Nano material-microorganism heterozygote as well as preparation method and hydrogen production application thereof

Abstract

The invention relates to the technical field of green hydrogen energy production, in particular to a nano material-microorganism heterozygote as well as a preparation method and hydrogen production application thereof, and the nano material-microorganism heterozygote is formed by electrostatic binding of escherichia coli and an Au-coated CeO2 nano material. The preparation method comprises the following steps: firstly, carrying out aerobic culture on escherichia coli under an aerobic condition, then carrying out anaerobic culture on a bacterial liquid obtained after aerobic culture under an anaerobic condition to obtain a bacterial liquid, resuspending the bacterial liquid in an anaerobic culture medium to obtain a bacterial liquid, then adding Au-coated CeO2, and incubating in an anaerobic atmosphere to obtain the nano material-microorganism heterozygote. According to the method, the hydrogen-producing bacteria escherichia coli and the nanometer material Au-coated CeO2 are combined through static electricity to form a heterozygote, under the mild illumination condition, the yield of hydrogen is very high, and the catalyst is easy to separate and can be recycled for multiple times.

Description

Technical Field

[0001] The present invention relates to the technical field of green hydrogen energy production, and more specifically to a nano material-microorganism hybrid and a preparation method and hydrogen production application thereof. Background Art

[0002] The information disclosed in this background technology section is only intended to enhance the understanding of the overall background of the invention, and should not necessarily be regarded as an admission or any form of suggestion that the information constitutes the prior art already known to a person skilled in the art.

[0003] As an important reducing agent, hydrogen has important applications in production and life. For example, it is used as a fuel for hydrogen-powered vehicles, used for cleaning and reduction in oil processing and metal production, used for welding and cutting in industrial production, and used to produce ammonia, methanol and other chemicals. These methods for preparing hydrogen usually have the disadvantages of complex processes, high energy consumption, and environmental pollution. Photocatalytic hydrogen production can utilize clean solar energy, reduce dependence on fossil fuels, and achieve energy sustainability. Photocatalytic hydrogen production is a green, environmentally friendly and safe method. In the process of photocatalytic hydrogen production, the catalyst generates electrons and holes under illumination. The electrons and holes participate in the reduction-oxidation reaction respectively, and at the same time face the problem of rapid recombination of photocatalyst electron-hole pairs, which makes it challenging to obtain hydrogen efficiently. Hydrogen-producing organisms have the ability to self-replicate and self-repair, have enzymes related to hydrogen production in their bodies, and have high selectivity for hydrogen. Therefore, the combination of photocatalysts and hydrogen-producing organisms to prepare a hybrid system and use photocatalysts to provide energy to hydrogen-producing organisms are of great significance for increasing hydrogen yield and utilizing hydrogen resources.

