Method for biological leaching of metal-containing solid waste

Abstract

The present application relates to the technical field of industrial metal-containing solid waste treatment and resource recovery, and discloses a method for biological leaching treatment of metal-containing solid waste. The method comprises the following steps: (1) culturing first aerobic sulfur bacteria in a first culture medium, and then adding metal ions for continuous culture; (2) separating the obtained culture solution to obtain bacterial cells and supernatant; (3) sequentially performing extraction and concentration on the supernatant to obtain a group signal chemical factor crude extract; (4) adding second aerobic sulfur bacteria and the group signal chemical factor crude extract into a second culture medium for culture, and then mixing the obtained bacterial solution with metal-containing solid waste. The method can greatly improve the content of extracellular polymeric substance of the biological leaching bacterial cells by metal exposure culture of aerobic sulfur bacteria, improve the metal tolerance of the bacterial strain, and further improve the biological leaching treatment efficiency of the metal-containing solid waste, while reducing the cost.

Description

Technical Field

[0001] This invention relates to the field of industrial metal-containing solid waste treatment and resource recycling technology, specifically to a method for bioleaching treatment of metal-containing solid waste. Background Technology

[0002] With the development of industrial production, the amount of industrial solid waste containing metals is increasing daily, especially from the metallurgical, power plant, coal chemical, and petrochemical industries. Currently, most industrial solid waste contains multiple metallic elements such as copper, lead, zinc, chromium, cadmium, arsenic, and mercury. Improper disposal can lead to it flowing into nearby rivers or seeping into the ground with rainwater, polluting the environment and human health. As environmental pollution intensifies, various departments have adopted increasingly stringent environmental protection measures, and industries that enable the harmless disposal and resource utilization of industrial solid waste will enter a golden age of development. At the same time, the investment value of downstream sectors such as industrial solid waste resource utilization equipment and resource recycling methods will become increasingly apparent.

[0003] Currently, the traditional treatment method for metal-containing solid waste is pyrometallurgical processing, but pyrometallurgical processes suffer from high energy consumption, high costs, and large carbon emissions. Hydrometallurgical processes, including chemical wet processes and bioleaching wet processes, are widely used due to their low energy consumption, low carbon emissions, and low costs. However, hydrometallurgical processes also face many drawbacks that need to be overcome. For example, chemical wet processes require a large amount of acid, resulting in high costs; bioleaching technology has long processing times, and the strains used in bioleaching often cannot overcome the significant toxicity of metals, leading to low throughput per batch of solid waste. These drawbacks limit the industrial application of bioleaching processes to some extent.

[0004] Quorum sensing (QS) is an interspecies communication process guided by specific chemical signals (i.e., quorum sensing signal molecules) among bacteria. It can guide microorganisms to collectively alter the physicochemical properties of their cells, thereby enhancing relevant biological performance. The EPS (experimental exogenous substance) content of bacteria can be increased by adding quorum signal molecules. Currently, methods to enhance quorum sensing to improve specific microbial performance all involve adding exogenous pure signal molecules, typically various homoserine lactones, resulting in high costs. Therefore, finding a low-cost method to add these signal molecules is crucial for the practical application of this technology. Summary of the Invention

[0005] The purpose of this invention is to overcome the problems of existing technologies that use bioleaching processes to treat metal-containing solid waste. These problems include the fact that bioleaching strains often cannot overcome the significant metal toxicity, resulting in low single-batch solid waste treatment capacity and limiting the industrial application of bioleaching. Furthermore, current methods for enhancing quorum sensing to improve specific microbial performance involve adding exogenous pure signaling molecules, which is costly. This invention provides a method for bioleaching treatment of metal-containing solid waste. This method involves exposing aerobic sulfur bacteria to metals and preparing a crude extract of quorum signaling chemical factors. Adding this crude extract to the bioleaching bacterial solution significantly increases the extracellular polymer content of the bioleaching bacterial cells, enhancing the strain's resistance to metal toxicity, thereby improving the bioleaching treatment efficiency of metal-containing solid waste while reducing costs.

[0006] To achieve the above objectives, the present invention provides a method for bioleaching treatment of metal-containing solid waste, the method comprising the following steps:

[0007] (1) The first aerobic sulfur bacteria were cultured in the first culture medium, and then metal ions were added to continue the culture.

[0008] (2) Separate the culture medium obtained in step (1) to obtain bacterial cells and a clear liquid containing quorum sensing signal factors;

[0009] (3) The clear liquid is extracted and concentrated sequentially to obtain a crude extract of the population signal chemical factor;

[0010] (4) The second aerobic sulfur bacteria and the crude extract of the population signal chemical factor were added to the second culture medium for cultivation. Then the bacterial solution was mixed with the metal-containing solid waste to leach out the metal in the metal-containing solid waste.

[0011] Preferably, in step (1), the first aerobic sulfur bacteria is a chemoautotrophic aerobic sulfur bacteria.

[0012] Preferably, the first aerobic sulfur bacteria is Thiobacillus thiooxidans and / or Thiobacillus thermophilus.

