Method and system for producing high-purity medium-chain fatty acid based on waste molasses two-stage regulation fermentation

Abstract

The invention discloses a method and a system for producing high-purity medium-chain fatty acid (MCFA) based on waste molasses two-stage regulation fermentation. Comprising the following steps: (1) diluting waste molasses, then adding a compound enzyme agent of pectinase, beta-glucanase and cellulase, and coupling microwave to assist treatment; (2) inoculating the pretreated waste molasses with a composite flora containing a clostridium kluyveri mutant strain and a Megasphaera elsdeni engineering bacterium, and carrying out a fermentation treatment on the waste molasses, wherein the composite flora contains the clostridium kluyveri mutant strain and the Megasphaera elsdeni engineering bacterium; (3) sequentially performing two-stage dynamic regulation and control fermentation on the waste molasses inoculated with the composite flora through a pH-stat control strategy to realize efficient conversion from butyric acid to hexanoic acid / octanoic acid; and (4) a hydrophobic hollow fiber membrane module is adopted to realize on-line removal of the product, and the inhibition effect of the product is relieved. According to the technology, the yield of the MCFA produced from the waste molasses reaches 0.45 g / g COD, the cost is reduced by 68% compared with that of a traditional substrate, and a new way is provided for recycling of sugar manufacturing waste.

Description

Technical Field

[0001] This invention belongs to the field of bioengineering technology, specifically relating to a method and system for producing high-purity medium-chain fatty acids based on two-stage controlled fermentation of waste molasses. Background Technology

[0002] Medium-chain fatty acids (MCFAs) are carboxylic acid compounds with carbon chain lengths of 6 to 12, including hexanoic acid (C6), octanoic acid (C8), and decanoic acid (C10). These compounds have significant applications in the food, pharmaceutical, chemical, and energy sectors due to their unique physicochemical properties. For example, hexanoic acid is a key precursor in the synthesis of fragrances and biodiesel, octanoic acid is widely used in antibacterial drugs and cosmetics, and decanoic acid is used to produce high-performance lubricants. Traditional MCFA production mainly relies on chemical synthesis or vegetable oil processing, but this involves high energy consumption, complex byproducts, and environmental pollution. In recent years, biosynthesis based on microbial fermentation has attracted attention due to its green and sustainable characteristics; however, existing technologies mostly use pure glucose or vegetable oil as substrates, with raw material costs accounting for 60% to 70%, severely restricting its industrial application.

[0003] The sugar industry is a major global agricultural product processing sector, generating approximately 45 million tons of waste molasses annually. Waste molasses is the residual mother liquor after sugar crystallization, with a complex composition including sucrose (35%–45%), fructose (10%–15%), glucose (5%–10%), as well as colloids, pigments, and ash (8%–12%), with a chemical oxygen demand (COD) as high as 90–120 g / L. Currently, about 65% of global waste molasses is used to produce alcohol or animal feed additives, but its economic value is low and it represents a waste of resources. For example, producing one ton of alcohol requires only about 4.5 tons of waste molasses, leaving the remaining organic matter unutilized. In recent years, publications such as Environ. Sci. Ecotechnol. 2025, 26:100568 have proposed technologies for producing hydrogen through fermentation of waste molasses, but the added value of the product is limited, and the fermentation process requires gradually increasing the load to achieve equilibrium, resulting in low production efficiency.

