Methods of recovering gold from waste streams

Abstract

The present invention relates to systems and methods for removing and recovering gold contaminants from wastewater and water streams. The invention also relates to systems and methods for recycling the recovered gold metals. The systems and methods involve a biofilm comprising a hydrogen oxidizer, an acidophile, and a microorganism that reduces gold that captures the reduced gold metal.

Description

METHODS OF RECOVERING GOLD FROM WASTE STREAMSCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of and priority to U.S. Provisional Application No. 63 / 500,900, filed May 8, 2023, the contents of which are incorporated herein by reference in its entirety.FIELD OF THE INVENTION

[0002] The invention relates to systems and methods for removing and recovering gold ions from waste streams or wastewater. The invention also relates to systems and recycling methods for recycling the recovered gold metal for subsequent applications.BACKGROUND OF THE INVENTION

[0003] Precious metals are widely applied in the various fields due to the distinct physical and chemical properties, such as catalytic activity, good electrical conductivity, and corrosion resistance. However, the concentration of precious metals in the Earth’s crust is extremely low, with most of them are below 0.01 ppm. Therefore, it is necessary to find recovery technologies for precious metals.

[0004] Many factors hinder the efficiency of processes that recover gold from wastewaters from the mining, ore-processing, and recycling industries. Hydro- and pyro-metallurgical processes are the conventional processes for recovering gold, but they are expensive and require significant amounts of labor and time. Often, these methods also generate a secondary waste stream and require the use of a series of toxic and expensive chemical agents.SUMMARY OF THE INVENTION

[0005] In one aspect, a system for removing and harvesting gold contaminant (for example, Au3+) from a fluid is disclosed. The system comprises a non-porous gas-transfer membrane, an inoculant comprising a biofilm-forming population of microorganisms that reduces the gold contaminant, and a hydrogen gas (H2) source. In some embodiments, the biofilm comprises at least one species selected from the group consisting of Tabrizicola, Pannonibacter, Azonexus, Aquimonas, and Stenotrophomonas . The non-porous gas-transfer membrane is a hollow-fiber membrane, and the biofilm is anchored to the outer surface of the hollow-fiber membrane. The H2 gas diffuses to the biofilm through the membrane. The microorganisms of the biofilm utilizeIH2 gas as the electron donor to reduce the gold contaminant. In some implementations, the system further comprises a means of harvesting the reduced gold from the biofilm. In some aspects, the means of harvesting the reduced gold from the biofilm harvests the biofilm (the biomass of the system). The reduced gold can be harvested by separation from the biomass.

[0006] In another aspect, a method for removing and harvesting gold contaminant (for example, Au3+) from a fluid is disclosed. Preferably the method includes harvesting gold from contaminated fluids, such as a waste stream or wastewater containing such contaminants. The methods comprise providing an aqueous system comprising a biofilm and a fluid comprising a gold contaminant and contacting the aqueous system with H2 gas. Preferably, the biofilm comprises a Ifc-oxidizing microorganism that reduces the gold contaminant, and the biofilm is anchored to a gas-transfer membrane. In some embodiments, the biofilm comprises at least one species selected from the group consisting of: Tabrizicola, Pannonibacter, Azonexus, Aquimonas, and Stenotrophomonas. During the reduction step, the microorganisms of the biofilm reduce the gold contaminant, using the H2 gas as the electron donor. In some implementations, the gold contaminant is preferably reduced to a solid form, most preferably in the form of a nanoparticle. The resultant solid gold is captured by the biofilm. The solid gold may be stored within the microorganisms of the biofilm and / or trapped within the matrix of extracellular polymeric substances of the biofilm. Accordingly, the reduced gold may be harvested by harvesting the biomass within the aqueous system, which comprises the biofilm.BRIEF DESCRIPTION OF THE DRAWINGS

[0007] The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.

[0008] FIG. 1 illustrates, in accordance with certain embodiments, a schematic of one bench-scale membrane biofilm reactor (MBfR) system.

[0009] FIG. 2 shows, in accordance with certain embodiments, a schematic of a cross section of a hollow membrane with Au-recovering biofilm in a MBfR system.

[0010] FIGs. 3A-3D depict exemplary performance profiles during stages 0-3of gold removal. pH values (FIG. 3A), NCh’-N removal efficiencies (FIG. 3B), Au recoveries (FIG. 3C), and NCh-to N2 and Au reduction fluxes (FIG. 3D) at steady states for all stages.

[0011] FIGs. 4A and 4B depict, in accordance with certain embodiments, relative abundances of bacterial communities at the phylum level (FIG. 4A) and dominant phylotypes at the genus level (FIG. 4B). Boxes in FIG. 4B reveal the selected genus. The scale bar presents the log scale of the relative abundance of each genus within a sample.

[0012] FIGs. 5A-5E depict, in accordance with certain embodiments, optical photos of color changes of different phases in the Au-biofilm reactor.

