Method for preparing butanediol from nanometer material CdS and CuS heterojunction heterozygote constructed based on genetically engineered bacteria and application of butanediol

Abstract

The invention provides a method for preparing butanediol from a nano-material CdS and CuS heterojunction heterozygote constructed based on genetically engineered bacteria and application of the butanediol, the preparation method comprises the following steps: S1, adding genetically modified vibrio natriureticus into a first culture medium for culturing to obtain a bacterial suspension; s2, collecting thalli of the bacterial suspension, transferring the thalli to a second culture medium, adding heavy metal salt, and culturing to obtain a nanometer material CdS and CuS heterojunction heterozygote constructed based on genetically engineered bacteria; s3, transferring the nanometer material CdS and CuS heterojunction heterozygote constructed based on the genetically engineered bacteria to a third culture medium, adding a culture solution for illumination culture, and collecting supernate to obtain butanediol. The nano material CdS and CuS heterojunction heterozygote constructed based on genetically engineered bacteria prepared by the invention can realize efficient transfer of photo-induced electrons from outside to inside of cells, so as to promote the utilization efficiency of light energy and improve the production efficiency of butanediol.

Description

Technical Field

[0001] The invention relates to the technical field of photocatalysis, and mainly to a method for preparing butanediol based on a nano material CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria and its application. Background Art

[0002] Traditional synthesis methods for chemicals such as butanediol consume a large amount of fossil fuels, and have disadvantages such as non-renewable raw materials, expensive catalysts, low conversion rates, and the generation of toxic and harmful gases and greenhouse gases during the production process. Sugar fermentation is a green way to replace fossil fuels to synthesize chemicals because of its advantages such as good biological selectivity, renewable raw materials, and reduced greenhouse gases. However, it also has the problems of low substrate utilization and high production costs.

[0003] The hybrid consists of two parts: microorganisms and semiconductors. Therefore, it has the advantages of both the efficient light-capturing ability of semiconductor photocatalytic materials and the high specificity, renewability and low cost of biocatalysts. Semiconductors generate photogenerated electrons under light and participate in the metabolic activities in microorganisms, converting light energy into chemical energy. Light energy enhances microbial metabolic synthesis and improves substrate conversion rate and product yield.

[0004] However, there are some problems with the current technical means of producing butanediol using hybrids, including: (1) The light utilization efficiency is not high enough: Although hybrids can utilize light energy, there is still room for improvement in the current light conversion efficiency. For example, the absorption range and conversion capacity of semiconductor materials to light are limited, and not all wavelengths of light can be effectively utilized. In addition, there is energy loss in the process of converting light energy into bioenergy and using it for butanediol synthesis, which ultimately leads to a small effect of light on butanediol production and no obvious advantage over dark conditions; (2) Single metal ion semiconductors cannot meet the demand: Although the hybrids formed by single metal ion semiconductors and microorganisms have a certain promoting effect on the production of butanediol, there is still room for improvement.

[0005] In summary, it is necessary to develop a new technical solution to solve the defects and shortcomings in the existing technology. Summary of the invention

[0006] The invention provides a method for preparing butanediol based on a heterojunction hybrid of nanomaterials CdS and CuS constructed by genetically engineered bacteria and an application thereof. The method for preparing butanediol based on a heterojunction hybrid of nanomaterials CdS and CuS constructed by genetically engineered bacteria utilizes heavy metal salts to construct a photocatalytic material-microorganism hybrid, uses light energy as a drive, uses safe and environmentally friendly microorganisms as carriers, and improves the conversion rate of a substrate and the synthesis efficiency of the product butanediol while converting organic carbon in wastewater at a high value, treating organic wastewater, and treating a variety of heavy metals.

[0007] One object of the present invention is to provide a method for preparing butanediol based on a heterojunction hybrid of nanomaterials CdS and CuS constructed by genetically engineered bacteria. The method for preparing butanediol based on a heterojunction hybrid of nanomaterials CdS and CuS constructed by genetically engineered bacteria comprises the following steps:

[0008] S1, adding the genetically modified Vibrio natrii to the first culture medium for cultivation to obtain a bacterial suspension;

[0009] S2, collecting the bacterial cells of the bacterial suspension, transferring them to a second culture medium, adding heavy metal salts, and culturing to obtain a nanomaterial CdS, CuS heterojunction hybrid constructed based on genetically engineered bacteria;

[0010] S3, transferring the nanomaterial CdS and CuS heterojunction hybrid constructed based on genetically engineered bacteria to a third culture medium, adding culture fluid for light culturing, collecting the supernatant, and obtaining butanediol;

[0011] The structure of the nanomaterial CdS and CuS heterojunction hybrid constructed based on genetically engineered bacteria is that CdS and CuS are coated on the surface of genetically modified Vibrio natrii.