[0013] Preferably, the first aerobic sulfur bacterium is *Acidithiobacillus thiooxidans*, with accession number GDMCC No: 63522.

[0014] Preferably, in step (1), the first culture medium contains 0.5-1.5 g / L of nitrogen source, 0.02-0.2 g / L of magnesium source, 0.05-0.25 g / L of calcium source, 2-4 g / L of KH2PO4 and 5-20 g / L of energy substance, wherein the nitrogen source is calculated as nitrogen element, the magnesium source as magnesium element, the calcium source as calcium element, and the energy substance is a sulfur source and / or ferrous source; the pH value of the first culture medium is 2-3.

[0015] Preferably, the first culture medium contains 2.5-3.5 g / L (NH4)2SO4, 2-4 g / L KH2PO4, 0.3-1 g / L MgSO4·7H2O, 0.15-0.3 g / L CaCl2, 0.005-0.015 g / L FeSO4 and 10-15 g / L sulfur powder.

[0016] Preferably, in step (1), the culture conditions include: a temperature of 25-40°C and a time of 2-6 days.

[0017] Preferably, in step (1), after the first aerobic sulfur bacteria have been cultured in the first culture medium, the bacterial density is 5 × 10⁻⁶. 8 cfu / mL or higher.

[0018] Preferably, in step (1), the conditions for adding metal ions and continuing cultivation include: a temperature of 25-40℃ and a time of 2-6 days.

[0019] Preferably, in step (1), the metal ion is Ni. 2+ and / or Cu 2+ .

[0020] Preferably, when the metal ion is Ni 2+ When metal ions are added to the first culture medium, the concentration is 0.1–0.5 g / L.

[0021] Preferably, when the metal ion is Cu 2+ At that time, the concentration of metal ions after being added to the first culture medium was 10-50 g / L.

[0022] Preferably, in step (2), the separation method is centrifugation or membrane filtration.

[0023] Preferably, in step (3), the process of sequentially extracting and concentrating the clear liquid includes: extracting the clear liquid with an extractant, and then concentrating the extracted extractant.

[0024] Preferably, the extractant is an organic solvent, and more preferably ethyl acetate.

[0025] Preferably, the volume of the extractant is 1 / 4 to 1 / 2 of the volume of the clear liquid.

[0026] Preferably, the volume of the extracted extractant is concentrated to 1 / 10 to 1 / 4 of its original volume.

[0027] Preferably, in step (3), the extraction process includes: adding the extractant to the clear liquid, stirring for 40-120 minutes, and then letting it stand for 20-50 minutes.

[0028] Preferably, in step (4), the second culture medium contains 0.5-1.5 g / L nitrogen source, 0.02-0.2 g / L magnesium source, 0.05-0.25 g / L calcium source, 2-4 g / L KH2PO4 and 5-20 g / L energy substance, wherein the nitrogen source is calculated as nitrogen element, the magnesium source is calculated as magnesium element, the calcium source is calculated as calcium element, and the energy substance is a sulfur source and / or ferrous source; the pH value of the second culture medium is 2-3.

[0029] Preferably, in step (4), the culture conditions include a temperature of 25-40°C and a time of 2-6 days.

[0030] Preferably, in step (4), after the second aerobic sulfur bacteria and the crude extract of the population signaling chemical factor are added to the second culture medium for cultivation, the viable bacterial count is 10. 9 cfu / mL or higher.

[0031] Preferably, in step (4), the process of adding the second aerobic sulfur bacteria and the crude extract of the population signaling chemical factor to the second culture medium for cultivation includes: first adding the second aerobic sulfur bacteria to the second culture medium for cultivation, and then adding 1 / 30 to 1 / 8 of the volume of the crude extract of the population signaling chemical factor to the resulting system for continued cultivation.

[0032] Preferably, in step (4), the second aerobic sulfur bacteria is a chemoautotrophic aerobic sulfur bacteria and / or the bacterial cells described in step (2).

[0033] More preferably, the chemoautotrophic aerobic sulfur bacteria are thiobacillus thiooxidans and / or thiobacillus thermophilus.

[0034] More preferably, the chemoautotrophic aerobic sulfur bacteria is *Acidithiobacillus thiooxidans*, with accession number GDMCC No: 63522.

[0035] Preferably, in step (4), the metal-containing solid waste contains at least one of Fe, Zn, Cr, Cd, Ni, V, La and Ce.

[0036] Preferably, the metal-containing solid waste is selected from at least one of waste catalysts, sludge, waste lithium-ion batteries, and fly ash from incineration.

