Microbial synthesis reaction system based on electrochemical energy supply and application thereof

Abstract

The application discloses a kind of microbial synthesis reaction system and its application based on electrochemical energy supply, the system is provided with anode chamber and cathode chamber, biomass derivative is configured in anode chamber and easily oxidized loses electron, the anode chamber is separated from cathode chamber by proton exchange membrane and allows H + Delivery, maintain system pH balance;In cathode chamber, NAD + Conversion is NADH by electrochemical reduction, utilizes H + Maintain neutral environment, NADH is transported to microorganism in cathode chamber by peristaltic pump, microorganism utilizes transport protein to uptake NADH, NADH drives metabolic synthesis, such as rare sugar, rare sugar alcohol, biodegradable plastic monomer, high-activity natural product, microbial oil, etc. + Target product is discharged from organism by transport carrier, backflow to cathode chamber by filter membrane, realizes cyclic utilization.

Description

Technical Field

[0001] This invention relates to advanced manufacturing technology in the field of microorganisms, specifically to a microbial synthesis reaction system based on electrochemical power and its application. Background Technology

[0002] Traditional biosynthetic technologies, such as microbial fermentation, rely on reducing carbon sources like glucose as energy carriers to drive the synthesis of biomolecules through metabolic pathways. However, existing technologies face significant bottlenecks:

[0003] 1. Low carbon efficiency: In traditional synthetic biology, each molecule of glucose is metabolized to produce 6 molecules of CO2, resulting in a carbon loss of about 30%, which exacerbates carbon emissions and resource waste.

[0004] 2. High energy dependence: It requires a large amount of sugar or reduced carbon sources derived from fossil fuels, which is costly and unsustainable.

[0005] 3. Limitations of the reaction system: Traditional electrochemical-biological coupling systems are inefficient, have poor catalyst stability, and are difficult to adapt to complex biosynthetic pathways. In addition, although synthetic chemistry can precisely control molecular structures, it relies on harsh conditions such as high pressure / high temperature, resulting in high energy consumption and pollution, making it difficult to meet the requirements of green manufacturing.

[0006] For example, patent CN107244658A discloses a method for synthesizing hydrogen peroxide using a microbial electrochemical system. This method relies on reducing carbon sources such as glucose as an energy source, generating ATP and NADH through glucose metabolism to drive the synthesis of the target product. In this method, each molecule of glucose produces 6 molecules of CO2, resulting in approximately 30% carbon loss, exacerbating carbon emissions and resource waste. Furthermore, the bioproduct is H2O2, which can be synthesized in large quantities and at low cost in current industrial processes. This method is a typical example of traditional carbon-source-powered biosynthetic technology.

[0007] For example, patent CN102340015A discloses a microbial electrochemical system capable of simultaneously generating electricity, producing hydrogen, and treating wastewater. This system utilizes a membrane technology to separate the pH gradient between the anode and cathode chambers, increasing the cathode hydrogen evolution potential while maintaining the activity of the anode-generating microorganisms oxidizing organic matter in the wastewater. This allows for the simultaneous treatment of organic matter at the anode and the generation of hydrogen at the cathode. Furthermore, due to the increased cathode potential, the reaction can occur spontaneously, outputting electrical energy. However, this solution requires a large amount of electricity derived from fossil fuels, has extremely low output, minimal hydrogen production significance, high cost, and is unsustainable. This solution is a typical example of a traditional electrochemical-biological coupling system with direct electrode power supply.

[0008] For example, the patent CN104328046B, concerning a device and method for reducing carbon dioxide to produce acetic acid using a microbial electrochemical system, discloses a scheme using neutral red, methylene blue, or cobalt-based catalysts to mediate extracellular electron transfer. In this scheme, the electrochemical-biological coupling system is inefficient, the synthetic electron carrier has poor compatibility with the biological system, lacks uptake and transport channels, and the catalyst has poor stability, is easily oxidized by oxygen, and is difficult to adapt to complex biosynthetic pathways, thus failing to meet the requirements of green manufacturing. This scheme belongs to the traditional non-biological interface type artificial electron carrier technology.

[0009] Based on the above problems, the inventors have conducted in-depth research on the mechanism of microbial synthesis reaction and electrochemical energy supply technology, and thus proposed an electrochemical energy-based microbial synthesis reaction system that can solve the above problems. Summary of the Invention

[0010] To overcome the aforementioned problems, the inventors conducted intensive research and designed a microbial synthetic reaction system based on electrochemical power and its application. This system is based on a deep integration of electrochemistry and synthetic biology, selecting biocompatible electron carriers, such as NAD+. + Using monoMOF as a heterogeneous catalyst, NADH is reduced to oxidized NAD+ at the cathode via electrochemical reduction, achieving high efficiency and selectivity. + The conversion to reduced NADH, essentially "molecule charging," replaces the traditional method of NADH production through sugar metabolism. This directly converts renewable electrical energy into bioavailable reducing power, NADH. Furthermore, transport proteins are constructed on the cell membranes of model organisms to better integrate with the biological system, avoiding carbon loss during glucose metabolism and achieving an energy "storage-release" cycle that adapts to the dynamic metabolic needs of microorganisms. Specifically, easily oxidized biomass derivatives that lose electrons are placed in the anode chamber, and a proton exchange membrane separates the anode and cathode chambers while allowing H2O to pass through. + The system is transferred to maintain pH balance; then, NAD+ is reduced electrochemically in the cathode chamber. + Convert to NADH, using H + Maintaining a neutral environment, a peristaltic pump delivers NADH from the cathode chamber to the microorganisms. The microorganisms utilize transport proteins to take up NADH, which drives the metabolic synthesis of target products such as rare sugars, rare sugar alcohols, biodegradable plastic monomers, highly active natural products, and microbial lipids. Simultaneously, NAD+ is also produced. + The organism is discharged through a transport carrier and returned to the cathode chamber through a filter membrane, thus achieving recycling; thereby completing the present invention.