[0004] Current biosynthetic MCFA technologies face four major bottlenecks: First, insufficient substrate pretreatment efficiency. Approximately 30%–40% of the organic matter in waste molasses exists in colloidal and polysaccharide forms, resulting in a carbon source utilization rate of less than 65% during direct fermentation. Conventional acid / alkali pretreatment easily produces inhibitors such as furfural, while single enzymatic hydrolysis is insufficient to completely decompose pectin and dextran. Second, strain tolerance deficiencies. The initial conductivity of waste molasses often exceeds 50 mS / cm, while traditional MCFA-producing strains (such as Clostridium kluyveri) experience a growth rate decrease of >60% when conductivity >30 mS / cm. Furthermore, osmotic stress caused by high sugar concentrations (>80 g / L) can lead to cell membrane damage and reduced metabolic activity. Third, inefficient fermentation process control. Existing technologies often employ a single-stage fermentation model (such as CN110734934A), making it difficult to balance acid production and carbon chain elongation. For example, butyric acid accumulation is dominant (>70%) under pH 5.5–6.0 conditions, but this acidic environment inhibits the activity of carbon chain elongation enzymes (such as acyl-CoA synthase), resulting in a hexanoic acid / octanoic acid conversion rate of less than 20%. Fourth, the product inhibition effect is significant. MCFA has strong surface activity, and concentrations exceeding 5 g / L can damage cell membrane structures, inducing cell autolysis. Existing technologies mostly employ intermittent feeding or end-effector extraction, but the former increases the risk of contamination, while the latter consumes large amounts of organic solvents (such as n-hexane) and has low separation efficiency (recovery rate <70%).

[0005] It is worth noting that although some studies have attempted to increase MCFA yield through genetic engineering of bacterial strains (such as overexpression of thioesterase genes) or optimization of fermentation parameters (such as adjusting the C / N ratio), these improvements are mostly limited to laboratory scale. For example, the literature (Bioresour. Technol. 2018, 247:12531257) reported a yield of only 528 mg / L of hexanoic acid from glucose using recombinant E. coli. Furthermore, existing technologies are not well-suited to the characteristics of waste molasses and lack systematic solutions for high-salt, high-osmotic-pressure environments. Therefore, developing an efficient, low-cost MCFA biosynthesis technology adapted to the characteristics of waste molasses has become a key challenge urgently needing breakthroughs in the field of industrial biotechnology. Summary of the Invention

[0006] The technical problem to be solved by this invention is to overcome the deficiencies and defects mentioned in the background art above, and to propose a method and system for producing high-purity medium-chain fatty acids based on two-stage controlled fermentation of waste molasses, so as to realize the efficient utilization of waste molasses and the production of medium-chain fatty acids.

[0007] To solve the above-mentioned technical problems, the technical solution proposed by this invention is as follows:

[0008] A method and system for producing high-purity medium-chain fatty acids based on two-stage controlled fermentation of waste molasses includes the following steps:

[0009] (a) Pretreatment of waste molasses: The waste molasses is successively diluted and conditioned, subjected to compound enzymatic hydrolysis, and microwave-assisted cell disruption.

[0010] (b) Inoculation with a complex microbial community: The pretreatment solution was inoculated with a complex microbial community containing a mutant strain of Clostridium kluyveri and engineered strains of Megasphaera elsdeni;

[0011] (c) Two-stage pH-controlled fermentation: The inoculated waste molasses is placed in a fermentation tank. The first stage is controlled at pH 5.8-6.2 to accumulate acid. When the butyric acid concentration reaches 8-12 g / L, the second stage is started at pH 7.0-7.5 to promote carbon chain elongation.

[0012] (d) Product separation: The product is removed in real time through a hollow fiber membrane module to maintain the MCFA concentration in the reactor ≤5g / L.

[0013] As a further improvement, the complex enzyme in step (a) comprises pectinase, β-glucanase, and cellulase in a ratio of 3:2:1, with a total enzyme activity ≥1500U / g substrate. The microwave treatment parameters are 800-1000W power with pulse treatment of 3s on / 10s off, and a total treatment time of 4-6min.

[0014] As a further improvement, the complex microbial community in step (b) consists of the Clostridium kluyveri mutant strain with accession number CCTCC CB 20081677 and the Megasphaera elsdenii engineered strain with accession number ATCC 25940 at a viable ratio of (3-4):1, wherein the Clostridium kluyveri strain carries recombinant plasmids that overexpress the phosphotransacetase gene (pta) and the thioesterase gene (tesB).