[0013] FIGs. 6A-6D depict, in accordance with certain embodiments, characterizations of the deep purple solution (FIGs. 6A and 6B) and the light purple solution (FIGs. 6C and 6D) taken from the reactors by TEM.

[0014] FIGs. 7A-7D depict, in accordance with certain embodiments, characterizations of the Au-biofilm by TEM. FIGs. 7A and 7B are TEM images of the Au-biofilm. FIG. 7C is a TEM image featuring the AuNPs. FIG. 7D shows diffraction patterns of AuNPs from FIG. 7C.

[0015] FIGs. 8A-8F depict, in accordance with certain embodiments, the XRD results (FIG. 8A) and XRD analysis (FIG. 8B) of a biofilm sample from a Au-MBfR. FIGs. 8C and 8D are exemplary STEM images. FIGs. 8E and 8F are high-angle dark-field HAADF-STEM images showing Au-element mapping.DETAILED DESCRIPTION OF THE INVENTION

[0016] Detailed aspects and applications of the invention are described below in the drawings and detailed description of the invention. Unless specifically noted, it is intended that the words and phrases in the specification and the claims be given their plain, ordinary, and accustomed meaning to those of ordinary skill in the applicable arts.

[0017] In the following description, and for the purposes of explanation, numerous specific details are set forth in order to provide a thorough understanding of the various aspects of the invention. It will be understood, however, by those skilled in the relevant arts, that the present invention may be practiced without these specific details. It should be noted that there are many different and alternative configurations, devices, and technologies to which the disclosed inventions may be applied. The full scope of the inventions is not limited to the examples that are described below.

[0018] The singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to “a step” includes reference to one or more of such steps.

[0019] As used herein, the term “denitrifying medium” refers to a growth medium comprising nitrate salt (NO3 ) and / or a nitrite salt (NCh ). As the denitrifying medium is a growth medium, it also comprises nutrients to support microbial growth. In some embodiments, the nitrate in the denitrifying medium is from an inorganic nitrate salt, for example, sodium nitrate or potassium nitrate. In some embodiments, the denitrifying medium comprises between 0.5 to 4.0 mM nitrate or nitrite, for example, 1.0 to 4.0 mM, about 0.5 mM, about 1.0, about 1.5 mM, about 2.0 mM, about 3.0 mM, or about 4 mM nitrate or nitrite. In other embodiments, the denitrifying medium comprises 10-70 mg NCh-N / L (orNO2-N / L), for example, 14-70 NO3- N / L (or NO2-N / L), 10-35 mg NO3-N / L (or NO2-N / L), 10-20 mg NO3-N / L (or NO2-N / L), preferably between 12-18 mg NO3-N / L (or NO2-N / L). For example, the denitrifying medium comprises 10 mg NO3-N / L (or NO2-N / L), 11 mg NO3-N / L (or NO2-N / L), 12 mg NCh-N / L (or NO2-N / L), 13 mg NO3-N / L (orNCh-N / L), 14 mgNCh-N / L (orNCh-N / L), 15 mg NCh-N / L (or NO2-N / L), 16 mg NO3-N / L (orNCh-N / L), 17 mgNCh-N / L (orNCh-N / L), 18 mg NCh-N / L (or NO2-N / L), 19 mg NO3-N / L (or NO2-N / L), or 20 mg NO3-N / L (or NO2-N / L). In preferred embodiments, the denitrifying medium comprises trace metals, for example a combination comprising at least one of the group consisting of: zinc, manganese, boron, cobalt, copper, nickel, molybdenum, and selenium.

[0020] As used herein, the term “sludge” refers to thick, soft, wet mud or a similar viscous mixture of liquid and solid components, especially the product of an industrial or refining process. For example, the process may be a wastewater reclamation process. Accordingly, the sludge may be from a wastewater reclamation plant. The sludge may also be anoxic.

[0021] Disclosed herein is an innovative technology for removing and recovering gold from wastewater and waste streams and recycling the recovered gold for subsequent uses, for example, those needing gold metal. The present invention addresses the current deficiencies in commercial application of bioreduction of gold contaminant in wastewater, for example, the presence of Au3+in water. The disclosed systems and methods efficiently remove gold contaminant from wastewater and also enable the harvest of the useful solid forms of gold (Au°). More specifically, the invention involves microbial reduction of Au3+contaminants wherein the Au3+-reducing microorganisms are in a biofilm.

[0022] In some embodiments, the biofilm comprises at least one genus selected from the group consisting of: Tabrizicola. Pannonibacter , Azonexus, Thermomonas, Hydrogenophaga, and Stenotrophomonas . In preferred embodiments, the biofilm comprises a Fh-oxidizing microorganism that reduces Au3+, and the biofilm is anchored to a membrane. In some aspects,the membrane does not have any pores in its wall. In certain preferred embodiments, the membrane comprises a nonporous hollow fiber; and, more specifically in these embodiments, the biofilm anchors to the outer surface of the hollow-fiber of the membrane. The H2 gas is diffused to the biofilm through the membrane. Thus, the H2 gas delivered into the inner surface of the hollow-fiber membrane.