[0012] Furthermore, the genetically modified Vibrio natriuresis is XG203A.

[0013] Furthermore, in the nanomaterial CdS and CuS heterojunction hybrid constructed by the genetically engineered bacteria, the content of CdS is 64wt%, and the content of CuS is 36wt%.

[0014] Furthermore, the heavy metal salt is selected from one or more of Cd salt and Cu salt.

[0015] Furthermore, the heavy metal salt is CdCl 2 , CuCl 2 .

[0016] Furthermore, the concentration of the Cd salt in the second culture medium is 0.005-0.015 mmol / L, and the concentration of the Cu salt in the second culture medium is 0.02-0.08 mmol / L.

[0017] Further, in step S1, the components of the first culture medium are broth peptone 20-30 g, NaCl 11-12 g, KCl 0.3-0.4 g, MgCl 2-3 g and distilled water 800-1200 mL.

[0018] Furthermore, in step S1, the culture temperature is 35-40°C and the rotation speed is 150-250 rpm.

[0019] Furthermore, in step S1, the OD of the bacterial suspension is600 It is 1.0-1.2.

[0020] Further, in step S2, the components of the second culture medium are NH 4 Cl 0.5-1.5g, NaCl 12-13g, Tris-HCl 10-11g, Thiamine HCl 330-350mg, glycerol 3-5ml, Casimino acid 0.5-1.5g, MgSO 4 0.2-0.3 g, CaSO 4 0.01-0.02g, KCl 0.3-0.4g, MgCl 2 2-3g, distilled water 800-1200mL, pH value is 7.

[0021] Furthermore, in step S2, the collection temperature is 10-20°C, the rotation speed is 4000-6000rpm, and the time is 4-6min.

[0022] Furthermore, in step S2, the culture temperature is 35-40°C, the rotation speed is 150-250 rpm, and the time is 20-28 hours.

[0023] Furthermore, in step S3, the components of the third culture medium are the same as those of the second culture medium.

[0024] Furthermore, in step S3, the culture temperature is 35-40°C, the rotation speed is 150-250 rpm, and the time is 20-28 hours.

[0025] Furthermore, in step S3, the transfer temperature is 10-20°C, the rotation speed is 4000-6000 rpm, and the time is 4-6 min.

[0026] Another object of the present invention is to provide the application of the method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria in the production of butanediol.

[0027] The present invention has the following beneficial effects:

[0028] The genetically modified Vibrio natrii can synergistically utilize a variety of heavy metals, sulfates, and organic matter to construct a hybrid, and under suitable lighting conditions, generate as many photogenerated electrons as possible to participate in bacterial metabolic activities, thereby improving the conversion rate of organic matter and the synthesis efficiency of the product butanediol. The genetically modified Vibrio natrii in the present invention can synergistically generate a variety of semiconductor types of hybrids with a variety of heavy metals, and the band gaps of the semiconductors CuS and CdS form a Z-type heterojunction, which reduces the recombination of photogenerated holes and photogenerated electrons, allowing more photogenerated electrons to enter the bacterial metabolic activities, and further improving the synthesis efficiency of the product butanediol.

[0029] The invention provides a method for preparing butanediol based on a heterojunction hybrid of nanomaterials CdS and CuS constructed by genetically engineered bacteria. The butanediol yield of the heterojunction hybrid of nanomaterials CdS and CuS constructed by genetically engineered bacteria is much higher than the butanediol yield of a single genetically modified engineered bacterium Sodium Vibrio under the same culture conditions, which is more than 7 times the yield of a single bacterium cultured under dark conditions and more than 10 times the yield of a single bacterium cultured under light conditions.

[0030] The butanediol production of the heterojunction hybrid of nanomaterials CdS and CuS constructed based on genetically engineered bacteria is also much higher than that of the hybrid constructed with a single nanosulfide heavy metal material under the same culture conditions. It is more than twice the production of the CdS-genetically modified engineering bacteria Sodium Vibrio nitrobenzene hybrid cultured under the same suitable light conditions, and more than 1 times the production of the CuS-genetically modified engineering bacteria Sodium Vibrio nitrobenzene hybrid cultured under the same suitable light conditions. This realizes the efficient transfer of photogenerated electrons from the extracellular to the intracellular, greatly improving the efficiency of light energy utilization and the efficiency of butanediol production. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 A scanning electron microscope image of the product of step S2 of Example 1 is shown.