[0004] Methods for producing hydrogen by hybrids have been reported. Ye Liqun et al. (Wang B, Zeng C, Chu KH, et al. Enhanced biological hydrogen production from Escherichia coli with surfaceprecipitated cadmium sulfide nanoparticles [J]. Advanced Energy Materials, 2017, 7 (20): 1700611.) constructed a whole-cell Escherichia coli-CdS hybrid, and optimized the CdS concentration through a series of experiments to construct and characterize the hybrid. After the hybrid was successfully constructed, Escherichia coli could easily induce endogenous [Ni-Fe] hydrogenase under anaerobic conditions, thus avoiding the cost of complex genetic modification, bacterial screening and enzyme purification. The authors explored the production of pyruvate, lactic acid fermentation, formate concentration, stimulation of hydrogenase activity, and NADH / NAD +Is the ratio the main reason for promoting biological hydrogen production? Under illumination, the hybrid produced 400 μmol more hydrogen than pure Escherichia coli within 3 hours, and the quantum efficiency under 470nm and 620nm monochromatic xenon lamp irradiation reached 7.93% and 9.59%, respectively. However, the electrons generated by the CdS nanomaterials located on the surface of the bacteria under light excitation are blocked by the bacterial membrane, resulting in low utilization of electrons involved in the bacterial hydrogen production pathway. Therefore, there is no strong promotion of the conversion of glucose into pyruvate and further production of hydrogen. Secondly, the CdS nanoparticles formed on the surface of Escherichia coli by in situ mineralization have a certain effect on the biological activity of Escherichia coli. Yuan Shouqi et al. (Jiang Z, Wang B, Jimmy CY, et al. AglnS2 / In2S3 heterostructure sensing of Escherichia coli for sustainable hydrogen production [J]. NanoEnergy, 2018, 46: 234-240.) prepared a heterojunction of AglnS2 and In2S3 to form an inorganic / whole cell hybrid with Escherichia coli. The AglnS2 / In2S3 junction is an excellent light absorbing material with faster conductivity than In2S3. In the study, the authors added an appropriate amount of In 3+In2S3 nanoparticles were biologically cultivated on the surface of Escherichia coli with cysteine, and AglnS2 nanoparticles were fixed on the surface of In2S3 by in situ ion exchange under mild conditions. When the hybrid is illuminated, both AglnS2 and In2S3 can generate photogenerated electrons and holes. The photogenerated electrons on the conduction band of AglnS2 will quickly transfer to the conduction band of In2S3, and finally transfer to Escherichia coli for microbial hydrogen production. At the same time, the photogenerated holes on the valence band of In2S3 will transfer to the valence band of AglnS2. The heterojunction can improve the utilization rate of photogenerated electrons participating in microbial hydrogen production by inhibiting the recombination of photogenerated electrons and holes, and the photogenerated electrons can be quickly transferred to microorganisms without electron transfer agents. However, the efficiency of the electrons generated by the heterojunction on the surface of bacteria under light excitation to pass through the bacterial membrane to enter the interior of the bacteria is low, and the hydrogen production improvement is low. Wang Shaojie et al. (Wang Y, Zhao Y, Wang S, et al. Visible-Light-Driven Enhanced BiohydrogenProduction by Photo-Biohybrid System Based on Photoelectron Transfer betweenIntracellular Photosensitizer Gold Nanoparticles and Clostridium butyricum[J]. ACS Sustainable Chemistry&Engineering, 2022, 11(1): 300-311.) combined gold nanoparticles as intracellular photosensitizers with Clostridium butyricum to construct a hybrid that efficiently produces hydrogen under visible light, with a quantum yield of up to 19.31%. Compared with dark-fermented Clostridium butyricum, the hybrid biohydrogen production increased by 88.74%. The gold nanoparticle-clostridium butyricum hybrid formed has the advantages of being simple, efficient, and not requiring complex and high-cost genetic modification. In addition, there is no problem of photogenerated electrons shuttling between cell membranes and consuming a lot of energy. Compared with Clostridium butyricum, the hybrid has an 88.74% increase in biohydrogen production. Under light irradiation, Au NPs can promote the expression of hydrogenases and key enzymes in the hydrogen production pathway of pyruvate formate lyase. In addition, the photoelectrons of intracellular Au NPs can be transferred to ferredoxin-coupled hydrogenases via electron transfer flavoproteins and FAD, thereby promoting biohydrogen production. Thus, the synergistic catalysis of efficient biohydrogen production between inorganic Au NPs and Clostridium butyricum based on photogenerated electron transfer is achieved.

[0005] Although the hybrid method of preparing hydrogen has made some progress, there are still problems. For example, the electrons generated by extracellular nanomaterials under illumination are difficult to enter the bacteria, and the number of electrons involved in the bacterial hydrogen production pathway is small. Under the illumination of nanomaterials, the holes generated undergo oxidation reactions, promote the production of reactive oxygen, and affect the normal physiological activities of bacteria. Summary of the invention

[0006] In order to address the deficiencies of the prior art, the purpose of the present invention is to provide a nanomaterial-microorganism hybrid and its preparation method and hydrogen production application, using hydrogen-producing bacteria Escherichia coli and nanomaterial Au@CeO2 to form a hybrid through electrostatic bonding. Under milder lighting conditions, the hydrogen yield is very high, and the catalyst is easy to separate and can be recycled multiple times.

[0007] In order to achieve the above object, the technical solution of the present invention is:

[0008] In a first aspect, the present invention provides a nanomaterial-microorganism hybrid, which is composed of electrostatically bonded Escherichia coli and Au@CeO2 nanomaterial.

[0009] Furthermore, the phosphate group of the E. coli is covalently bonded to the Au@CeO2 nanomaterial to form a cerium-phosphate group electronic bridge;

[0010] Preferably, the Escherichia coli is Escherichia coli bio-52502.

[0011] In a second aspect, the present invention provides a method for preparing the above-mentioned nanomaterial-microorganism hybrid, comprising the following steps:

[0012] (1) first aerobically culturing Escherichia coli under aerobic conditions, and then anaerobic culturing the aerobic cultured bacterial solution under anaerobic conditions to obtain a bacterial solution;

[0013] (2) Resuspending the bacterial solution in an anaerobic culture medium to obtain a bacterial solution, adding Au@CeO2, and incubating under an anaerobic atmosphere to obtain the obtained solution.