[0037] Compared with the prior art, the present invention has at least the following beneficial effects:

[0038] (1) In the prior art, the synthesis of extracellular polymers (EPS) of microorganisms is generally promoted under microbial stress conditions, which will inevitably affect the growth of microorganisms and thus the efficiency of microbial treatment. The method provided by the present invention to improve the efficiency of solid waste bioleaching treatment by increasing the content of extracellular polymers of microorganisms through quorum sensing effect utilizes the principle of quorum effect of microorganisms. The quorum effect chemical factor signal is prepared and extracted in advance, so that the stress conditions for secretion of quorum signal and the promotion of EPS synthesis are separated, avoiding the inhibition of microbial growth caused by promoting EPS synthesis. That is, firstly, the bioleaching bacterial solution is cultured under metal exposure to promote the synthesis of quorum sensing signal molecules. Then, an extractant is added to extract the culture supernatant, and the quorum sensing signal molecule crude extract is prepared after concentration. Then, in the amplification culture stage of the bioleaching bacterial solution, the crude extract of quorum sensing signal molecules is added to significantly increase the content of extracellular polymers of the strain cells without metal stress. Then, the bacterial solution with high content of extracellular polymers is used for bioleaching treatment of solid waste containing metal. The extracellular polymers of the strain can significantly improve the strain's resistance and tolerance to metal toxicity, thereby significantly improving the bioleaching efficiency of the strain.

[0039] (2) Currently, the methods for promoting the resistance of microorganisms to adversity by utilizing the principle of quorum effect all involve adding exogenous pure quorum effect signaling factors, which results in high technical costs. This invention uses crude extracts of signaling factors to promote the synthesis of EPS, which is low in cost, easy to operate, and has significant practical advantages. Detailed Implementation

[0040] The following provides a detailed description of specific embodiments of the present invention. It should be understood that the specific embodiments described herein are for illustrative and explanatory purposes only and are not intended to limit the scope of the invention.

[0041] The endpoints and any values ​​of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0042] The method for bioleaching treatment of metal-containing solid waste provided by the present invention includes the following steps:

[0043] (1) The first aerobic sulfur bacteria were cultured in the first culture medium, and then metal ions were added to continue the culture.

[0044] (2) Separate the culture medium obtained in step (1) to obtain bacterial cells and a clear liquid containing quorum sensing signal factors;

[0045] (3) The clear liquid is extracted and concentrated sequentially to obtain a crude extract of the population signal chemical factor;

[0046] (4) The second aerobic sulfur bacteria and the crude extract of the population signal chemical factor were added to the second culture medium for cultivation. Then the bacterial solution was mixed with the metal-containing solid waste to leach out the metal in the metal-containing solid waste.

[0047] In the method described in this invention, the first aerobic sulfur bacteria is not limited and can be various types of chemoautotrophic aerobic sulfur bacteria. In some embodiments, the first aerobic sulfur bacteria is *Acidithiobacillus thiooxidans* and / or *Acidithiobacillus thermophilus*. In a preferred embodiment, in order to improve the removal efficiency of metals from metal-containing solid waste, the first aerobic sulfur bacteria is *Acidithiobacillus thiooxidans*, which was isolated from activated sludge by the applicant, and this strain was deposited at the Guangdong Provincial Center for Microbial Culture Collection on June 2, 2023, with accession number GDMCC No: 63522.

[0048] In this invention, the viable cell count of the first aerobic sulfur bacteria before culturing is 10. 8 The concentration of cfu / mL or higher is required. Before use, the first aerobic sulfur bacteria strain is activated, and then the seed culture of the first aerobic sulfur bacteria is added to the first culture medium.

[0049] In this invention, the preparation method of the first culture medium includes: adding nitrogen source, magnesium source, calcium source, KH2PO4 and energy substance to deionized water, and then adjusting the pH value to 2-3 using sulfuric acid solution.

[0050] In some embodiments, in step (1), the first culture medium contains 0.5-1.5 g / L of nitrogen source, 0.02-0.2 g / L of magnesium source, 0.05-0.25 g / L of calcium source, 2-4 g / L of KH2PO4, and 5-20 g / L of energy substance, wherein the nitrogen source is calculated as nitrogen element, the magnesium source as magnesium element, and the calcium source as calcium element, and the energy substance is a sulfur source and / or a ferrous source; the pH value of the first culture medium is 2-3. The nitrogen source, magnesium source, and calcium source can be used in the form of salts.

[0051] In some preferred embodiments, the first culture medium contains 2.5-3.5 g / L (NH4)2SO4, 2-4 g / L KH2PO4, 0.3-1 g / L MgSO4·7H2O, 0.15-0.3 g / L CaCl2, 0.005-0.015 g / L FeSO4 and 10-15 g / L sulfur powder.

[0052] In this invention, the conditions for culturing the first aerobic sulfur bacteria in the first culture medium can be appropriately selected according to the actual purpose. In some embodiments, in step (1), the culture conditions include: a temperature of 25-40°C and a time of 2-6 days.

[0053] In this invention, in order to effectively promote the synthesis of population-sensitive signaling factors during subsequent cultivation under metal exposure, in a preferred embodiment, the bacterial density of the first aerobic sulfur bacteria after cultivation in the first culture medium can be 5 × 10⁻⁶. 8 cfu / mL or higher.

[0054] In the method described in this invention, after adding an appropriate amount of metal ions to the first culture medium, the first aerobic sulfur bacteria can grow normally and reach the exponential growth phase. Specifically, after adding metal ions, the growth inhibition rate of the metal ions on the first aerobic sulfur bacteria in the first culture medium is 30-70%, preferably 40-60%.