[0032] Figure 2 The EDS graph of the product of step S2 of Example 1 is shown.

[0033] Figure 3 The scanning electron microscope image of the product of Comparative Example 1 is shown.

[0034] Figure 4 The butanediol concentrations of Example 1, Comparative Example 2, Comparative Example 3, and Comparative Example 4 are shown.

[0035] Figure 5 The butanediol concentrations of Example 1, Comparative Example 3, Comparative Example 5, and Comparative Example 6 are shown. DETAILED DESCRIPTION

[0036] In order to more clearly illustrate the technical solution of the present invention, the following examples are listed. Unless otherwise stated, the raw materials, reactions and post-treatment methods shown in the examples are common raw materials on the market and technical methods well known to those skilled in the art.

[0037] The words "preferred", "preferably", "more preferably", etc. in the present invention refer to embodiments of the present invention that can provide certain beneficial effects in certain circumstances. However, other embodiments may also be preferred under the same circumstances or other circumstances. In addition, the description of one or more preferred embodiments does not imply that other embodiments are not applicable, nor is it intended to exclude other embodiments from the scope of the present invention.

[0038] It should be understood that, except in any operating examples, or where otherwise indicated, all numbers indicating, for example, the amounts of ingredients used in the specification and claims should be understood to be modified in all cases by the term "about". Therefore, unless indicated to the contrary, the numerical parameters set forth in the following specification and the appended claims are approximate values ​​that vary depending on the desired properties to be obtained by the present invention.

[0039] The genetically modified Vibrio natrii of the present invention is XG203A, see Pi, S., Yang, W., Feng, W., Yang, R., Chao, W., Cheng, W., Cui, L., Li, Z., Lin, Y., Ren, N., Yang, C., Lu, L.*, Gao, Xiang. Solar-driven waste-to-chemical conversion by wastewater-derived semiconductor biohybrids. Nature Sustainability. 2023, 6, 1673-1684.

[0040] The following raw materials are used in the embodiments of the present invention:

[0041] The first culture medium consisted of: broth peptone 25 g, NaCl 11.9 g, KCl 0.313 g, MgCl 2.2 g and distilled water 1000 mL.

[0042] The second and third culture media are composed of: NH 4 Cl 1g, NaCl 12.4g, Tris-HCl 10.99g, ThiamineHCl 340mg, glycerol 4ml, Casimino acid 1g, MgSO 4 0.2408 g, CaSO 40.0136g, KCl 0.313g, MgCl 2 2.2g, distilled water 1000mL, pH 7.

[0043] Example 1

[0044] A method for preparing butanediol based on a nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria comprises the following steps:

[0045] S1. Inoculate the genetically modified Vibrio natrii into the first culture medium and culture at 37°C and 200 rpm until the OD 600 =1.0, and bacterial suspension was obtained;

[0046] S2. Centrifuge at 15°C and 5000 rpm for 5 min to collect the bacterial cells in the bacterial suspension, pipette the cells into the second culture medium, and add CdCl 2 , CuCl 2 , so that CdCl 2 , CuCl 2 The concentrations in the second culture medium were 0.01 mmol / L and 0.05 mmol / L, respectively. The culture was shaken at 37°C and 200 rpm for 24 hours. It was observed that the liquid in the second culture medium turned yellow-black, and a heterojunction hybrid of nanomaterials CdS and CuS constructed based on genetically engineered bacteria was obtained.

[0047] S3. Centrifuge the CdS and CuS heterojunction hybrids of nanomaterials constructed based on genetically engineered bacteria at 15°C and 5000 rpm for 5 min, pipette them into the third culture medium, add glucose, L-cysteine ​​hydrochloride and flavin mononucleotide to make their concentrations in the third culture medium 5 g / L, 1 mmol / L and 5 μmol / L, respectively, culture them at 37°C, 200 rpm and light with shaking for 24 h, centrifuge and collect the supernatant to obtain butanediol.

[0048] Figure 1 A scanning electron microscope image of the product of step S2 of Example 1 is shown.

[0049] Figure 2 The EDS graph of the product of step S2 of Example 1 is shown.

[0050] Table 1 shows the element content test results of the product of step S2 of Example 1.

[0051] Table 1 Element content test results of the product of step S2 of Example 1

[0052]

[0053]

[0054] Depend on Figure 1 It can be seen that irregular granular substances are attached to the surface of Example 1; Figure 2 As can be seen from Table 1, the particles attached to the surface of Example 1 contain Cd, Cu and S elements, that is, synthesized CdS / CuS nanoparticles.