[0014] Furthermore, in step (1), the Escherichia coli is Escherichia coli bio-52502;

[0015] The aerobic culture medium is LB broth, and the aerobic culture time is 3-4 hours; the anaerobic culture medium is LB broth, glucose and cysteine, and the anaerobic culture time is 2-3 hours.

[0016] Furthermore, in step (2), the anaerobic culture medium is LB broth, glucose and cysteine.

[0017] Further, in step (2), the bacterial solution is resuspended in anaerobic medium to obtain an OD value of the bacterial solution. 600The value is 1.6-1.8;

[0018] The Au@CeO2 is 27.89-55.79 nmol, preferably 41.84 nmol.

[0019] Furthermore, in step (2), the incubation time is 10-15 hours, preferably 10 hours.

[0020] In a third aspect, the present invention provides a method for photocatalytic hydrogen production using the above-mentioned nanomaterial-microorganism hybrid, comprising the following steps:

[0021] The nanomaterial-microorganism hybrid is centrifuged, and then the precipitate is dispersed in a photoreaction container containing an anaerobic culture medium, sealed and filled with inert gas, stirred, and the reaction temperature is 20°C to 35°C. Under light conditions, the reaction time is 3-36 hours, and the reaction produces hydrogen.

[0022] Furthermore, the centrifugal speed is 6500-7000 rpm, and the time is 3-4 min;

[0023] The OD of the nanomaterial-microorganism hybrid dispersed into the anaerobic culture medium 600 The value is 1.8-2.0;

[0024] The anaerobic culture medium contains LB broth, glucose and cysteine;

[0025] The inert gas is nitrogen.

[0026] In a fourth aspect, the present invention provides the use of the above-mentioned nanomaterial-microorganism hybrid in photocatalytic hydrogen production.

[0027] One or more embodiments of the present invention have at least the following beneficial effects:

[0028] (1) The present invention utilizes hydrogen-producing bacteria Escherichia coli and nanomaterial Au@CeO2 to form a hybrid through electrostatic bonding, with a clear hydrogen production metabolic pathway, which reduces the rapid recombination of electron-hole pairs in semiconductor nanomaterials; and the microorganisms have the ability to self-repair and self-repair, and the formed catalyst is easy to separate and can be recycled multiple times. It conforms to the concept of green environmental protection, has mild reaction conditions, can achieve very high conversion efficiency at room temperature, and has very high hydrogen selectivity.

[0029] (2) The present invention uses glucose, which is abundant, cheap and easily available, as a carbon source and does not add expensive photocatalysts, which greatly reduces the cost and provides a new green method for the preparation of hydrogen.

[0030] (3) The hydrogen production method of the present invention is simple, and the hydrogen production rate of the nanomaterial-microorganism hybrid is 5.173 times that of pure Escherichia coli, which is conducive to industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] The accompanying drawings in the specification, which constitute a part of the present invention, are used to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations on the present invention.

[0032] Figure 1 This is a TEM image of the nanomaterial-microorganism hybrid prepared in Example 1 of the present application.

[0033] Figure 2 This is a confocal image of the nanomaterial-microorganism hybrid prepared in Example 1 of the present application. DETAILED DESCRIPTION

[0034] It should be noted that the following detailed descriptions are illustrative and are intended to provide further explanation of the present application. Unless otherwise specified, all technical and scientific terms used in the present application have the same meanings as those commonly understood by those skilled in the art to which the present application belongs.

[0035] It should be noted that the terms used herein are only for describing specific embodiments and are not intended to limit the exemplary embodiments according to the present application. As used herein, unless the context clearly indicates otherwise, the singular form is also intended to include the plural form. In addition, it should be understood that when the terms "comprise" and / or "include" are used in this specification, it indicates the presence of features, steps, operations, devices, components and / or combinations thereof.

[0036] As described in the background technology, although some progress has been made in the method of preparing hydrogen, there are still problems such as harsh reaction conditions, high raw material prices, easy recombination of electrons and holes in semiconductors under light excitation, and low hydrogen yield. Therefore, the present invention proposes a nanomaterial-microorganism hybrid and its preparation method and hydrogen production application. In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail in conjunction with specific embodiments.