[0011] Specifically, the purpose of this invention is to provide a microbial synthesis reaction system based on electrochemical energy supply, the system comprising an anode chamber and a cathode chamber separated by a proton exchange membrane 1; an anode 2 is disposed in the anode chamber and a cathode 3 is disposed in the cathode chamber;

[0012] An anode flow channel 4 is provided in the anode chamber, and the anode flow channel 4 is filled with an anode substrate, through which electrons are provided;

[0013] Outside the anode and cathode chambers, the anode 2 and cathode 3 are connected in series via a power source to form a circuit;

[0014] A cathode flow channel 5 is provided in the cathode chamber, and the cathode flow channel 5 is connected to the reactor 7 via a peristaltic pump 6; an electron carrier is provided in the cathode flow channel 5 and the reactor 7, and the cathode 3 is capable of continuously generating the electron carrier;

[0015] Microorganisms are also arranged in the reactor 7. These microorganisms take up electron carriers through transport proteins and then use the electron carriers to drive the microbial metabolism to synthesize the target product.

[0016] The proton exchange membrane 1 allows H + This process transfers nutrients to maintain the pH balance of the system and prevents acid-base imbalance from inhibiting microbial activity.

[0017] The electronic carrier is biocompatible;

[0018] Preferably, the electron carrier is selected from one or a mixture of more than one of NADH, NADH artificial analogs, FADH2, and FADH2 artificial analogs;

[0019] Preferably, when the electron carrier is NADH, the microorganism oxidizes NADH to NAD during the metabolic synthesis of the target product. + The cathode 3 continuously generates NADH based on NAD+;

[0020] When the electron carrier is FADH2, the microorganisms oxidize FADH2 to FAD when synthesizing the target product through metabolism, and the cathode 3 continuously generates FADH2 based on FAD.

[0021] The anode 2 and cathode 3 are both obtained by loading a monoMOF catalyst on a substrate; the thickness of the film on the anode 2 and cathode 3 is 2-20 nm.

[0022] Preferably, the substrate is selected from any one or a mixture of carbon / graphite substrate, metal substrate, ITO substrate, and silicon substrate;

[0023] Preferably, the metal substrate is selected from any one of titanium, gold, silver, copper, nickel, platinum, cobalt, and stainless steel.

[0024] Among them, the directional assembly of monoMOF catalysts is achieved on substrates such as carbon and metals by in-situ growth of monoMOF or chemical modification grafting.

[0025] The anode substrate is capable of being oxidized and losing electrons;

[0026] Preferably, the anode substrate is a biomass derivative; more preferably, the anode substrate is 5-hydroxymethylfurfural (HMF), which loses electrons to convert into 2,5-furandicarboxylic acid (FDCA) and is discharged from the outlet of the anode channel 4.

[0027] More preferably, the industrial wastewater containing cellulose or starch is pretreated before being fed into the anode channel as an anode substrate.

[0028] The reactor 7 is either a batch fermenter or a microfluidic continuous flow reactor.

[0029] Preferably, a filter membrane 8 is provided between the reactor 7 and the peristaltic pump 6. This filter membrane 8 allows electron carriers to pass through and is capable of intercepting microorganisms.

[0030] The power source includes solar panels or wind turbines, and also includes energy storage batteries.

[0031] This invention also provides an application of the electrochemically powered microbial synthesis reaction system described above in the production of high-value chemicals, including fragrances, amino acids, rare sugars, rare sugar alcohols, biodegradable plastic monomers, highly active natural products, and microbial oils.

[0032] The fragrances include vanillin, raspberry ketone, naringin, rose essential oil, artemisia annua oil, menthol, γ-decanolide, 2-acetylpyrrolidone, benzaldehyde, ionone, citronellol, β-caryophyllene, and algal β-ionone.

[0033] The amino acids include L-monosodium glutamate, L-lysine, L-threonine, L-tryptophan, L-arginine, L-proline, L-cysteine, L-valine, γ-aminobutyric acid, L-theanine, L-seleno-methylselenocysteine, and ε-polylysine.

[0034] The rare sugars include D-allulose, D-tagatose, L-ribose, D-allulose, L-xylulose, D-minobiose, L-fucose, D-isomalulose, L-rhamnose, D-mannohepanisose, D-allulose, and L-lysolose.