[0015] As a further improvement, in step (c), the first stage maintains an oxidation-reduction potential (ORP) of -350 to -300 mV, a temperature of 37 ± 0.5 °C, and a stirring rate of 120 to 150 rpm; in the second stage, the ORP is increased to -280 to -250 mV, the temperature is adjusted to 39 ± 0.5 °C, and the stirring rate is reduced to 80 to 100 rpm.

[0016] As a further improvement, the hollow fiber membrane module described in step (d) is made of polytetrafluoroethylene with a pore size of 0.1 to 0.3 μm, a membrane area to fermentation broth volume ratio of 0.8 to 1.2 m² / m³, and a transmembrane pressure difference controlled at 15 to 25 kPa.

[0017] As a further improvement, the membrane module outlet is connected to a product enrichment device, which includes activated carbon and anion exchange resin connected in series. The resin type is Amberlite IRA-900, the operating temperature is 40-45°C, and the flow rate is 2-3 BV / h.

[0018] As a further improvement, the composition of volatile fatty acids is monitored in real time during fermentation. When the molar ratio of hexanoic acid / octanoic acid is detected to be lower than 1.5, a mixed solution of ethanol and acetic acid (volume ratio 2:1) is automatically added at a rate of 0.5 to 1.5 g / L·h.

[0019] As a further improvement, a trace element enhancement solution was added after the pretreatment stage, with the following composition: FeSO4·7H2O 0.2~0.4g / L, CoCl2·6H2O 0.05~0.1g / L, Na2MoO4·2H2O 0.03~0.06g / L, and NiCl2·6H2O 0.01~0.03g / L.

[0020] As a further improvement, an ultrasonic auxiliary module was installed in the fermenter and started during the second stage of fermentation. The ultrasonic frequency was 28kHz, the power density was 15-20W / L, and the action mode was to apply a 2min pulse every 30min.

[0021] This invention also provides a system for two-stage controlled fermentation of waste molasses to produce high-purity medium-chain fatty acids, comprising:

[0022] Pretreatment unit: integrates an enzymatic hydrolysis reaction vessel and a microwave irradiation chamber;

[0023] Fermentation unit: A two-stage fermenter equipped with a pH / ORP control module and an online monitoring probe;

[0024] Membrane separation-product enrichment unit: This unit includes a cross-flow membrane module and a resin adsorption tower.

[0025] Central control system: Equipped with a central control system based on machine learning algorithms, used to adjust operating parameters in real time.

[0026] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0027] (1) A three-stage pretreatment process of "chemical conditioning-compound enzymatic hydrolysis-microwave cell disruption" was developed, which broke through the bottleneck that the traditional pretreatment method is not efficient enough in decomposing complex components (pectin, dextran, colloids) in waste molasses, resulting in a loss rate of fermentable carbon source of more than 35%.

[0028] (2) The Clostridium kluyveri ATCC 8527 mutant strain and Megasphaera elsdenii ATCC 25940 engineered strain were constructed to be directionally resistant to high salinity. The Clostridium kluyveri strain was modified to overexpress the phosphate transacetase gene (pta) and thioesterase gene (tesB) to enhance its survival rate under high salinity conditions.

[0029] (3) Using a two-stage dynamic regulation fermentation process, the environmental parameters are precisely controlled to achieve the temporal switching of metabolic pathways, solving the problem that traditional single-stage fermentation is difficult to balance the acid production and carbon chain elongation process, resulting in an excessively high proportion of butyric acid (>70%).

[0030] (4) A hydrophobic PTFE hollow fiber membrane module was designed to separate products in real time and solve the problem of product inhibition.