[0037] The present invention provides a method for preparing a nanomaterial-microorganism hybrid, comprising the following steps:

[0038] (1) First, Escherichia coli is cultured aerobically under aerobic conditions for 3-4 hours, and then the aerobic cultured bacterial solution is cultured anaerobically under anaerobic conditions for 2-3 hours;

[0039] The Escherichia coli is Escherichia coli bio-52502; the aerobic culture medium is LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH=7); the anaerobic culture medium is LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH=7), 5 g / L glucose and 0.5 g / L cysteine.

[0040] (2) Resuspend the bacterial solution in 50 mL of anaerobic culture medium to obtain a bacterial solution, then add 27.89-55.79 nmol of Au@CeO2, and incubate for 10-15 hours under anaerobic atmosphere to obtain the obtained solution.

[0041] The anaerobic culture medium is LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH=7), 5 g / L glucose and 0.5 g / L cysteine;

[0042] The bacterial solution was resuspended in anaerobic medium to obtain the OD value of the bacterial solution. 600 The value is 1.6-1.8;

[0043] The preferred concentration of Au@CeO2 is 41.84 nmol, and the preferred incubation time is 10 h.

[0044] The present invention combines Escherichia coli (Escherichia coli Castellani & Chalmers) and Au@CeO2 nanomaterials through electrostatic bonding to form a hybrid, and the phosphate group of Escherichia coli and the Au@CeO2 nanomaterials are covalently bonded to form a cerium-phosphate group electronic bridge, so that the hydrogen production metabolic pathway is clear and the rapid recombination of electron-hole pairs in semiconductor nanomaterials is reduced; and the organisms have self-replication and self-repair capabilities, and the formed catalyst is easy to separate and can be recycled for many times.

[0045] The present invention also provides a method for photocatalytic hydrogen production by the above-mentioned nanomaterial-microorganism hybrid, comprising the following steps:

[0046] The nanomaterial-microorganism hybrid is centrifuged, and then the precipitate is dispersed in a photoreaction container containing 50 ml of anaerobic culture medium, and the container is sealed and filled with inert gas, stirred, and the reaction temperature is 20°C to 35°C, and the reaction time is 3-12 hours under the illumination condition of a xenon lamp (λ>420nm), and the reaction produces hydrogen;

[0047] Wherein, the centrifugal speed is 6500-7000rpm, and the time is 3-4min;

[0048] The OD of the nanomaterial-microorganism hybrid dispersed into the anaerobic culture medium 600 The value is 1.8-2.0;

[0049] The anaerobic culture medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH=7), 5 g / L glucose and 0.5 g / L cysteine; the inert gas was nitrogen.

[0050] The reaction mechanism of the nanomaterial-microorganism hybrid to prepare hydrogen is that the oxidized nicotinamide adenine dinucleotide (NAD + ), flavin mononucleotide (FMN), and nicotinamide adenine dinucleotide phosphate (FAD) as electron acceptors. These electron acceptors participate in the redox reaction in microorganisms, which is crucial for promoting the transfer of photogenerated electrons on the surface of nanomaterials to microorganisms and improving the efficiency of hybrid systems for photocatalytic reduction and hydrogen production. Au@CeO2 nanomaterials can covalently bind to the phosphate group of electron acceptors. Under light, Au@CeO2 nanomaterials generate electrons and transfer them to the electron acceptor NAD + , FMN, FAD, promote the generation of NADH, FMNH2, FADH2, and the conduction band position of Au@CeO2 nanomaterials meets the requirements of photogenerated electrons to NAD + , FMN, and FAD, promoting the conversion of glucose into pyruvate and then into formic acid in Escherichia coli, and under the action of formate dehydrogenase, promoting the production of hydrogen.

[0051] Example 1

[0052] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 41.84 nmol Au@CeO2 was added, and the mixture was incubated for 15 h under anaerobic atmosphere to form a hybrid.

[0053] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 3 hours. The product was detected by gas chromatography. The hydrogen production rate was 24.74mmol / h / g dcw .

[0054] Example 2

[0055] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 The value was 1.6, 55.79 nmol Au@CeO2 was added, and the mixture was incubated for 15 h under anaerobic atmosphere to form a hybrid.

[0056] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 3 hours. The product was detected by gas chromatography, and the hydrogen production rate was 17.76mmol / h / g dcw .

[0057] Example 3

[0058] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 27.89 nmol Au@CeO2 was added, and the mixture was incubated for 15 h under anaerobic atmosphere to form a hybrid.