[0035] The rare sugar alcohols include allitol, erythritol, xylitol, allol, maltitol, sorbitol, lactitol, isomaltitol, mannitol, ribitol, tagatitol, galactitol, and erythritol;

[0036] The biodegradable plastic monomers include lactide, succinic acid, 5-hydroxymethylfurfural, 1,4-butanediol, FDCA, lactic acid, adipic acid, 3-hydroxypropionic acid, ε-caprolactone, glycolide, isosorbide, and dimethyl terephthalate.

[0037] The highly active natural products include fucoidan sulfate, rare ginsenoside CK, paclitaxel, astaxanthin, lycopene, steviol glycosides, mogrosides, artemisinin, ginkgolide B, ganoderic acid, huperzine A, and scutellarin.

[0038] The microbial oils include Schizochytrium DHA algal oil, Yersinia lipolytica ARA oil, Rhodotorula glutinis γ-linolenic acid, Chlorella EPA algal oil, M. algae arachidonic acid, Chlorella elliptica astaxanthin oil, Stardew yeast microbial butter, Cryptodinium docosahexaenoic acid, Rhodotorula lycopene lycopene oil, Rhodotorula glutinis β-carotene oil, Microcystis squalene oil, and marine bacterial wax esters.

[0039] The beneficial effects of this invention include:

[0040] (1) The microbial synthesis reaction system based on electrochemical energy supply and its application provided by the present invention uses monoMOF as a heterogeneous electrocatalyst to efficiently and selectively synthesize oxidized NAD3. + It converts renewable electrical energy into reduced NADH, directly transforming it into bioavailable NADH with reducing power. Furthermore, it constructs transport proteins on the cell membranes of model organisms to better match the biological system, avoiding carbon loss during glucose metabolism and significantly improving carbon efficiency. It also realizes an energy "storage-release" cycle, adapting to the dynamic metabolic needs of microorganisms.

[0041] (2) According to the microbial synthesis reaction system based on electrochemical energy supply and its application provided by the present invention, the anode chamber and cathode chamber separated by a proton exchange membrane in the system solve the compatibility problem between electrolyte and fermentation broth;

[0042] (3) According to the microbial synthesis reaction system based on electrochemical energy provided by the present invention, the NADH delivery rate is controlled by a peristaltic pump and synchronized; in addition, NADH is the core reducing power of microorganisms, which supports multi-step product synthesis, making the system adaptable to complex reaction paths, thereby making the system universal.

[0043] (4) The microbial synthesis reaction system based on electrochemical energy supply and its application provided by the present invention, wherein NAD+ is used in the system. + The recycling process reduces carrier consumption, extends the lifespan of the platinum-based catalyst to over 100 hours, improves system stability, and lowers the economic cost of using the system, giving it a brighter market prospect.

[0044] (5) According to the microbial synthesis reaction system based on electrochemical power supply and its application provided by the present invention, the power source in the system can be set as a solar panel or a wind turbine, which, combined with an energy storage battery, can solve the problem of intermittent renewable energy, realize the sustainable supply of energy at all times, and reduce the dependence on traditional carbon sources.

[0045] (6) According to the microbial synthesis reaction system based on electrochemical energy provided by the present invention and its application, the anode substrate can be replaced with industrial wastewater, such as papermaking wastewater containing organic matter such as cellulose and starch, so that the anode can be used to treat wastewater at the same time. Attached Figure Description

[0046] Figure 1 This diagram shows the overall structure of the microbial synthesis reaction system based on electrochemical power supply in this application;

[0047] Figure 2 The hydrogen NMR spectrum of Me2DBB is shown;

[0048] Figure 3 The 1H NMR spectrum of Me2DBB-RhCp*Cl is shown.

[0049] Figure 4 The hydrogen nuclear magnetic resonance spectrum of H2DBB-RhCp*Cl is shown.

[0050] Figure 5 Hf is shown 12 Electron micrograph of -DBB-RhCp*ClmonoMOF;

[0051] Figure 6 The image shows an SEM image of Rh monoMOF grown in situ on a titanium felt, i.e., Rh monoMOF-Ti substrate.

[0052] Figure 7 The image shows a SEM image of Rh monoMOF grown in situ on carbon paper, i.e., Rh monoMOF-C substrate.

[0053] Figure 8 The UV standard concentration curve of NADH in the experimental example is shown.

[0054] Figure 9 and Figure 10Electrochemical NAD of H2DBB-RhCp*Cl ligands is shown + Reconstructing the test results diagram;

[0055] Figure 11 and Figure 12 Electrochemical NAD of Rh monoMOF-C is shown. + Reconstructing the test results diagram;

[0056] Figure 13 and Figure 14 Electrochemical NAD of Rh monoMOF-Ti is shown + Reconstructing the test results diagram;

[0057] Figure 15 A schematic diagram showing the expression of NTT4 on the membrane surface of Escherichia coli;

[0058] Figure 16 This illustrates a plasmid system for the production of allicin from Escherichia coli.

[0059] Figure 17 The results of SDS-PAGE staining and Western blot analysis of purified Escherichia coli NTT4 are shown.

[0060] Figure 18 The image shows Coomassie smear staining of SDS-PAGE purified from DPE and RDH.

[0061] Explanation of reference numerals in the attached figures

[0062] 1-Proton exchange membrane, 2-Anode, 3-Cathode, 4-Anode channel, 41-Channel inlet, 42-Channel outlet, 5-Cathode channel, 6-Peristaltic pump, 7-Reactor, 8-Filter membrane. Detailed Implementation

[0063] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments. Through these descriptions, the features and advantages of the present invention will become clearer and more apparent.