[0023] In certain implementations, the methods comprise providing an aqueous system comprising a biofilm and a fluid comprising a gold contaminant and contacting the aqueous system with hydrogen gas. Preferably, the biofilm comprises a fh-oxidizing microorganism that reduces Au3+, and the biofilm is anchored to a membrane. The method further comprises reducing the Au3+, wherein the reduced gold is captured in the biofilm. The method also may comprise harvesting the reduced gold from the biofilm.

[0024] In some implementations, the methods of the invention comprise producing the biofilm that reduces Au3+contaminant. In such embodiments, the methods of the invention comprise providing an anoxic aqueous system comprising a hollow-fiber membrane; inoculating the hollow-fiber membrane with sludge to establish biofilm on the hollow-fiber membrane; providing a fluid comprising Au3+contaminant to the biofilm; contacting the aqueous system with hydrogen gas; and reducing the Au3+contaminant, wherein the reduced gold metal (Au°) is captured in the biofilm. As nitrate often is present in waste stream or wastewater containing Au3+contaminant, the fluid comprising Au3+contaminant, in some embodiments, further comprises other contaminants, for example nitrate. In some embodiments, the sludge used to inoculate the aqueous system is anoxic. For example, the anoxic sludge is collected from a wastewater reclamation plant. In some aspects, the method further comprises providing a fluid comprising nitrate (1-4 mM) to the inoculated hollow-fiber membrane to establish the biofilm on the hollow-fiber membrane. In some implementations, the fluid comprising Au3+contaminant has a concentration of 100-200 mg / L Au3+contaminant. In some aspects, the fluid comprising Au3+contaminant has a concentration of 100 mg Au3+ / L.

[0025] In other implementation, the methods comprise providing an anoxic aqueous system comprising a hollow-fiber membrane; inoculating the hollow-fiber membrane with anoxic sludge to establish a biofilm-forming population of microorganisms on the hollow-fiber membrane; and providing the inoculated aqueous system with a denitrifying medium until the biofilm-forming population of microorganisms forms a biofilm. The resulting biofilm is capable of reducing Au3+contaminant. The biofilm also is capable of capturing the resultant reduced Au° solids. Hydrogen gas should be provided to the aqueous system during theestablishment of the biofilm. The methods further comprise providing a fluid comprising Au3+contaminant to the biofilm and reducing the Au3+contaminant. In some implementations, methods additionally comprise harvesting the reduced gold from the biofilm.

[0026] In some embodiments, the biofilm may be grown using a denitrifying medium. Accordingly, the methods of the invention may comprise providing an anoxic aqueous system comprising a hollow-fiber membrane; inoculating the hollow-fiber membrane with anoxic sludge to establish biofilm on the hollow-fiber membrane; providing the inoculated aqueous system with a denitrifying medium until the biofilm is formed; providing a fluid comprising Au3+contaminant to the biofilm; contacting the aqueous system with hydrogen gas; and reducing the Au3+contaminant wherein the reduced Au metal is captured in the biofilm.

[0027] In some aspects, the denitrifying medium comprises trace metals. Thus, in some implementations, the denitrifying medium may be supplemented with a trace metal solution. In some embodiments, the trace metal solution comprises, in mg / L, ZnSCh 7H2O 100, MnCh-4 H2O 30, H3BO3 300, C0CI2 6 H2O 200, CuCh 2 H2O 10, NiCh 6 H2O 10, Na2MoO42 H2O 30, and Na2SeO3 30. In such embodiments, the denitrifying medium preferably comprises between 0.5-1.5 mL trace metal stock solution to 1 L of the denitrifying medium, preferably between 0.5-1.0 mL or between 0.75-1.0 mL trace metal stock solution to 1 L of the denitrifying medium.

[0028] In some aspects, the inoculant comprises a hydrogen-oxidizing microorganism, an acidophilic microorganism, and a microorganism that reduces Au3+. In other implementations of the method, the biofilm capable of reducing Au3+contaminant and of capturing the resultant reduced Au is already established. In some aspects, the inoculant comprises at least one Pseudomonadota species. In some aspects, the fluid comprising Au3+contaminant supports the growth of the biofilm-forming population of microorganisms and may be used to establish a biofilm without a separate denitrifying medium.

[0029] Thus, a system for removing and harvesting Au3+contaminant from a fluid is disclosed. In one aspect, the system comprises a biofilm anchored to a membrane and a hydrogen gas source. The biofilm comprises a hydrogen-oxidizing bacterium, an acidophilic bacterium, and a bacterium that reduces Au3+. In a preferred embodiment, the biofilm comprises a microorganism that reduces Au3+. In some aspects, the biofilm comprises at least one genus selected from the group consisting of: Tabrizicola. Pannonibacler. Azonexus, Thermomonas, Hydrogenophaga, and Stenotrophomonas . In another aspect, the system comprises a gas-transfer membrane; an inoculant comprising a biofilm-forming population ofmicroorganisms; and a hydrogen-gas source. The biofilm-forming population of microorganisms comprises a hydrogen-oxidizing microorganism, an acidophilic microorganism, and a microorganism that reduces Au3+.