[0055] Example 2

[0056] A method for preparing butanediol based on a nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria comprises the following steps:

[0057] S1. Inoculate the genetically modified Vibrio natrii into the first culture medium and culture at 37°C and 200 rpm until the OD 600 =1.2, and obtain bacterial suspension;

[0058] S2. Centrifuge at 15°C and 5000 rpm for 5 min to collect the bacterial cells in the bacterial suspension, pipette the cells into the second culture medium, and add CdCl 2 , CuCl 2 , so that CdCl 2 , CuCl 2 The concentrations in the second culture medium were 0.01 mmol / L and 0.05 mmol / L, respectively. The culture was shaken at 37°C and 200 rpm for 24 hours. It was observed that the liquid in the second culture medium turned yellow-black, and a heterojunction hybrid of nanomaterials CdS and CuS constructed based on genetically engineered bacteria was obtained.

[0059] S3. Centrifuge the CdS and CuS heterojunction hybrids of nanomaterials constructed based on genetically engineered bacteria at 15°C and 5000 rpm for 5 min, pipette them into the third culture medium, add glucose, L-cysteine ​​hydrochloride and flavin mononucleotide to make their concentrations in the third culture medium 5 g / L, 1 mmol / L and 5 μmol / L, respectively, culture them at 37°C, 200 rpm and light with shaking for 24 h, centrifuge and collect the supernatant to obtain butanediol.

[0060] Example 3

[0061] A method for preparing butanediol based on a nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria comprises the following steps:

[0062] S1. Inoculate the genetically modified Vibrio natrii into the first culture medium and culture at 37°C and 200 rpm until the OD 600 =1.1, and obtain bacterial suspension;

[0063] S2. Centrifuge at 15°C and 5000 rpm for 5 min to collect the bacterial cells in the bacterial suspension, pipette the cells into the second culture medium, and add CdCl 2 , CuCl 2 , so that CdCl 2 , CuCl 2 The concentrations in the second culture medium were 0.01 mmol / L and 0.05 mmol / L, respectively. The culture was shaken at 37°C and 200 rpm for 24 hours. It was observed that the liquid in the second culture medium turned yellow-black, and a heterojunction hybrid of nanomaterials CdS and CuS constructed based on genetically engineered bacteria was obtained.

[0064] S3. Centrifuge the CdS and CuS heterojunction hybrids of nanomaterials constructed based on genetically engineered bacteria at 15°C and 5000 rpm for 5 min, pipette them into the third culture medium, add glucose, L-cysteine ​​hydrochloride and flavin mononucleotide to make their concentrations in the third culture medium 5 g / L, 1 mmol / L and 5 μmol / L, respectively, culture them at 37°C, 200 rpm and light with shaking for 24 h, centrifuge and collect the supernatant to obtain butanediol.

[0065] Comparative Example 1

[0066] Comparative Example 1 is prepared in the same manner as Example 1, except that CdCl is not added in step S2. 2 , CuCl 2 And delete step S3.

[0067] Figure 3 The scanning electron microscope image of the product of Comparative Example 1 is shown.

[0068] Depend on Figure 3 It can be seen that the surface of Comparative Example 1 is smooth and no particles are observed.

[0069] Comparative Example 2

[0070] Comparative Example 2 is prepared in the same manner as in Example 1, except that CdCl is not added in step S2. 2 , CuCl 2 And the lighting condition in step S3 is replaced by the dark condition.

[0071] Comparative Example 3

[0072] Comparative Example 3 is prepared in the same manner as in Example 1, except that CdCl is not added in step S2. 2 , CuCl 2 .

[0073] Comparative Example 4

[0074] The preparation method of Comparative Example 4 is the same as that of the embodiment, except that the illumination condition of step S3 is replaced by dark condition.

[0075] Comparative Example 5

[0076] Comparative Example 5 is prepared in the same manner as in Example 1, except that: CuCl is not added in step S2. 2 .

[0077] Comparative Example 6

[0078] Comparative Example 6 is prepared in the same manner as in Example 1, except that CdCl is not added in step S2. 2 And CuCl 2 The concentration in the second culture medium was 0.1 mmol / L.

[0079] Test Example 1

[0080] The butanediol concentrations of Example 1, Comparative Example 2, Comparative Example 3, and Comparative Example 4.

[0081] Test Method

[0082] The butanediol concentrations of Example 1, Comparative Example 2, Comparative Example 3, and Comparative Example 4 were measured.

[0083] Test Results

[0084] Figure 4 The butanediol concentrations of Example 1, Comparative Example 2, Comparative Example 3, and Comparative Example 4 are shown.