[0064] The term “exemplary” as used herein means “serving as an example, embodiment, or illustration.” Any embodiment illustrated herein as “exemplary” is not necessarily to be construed as superior to or better than other embodiments. Although various aspects of embodiments are shown in the accompanying drawings, the drawings are not necessarily drawn to scale unless specifically indicated otherwise.

[0065] This invention provides a microbial synthesis reaction system based on electrochemical energy supply, such as... Figure 1 As shown, the system includes an anode chamber and a cathode chamber separated by a proton exchange membrane 1; an anode 2 is disposed in the anode chamber and a cathode 3 is disposed in the cathode chamber;

[0066] An anode flow channel 4 is provided in the anode chamber, and the anode flow channel 4 is filled with an anode substrate, through which electrons are provided;

[0067] Outside the anode and cathode chambers, the anode 2 and cathode 3 are connected in series by a power source to form a circuit, thereby providing a path for the directional flow of electrons and continuously supplying electrons to the cathode;

[0068] A cathode flow channel 5 is provided in the cathode chamber, and the cathode flow channel 5 is connected to the reactor 7 via a peristaltic pump 6. An electron carrier is provided in the cathode flow channel 5 and the reactor 7, and the cathode 3 can continuously generate the electron carrier. That is, the electron carrier that has lost electrons is transported to the cathode chamber by the peristaltic pump 6, regains electrons in the cathode chamber and becomes an electron carrier again, and is then pumped into the reactor 7 by the peristaltic pump, thereby realizing the recycling of the electron carrier.

[0069] Microorganisms are also arranged in the reactor 7. These microorganisms take up electron carriers through transport proteins and then use the electron carriers to drive the microbial metabolism to synthesize the target product.

[0070] In a preferred embodiment, the proton exchange membrane 1 allows H... + The transfer of these substances maintains the pH balance of the system, preventing acid-base imbalance from inhibiting microbial activity. While preventing electrochemical reagents from entering reactor 7, the pH value in reactor 7 is maintained at 6.5-7.5, providing a suitable acid-base environment for microbial survival.

[0071] In a preferred embodiment, the electron carrier is biocompatible and can meet the needs of specific microorganisms through electrochemical reduction;

[0072] Preferably, the electron carrier is selected from one or a mixture of more than one of NADH, NADH artificial analogs, FADH2, and FADH2 artificial analogs; preferably NADH or FADH2.

[0073] When the electron carrier is NADH, the microorganism oxidizes NADH to NAD during the metabolic synthesis of the target product. + The cathode 3 is continuously based on NAD + Generate NADH;

[0074] When the electron carrier is FADH2, the microorganisms oxidize FADH2 to FAD when synthesizing the target product through metabolism, and the cathode 3 continuously generates FADH2 based on FAD.

[0075] Taking NADH as an example, the working process of electron carriers is explained as follows:

[0076] Select biocompatible NAD +Using NADH as an electron carrier and monoMOF as a heterogeneous catalyst, oxidized NAD+ is efficiently and selectively reduced at the cathode via electrochemical reduction. + The process of converting NADH into reduced NADH, known as "molecular charging," replaces the traditional method of producing NADH through sugar metabolism. In this application, renewable electrical energy is directly converted into biologically usable reducing power (NADH), and transport proteins are constructed on the cell membranes of model organisms to better match the biological system and avoid carbon loss from glucose metabolism. This achieves an energy "storage-release" cycle that adapts to the dynamic metabolic needs of microorganisms.

[0077] NAD in this application + The full name is nicotinamide adenine dinucleotide (oxidized coenzyme I); the full name of NADH in this application is reduced nicotinamide adenine dinucleotide (reduced coenzyme I).

[0078] The construction of transport proteins on the cell membranes of model organisms described in this application can be based on existing technologies. For example, NTT4 is expressed on the surface of E. coli membranes to achieve NAD(H) transport. A schematic diagram of NTT4 expression on the surface of E. coli membranes is shown below. Figure 15 As shown, the plasmid system of Escherichia coli for the production of allicin is as follows: Figure 16 As shown; Coomassie smear staining and Western blot results of SDS-PAGE of purified E. coli NTT4 are as follows. Figure 17 As shown; Coomassie staining of SDS-PAGE purified from DPE and RDH as shown. Figure 18 As shown.

[0079] In a preferred embodiment, both the anode 2 and the cathode 3 are obtained by loading a monoMOF catalyst onto a substrate; the thickness of the films on the anode 2 and the cathode 3 is 2-20 nm.

[0080] Preferably, the substrate is selected from any one or a mixture of carbon / graphite substrate, metal substrate, ITO substrate, and silicon substrate;

[0081] Preferably, the metal substrate is selected from any one of titanium, gold, silver, copper, nickel, platinum, cobalt, and stainless steel.

[0082] Preferably, the monoMOF catalyst is oriented assembled on a substrate such as carbon or metal by in-situ growth of monoMOF or by chemical modification grafting.