[0030] During the reduction step, the microorganisms of the biofilm reduce the Au3+contaminant, using the H2 gas as the electron donor, preferably to a solid form of Au, most preferably in the form of a nanoparticle. The resultant solid Au° metal is captured by the biofilm. The solid Au metal may be stored within the microorganisms of the biofilm and / or trapped within the matrix of extracellular polymeric substances of the biofilm. Accordingly, the reduced Au° may be harvested by harvesting the biomass within the aqueous system, which comprises the biofilm.

[0031] In the methods of the invention, microbial reduction of Au3+contaminant uses H2 gas as the electron donor. Compared to organic electron donors, H2 is an advantageous electron donor due to its non-toxicity, lower biomass yield, lower cost, and negative CO2 release. As adding organics to stimulate microbial activities causes membrane fouling and organic residuals in the effluent, using H2 is particularly promising when recovering Au3+from mining waste streams containing high level of soluble Au3+contaminant, but minimal organics. Accordingly, H2 is a particularly promising electron donor when the level of soluble Au3+in a fluid comprising a Au3+contaminant is greater than 100 mg / L and the level of organics is less than 10 mg / L dissolved organic carbon. In some embodiment, the level of soluble Au3+may be greater than 110 mg / L, 120 mg / L, 130 mg / L, 140 mg / L, 150 mg / L, 160 mg / L, 170 mg / L, 180 mg / L, 190 mg / L, or 200 mg / L.

[0032] The H2-gas source can be any reliable source of H2 gas, for example, a gas tank having H2 gas. In some aspects, the gas tank comprises pure H2 gas, which may be purchased directly. The Fb-gas source also may be a hydrogen generator, for example an electrolyzer or a methane reformer that generates H2 gas on-site. The H2 gas is delivered into the hollow fiber such that the H2 gas diffuses to the biofilm through the membrane. Accordingly, in some aspects, gas pressure regulator is used to regulates the pressure of H2 gas provided from the H2-gas source to the membrane. For example, H2 gas is delivered into the hollow fiber such that the H2 gas diffuses to the biofilm through the membrane. The microorganisms of the biofilm utilize H2 gas as the electron donor to reduce the Au3+contaminant. The reduced Au is also captured in the biofilm. In preferred embodiments, the gas regulator is capable of supplying H2 at between 18.0-28.0 psig, preferably between 19.0-28.0 psig, between 20.0-28.0 psig, between 19.0-24.0 psig, between 20.0-24.0 psig, between 20.0-23.0 psig, between 19.0-20.0 psig, between 18.0-20.0 psig, between 20.0-21.0 psig, between 21.0-22.0 psig, between 22.0-23.0 psig, between 23.0-24.0 psig, between 24.0-25.0 psig, between 25.0-26.0 psig, between 26.0-27.0 psig, between 27.0-28.0 psig, about 19.8 psig, about 21.8 psig, about 23.0 psig, about 23.8 psig, or about 27.2 psig. The microorganisms of the biofilm utilize H2 gas as the electron donor to reduce the Au3+contaminant. The reduced Au solid also is captured in the biofilm.

[0033] In some implementations of the methods of the invention, the hollow-fiber membrane comprises hollow fibers having an outer diameter of 200-300 pm, preferably 200- 280 pm, for example 200 pm or 280 pm. The inner diameter of the hollow fibers of the hollowfiber membrane may be 100-110 pm. In some aspects, the cross-sectional area of the hollow fibers of the hollow-fiber membrane is 31,000-66,000 pm2, for example 31,000-36,000 pm2, 36,000-41,000 pm2, 41,000-46,000 pm2, 46,000-51,000 pm2, 51,000-56,000 pm2, 56, GOO- 61, 000 pm2, 61,000-66,000 pm2, or preferably 31,000-32,000 pm2, 61,000-62,000 pm2, 61,000-61,500 pm2, 61,500-62,000 pm2, or more preferably 31,400 pm2or 61,544 pm2. The wall thickness of the hollow fibers of the hollow-fiber membrane may be 50-70 pm, for example between 50-55 pm, 55-60 pm, 60-65 pm, 65-70 pm, or preferably between 55 pm, 55 pm, or 67 pm. In some embodiments, the hollow-fiber membrane is made of composite material, polyester material, or polypropylene material. For example, the hollow-fiber membrane comprises composite hollow fiber, a polyester hollow fiber, or a polypropylene hollow fiber. In some embodiments, the hollow fiber is nonporous. For example, the hollowfiber membrane does not comprise any pore in its wall. The membrane of the system comprises a hollow fiber in preferred embodiments. In some aspects, the biofilm is anchored to the outer surface of the hollow fiber of the membrane. In some embodiments, the membrane comprises polypropylene fibers and has a permeability of 1.8 x 107m3H2 • m membrane thickness / m2hollow fiber surface area • d • bar at standard temperature and pressure. In some aspects, the membrane of the system is nonporous, for example, the membrane lacks pores in its walls.