[0031] The method and system for producing high-purity medium-chain fatty acids (MCFAs) based on two-stage controlled fermentation of waste molasses, as described in this invention, have the following advantages: First, significantly increased yield. Using this technology to treat sugarcane waste molasses (COD 45 g / L), the total MCFA yield reaches 32.7 g / L within 120 hours, including hexanoic acid (58%), octanoic acid (29%), and decanoic acid (13%), with a product yield of 0.45 g / g COD. Second, outstanding economic and environmental benefits. The COD removal rate is 91.2%, the unit product cost is reduced by 68%, and there is no need to add expensive exogenous inducers or organic solvents. This invention provides a novel technical path for the high-value utilization of sugar industry waste, with significant economic and environmental benefits. Attached Figure Description

[0032] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the accompanying drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0033] Figure 1 This is a process flow diagram of the present invention;

[0034] Figure 2 This is a schematic diagram of the system structure of the present invention;

[0035] Figure 3 This is a flowchart illustrating the construction process of the functional microbial community of the present invention;

[0036] Figure 4 This is a schematic diagram of the membrane module structure of the present invention. Detailed Implementation

[0037] To facilitate understanding of the present invention, the present invention will be described more fully and in detail below with reference to the accompanying drawings and preferred embodiments, but the scope of protection of the present invention is not limited to the following specific embodiments.

[0038] Unless otherwise defined, all technical terms used herein have the same meaning as commonly understood by those skilled in the art. The technical terms used herein are for the purpose of describing particular embodiments only and are not intended to limit the scope of the invention.

[0039] Unless otherwise specified, all raw materials, reagents, instruments and equipment used in this invention can be purchased from the market or prepared by existing methods.

[0040] refer to Figure 1 In some specific embodiments, the method for producing high-purity medium-chain fatty acids from waste molasses through two-stage controlled fermentation according to the present invention includes the following steps:

[0041] (a) Pretreatment of waste molasses:

[0042] Waste molasses was successively diluted and conditioned, subjected to compound enzymatic hydrolysis, and microwave-assisted cell wall disruption.

[0043] Molasses waste is a byproduct of the sugar industry. It is a thick liquid left over after sugar is extracted from sugarcane or sugar beets. It is rich in sugars, mainly sucrose, glucose, and cellulose.

[0044] In some embodiments, waste molasses is diluted to COD = 20-50 g / L.

[0045] In some embodiments, the added compound enzyme is pectinase, β-glucanase, and cellulase in a ratio of 3:2:1, with a total enzyme activity ≥1500U / g substrate.

[0046] In some embodiments, the microwave processing parameters are pulse processing with a power of 800-1000W, which is turned on for 3 seconds and off for 10 seconds, with a total processing time of 4-6 minutes.

[0047] In some embodiments, a trace element enhancement solution is added after the pretreatment stage, the composition of which is: FeSO4·7H2O 0.2~0.4g / L, CoCl2·6H2O 0.05~0.1g / L, Na2MoO4·2H2O 0.03~0.06g / L, NiCl2·6H2O 0.01~0.03g / L.

[0048] In some embodiments, an ultrasonic auxiliary module is installed in the fermenter and activated during the second stage of fermentation. The ultrasonic frequency is 28 kHz, the power density is 15-20 W / L, and the action mode is to apply a 2-minute pulse every 30 minutes.

[0049] Due to the combined enzymatic decomposition by pectinase, β-glucanase, and cellulase, as well as microwave-assisted cell wall disruption, the pretreated waste molasses mainly contains glucose, fructose, and sucrose (accounting for more than 80% of the fermentable carbon source), along with small amounts of organic acids, glycerol, and inorganic ions. Through three-stage pretreatment, non-fermentable polysaccharides / colloids are converted into small-molecule carbon sources, providing highly active substrates for the subsequent two-stage fermentation.

[0050] (b) Inoculation with a complex microbial community:

[0051] The pretreatment solution was inoculated with a complex of bacteria containing a mutant strain of Clostridium kluyveri and an engineered strain of Megasphaera elsdeni.

[0052] In some embodiments, the Clostridium kluyveri mutant strain with accession number ATCC 8527 and the Megasphaera elsdenii engineered strain with accession number ATCC25940 are composed of viable strains at a ratio of (3-4):1, wherein the Clostridium kluyveri strain carries recombinant plasmids that overexpress the phosphotransacetase gene (pta) and the thioesterase gene (tesB).