[0083] In a preferred embodiment, the monoMOF catalyst is selected as Hf. 12 -DBB-RhCp*ClmonoMOF, its synthesis process is as follows: where the Chinese name corresponding to DBB-RhCp*Cl is [chlorine-η] 5-pentamethylcyclopentadienyl-(4,4'-bis(4-carboxyphenyl)-2,2'-bipyridine)rhodium(III)]+;

[0084] Step 1, synthesize Me2DBB; the synthesis route is as follows:

[0085]

[0086] The specific synthesis process is as follows:

[0087] 5,5'-dibromo-2,2'-bipyridine (4.0 g, 12.75 mmol) and 4-(methoxycarbonyl)phenylboronic acid (6.88 g, 38.25 mmol) were dissolved in tetrahydrofuran (THF, 103 mL); the system was deoxygenated for 15 minutes.

[0088] Add an aqueous solution (21 mL) containing K2CO3 (8.8 g, 63.67 mmol) and Pd(PPh3)4 catalyst (1.5 g, 1.30 mmol);

[0089] Seal the reaction vessel and stir the reaction at 80°C for 3 days.

[0090] After cooling to room temperature, the solid was collected by filtration; it was washed successively with THF and deionized water to obtain the pure product Me2DBB. The 1H NMR spectrum of Me2DBB is shown below. Figure 2 As shown in the image.

[0091] Step 2, synthesize Me2DBB-RhCp*Cl; the synthetic route is as follows:

[0092]

[0093] The specific synthesis process is as follows:

[0094] Add [RhCp*Cl]2Cl2 (1.0 mmol), Me2DBB (0.85 g, 2.0 mmol), methanol (25 mL), and chloroform (25 mL) to a 200 mL thick-walled reaction tube.

[0095] The reaction tube was degassed with N2, sealed, and heated and stirred at 120°C for 2 days.

[0096] After cooling to room temperature, the solvent was removed under reduced pressure to obtain pure Me2DBB-RhCp*Cl product as an orange solid (1.92 g, 2.0 mmol, 100%).

[0097] If the product is impure, the solid can be sonicated with ethanol and then filtered to remove any remaining impurities. The 1H NMR spectrum of this Me2DBB-RhCp*Cl is as follows: Figure 3 As shown in the image.

[0098] Step 3, synthesize H2DBB-RhCp*Cl; the synthesis route is as follows:

[0099]

[0100] The specific synthesis process is as follows:

[0101] 100 mg Me2DBB-Rh was added to 15 mL of a 1:1 mixture of MeOH and THF. The reaction solution was pale orange-yellow. The ambient temperature during the above process was room temperature, and the reaction time was 6 h.

[0102] Add 5 mL of LiOH aqueous solution to the reaction solution. The amount of LiOH in the aqueous solution is 10 mg. After half an hour of reaction, the reaction solution becomes turbid.

[0103] The reaction was carried out for 4 hours, cooled to room temperature, and monitored by TLC. The reaction was basically complete. The solution was then evaporated to dryness under vacuum to obtain H2DBB-RhCp*Cl.

[0104] 1 H NMR sample dissolved in deuterated DMSO for NMR characterization, as follows: Figure 4 As shown in the image.

[0105] Step 4, synthesize Hf 12 -Rh;

[0106] The specific synthesis process is as follows:

[0107] Add the following to a 5 mL glass bottle: 0.5 mL HfCl4 solution (concentration 2.0 mg / mL, solvent: N,N-dimethylformamide / DMF), 0.5 mL H2DBB-Rh solution (concentration 3.6 mg / mL, solvent: DMF), 2 μL trifluoroacetic acid (TFA), and 5 μL deionized water; H2DBB-Rh in this application is an abbreviation of H2DBB-RhCp*Cl.

[0108] Place the mixture in an 80℃ oven and let it stand for 24 hours to react.

[0109] The resulting orange precipitate Hf was collected by centrifugation. 12 -Rh, the precipitate was washed successively with DMF and ethanol. In this application, Hf 12 -Rh is equivalent to Hf 12 -DBB-RhCp*Cl is an abbreviation for MonoMOF;

[0110] Based on the recovery rate of hafnium, the yield was determined to be 56% by ICP-MS.

[0111] Figure 5 Hf is shown in 12Electron micrograph of -DBB-RhCp*Cl monoMOF (abbreviated as Rh monoMOF).

[0112] In a preferred embodiment, this application achieves the highest close packing of active sites per unit area of ​​the point electrode by assembling molecular catalysts into periodic monoMOF structures. How the monoMOF binds to the electrode is a key issue; commonly used Nafion... TM The mixed drop casting method easily causes coating of active sites, which can easily detach from the electrode. Therefore, this application grows monoMOF in situ on a carbon substrate or metal substrate to achieve directional assembly of the catalyst.

[0113] Figure 6 The image shows a SEM image of H2DBB-RhCp*ClmonoMOF (Rh monoMOF for short) grown in situ on a titanium felt substrate.

[0114] Figure 7 The image shows a SEM image of H2DBB-RhCp*Cl monoMOF grown in situ on a carbon paper substrate.

[0115] In a preferred embodiment, the anode substrate is capable of oxidizing and losing electrons;

[0116] Preferably, the anode substrate is a biomass derivative; more preferably, the anode substrate is 5-hydroxymethylfurfural (HMF), which loses electrons to convert into 2,5-furandicarboxylic acid (FDCA) and is discharged from the outlet of the anode channel 4.