[0055] In this invention, the metal ion can be a toxic metal ion commonly found in the art. In some embodiments, the metal ion in step (1) is Ni. 2+ and / or Cu 2+ In some more specific embodiments, when the metal ion is Ni... 2+ When metal ions are added to the first culture medium, the concentration can be 0.1–0.5 g / L. In some more specific embodiments, when the metal ion is Cu... 2+ When metal ions are added to the first culture medium, the concentration can be 10–50 g / L.

[0056] In this invention, the conditions for continued cultivation are not particularly limited, as long as the synthesis of population-sensitive signaling factors can be guaranteed. In some embodiments, the conditions for continued cultivation with the addition of metal ions include: a temperature of 25-40°C and a time of 2-6 days.

[0057] In this invention, the quorum sensing signal factor contained in the separated clear liquid is a conventional choice in the art, such as various homoserine lactones.

[0058] In the method described in this invention, the method of separating the culture medium obtained in step (1) is not limited, as long as the bacterial cells can be separated. In some embodiments, the separation method in step (2) can be centrifugation or membrane filtration.

[0059] In one embodiment, step (3), the process of sequentially extracting and concentrating the clarified liquid, includes: extracting the clarified liquid with an extractant, transferring the quorum sensing signal factor in the clarified liquid to the extractant, and then concentrating the extracted extractant. The extractant can be a common, non-toxic organic solvent in the art, preferably ethyl acetate; through the extraction operation, the quorum sensing signal factor in the clarified liquid is transferred to the extractant. In a more specific embodiment, the extraction process in step (3) includes: adding the extractant to the clarified liquid, stirring for 40-120 min, and then allowing it to stand for 20-50 min. The extraction can be performed in an extractor, with a stirring speed of 500-1000 rpm.

[0060] In this invention, when the extractant is added to the clear liquid, the ratio of the amount of extractant to the amount of clear liquid is such that the volume of the extractant is 1 / 4 to 1 / 2 of the volume of the clear liquid.

[0061] In some embodiments, the volume of the extracted extractant can be concentrated to 1 / 10 to 1 / 4 of its original volume. In the method described in this invention, the concentration operation can be performed in a rotary evaporator. The concentration temperature can be 30-50°C.

[0062] (4) The second aerobic sulfur bacteria and the crude extract of the population signal chemical factor were added to the second culture medium for cultivation. Then the bacterial solution was mixed with the metal-containing solid waste to leach out the metal in the metal-containing solid waste.

[0063] In this invention, the second aerobic sulfur bacteria mentioned in step (4) can be a new aerobic sulfur bacteria, or a mixture of a new aerobic sulfur bacteria and the bacterial cells isolated in step (2). Specifically, the second aerobic sulfur bacteria is a chemoautotrophic aerobic sulfur bacteria and / or the bacterial cells mentioned in step (2); the second aerobic sulfur bacteria can be the same as or different from the first aerobic sulfur bacteria. In some embodiments, the chemoautotrophic aerobic sulfur bacteria is *Acidithiobacillus thiooxidans* and / or *Acidithiobacillus thermophilus*. In some preferred embodiments, the chemoautotrophic aerobic sulfur bacteria is *Acidithiobacillus thiooxidans*, with accession number GDMCC No: 63522.

[0064] In this invention, the composition of the second culture medium in step (4) is similar to that of the first culture medium. In some embodiments, the second culture medium contains 0.5-1.5 g / L nitrogen source, 0.02-0.2 g / L magnesium source, 0.05-0.25 g / L calcium source, 2-4 g / L KH2PO4, and 5-20 g / L energy substance, wherein the nitrogen source is calculated as nitrogen element, the magnesium source as magnesium element, the calcium source as calcium element, and the energy substance is a sulfur source and / or a ferrous source; the pH value of the second culture medium is 2-3.

[0065] In the method described in this invention, in order to improve the treatment effect of the bacterial solution on metal-containing solid waste, in a preferred embodiment, in step (4), after the second aerobic sulfur bacteria and the crude extract of the population signaling chemical factor are added to the second culture medium for cultivation, the viable count of the bacteria can be up to 10. 9 cfu / mL or higher.

[0066] In some embodiments, the process of adding the second aerobic sulfur bacteria and the crude extract of the population signaling chemical factor to the second culture medium in step (4) includes: first adding the second aerobic sulfur bacteria to the second culture medium for culture, and then adding 1 / 30 to 1 / 8 of the volume of the crude extract of the population signaling chemical factor to the resulting system for further culture.

[0067] In some embodiments, the conditions for culturing and continuing culturing in step (4) can be performed according to conventional practices in the art. In some embodiments, the conditions for culturing and continuing culturing are independent of each other, including a temperature of 25-40°C and a time of 2-6 days.

[0068] The metals contained in the metal-containing solid waste treated by the method of the present invention can be various metals commonly found in the art, especially transition metals. Specifically, in step (4), the metal-containing solid waste may contain at least one of Fe, Zn, Cr, Cd, Ni, V, La and Ce.

