General Microbiological Culture Collection Center (CGMCC) on March 13, 2002, with accession number CGMCC NO.0727; the acidophilic algae is *Chlorella pseudoepiphyllum*. Parachlorella sp. AH02, deposited at the China General Microbiological Culture Collection Center (CGMCC) on June 5, 2023, with accession number CGMCC NO.40698; N2, added nutrients; N3 involves placing acidic mine wastewater under light conditions to induce iron biomineralization, followed by static sedimentation to obtain treated wastewater. N4 is added to the treated wastewater to adjust the pH and obtain the final effluent.

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