[0117] Preferably, the anode channel 4 is provided with a channel inlet 41 and a channel outlet 42. The anode substrate, 5-hydroxymethylfurfural (HMF), is injected into the anode channel through the channel inlet 41. After losing electrons, the resulting 2,5-furandicarboxylic acid (FDCA) leaves the anode channel 4 through the channel outlet 42.

[0118] More preferably, the industrial wastewater containing cellulose or starch is pretreated before being fed into the anode channel as an anode substrate. The pretreatment can take various forms, such as biological fermentation, hydrolysis, or acid treatment, to convert the cellulose or starch into biomass derivatives.

[0119] In a preferred embodiment, the reactor 7 is a batch fermenter or a microfluidic continuous flow reactor; the appropriate reactor type can be selected according to the production efficiency of the target product.

[0120] Preferably, a filter membrane 8 is provided between the reactor 7 and the peristaltic pump 6. The filter membrane 8 allows electron carriers to pass through and can intercept microorganisms, thereby ensuring that the electron carriers can be recycled.

[0121] In this application, under the action of the peristaltic pump 6, the flow rate in the cathode channel 5 can be arbitrarily adjusted between 1-50 mL / min, thereby ensuring that the supply rate of the electron carrier matches the production rate of the target product, avoiding excessive or insufficient reducing power, and enabling the reaction system to adapt to the production needs of different types of target products.

[0122] In a preferred embodiment, the power source includes a solar panel or a wind turbine, and also includes an energy storage battery. In this application, solar panels or wind turbines can replace fossil fuel-derived electricity, achieving a sustainable energy supply and reducing dependence on traditional carbon sources. By configuring energy storage batteries, such as lithium batteries, the intermittency of renewable energy sources can be addressed, for example, by supplementing power supply at night when there is no wind, ensuring the continuous biosynthesis in the reaction system.

[0123] In a preferred embodiment, this application provides the application of the electrochemically powered microbial synthesis reaction system described above in the production of high-value chemicals.

[0124] The high-value chemicals include fragrances, amino acids, rare sugars, rare sugar alcohols, biodegradable plastic monomers, highly active natural products, and microbial oils.

[0125] Preferably, the fragrance includes vanillin, raspberry ketone, naringin, rose essential oil, artemisia oil, menthol, γ-decanolide (milky aroma), 2-acetylpyrrolidone (popcorn aroma), benzaldehyde (bitter almond aroma), ionone (floral and woody aroma), citronellal (citrus aroma), β-caryophyllene (spicy aroma), and algal β-ionone (marine aroma).

[0126] The amino acids include monosodium glutamate (MSG), L-lysine, L-threonine, L-tryptophan, L-arginine, L-proline, L-cysteine, L-valine, γ-aminobutyric acid (GABA), L-theanine, L-seleno-methylselenocysteine ​​(anti-cancer activity), and ε-polylysine (preservative).

[0127] The rare sugars include D-allulose, D-tagatose, L-ribose, D-allulose, L-xylulose, D-minobiose, L-fucose, D-isomalulose, L-rhamnose, D-mannohepanisose, D-allulose, and L-lysolose.

[0128] The rare sugar alcohols include allitol, erythritol, xylitol, allol, maltitol, sorbitol, lactitol, isomaltitol, mannitol, ribitol, tagatitol, galactitol, and erythritol;

[0129] The biodegradable plastic monomers include lactide, succinic acid, 5-hydroxymethylfurfural (HMF), 1,4-butanediol (BDO), FDCA (2,5-furandicarboxylic acid), lactic acid, adipic acid, 3-hydroxypropionic acid, ε-caprolactone, glycolide, isosorbide, and dimethyl terephthalate.

[0130] The highly active natural products include fucoidan sulfate, rare ginsenoside CK, paclitaxel, astaxanthin, lycopene, steviol glycosides, mogrosides, artemisinin, ginkgolide B, ganoderic acid, huperzine A, and scutellarin.

[0131] The microbial oils include Schizochytrium DHA algal oil, Yersinia lipolytica ARA oil, Rhodotorula glutinis γ-linolenic acid, Chlorella EPA algal oil, M. algae arachidonic acid, Chlorella elliptica astaxanthin oil, Stardew yeast microbial butter, Cryptodinium docosahexaenoic acid, Rhodotorula lycopene lycopene oil, Rhodotorula glutinis β-carotene oil, Microcystis squalene oil, and marine bacterial wax esters.

[0132] The technical effects that the technical solution of this application can produce in practical applications are further described in detail below;

[0133] (1) Break through traditional bottlenecks and achieve efficient and green manufacturing:

[0134] Improved carbon efficiency: CO2 emissions reduced by >30%, with a reduction of 0.3 tons of CO2 emissions for every ton of MSG produced;

[0135] Improved product conversion rate: The conversion rate of the target product is increased by >30%, such as the conversion rate of glutamic acid from 60% to over 80%;

[0136] Enhanced system stability: Stable operation time > 48 hours, compared to 12-24 hours for the traditional system, representing a 100% improvement in stability;

[0137] Complex path adaptation: Supports the production of complex products such as antibiotics and vaccines. For example, penicillin production efficiency can be increased by 40%.