[0053] Clostridium kluyveri ATCC 8527 was purchased from the American Type Culture Collection (ATCC).

[0054] Megasphaera elsdeni ATCC 25940, purchased from the American Type Culture Collection (ATCC).

[0055] (c) Two-stage pH-controlled fermentation:

[0056] The inoculated waste molasses was placed in a fermentation tank, and the pH was controlled at 5.8-6.2 in the first stage to accumulate acid. When the butyric acid concentration reached 8-12 g / L, the second stage was started at pH 7.0-7.5 to promote carbon chain elongation.

[0057] In some embodiments, the first stage maintains an oxidation-reduction potential (ORP) of -350 to -300 mV, a temperature of 37 ± 0.5 °C, and a stirring rate of 120 to 150 rpm; the second stage increases the ORP to -280 to -250 mV, adjusts the temperature to 39 ± 0.5 °C, and reduces the stirring rate to 80 to 100 rpm.

[0058] (d) Product separation:

[0059] The product is removed in real time using a hollow fiber membrane module, maintaining the MCFA concentration in the reactor at ≤5g / L.

[0060] In some embodiments, the hollow fiber membrane module is made of polytetrafluoroethylene with a pore size of 0.1–0.3 μm, a membrane area to fermentation broth volume ratio of 0.8–1.2 m² / m³, and a transmembrane pressure difference controlled at 15–25 kPa.

[0061] In some embodiments, the membrane module outlet is connected to a product enrichment device, which includes activated carbon and anion exchange resin connected in series. The resin type is Amberlite IRA-900, the operating temperature is 40-45°C, and the flow rate is 2-3 BV / h.

[0062] In some embodiments, the composition of volatile fatty acids is monitored in real time during fermentation. When the molar ratio of hexanoic acid to octanoic acid is detected to be less than 1.5, a mixed solution of ethanol and acetic acid (volume ratio 2:1) is automatically added at a rate of 0.5 to 1.5 g / L·h.

[0063] refer to Figure 2 In some specific embodiments, the system for producing high-purity medium-chain fatty acids through two-stage controlled fermentation of waste molasses according to the present invention includes:

[0064] Pretreatment unit: integrates an enzymatic hydrolysis reaction vessel and a microwave irradiation chamber;

[0065] Fermentation unit: A two-stage fermenter equipped with a pH / ORP control module and an online monitoring probe;

[0066] Membrane separation-product enrichment unit: This unit includes a cross-flow membrane module and a resin adsorption tower.

[0067] Central control system: Equipped with a central control system based on machine learning algorithms, used to adjust operating parameters in real time.

[0068] This invention utilizes a multi-stage pretreatment, two-stage dynamically regulated fermentation, complex microbial inoculation, and PTFE membrane in-situ separation technology to efficiently convert waste molasses into medium-chain fatty acids (MCFA), providing a green and high-value utilization pathway for sugar production waste. The core technology improvements and implementation methods are as follows:

[0069] Example 1: Pretreatment Process Optimization

[0070] Sugarcane molasses (85°Bx, COD 98.6 g / L) was diluted to COD 24.7 g / L at a 1:4 volume ratio, and the pH was adjusted to 4.8. A compound enzyme preparation (pectinase 1200 U / g, β-glucanase 800 U / g, cellulase 400 U / g) was added, and enzymatic hydrolysis was performed at 52℃ for 2.5 h. The solution was then transferred to a microwave reactor and pulsed at 900 W (3 s on / 10 s off, total time 5 min). After treatment, the reducing sugar content was measured to be 68.2 g / L, an increase of 41.3% compared to the untreated group (48.3 g / L), and the furfural content was only 0.12 g / L. The pretreated solution was centrifuged (8000 rpm, 10 min) to remove precipitate before being used for subsequent fermentation.