[0059] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 3 hours. The product was detected by gas chromatography, and the hydrogen production rate was 9.79mmol / h / g dcw .

[0060] Example 4

[0061] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 41.84 nmol Au@CeO2 was added, and the mixture was incubated for 10 h under anaerobic atmosphere to form a hybrid.

[0062] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 3 hours. The product was detected by gas chromatography, and the hydrogen production rate was 32.96mmol / h / g dcw .

[0063] Example 5

[0064] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 41.84 nmol Au@CeO2 was added, and the mixture was incubated for 10 h under anaerobic atmosphere to form a hybrid.

[0065] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 6 hours. The product was detected by gas chromatography, and the hydrogen production rate was 35.14mmol / h / g dcw .

[0066] Example 6

[0067] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 41.84 nmol Au@CeO2 was added, and the mixture was incubated for 10 h under anaerobic atmosphere to form a hybrid.

[0068] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 9 hours. The product was detected by gas chromatography, and the hydrogen production rate was 40.43mmol / h / g dcw .

[0069] Example 7

[0070] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 41.84 nmol Au@CeO2 was added, and the mixture was incubated for 10 h under anaerobic atmosphere to form a hybrid.

[0071] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was illuminated by a xenon lamp (λ>420nm) for 12 hours. The product was detected by gas chromatography, and the hydrogen production rate was 42.06mmol / h / g dcw .

[0072] Comparative Example 1

[0073] The difference from Example 1 is that the photocatalyst is only Escherichia coli bio-52502, and its hydrogen production rate is 0.42 mmol / h (the calculation process of its hydrogen production rate is not divided by dry weight).

[0074] Comparative Example 2

[0075] The difference from Example 1 is that the photocatalyst is only Au@CeO2, and its hydrogen production rate is 8.7mmol / h / g dcw .

[0076] Comparative Example 3

[0077] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 41.84 nmol Au@CeO2 was added, and the mixture was incubated for 5 h under anaerobic atmosphere to form a hybrid.

[0078] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 3 hours. The product was detected by gas chromatography, and the hydrogen production rate was 8.58mmol / h / g dcw .

[0079] Comparative Example 4

[0080] First, Escherichia coli bio-52502 was cultured aerobically for 3 h under aerobic conditions (the aerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7)), and then the aerobic cultured bacterial solution was cultured anaerobically for 2 h under anaerobic conditions (LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the bacterial solution was resuspended in 50 mL anaerobic culture medium (the anaerobic culture medium was LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose and 0.5 g / L cysteine), and the OD of the bacterial solution was obtained. 600 value was 1.6, 41.84 nmol Au@CeO2 was added, and the mixture was incubated for 20 h under anaerobic atmosphere to form a hybrid.

[0081] The nanomaterial-microorganism hybrid was centrifuged at 6000 rpm for 3 min, and then the precipitate was dispersed into a photoreaction container containing 50 ml of anaerobic medium (the anaerobic medium contained LB broth (5 g / L yeast extract, 10 g / L tryptone and 10 g / L sodium chloride, pH = 7), 5 g / L glucose, and 0.5 g / L cysteine). The nanomaterial-microorganism hybrid was dispersed into the anaerobic medium at an OD of 600 The value was 1.8. After nitrogen replacement for 30 minutes, the product was irradiated with a xenon lamp (λ>420nm) for 3 hours. The product was detected by gas chromatography, and the hydrogen production rate was 8.49mmol / h / g dcw .

[0082] By comparing Examples 1-7 with Comparative Examples 1 and 2, we can see the advanced nature of the nanomaterial-microorganism hybrid of the present invention: compared with Escherichia coli bio-52502 (Comparative Example 1) and Au@CeO2 (Comparative Example 2), the hydrogen production rate of the hybrid of the present invention is significantly improved.

[0083] By comparing Examples 1 and 5 with Comparative Examples 3 and 4, it can be seen that the incubation time under anaerobic atmosphere when Escherichia coli bio-52502 is combined with Au@CeO2 affects the hydrogen production rate of the hybrid. Compared with incubation under anaerobic atmosphere for 5 hours (Comparative Example 3) and incubation under anaerobic atmosphere for 20 hours (Comparative Example 4), when the incubation time under anaerobic atmosphere is 10 hours and 15 hours, the hybrid of the present invention has a higher hydrogen production rate. Further comparing Example 1 and Example 5, the optimal incubation time of the present invention is 10 hours.