[0069] In this invention, the metal-containing solid waste can be any conventional choice in the art, as long as it contains a metal element. In some embodiments, the metal-containing solid waste is selected from at least one of spent catalysts, sludge, spent lithium-ion batteries, and fly ash from incineration.

[0070] This invention proposes a method to improve the efficiency of solid waste bioleaching treatment by increasing the extracellular polymeric substances (EPS) content of bacterial strains. Utilizing the principle of quorum sensing, a crude extract of a quorum sensing chemical factor that promotes EPS is added to the culture medium of the bioleaching strain. This significantly enhances the synthesis of EPS by the strain cells without metal stress, thereby increasing the metal resistance of the bioleaching strain cells without affecting cell growth. Using bioleaching strains with increased EPS content for the bioleaching treatment of metal-containing solid waste can significantly improve the treatment efficiency.

[0071] The following examples further illustrate the method for bioleaching treatment of metal-containing solid waste according to the present invention. These examples are implemented based on the technical solution of the present invention, providing detailed implementation methods and specific operating procedures; however, the scope of protection of the present invention is not limited to the following examples.

[0072] Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods in the art. Unless otherwise specified, the experimental materials used in the following embodiments are commercially available.

[0073] Example 1

[0074] This embodiment illustrates the preparation process of crude extract of population signaling chemical factors.

[0075] The bacterial strain used in this embodiment is *Acidithiobacillus thiooxidans*, which was isolated from activated sludge by the applicant and deposited at the Guangdong Provincial Center for Microbial Culture Collection on June 2, 2023, with accession number QD16N 63522.

[0076] The specific process includes:

[0077] S1. Prepare a mixture according to the following ratio: (NH4)2SO4 3g / L, KH2PO4 3g / L, MgSO4·7H2O 0.5g / L, CaCl2 0.25g / L, FeSO4 0.01g / L and sulfur powder 10g / L. Add (NH4)2SO4, KH2PO4, MgSO4·7H2O, CaCl2, FeSO4 and sulfur powder to deionized water. Then adjust the pH of the resulting mixture to 3.0 with sulfuric acid solution. After sterilization, obtain the biological leaching liquid culture medium (first culture medium).

[0078] S2. After activating the *Acidithiobacillus thiooxidans* strain, add the *Acidithiobacillus thiooxidans* seed culture to the bioleaching liquid culture medium obtained in step S1, then place it in a 1000mL Erlenmeyer flask (working volume 500mL), and incubate in a shaking incubator at 135rpm and 37℃; incubate for 3 days to achieve a bacterial density of 5×10⁻⁶. 8 cfu / mL or higher;

[0079] S3. Add NiCl2 to the bacterial solution obtained in step S2, so that Ni 2+ The concentration in the bacterial culture was 0.3 g / L, and it was incubated again at 37°C for 3 days.

[0080] S4. The culture medium after metal exposure culture was separated and filtered using a 0.22 μm filter membrane to obtain bacterial cells and filtrate containing quorum sensing signal factors; the bacterial cells retained on the filter membrane were stored at 4 °C, and the filtrate was used for quorum sensing signal factor extraction.

[0081] S5. Add 200 mL of ethyl acetate to approximately 480 mL of filtrate and extract using an extractor at 700 rpm. Stir for 60 min and then let stand for 30 min. After extraction, separate the liquid phases to obtain the extract containing the quorum sensing signal factor.

[0082] S6. Use a rotary evaporator to concentrate about 200 mL of the extract by volume, adjust the rotary evaporation temperature to 40℃, and concentrate to a volume of 40 mL to obtain the crude extract of population signaling chemical factors (QSE).

[0083] Example 2

[0084] This example illustrates the promoting effect of the crude extract of population signaling factor (QSE) prepared in Example 1 on the synthesis of extracellular polymeric substances (EPS) in the strain.

[0085] The specific process includes:

[0086] (1) Prepare a mixture of (NH4)2SO4 3g / L, KH2PO4 3g / L, MgSO4·7H2O 0.5g / L, CaCl2 0.25g / L, FeSO4 0.01g / L and sulfur powder 10g / L. Add (NH4)2SO4, KH2PO4, MgSO4·7H2O, CaCl2, FeSO4 and sulfur powder to deionized water. Then adjust the pH of the resulting mixture to 3 with sulfuric acid solution. After sterilization, obtain the biological leaching liquid culture medium (second culture medium).

[0087] (2) After activating the *Acidithiobacillus thiooxidans* strain, add the *Acidithiobacillus thiooxidans* seed culture to the bioleaching liquid culture medium obtained in step S1, then place it in a 1000mL Erlenmeyer flask (working volume 500mL), and incubate in a shaking incubator at 135rpm and 37℃; incubate for 3 days to achieve a bacterial density of 5×10⁻⁶. 8 cfu / mL or higher;

[0088] (3) Experimental group: 10 mL of the crude extract of population signaling factor (QSE) prepared in Example 1 was added to 100 mL of bacterial culture obtained in step (2), and cultured again at 37℃ for 72 h; at the same time, only steps (1)-(2) were performed, and the bacterial culture without adding the crude extract of population signaling factor (QSE) prepared in Example 1 was set as the control group.