[0071] Example 2: Two-stage dynamic fermentation

[0072] Pretreated waste molasses (COD 45 g / L) was pumped into a 15 L fermenter and inoculated with a complex of Clostridium kluyveri SZD-M211 and Megasphaera elsdenii GE-7 bacteria (viable count ratio 3:1). First stage: pH 6.0±0.1, temperature 37℃, ORP -320 mV, and stirring speed 130 rpm were maintained. After 24 h, the butyric acid concentration reached 10.8 g / L. Second stage: pH 7.2±0.1, temperature 39℃, ORP -260 mV were maintained, the stirring speed was reduced to 90 rpm, and the PTFE membrane separation system (membrane area 12 m², transmembrane pressure difference 20 kPa) was simultaneously started. By 120 h, the total MCFA concentration reached 31.4 g / L (hexanoic acid 18.2 g / L, octanoic acid 9.1 g / L, decanoic acid 4.1 g / L), and the COD removal rate was 91.5%.

[0073] Example 3: Enrichment of membrane separation coupled products

[0074] Based on Example 2, the MCFA mixture from the membrane module outlet was introduced into activated carbon (temperature 42°C, flow rate 2.5 BV / h) to remove pigments, and then selectively adsorbed with Amberlite IRA-900 resin (flow rate 2 BV / h). The eluent was distilled under reduced pressure to obtain an MCFA mixture with a purity >95%, with a total recovery rate of 96.7%. The residual sugar (1.8 g / L) was recycled back to the fermenter.

[0075] Comparative Example 1: Traditional Single-Stage Fermentation

[0076] Using the same pretreated waste molasses (COD 45 g / L), unmodified wild-type Clostridium kluyveri strain was inoculated, with single-stage control at pH 6.0 and temperature 37℃. After 120 h, the butyric acid concentration reached 22.3 g / L, but the total MCFA yield was only 7.6 g / L (hexanoic acid 4.2 g / L, octanoic acid 2.1 g / L), the COD removal rate was 68.4%, and the fermentation broth showed cell autolysis due to product inhibition (viable cell count decreased by 72%).

[0077] Comparative Example 2: Membrane-free separation system

[0078] The process of Example 2 was repeated, but the membrane separation module was turned off. When the MCFA concentration rose to 14.5 g / L, the cell activity decreased sharply (the specific growth rate dropped from 0.19 h⁻¹ to 0.06 h⁻¹), and the final MCFA yield stagnated at 18.3 g / L, with the fermentation cycle extended to 168 h.

[0079] Table 1 shows a comparison of the effects of each embodiment and the comparative example.

[0080] Table 1 Comparison and analysis of the effects of each embodiment

[0081]

[0082] Note: The yield of Example 3 is the same as that of Example 2. Since they share the same fermentation system, the only difference is in the purity and recovery rate of the product.

[0083] This invention, through Examples 1-3, verifies the synergistic advantages of multi-stage pretreatment, two-stage dynamic control, and membrane separation coupling processes. Compared to Comparative Example 1 (traditional single-stage process), Example 2 shows a 313% increase in MCFA yield and an improved cell activity retention rate of 89.4%; while Comparative Example 2 indicates that the absence of the in-situ separation module leads to product inhibition, reducing yield by 41.7%. Example 3 further demonstrates that combining an activated carbon-resin enrichment system can increase product purity to over 95%, while simultaneously achieving raw material recycling. This technical solution significantly outperforms existing technologies in terms of product yield (0.45 g / g COD), energy consumption (<15 kWh / kg-MCFA), and environmental friendliness (COD removal rate >91%), demonstrating its industrial-scale application value.

[0084] The above are merely preferred embodiments of the present invention and are not intended to limit the present invention in any way. Therefore, any simple modifications, equivalent changes, and alterations made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention should fall within the protection scope of the present invention.