[0085] Depend on Figure 4 It can be seen that the butanediol concentration of Example 1 is about 7 times more than that of Comparative Example 2, more than 10 times more than that of Comparative Example 3, and more than 3 times more than that of Comparative Example 4, which illustrates the excellent performance of the nanomaterial CdS, CuS heterojunction hybrid constructed based on genetically engineered bacteria prepared in Example 1, which can efficiently capture light energy and utilize the specific conversion of bacteria, significantly improve the energy conversion efficiency and thus increase the butanediol yield.

[0086] Test Example 2

[0087] The butanediol concentrations of Example 1, Comparative Example 3, Comparative Example 5, and Comparative Example 6.

[0088] Test Method

[0089] The butanediol concentrations of Example 1, Comparative Example 3, Comparative Example 5, and Comparative Example 6 were measured.

[0090] Test Results

[0091] Figure 5 The butanediol concentrations of Example 1, Comparative Example 3, Comparative Example 5, and Comparative Example 6 are shown.

[0092] Depend on Figure 5 It can be seen that the butanediol concentration of the CdS and CuS heterojunction hybrid nanomaterials prepared in Example 1 based on genetically engineered bacteria is higher than that of the nanomaterials without adding CdCl 2 , CuCl 2 The improvement of Comparative Example 3 is the largest, and is also significantly higher than Comparative Examples 5 and 6, indicating that Example 1 has a more excellent performance.

[0093] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above and that the present invention can be implemented in other specific forms without departing from the spirit or essential features of the present invention. Therefore, the embodiments should be considered exemplary and non-restrictive in all respects, and the scope of the present invention is defined by the appended claims rather than the above description, and it is intended that all changes falling within the meaning and scope of the equivalent elements of the claims be included in the present invention.

[0094] In addition, it should be understood that although the present specification is described according to implementation modes, not every implementation mode contains only one independent technical solution. This description of the specification is only for the sake of clarity. Those skilled in the art should regard the specification as a whole. The technical solutions in each embodiment may also be appropriately combined to form other implementation modes that can be understood by those skilled in the art.

Claims

1. A method for preparing butanediol based on a nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria, characterized in that: The method for preparing the nanomaterial CdS and CuS heterojunction hybrid based on genetically engineered bacteria comprises the following steps: S1, adding the genetically modified Vibrio natrii to the first culture medium for cultivation to obtain a bacterial suspension; S2, collecting the bacterial cells of the bacterial suspension, transferring them to a second culture medium, adding heavy metal salts, and culturing to obtain a nanomaterial CdS, CuS heterojunction hybrid constructed based on genetically engineered bacteria; S3, transferring the nanomaterial CdS and CuS heterojunction hybrid constructed based on genetically engineered bacteria to a third culture medium, adding culture fluid for light culturing, collecting the supernatant, and obtaining butanediol; The structure of the nanomaterial CdS and CuS heterojunction hybrid constructed based on genetically engineered bacteria is that CdS and CuS are coated on the surface of genetically modified Vibrio natrii.

2. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 1, characterized in that: The heavy metal salt is selected from one or more of Cd salt and Cu salt.

3. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 2, characterized in that: The concentration of the Cd salt in the second culture medium is 0.005-0.015 mmol / L, and the concentration of the Cu salt in the second culture medium is 0.02-0.08 mmol / L.

4. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 1, characterized in that: In step S1, the culture temperature is 35-40°C and the rotation speed is 150-250 rpm.

5. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 1, characterized in that: In step S1, the OD of the bacterial suspension 600 It is 1.0-1.

2.

6. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 1, characterized in that: In step S2, the collection temperature is 10-20°C, the rotation speed is 4000-6000rpm, and the time is 4-6min.

7. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 1, characterized in that: In step S2, the culture temperature is 35-40°C, the rotation speed is 150-250 rpm, and the time is 20-28 hours.

8. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 1, characterized in that: In step S3, the culture temperature is 35-40°C, the rotation speed is 150-250 rpm, and the time is 20-28 hours.

9. The method for preparing butanediol based on the nanomaterial CdS and CuS heterojunction hybrid constructed by genetically engineered bacteria according to claim 1, characterized in that: In step S3, the transfer temperature is 10-20°C, the rotation speed is 4000-6000 rpm, and the time is 4-6 min.

10. Use of the method for preparing butanediol based on the nanomaterial CdS, CuS heterojunction hybrid constructed by genetically engineered bacteria as claimed in any one of claims 1 to 9 in butanediol production.

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