[0034] In some aspects, establishing a biofilm capable of reducing Au3+contaminant and capturing reduced Au involves culturing the inoculated hollow-fiber membrane with the denitrifying medium or the fluid containing Au3+contaminant for at least three weeks. Accordingly, the denitrifying medium is provided to the inoculated aqueous system for at least three weeks, for example three to four weeks. In some implementations, the denitrifying medium is provided to the inoculated aqueous system for longer periods, such as 160 days or six months.

[0035] The system of the invention may further comprise a growth medium. The growth medium stimulates sufficient microbial growth to establish and / or maintain the biofilm. In some embodiments, the growth medium comprises NH4Q, K2HPO4, CaCh, MgCh, and FeCh. The growth medium may further comprise a nitrate salt (NO3 ) or a nitrite salt (NO2 ), which serves as the electron acceptor. In some aspects, the denitrifying medium comprises trace metals. Thus, in some implementations, the denitrifying medium may be supplemented with a trace metal solution. In some embodiments, the trace metal solution comprises, in mg / L, ZnSO47H2O 100, MnCh-4 H2O 30, H3BO3 300, C0CI2 6 H2O 200, CUC12-2 H2O 10, NiCh-6 H2O 10, Na2MoO4 2 H2O 30, and Na2SeO3 30. In such embodiments, the denitrifying medium preferably comprises between 0.5-1.5 mL trace metal stock solution to I L of the denitrifying medium, preferably between 0.5-1.0 mL or between 0.75-1.0 mL trace metal stock solution to 1 L of the denitrifying medium.

[0036] The aqueous system comprising the biofilm is preferably an anaerobic system. However, in some implementations, the aqueous system may also be aerobic. Accordingly, the sludge need not be anaerobic. In fact, the aqueous system may be inoculated using wastewater / waste stream.

[0037] As wastewater or waste stream containing Au3+contaminant typically is acidic, the fluid containing Au3+contaminant used in the methods of the invention may be acidic. For example, the pH of the fluid containing the Au3+contaminant may have a pH range of 2-8, preferably a pH range of 3-5. In some implementations, the pH of the fluid containing the Au3+contaminant is less than 7, for example a pH of 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, or 4.5. In other aspects, the fluid containing the Au3+contaminant may also have a circumneutral pH, such as a pH range of 6.5-7.5, for example a pH of 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, or 7.5. In still other aspects, the fluid containing the Au3+contaminant may also have a basic pH, such as a pH range of 8.0-9.0, for example a pH of 8.0, 8.1, 8,2, 8.3, 8.4, 8,5, 8.6, 8.7, 8.8, 8.8, or 9.0.Examples

[0038] The present invention is further illustrated by the following examples that should not be construed as limiting. The contents of all references, patents, and published patent applications cited throughout this application, as well as the Figures, are incorporated herein by reference in their entirety for all purposes.Example 1. Set-up of the MBfR

[0039] The concepts underlying the Eb-based MBfR for removing and recovering Au are shown in FIG. 1. An EE-MBfRs was set up to test the recovery of Au. The MBfR system had dual tubes containing a 60-mL working volume, one containing a bundle of 50 hollow-fiber membranes with both ends connected to the EE gas pipeline, and the other containing a “coupon” bundle of 10 identical membranes with only one end connected to the EE gas pipeline. Two tubes were connected using Viton tubing, with one peristaltic pump introducing the influent medium at the foot of the reactor and the effluent exiting on the top. A recirculation pump was used to mix the liquid thoroughly.

[0040] EE gas was delivered directly to a bacterial biofilm present on the outer surface of non-porous gas-transfer membranes. EE gas, fed to the interior (lumen) of hollow-fiber membranes at a controlled pressure, diffused through the membrane walls at a rate determined by the demand generated within the biofilm and in stoichiometric proportion to the supply of the electron acceptors (Au3+) in the water. The EE-delivery capacity to the biofilm was adjusted to meet the stoichiometric demand, which eliminated under- and over-dosing.

[0041] FIG. 2 illustrates the mechanisms for reduction of Au3+to Au° and the capture of Au° solids in the biofilm matrix.Example 2, Continuous evaluation of gold removal and recovery in the MBfR

[0042] Table 1 summarizes the operational conditions for continuous test of gold recovery. To obtain a gold-reducing biofilm, the system was inoculated with a diverse inoculum, such as activated sludge collected from a wastewater treatment plant. Because many gold-reducing bacteria also reduce nitrate (NO3 ), a robust biofilm capable of complete nitrate reduction (to N2 gas) was obtained initially by continuously feeding a synthetic medium, containing 14-70 mg-N / L (4 mM) nitrate as the sole electron acceptor (Stage 0).