[0138] (2) Reduce costs and improve the feasibility of industrialization:

[0139] Reduced raw material costs: Reduced reliance on sugar and lower renewable energy costs are expected to reduce raw material costs by 20%-30%; MSG raw material costs will decrease from 1.2 yuan / kg to 0.8-0.9 yuan / kg.

[0140] Increased production efficiency: The continuous flow reactor and rate-matched design increase production efficiency by more than 50%; amino acid output per unit time increases from 1 ton / day to 1.5 tons / day.

[0141] Good industrial compatibility: It is compatible with existing fermentation equipment, reducing the transformation cost by 40%-50%; the transformation cost of a certain monosodium glutamate factory was reduced from 20 million yuan to 10-12 million yuan.

[0142] (3) Leading zero-carbon biomanufacturing and driving the green revolution:

[0143] Zero-carbon conversion: using renewable energy to achieve zero-carbon manufacturing from electricity to biomolecules, such as converting solar energy into monosodium glutamate and antibiotics;

[0144] Resource recycling: The anode uses industrial wastewater or biomass derivatives to realize waste resource utilization, and 0.5 tons of target product are generated by treating 1 ton of papermaking wastewater;

[0145] CO2 resource utilization: Combined with CO2 capture technology, industrial waste gas CO2 is converted into biomolecules. For every ton of acetic acid produced, 0.7 tons of CO2 are captured, reducing greenhouse gas emissions.

[0146] Experimental Example

[0147] Platinum sheet was used as counter electrode and silver / silver chloride as reference electrode;

[0148] Electrodes obtained by in-situ growth of the experimental material on carbon paper or titanium felt are directly used as working electrodes; some of the experimental materials used as control materials are directly drop-cast onto carbon paper or titanium felt using the Nafion ink mixing and preparation method.

[0149] Electrolyte: 100 mM Tris-HCl buffer (pH 7.2); Substrate concentration: 1 mM NAD + The entire process was conducted under nitrogen protection. The product NADH was detected using a quantitative ultraviolet (UV) method. The UV standard concentration curve for NADH is shown below. Figure 8 As shown in the image.

[0150] Experimental Example 1

[0151] Performance testing of H2DBB-RhCp*Cl ligands; i.e., the experimental substance is H2DBB-RhCp*Cl.

[0152] Because the ligands have poor solubility, H2DBB-RhCp*Cl was directly drop-cast onto carbon paper using the Nafion ink mixing method to serve as the working electrode.

[0153] The test results obtained are as follows Figure 9 and Figure 10 As shown,

[0154] Figure 9The images show the UV-Vis spectra of the electrolyte before and after testing. Before testing, there was virtually no NADH absorption peak in the electrolyte; after testing, the electrolyte showed a significant NADH absorption intensity. The NADH concentration was obtained from the standard curve, and the average ligand-reduced NAD content was subsequently calculated. + The Faraday efficiency is approximately 50% (e.g., Figure 10 As shown in the figure, this demonstrates the effective catalytic ability of the ligand.

[0155] Experimental Example 2

[0156] Quantitative analysis was performed using ICP-MS. H2DBB-RhCp*Cl (RhDBB-Nafion-C), Rh monoMOF (Rh monoMOF-Nafion-C), and Rh monoMOF (Rh monoMOF-C) grown in situ on carbon paper at the same Rh site were used for NAD measurement. + Restore performance test.

[0157] The test results obtained are as follows Figure 11 and Figure 12 As shown;

[0158] Figure 11 The images show the UV-Vis spectra of the electrolyte before and after testing with different materials. Before electrolyte testing, there was virtually no NADH absorption peak. The NADH concentration was obtained from the standard curve, and subsequently calculated to determine the reduced NAD content of the RhmonoMOF in situ grown on carbon paper. + The Faraday efficiency is approximately 12%, and the reduction of NAD by RhDBB-Nafion-C is... + The Faraday efficiency is approximately 48% (e.g., Figure 12 (As shown). In this application, Rh monoMOF is short for DBB-RhCp*ClMonoMOF.

[0159] Experimental Example 3

[0160] Rh monoMOF grown in situ on titanium felt and titanium felt without catalyst were used for NAD. + Restore performance test.

[0161] The test results obtained are as follows Figure 13 and Figure 14 As shown;

[0162] Figure 13 The images show the UV-Vis spectra of the electrolyte before and after testing with different materials. Before electrolyte testing, there was virtually no NADH absorption peak. The NADH concentration was obtained from the standard curve, and subsequently calculated to determine the reduced NAD content of the RhmonoMOF in situ grown on the carbon felt. +The Faraday efficiency is approximately 38%, and the unsupported titanium felt has virtually no catalytic effect.

[0163] The above experiments show that in-situ growth of monoMOF catalysts exhibits better performance than drop-coating, demonstrating that the loading method affects subsequent catalytic performance. Materials loaded on different substrates exhibit different reaction properties, with current results indicating that metal substrates perform better.

[0164] The present invention has been described above with reference to preferred embodiments; however, these embodiments are merely exemplary and illustrative. Various substitutions and modifications can be made to the present invention based on these embodiments, all of which fall within the scope of protection of the present invention.