Claims

1. A method for producing medium-chain fatty acids (MCFA) through anaerobic fermentation of waste molasses, characterized in that... Includes the following steps: (a) Pretreatment of waste molasses: The waste molasses is successively diluted and conditioned, subjected to compound enzymatic hydrolysis, and microwave-assisted cell disruption. (b) Inoculation with a complex microbial community: The pretreatment solution was inoculated with a complex microbial community containing a mutant strain of Clostridium kluyveri and engineered strains of Megasphaera elsdeni; (c) Two-stage pH-controlled fermentation: The inoculated waste molasses is placed in a fermentation tank. The first stage is controlled at pH 5.8-6.2 to accumulate acid. When the butyric acid concentration reaches 8-12 g / L, the second stage is started at pH 7.0-7.5 to promote carbon chain elongation. (d) Product separation: The product is removed in real time through a hollow fiber membrane module to maintain the MCFA concentration in the reactor ≤5g / L.

2. The method according to claim 1, characterized in that, The complex enzyme in step (a) comprises pectinase, β-glucanase, and cellulase in a ratio of 3:2:1, with a total enzyme activity ≥1500U / g substrate. The microwave treatment parameters are 800-1000W power with pulse treatment of 3s on / 10s off, and a total treatment time of 4-6min.

3. The method according to claim 1, characterized in that, The complex microbial community described in step (b) consists of the Clostridium kluyveri mutant strain with accession number ATCC8527 and the Megasphaera elsdenii engineered strain with accession number ATCC 25940 at a viable ratio of (3-4):

1. The Clostridium kluyveri strain carries recombinant plasmids that overexpress the phosphotransacetase gene (pta) and the thioesterase gene (tesB).

4. The method according to claim 1, characterized in that, In step (c), the first stage maintains the oxidation-reduction potential (ORP) at -350 to -300 mV, the temperature at 37 ± 0.5 °C, and the stirring speed at 120 to 150 rpm; in the second stage, the ORP is increased to -280 to -250 mV, the temperature is adjusted to 39 ± 0.5 °C, and the stirring speed is reduced to 80 to 100 rpm.

5. The method according to claim 1, characterized in that, The hollow fiber membrane module described in step (d) is made of polytetrafluoroethylene with a pore size of 0.1 to 0.3 μm, a membrane area to fermentation liquid volume ratio of 0.8 to 1.2 m2 / m3, and a transmembrane pressure difference controlled at 15 to 25 kPa.

6. The method according to claim 5, characterized in that, The membrane module outlet is connected to a product enrichment device, which includes activated carbon and anion exchange resin connected in series. The resin type is Amberlite IRA-900, the operating temperature is 40-45℃, and the flow rate is 2-3 BV / h.

7. The method according to claim 1, characterized in that, During fermentation, the composition of volatile fatty acids is monitored in real time. When the molar ratio of hexanoic acid to octanoic acid is detected to be less than 1.5, a mixed solution of ethanol and acetic acid (volume ratio 2:1) is automatically added at a rate of 0.5 to 1.5 g / L·h.

8. The method according to claim 1, characterized in that, After the pretreatment stage, a trace element enhancement solution was added, with the following composition: FeSO4·7H2O 0.2~0.4g / L, CoCl2·6H2O 0.05~0.1g / L, Na2MoO4·2H2O 0.03~0.06g / L, and NiCl2·6H2O 0.01~0.03g / L.

9. The method according to claim 1, characterized in that, An ultrasonic auxiliary module was installed in the fermenter and started during the second stage of fermentation. The ultrasonic frequency was 28kHz, the power density was 15-20W / L, and the action mode was to apply a 2min pulse every 30min.

10. A dedicated fermentation system for implementing the method according to any one of claims 1 to 9, characterized in that, include: Pretreatment unit: integrates an enzymatic hydrolysis reaction vessel and a microwave irradiation chamber; Fermentation unit: A two-stage fermenter equipped with a pH / ORP control module and an online monitoring probe; Membrane separation-product enrichment unit: This unit includes a cross-flow membrane module and a resin adsorption tower. Central control system: Equipped with a central control system based on machine learning algorithms, used to adjust operating parameters in real time.

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