[0043] Once a robust biofilm was formed, Au3+was added at a concentration of 1.3 mM (100 mg / L) to enrich for Au3+-reducing bacteria in the biofilm. The system was operated continuously for about one month with liquid samples collected routinely to monitor nitrate and Au3+in the effluent. Other monitored parameters were pH and alkalinity.

[0044] Key operating conditions that were varied systematically were the influent Au3+concentration (mg / L) and Au3+surface loading rate (i.e., g / m2-day). Held constant were H2 pressure, which determines the H2-delivery capacity (i.e., g H2 / m2-day), and the hydraulicretention time (HRT). Table 1 summarized the operating condition for stage 0 (no addition of Au3+) and stages 1, 2, and 3, which had stepwise increases in Au3+input.Table 1. Operational conditions of the MBfR for four stages of continuous tests

[0045] FIGs. 3 A-3D show the effluent pH (FIG. 3 A), removal ratio of NCh' (FIG. 3B), Au recovery as Au° (FIG. 3C), and the removal fluxes for NOf and Au3+(FIG. 3D) across the 4 stages of operation. Table 2 summarizes the performance of the MBfR for stages 0 through 4. Throughout Stage 0, in which the influent contained 4 mM NCh’-N and no Au3+, NCh’-N removal stabilized at 98%. In Stage 1, when 100 mg / L Au3+was added to the influent along with 4 mM NCh’-N, effluent Au3+steadily declined to average of 2 mg / L (97% removal). The removal of NCh’-N reached steady state with an average removal >99% in stage 1. In Stage 2, with 200 mg / L Au(III) in the influent, Au removal was 95% at steady state. In Stage 3, with 500 mg / L Au(III) in the influent, Au removal reached 99% at steady state. The pH declined from 7.9 in stage 0 to 6.2 in stage 3 due to acid production from Au3+reduction, but the MBfR biofilm was capable of increasingly complete reduction of Au(III). NOf reduction declined modestly in stages 2 and 3 but remained greater than 88%. NCh’-N was occasionally detected throughout the entire process, but its concentration was small. The results in FIGs. 3A-3D confirm that the denitrifying biofilm was capable of removing Au3+from the water stream.Table 2. Average performance results at the steady states in the four phases during a continuous-flow operation of the MBfRExample 3, Characterizing the biofilm that reduced Au3+

[0046] The inoculum was sludge collected from the anoxic zone in the Mesa Northwest Wastewater Reclamation Plant (Mesa, AZ). The medium applied in this study was prepared by dissolving 1.5 mmol K2HPO4, 0.002 mmol CaCh, 0.01 mmol FeCh, 0.002 mmol MgSCh, and 1 mL trace metal stock solution (Table 3) in 1-L deionized water. 4 mM NO3-N was added in stage 0 as the sole electron acceptor and nitrogen source for around 30 days to obtain a mature nitrate-reducing biofilm (Stage 0). The influent flow rate was fixed at 0.08 ± 0.01 mL / min (HRT = 12 h), and the H2 pressure was fixed at 10 psig (i.e., 1.68 atm absolute pressure) for all continuous experiments. Once the robust biofilm is formed, media containing 100 mg Au3+ / L were supplied to test if the biofilm in the MBfRs can reduce gold anions (Stage 1-1).

[0047] The microbial community structure and composition were analyzed in each stage to understand the factors affecting the functional bacteria. At the end of each stage, biofilm samples were collected for DNA extraction, and the collected DNA was sequenced. The microorganisms responsible for Au3+reduction were bacteria resistant to heavy metals.Table 3. The trace minerals solution preparation

[0048] FIGs. 4A and 4B present the microbial community structures for stage 0 (without Au) and stages 1 - 3 (with Au). The phylum Pseudomonadota, consisting of a diverse group of denitrifiers, constituted over 80% of the sequences in all stages. The genus Thauera, a typical denitrifier in wastewater bio-treatment but not proficient in metal reduction, was significantlysuppressed by Au3+addition, from 34% in Stage 0 to 0.2% after Au3+addition. Instead, denitrifying genera capable of reducing Au3+and contributing to AuNPs synthesis, such as Stenotrophomonas and Hydrogenophaga, were increased, averaging, respectively, 16% and 24% in Stages 1-3, up from 0.3% and 0.1% in Stage 0. Additionally, denitrifiers with a broad spectrum of electron-donor substrates, including Pannonibacter and Thermomonas. also were enriched: from 3% and 0.07% in stage 0 to averages of 9% and 14% in Au(III)-fed stages, respectively. Thus, the recruitment of denitrifiers known to have multifunctionality overcame any inhibition by Au or low pH. Consequently, the MBfR’s biofilm achieved effective NOf and Au(III) reductions.

[0049] In summary, Tabrizicola. Pannonibacter, and Azonexus are the genera most likely responsible for Au3+reduction.Example 4, Au precipitated as nanoparticles and was captured by the biofilm

[0050] After reduction of Au3+, Au° precipitated as solids inside the biofilm, where they were retained and stabilized as nanoparticles [FIG. 2], In a commercial-scale MBfR, the nanoparticles in the biofilm would be recovered by capturing detached biomass during regular backwashing events that are used to manage the biofilm on the membranes.