Claims

1. A microbial synthesis reaction system based on electrochemical energy supply, characterized in that, The system includes an anode chamber and a cathode chamber separated by a proton exchange membrane (1); an anode (2) is disposed in the anode chamber and a cathode (3) is disposed in the cathode chamber. An anode flow channel (4) is provided in the anode chamber, and an anode substrate is filled in the anode flow channel (4) to provide electrons; Outside the anode chamber and cathode chamber, the anode (2) and cathode (3) are connected in series by a power source to form a circuit; A cathode flow channel (5) is provided in the cathode chamber, and the cathode flow channel (5) is connected to the reactor (7) via a peristaltic pump (6); an electron carrier is provided in the cathode flow channel (5) and the reactor (7), and the cathode (3) is able to continuously generate the electron carrier; Microorganisms are also arranged in the reactor (7), which take up electron carriers through transport proteins and then drive the microorganisms to metabolize and synthesize the target product through electron carriers; The electron carrier is selected from one or a mixture of NADH and NADH artificial analogs; Both the anode (2) and the cathode (3) were obtained by loading a monoMOF catalyst onto a substrate; The anode substrate is 5-hydroxymethylfurfural (HMF).

2. The microbial synthesis reaction system based on electrochemical energy supply according to claim 1, characterized in that, When the electron carrier is NADH, the microorganism oxidizes NADH to NAD during the metabolic synthesis of the target product. + The cathode (3) is continuously based on NAD + Generate NADH.

3. The microbial synthesis reaction system based on electrochemical energy supply according to claim 1, characterized in that, The thickness of the film on the anode (2) and cathode (3) is 2-20 nm; The substrate is selected from any one or a mixture of carbon / graphite substrate, metal substrate, ITO substrate, and silicon substrate; The metal substrate is selected from any one of titanium, gold, silver, copper, nickel, platinum, cobalt, and stainless steel.

4. The microbial synthesis reaction system based on electrochemical energy supply according to claim 3, characterized in that, The directional assembly of monoMOF catalysts can be achieved by in-situ growth or chemical modification of monoMOFs onto carbon or metal substrates.

5. The microbial synthesis reaction system based on electrochemical energy supply according to claim 1, characterized in that, The anode substrate loses electrons and is converted into 2,5-furandicarboxylic acid (FDCA), which is discharged from the outlet of the anode channel (4).

6. The microbial synthesis reaction system based on electrochemical energy supply according to claim 1, characterized in that, The reactor (7) is a batch fermenter or a microfluidic continuous flow reactor; A filter membrane (8) is placed between the reactor (7) and the peristaltic pump (6). The filter membrane (8) allows electron carriers to pass through and is able to intercept microorganisms.

7. The microbial synthesis reaction system based on electrochemical energy supply according to claim 1, characterized in that, The power source includes solar panels or wind turbines, and also includes energy storage batteries.

8. The application of a microbial synthesis reaction system based on electrochemical energy as described in any one of claims 1 to 7 in the production of high-value chemicals, characterized in that, The high-value chemicals include fragrances, amino acids, rare sugars, rare sugar alcohols, biodegradable plastic monomers, highly active natural products, and microbial oils.

9. The application of the electrochemically powered microbial synthesis reaction system according to claim 8 in the production of high-value chemicals, characterized in that, The fragrances include vanillin, raspberry ketone, naringin, rose essential oil, artemisia annua oil, menthol, γ-decanolide, 2-acetylpyrrolidone, benzaldehyde, ionone, citronellol, β-caryophyllene, and algal β-ionone. The amino acids include L-monosodium glutamate, L-lysine, L-threonine, L-tryptophan, L-arginine, L-proline, L-cysteine, L-valine, γ-aminobutyric acid, L-theanine, L-seleno-methylselenocysteine, and ε-polylysine. The rare sugars include D-allulose, D-tagatose, L-ribose, D-allulose, L-xylulose, D-minobiose, L-fucose, D-isomalulose, L-rhamnose, D-mannohepanisose, D-allulose, and L-lysolose. The rare sugar alcohols include allitol, erythritol, xylitol, allol, maltitol, sorbitol, lactitol, isomaltitol, mannitol, ribitol, tagatitol, galactitol, and erythritol; The biodegradable plastic monomers include lactide, succinic acid, 5-hydroxymethylfurfural, 1,4-butanediol, FDCA, lactic acid, adipic acid, 3-hydroxypropionic acid, ε-caprolactone, glycolide, isosorbide, and dimethyl terephthalate. The highly active natural products include fucoidan sulfate, rare ginsenoside CK, paclitaxel, astaxanthin, lycopene, steviol glycosides, mogrosides, artemisinin, ginkgolide B, ganoderic acid, huperzine A, and scutellarin. The microbial oils include Schizochytrium DHA algal oil, Yersinia lipolytica ARA oil, Rhodotorula glutinis γ-linolenic acid, Chlorella EPA algal oil, M. algae arachidonic acid, Chlorella elliptica astaxanthin oil, Stardew yeast microbial butter, Cryptodinium docosahexaenoic acid, Rhodotorula lycopene lycopene oil, Rhodotorula glutinis β-carotene oil, Microcystis squalene oil, and marine bacterial wax esters.

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