[0089] After the culture is completed, the number of viable bacteria in the bacterial solution is measured to be within 10. 9 The concentration of cfu / mL was above 100, and the EPS content of the bacterial cells was determined.

[0090] The method for determining EPS content is as follows:

[0091] EPS can be classified into soluble EPS (S-EPS), loosely bound EPS (L-EPS), and tightly bound EPS (T-EPS), and is extracted using different centrifugal forces. A 50 mL bacterial culture sample is centrifuged at 2500 g for 15 min to separate the supernatant and precipitate; the supernatant is S-EPS. 50 mL of 0.05% NaCl solution is added to the precipitate, and the mixture is shaken to suspend it. The precipitate is then centrifuged at 5000 g for 15 min, and the supernatant is L-EPS. Another 50 mL of 0.05% NaCl solution is added to the precipitate, and the mixture is shaken and heated in a 60°C water bath for 30 min. Finally, the mixture is centrifuged at 10000 g for 20 min, and the resulting supernatant is T-EPS.

[0092] The polysaccharide content in the supernatant was determined using the phenol-sulfuric acid method, and the protein content was determined using the BCA protein kit. The EPS content was expressed as the sum of the polysaccharide and protein contents.

[0093] The results showed that after adding QSE, the total EPS content of the strains reached 232.8 mg / L; under the same test conditions, the total EPS content in the control group without QSE was 157.2 mg / L, and the EPS content of the strains increased by 48.09% under the action of QSE.

[0094] Example 3

[0095] This example illustrates the promoting effect of the crude extract of population signaling chemical factor QSE prepared in Example 1 on the efficiency of bioleaching and demetallization treatment of waste catalyst.

[0096] This embodiment deals with metal-containing spent catalyst from an oil refinery. The spent catalyst is solid waste generated by an oil refinery, with metal content of 2.8 mg / L Ni, 5.6 mg / L V, 12.9 mg / L La, and 14.5 mg / L Ce.

[0097] The specific process includes:

[0098] Prepare bacterial solutions (including experimental and control groups) according to steps (1)-(3) in Example 2, then add 50g of waste catalyst to the bacterial solutions for demetallization treatment. After 7 days, take the treated waste catalyst to determine the amount of metal remaining and calculate the removal efficiency.

[0099] The results showed that with the addition of QSE, the removal rates of Ni, V, La, and Ce in the spent catalyst reached 36.7%, 41.6%, 59.4%, and 70.2%, respectively, and the bacterial cell density in the bacterial solution increased from the initial 5 × 10⁻⁶. 8 The cfu / mL was increased to 1.2 × 10⁻⁶. 9 cfu / mL.

[0100] In contrast, without the addition of QSE, the removal rates of Ni, V, La, and Ce in the spent catalyst were 9.6%, 14.2%, 32.6%, and 33.8%, respectively, and the bacterial cell density in the bacterial solution increased from the initial 5 × 10⁻⁶. 8 The cfu / mL concentration decreased to 2.3 × 10⁻⁶ at the end of the culture. 8 cfu / mL.

[0101] The results showed that adding QSE to promote EPS synthesis in bacterial cells significantly improved the resistance of the bacteria to the toxicity of removed metals and maintained the bioleaching performance of the bacteria. In contrast, in the treatment group without QSE, the strains had weak resistance to metal toxicity and could not maintain normal growth, resulting in low bioleaching efficiency.

[0102] Example 4

[0103] The same method as in Examples 1-3 was used, except that different aerobic sulfur bacteria were used.

[0104] This embodiment illustrates the process of preparing a crude extract of population signaling chemical factors using *Acidithiobacillus caldus* and improving *Acidithiobacillus* EPS. The strain used in this embodiment is *Acidithiobacillus caldus*, purchased from the China General Microbiological Culture Collection Center (CGMCC), accession number 1.15711.

[0105] Crude extracts of the population signaling factor and the synthesis of extracellular polymeric substances (EPS) of the promoting strain were prepared in the same manner as in Examples 1 and 2, and the content of EPS in *Thiobacillus thermophilus* in Example 2 was determined. The same spent catalyst as in Example 3 was treated with *Thiobacillus thermophilus* with and without the crude extract, in the same manner as in Example 3. The results are as follows:

[0106] The results showed that after adding QSE, the total EPS content of the strain reached 245.2 mg / L; under the same test conditions, the total EPS content in the control group without QSE was 168.5 mg / L, and the EPS content of Thiobacillus thermophilus increased by 45.52% under the action of QSE.

[0107] With the addition of QSE, the removal rates of Ni, V, La, and Ce in the spent catalyst reached 37.2%, 40.3%, 61.1%, and 63.4%, respectively, and the bacterial cell density in the bacterial solution increased from the initial 5 × 10⁻⁶. 8 The cfu / mL was increased to 1.4 × 10⁻⁶. 9 cfu / mL.