[0051] The size and morphology of Au nanoparticles were examined at each stage. At the end of each stage, solid samples of the biofilm are collected by cutting off two 0.5 to 1-cm fiber pieces. After fixation of these samples, they were examined using JEM-ARM200F scanning transmission electron microscopy (STEM) for imaging, crystallite diffraction, and latticefringe fingerprinting. X-ray photoelectron spectroscopy (XPS) of the fibers was carried out using a PHI Quantera SXM (ULVAC-PHI. Inc) with an Al source (a focused beam of 1.5 kV, 25 W) to investigate the valent states of Au and Ag. X-ray powder diffraction (XRD) was conducted with a Philips X’Pert Pro equipped with a Cu Ka radiation source (1.540598 A) to examine the facet of Au / AgNPs. It is reported that pH will affect the size of the formation of Au / AgNPs. a. Reduced-gold particles

[0052] After the addition of Au(III) to the influent, over 97% of the Au(III) was successfully removed (FIGs. 3A and 3B). As a result, the color of the liquid in the reactor changed from transparent (stage 0, not shown) to a deep purple hue (FIG. 5 A. Stage 1), which then transitioned to a light purple (FIG. 5D) before returning to a transparent state (FIG. 5E).Gold / rust-red precipitates attached to the biofilm, as shown in FIGs. 5B and 5C. Furthermore, gold foil was observed following one-month of operation, indicating the accumulation of AuNPs (FIG. 5E).

[0053] The purple color in the liquid was verified by STEM to be caused by AuNPs suspended in the liquid (FIGs. 5A and 5D). Due to surface plasmon resonance, Au nanoparticles of differing concentrations and particle sizes produce colors when observed with the naked eye. The deep purple comes from a high concentration of large AuNPs (FIGs. 6A and 6B), whereas the light purple is from fewer and smaller AuNPs (FIGs. 6C and 6D).

[0054] Due to the precipitation of AuNPs, the smooth rod-like microbial cells in the biofilm were surrounded by small solid particles (FIGs. 7A and 7B). Closer examination (FIG. 7C) shows that the NPs had a size of about 4 nm and exhibited aggregation. The lattice fringes shown in FIG. 7D indicate that the dominant plane for AuNPs is (1 1 1).

[0055] Biofilm-based XPS results revealed that the majority of Au was zero-valent, indicating the absence of unstable intermediate valency states of Au (FIG. 8A). The XRD results demonstrated the association of Au° in the (1 1 1), (2 0 0), (2 2 0), (3 1 1), and (2 2 2) planes with the biofilm (FIG. 8B).

[0056] The TEM results (FIGs. 8C-8D) depict smooth, rod-shaped bacteria within the biofilm, while nanoparticles smaller than 1 pm were visible on the outer surface of the biofilm. These observations confirm enzyme-medicated Au(III) reduction to AuNPs with higher than 90% efficiency.

[0057] High-angle dark-field STEM (HAADF-STEM) images and corresponding elemental distribution of Au STEM-EDX maps, depicted in FIGs. 8E-8F, also demonstrated complete coverage of AuNP on the biofilm.

[0058] In summary, this study confirms that Au3+was reduced to AuNPs and could be recovered in the biofilm.References1. Gericke, M.; Pinches, A., Biological synthesis of metal nanoparticles. Hydrometallurgy 2006, 83, (1-4), 132-140.2. Tarhriz, V.; Akbari, Z.; Dilmaghani, A.; Hamidi, A.; Hejazi, M. A.; Hejazi, M. S., Bioreduction of iron and biosorption of heavy metals (Ni 2+, Co 2+, Pb 2+) by a novel environmental bacterium, Tabrizicola aquatica RCRI19 t. Asian Journal of Water, Environment and Pollution 2019, 16, (3), 73-81.3. Liao, Q.; Tang, J.; Wang, H.; Yang, W.; He, L.; Wang, Y.; Yang, Z., Dynamic proteome responses to sequential reduction of Cr (VI) and adsorption of Pb (II) by Pannonibacter phragmitetus BB. Journal of hazardous materials 2020, 386, 121988.4. Amer, A.; Kim, Y., Isolation of Pb (Il)-reducing bacteria and demonstration of biological Pb (II) reduction to metallic Pb. Journal of Hazardous Materials 2022, 423, 126975.5. Wu, S.; Li, T.; Xia, X.; Zhou, Z.; Zheng, S.; Wang, G., Reduction of tellurite in Shinella sp. WSJ-2 and adsorption removal of multiple dyes and metals by biogenic tellurium nanorods. International Biodeterioration & Biodegradation 2019, 144, 104751.