[0084] By comparing Examples 1-3, it can be seen that the amount of Au@CeO2 added affects the hydrogen production rate of the hybrid. When the amount of Au@CeO2 added is 41.84 nmol, the hybrid has a higher hydrogen production rate. Therefore, the optimal amount of Au@CeO2 added in the present invention is 41.84 nmol.

[0085] By comparing Example 5 and Examples 7-9, it can be seen that under the optimal incubation time of 10 h and the optimal amount of nanomaterial of 41.84 nmol, the hydrogen production rate gradually increases with the extension of the light reaction time.

[0086] In summary, the present invention utilizes hydrogen-producing bacteria Escherichia coli and nanomaterial Au@CeO2 to form a hybrid through electrostatic bonding, thereby improving the efficiency of transferring photogenerated electrons from the surface of the nanomaterial to the microorganism, shortening the transfer distance between photogenerated electrons and electron acceptors NAD+, FAD, and FMN in microbial cells, increasing the hydrogen production efficiency, and giving full play to the advantages of the hybrid system in photocatalytic reduction hydrogen production. The catalyst is easy to separate and can be recycled multiple times.

[0087] Finally, it should be noted that the above is only a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the aforementioned embodiments, it is still possible for those skilled in the art to modify the technical solutions described in the aforementioned embodiments, or to perform equivalent replacements on parts thereof. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention. Although the above describes the specific implementation modes of the present invention in conjunction with the accompanying drawings, it is not intended to limit the protection scope of the present invention. Those skilled in the art should understand that, based on the technical solution of the present invention, various modifications or deformations that can be made by those skilled in the art without creative labor are still within the protection scope of the present invention.

Claims

1. A nanomaterial-microorganism hybrid, characterized in that: It is composed of Escherichia coli and Au@CeO2 nanomaterials electrostatically bonded.

2. The nanomaterial-microorganism hybrid according to claim 1, characterized in that: The phosphate group of the Escherichia coli and the Au@CeO2 nanomaterial are covalently bonded to form a cerium-phosphate group electronic bridge; Preferably, the Escherichia coli is Escherichia coli bio-52502.

3. A method for preparing the nanomaterial-microorganism hybrid according to claim 1, characterized in that: The following steps are involved: (1) first aerobically culturing Escherichia coli under aerobic conditions, and then anaerobic culturing the aerobic cultured bacterial solution under anaerobic conditions to obtain a bacterial solution; (2) resuspending the bacterial solution in an anaerobic culture medium to obtain a bacterial solution, then adding Au@CeO2 nanomaterials, and incubating under an anaerobic atmosphere to obtain the obtained solution.

4. The preparation method according to claim 3, characterized in that: In step (1), the Escherichia coli is Escherichia coli bio-52502; The aerobic culture medium is LB broth, and the aerobic culture time is 3-4 hours; the anaerobic culture medium is LB broth, glucose and cysteine, and the anaerobic culture time is 2-3 hours.

5. The preparation method according to claim 3, characterized in that: In step (2), the anaerobic culture medium is LB broth, glucose and cysteine.

6. The preparation method according to claim 3, characterized in that: In step (2), the bacterial solution is resuspended in anaerobic medium to obtain an OD value of the bacterial solution. 600 The value is 1.6-1.8; The Au@CeO2 is 27.89-55.79 nmol, preferably 41.84 nmol.

7. The preparation method according to claim 3, characterized in that: In step (2), the incubation time is 10-15 hours, preferably 10 hours.

8. A method for photocatalytic hydrogen production using the nanomaterial-microorganism hybrid according to claim 1, characterized in that: The following steps are involved: The nanomaterial-microorganism hybrid is centrifuged, and then the precipitate is dispersed in a photoreaction container containing an anaerobic culture medium, sealed and filled with inert gas, stirred, and the reaction temperature is 20°C to 35°C. Under light conditions, the reaction time is 3-36 hours, and the reaction produces hydrogen.

9. The method for photocatalytic hydrogen production using nanomaterial-microorganism hybrids according to claim 8, characterized in that: The centrifugal speed is 6500-7000 rpm, and the time is 3-4 min; The OD of the nanomaterial-microorganism hybrid dispersed into the anaerobic culture medium 600 The value is 1.8-2.0; The anaerobic culture medium contains LB broth, glucose and cysteine; The inert gas is nitrogen.

10. Use of the nanomaterial-microorganism hybrid as claimed in claim 1 in photocatalytic hydrogen production.

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