[0108] In contrast, without the addition of QSE, the removal rates of Ni, V, La, and Ce in the spent catalyst were 12.4%, 13.8%, 40.3%, and 41.6%, respectively, and the bacterial cell density in the bacterial solution increased from the initial 5 × 10⁻⁶. 8 The cfu / mL concentration decreased to 4.0 × 10⁻⁶ at the end of the culture. 8 cfu / mL.

[0109] The results showed that the addition of QSE promoted the synthesis of EPS in Thiobacillus thermophilus cells, significantly improved the resistance of Thiobacillus thermophilus to metal toxicity compared with the case without the addition of QSE, and maintained the growth of the cells and the performance of bioleaching treatment.

[0110] Example 5

[0111] This embodiment illustrates the promoting effect of the crude extract of population signaling factor QSE prepared in Example 1 on the efficiency of bioleaching and metal removal treatment of residual sludge, and compares the effect of adding and not adding QSE on the sludge treatment efficiency of Thiobacillus thiooxidans.

[0112] This embodiment deals with residual sludge rich in various metals. This sludge is solid waste generated by an activated sludge plant and treated at high temperature. The metal content is Fe (72340 mg / kg), Zn (815 mg / kg), Cr (225 mg / kg), and Ni (73.5 mg / kg).

[0113] The specific process includes:

[0114] Prepare bacterial solution (including experimental group and control group) according to steps (1)-(3) in Example 2, then add 100g of sludge to the bacterial solution and treat for 96h. Take the treated sludge to determine the metal residue and calculate the removal efficiency.

[0115] The results showed that with the addition of QSE, the removal rates of Fe, Zn, Cr and Ni in the sludge reached 71.1%, 62.4%, 73.5% and 84.3%, respectively.

[0116] In contrast, without the addition of QSE, the removal rates of Fe, Zn, Cr and Ni in the sludge were 45.5%, 39.3%, 47.8% and 37.7%, respectively.

[0117] Example 6

[0118] This embodiment illustrates the promoting effect of the crude extract of population signaling factor QSE prepared in Example 1 on the efficiency of bioleaching demetallization treatment of waste lithium-ion batteries, and compares the effect of adding and not adding QSE on the efficiency of sludge treatment by Thiobacillus thiooxidans.

[0119] This embodiment deals with waste lithium-ion batteries, which were taken from a battery factory in Qingdao and processed through unloading, disassembly and crushing. The batteries contained 351,800 mg / kg of cobalt.

[0120] The specific process includes:

[0121] Prepare bacterial solution (including experimental group and control group) according to steps (1)-(3) in Example 2, then add 50g of waste battery, treat for 120h, take the treated waste battery, measure the amount of metal remaining, and calculate the removal efficiency.

[0122] The test results showed that the removal rate of cobalt reached 77.5% with the addition of QSE; in contrast, the removal rate was 25.8% without the addition of QSE.

[0123] Example 7

[0124] This embodiment illustrates the promoting effect of the crude extract of population signaling factor QSE prepared in Example 1 on the efficiency of bioleaching and metal removal treatment of incinerated waste fly ash, and compares the effect of adding and not adding QSE on the efficiency of thiobacillus thiooxidans in treating sludge.

[0125] This embodiment deals with incinerated waste fly ash, taken from a waste treatment plant in Qingdao City. It is rich in Ni, Cu, Zn and Cd, with contents of 21.8 mg / kg, 360.4 mg / kg, 3440.4 mg / kg and 75.2 mg / kg, respectively.

[0126] The specific process includes:

[0127] Prepare bacterial solution (including experimental group and control group) according to steps (1)-(3) in Example 2, then add 100g of incinerated waste fly ash to the bacterial solution, treat for 96h, take the treated fly ash, determine the amount of metal residue, and calculate the removal efficiency.

[0128] The results showed that the removal rates of Ni, Cu, Zn, and Cd reached 70.8%, 65.2%, 78.6%, and 82.8%, respectively, with the addition of QSE.

[0129] In comparison, the removal rates of Ni, Cu, Zn, and Cd without the addition of QSE were 37.8%, 49.2%, 53.6%, and 53.9%, respectively.

[0130] The preferred embodiments of the present invention have been described in detail above; however, the present invention is not limited thereto. Within the scope of the inventive concept, various simple modifications can be made to the technical solutions of the present invention, including combinations of various technical features in any other suitable manner. These simple modifications and combinations should also be considered as the content disclosed in the present invention and are all within the protection scope of the present invention.

Claims

1. A method for biological leaching treatment of metal-containing solid waste, characterized in that, The method includes the following steps: (1) The first aerobic sulfur bacteria were cultured in the first culture medium, and then metal ions were added to continue the culture. (2) Separate the culture medium obtained in step (1) to obtain bacterial cells and a clear liquid containing quorum sensing signal factors; (3) The clear liquid is extracted and concentrated sequentially to obtain a crude extract of the population signal chemical factor; (4) The second aerobic sulfur bacteria and the crude extract of the population signal chemical factor were added to the second culture medium for cultivation. Then the bacterial solution was mixed with the metal-containing solid waste to leach out the metal in the metal-containing solid waste.