Claims

CLAIMSWhat is claimed:

1. A system for removing and harvesting gold from a fluid, the system comprising: a gas-transfer membrane; an inoculant comprising a biofilm-forming population of microorganisms comprising a hydrogen oxidizer, an acidophile, and a microorganism that reduces gold; and a hydrogen-gas source, wherein: the biofilm-forming population of microorganisms forms a biofilm, wherein the biofilm is anchored to the gas-transfer membrane, and the fluid comprises gold and the biofilm utilizes hydrogen gas to reduce the gold thereby capturing reduced gold metal in the biofilm.

2. The system of claim 1, wherein the biofilm-forming population of microorganisms comprises at least one genus consisting of: Tabrizicola. Pannonibacter , and Azonexus .

3. The system of claim 1, wherein the biofilm-forming population of microorganisms comprises at least one genus selected from the group consisting of: Tabrizicola, Pannonibacter , Azonexus, Thermomonas, Hydrogenophaga, and Stenotrophomonas .

4. The system of any one of claims 1-3, wherein the membrane is a hollow-fiber membrane.

5. The system of any one of claims 1-3, wherein the hydrogen-gas source is selected from the group consisting of hydrogen generator and a hydrogen gas tank, the system further comprises a gas pressure regulator, wherein the gas pressure regulator regulates the flow of H2 gas from the hydrogen gas source to the membrane.

6. The system of any one of claims 1-3, further comprising a growth medium, the growth medium comprising at least one of an electron acceptor and a nutrient, wherein the electron acceptor is selected from the group consisting of: a nitrate salt (NO3 ), and a nitrite salt (NCh’ ), wherein the nutrient comprises at least one selected from the group consisting of: NH4Q, K2HPO4, CaCl2, MgCh, FeCh, MgSCU, ZnSCU, MnCh, H3BO3, C0CI2, CuCh, NiCh, Na2MoO4, and Na2SeO3.

7. A method of removing gold contaminant from a fluid and harvesting a gold metal, the method comprising:contacting a fluid comprising gold to a hydrogen gas and a biofilm anchored to a gastransfer membrane, the biofilm anchored to the membrane comprising a hydrogen-oxidizing microorganism, an acidophilic microorganism, and a microorganism that reduces gold; capturing the reduced gold metal in the biofilm; and harvesting the reduced gold metal from the biofilm.

8. The method of claim 7, wherein the biofilm comprises a genus of microorganism that is hydrogen-oxidizing and reduces gold.

9. The method of claim 8, wherein the biofilm comprises at least one Pseudomonadota species.

10. The method of claim 8, wherein gold contaminant is removed and gold metal is harvested, the biofilm comprises at least one genus selected from the group consisting of Tabrizicola, Pannonibacter , Azonexus, Thermomonas, Hydrogenophaga, and Stenotrophomonas .

11. The method of claim 7, the method further comprising: providing an aqueous system, wherein the aqueous system comprises the hollow-fiber membrane; inoculating the hollow-fiber membrane with sludge to establish a biofilm-forming population of microorganisms on the hollow-fiber membrane, wherein the biofilm-forming population of microorganisms forms the biofilm anchored to the membrane; and providing the inoculated aqueous system with a fluid comprising gold contaminant, wherein the fluid comprising gold contaminant establishes the biofilm and supports the growth of the biofilm.

12. The method of claim 11, wherein the sludge is anoxic.

13. The method of claim 11, wherein the wherein the biofilm-forming population of microorganisms comprises Tabrizicola and / or Pannonibacter species.

14. The method of claim 11, wherein the biofilm-forming population of microorganisms comprises at least one species selected from the group consisting of Tabrizicola, Pannonibacter, Azonexus, Aquimonas, and Stenotrophomonas.

15. The method of any one of claims 7-14, wherein the fluid comprising gold contaminants further comprises an electron acceptor selected from the group consisting of a nitrate salt (NO:f ) and a nitrite salt (NCh ).

16. The method of any one of claims 7-14, the method further comprising:providing the inoculated aqueous system with a denitrifying medium until the biofilmforming population of microorganisms forms the biofilm, wherein the denitrifying medium comprises an electron acceptor selected from the group consisting of a nitrate salt (NO3 ) and a nitrite salt (NO2 ).

17. The method of any one of claims 7-14, wherein the reduced gold is reduced to form a nanoparticle.

18. The method of any one of claims 7-14, wherein the fluid comprising gold contaminant has a pH range selected from the ranges in the group consisting of: 2-8, 8-9, 6.5-7.5, less than 7, 3-5, and 3-4.

19. The system of claim 4 or the method of any one of claims 11-14, wherein the hollowfiber membrane does not comprise any pores in its wall.

20. The system of claim 4 or the method of any one of claims 11-14, wherein the hollowfiber membrane comprises fibers with an outer diameter of about 200 pm, an inner diameter of about 100-110 pm, and a wall thickness of about 50-55 pm.

21. The method of any one of claims 7-14, wherein the reduced gold is in a solid state.

22. The method of any one of claims 7-14, wherein the aqueous system is an anaerobic aqueous system.

Images