2. The method according to claim 1, characterized in that, In step (1), the first aerobic sulfur bacteria is a chemoautotrophic aerobic sulfur bacteria; Preferably, the first aerobic sulfur bacteria is thiobacillus thiooxidans and / or thiobacillus thermophilus; Preferably, the first aerobic sulfur bacterium is *Acidithiobacillus thiooxidans*, with accession number GDMCC No: 63522.

3. The method according to claim 1 or 2, characterized in that, In step (1), the first culture medium contains 0.5-1.5 g / L nitrogen source, 0.02-0.2 g / L magnesium source, 0.05-0.25 g / L calcium source, 2-4 g / L KH2PO4 and 5-20 g / L energy substance, wherein the nitrogen source is calculated as nitrogen element, the magnesium source as magnesium element, the calcium source as calcium element, and the energy substance is a sulfur source and / or ferrous source; the pH value of the first culture medium is 2-3; Preferably, the first culture medium contains 2.5-3.5 g / L (NH4)2SO4, 2-4 g / L KH2PO4, 0.3-1 g / L MgSO4·7H2O, 0.15-0.3 g / L CaCl2, 0.005-0.015 g / L FeSO4 and 10-15 g / L sulfur powder.

4. The method according to any one of claims 1-3, characterized in that, In step (1), the culture conditions include a temperature of 25-40℃ and a time of 2-6 days.

5. The method according to any one of claims 1-4, characterized in that, In step (1), after the first aerobic sulfur bacteria have been cultured in the first culture medium, the bacterial density is 5 × 10⁻⁶. 8 cfu / mL or higher.

6. The method according to any one of claims 1-5, characterized in that, In step (1), the conditions for adding metal ions and continuing the culture include: a temperature of 25-40℃ and a time of 2-6 days.

7. The method according to any one of claims 1-6, characterized in that, In step (1), the metal ion is Ni. 2+ and / or Cu 2+ ; Preferably, when the metal ion is Ni 2+ At that time, the concentration of metal ions after being added to the first culture medium was 0.1–0.5 g / L; Preferably, when the metal ion is Cu 2+ At that time, the concentration of metal ions after being added to the first culture medium was 10-50 g / L.

8. The method according to any one of claims 1-7, characterized in that, In step (2), the separation method is centrifugation or membrane filtration.

9. The method according to any one of claims 1-8, characterized in that, In step (3), the process of sequentially extracting and concentrating the clear liquid includes: extracting the clear liquid with an extractant, and then concentrating the extracted extractant; Preferably, the extractant is an organic solvent, and more preferably ethyl acetate; Preferably, the volume of the extractant is 1 / 4 to 1 / 2 of the volume of the supernatant; Preferably, the volume of the extracted extractant is concentrated to 1 / 10 to 1 / 4 of its original volume.

10. The method according to claim 9, characterized in that, In step (3), the extraction process includes: adding the extractant to the clear liquid, stirring for 40-120 min, and then letting it stand for 20-50 min.

11. The method according to any one of claims 1-10, characterized in that, In step (4), the second culture medium contains 0.5-1.5 g / L nitrogen source, 0.02-0.2 g / L magnesium source, 0.05-0.25 g / L calcium source, 2-4 g / L KH2PO4 and 5-20 g / L energy substance, wherein the nitrogen source is calculated as nitrogen element, the magnesium source is calculated as magnesium element, the calcium source is calculated as calcium element, and the energy substance is sulfur source and / or ferrous source; the pH value of the second culture medium is 2-3.

12. The method according to any one of claims 1-11, characterized in that, In step (4), the culture conditions include a temperature of 25-40°C and a time of 2-6 days.

13. The method according to any one of claims 1-12, characterized in that, In step (4), after the second aerobic sulfur bacteria and the crude extract of the population signaling chemical factor are added to the second culture medium and cultured, the viable bacterial count is 10. 9 cfu / mL or higher; Preferably, in step (4), the process of adding the second aerobic sulfur bacteria and the crude extract of the population signaling chemical factor to the second culture medium for cultivation includes: first adding the second aerobic sulfur bacteria to the second culture medium for cultivation, and then adding 1 / 30 to 1 / 8 of the volume of the crude extract of the population signaling chemical factor to the resulting system for continued cultivation.

14. The method according to any one of claims 1-13, characterized in that, In step (4), the second aerobic sulfur bacteria is a chemoautotrophic aerobic sulfur bacteria and / or the bacterial cells described in step (2); More preferably, the chemoautotrophic aerobic sulfur bacteria are thiobacillus thiooxidans and / or thiobacillus thermophilus; More preferably, the chemoautotrophic aerobic sulfur bacteria is *Acidithiobacillus thiooxidans*, with accession number GDMCC No: 63522.

15. The method according to any one of claims 1-14, characterized in that, In step (4), the metal-containing solid waste contains at least one of Fe, Zn, Cr, Cd, Ni, V, La and Ce; Preferably, the metal-containing solid waste is selected from at least one of waste catalysts, sludge, waste lithium-ion batteries, and fly ash from incineration.