A system for preparing biomass monomers by co-cultivation of microorganisms

Abstract

A system for co-cultivating microorganisms to prepare biomass monomers includes a system using biomass as a substrate and co-culturing biomass-degrading microorganisms with target monomer-generating microorganisms to prepare target monomers through special equipment process control. The system is characterized by: using rumen microorganisms and target product microorganisms to co-cultivate, and by controlling equipment that can separate products in situ, the system achieves symbiosis between rumen microorganisms and target product microorganisms and highly efficient preparation of target products.

Description

Technical Field

[0001] The present invention relates to a biomass monomer preparation system, in particular to a system for preparing biomass monomers by co-culturing microorganisms. Background Art

[0002] In recent years, a variety of methods and systems have emerged for producing biomass monomers using biological methods using non-food biomass as raw materials. Some use enzyme preparations to saccharify the biomass and then integrate it with yeast for fermentation, but the cost of enzyme preparations is too high. Others use aerobic microorganisms and yeast for integrated fermentation to produce bioethanol, but aerobic fermentation yields are relatively low. Existing microbial fermentation methods for producing biomass monomers also have problems such as the need for biomass sterilization, high energy consumption, batch fermentation, the inability to perform continuous fermentation, significant antagonistic effects, and low fermentation efficiency. Summary of the Invention

[0003] The purpose of the invention is to provide a system for the continuous preparation of biomass monomers at low cost and high efficiency by controlling the process of special equipment to achieve co-cultivation of rumen microorganisms and microorganisms that produce material monomers, and to prepare biomass monomers by controlling the equipment that can separate products in situ.

[0004] The system for preparing biomass monomers through co-cultivation of microorganisms described in the present invention includes using biomass as a substrate, and co-culturing biomass-degrading microorganisms with target monomer-generating microorganisms to prepare target monomers through special equipment process control. The co-cultivation of rumen microorganisms with target product microorganisms and the preparation of target monomers through equipment that can separate products in situ are used to control the co-cultivation of rumen microorganisms with target product microorganisms.

[0005] As described above, before collecting rumen microorganisms, live intensive culture is carried out, ruminants are selected, biomass to be used is selected as carbon source, and the experimental animals are fed, and the addition of concentrated feed is gradually reduced. The target monomer microorganisms to be co-cultured are fed, mainly ethanol-producing microorganisms, and the experimental animals are fed for more than one month.

[0006] The microbial purification method includes sucking the rumen contents of ruminants by a negative pressure stomach tube method, anaerobic rolling tube, purifying rumen microorganisms, constructing a small-scale continuous culture and membrane separation device in an anaerobic incubator to amplify the rumen fungi culture, and gradually adding ethanol-producing microbial metabolites to the culture medium after a period of culture to perform co-culture acclimation culture. The purified rumen microorganisms include cellulose-decomposing bacteria such as Fibrobactersuccinogenes, Ruminococcus, Buutyrivibriofibrisolvens, and Clostridium; the fiber-decomposing fungi include Caecomyes monocentric bacteria, single-flagellated zoospores, spherical, branched rhizoids C.communis and C.equi; Cyllamyces multicentric bacteria, multi-flagellated zoospores, spherical, branched spore stalks Cy.Aberensis; Neocallimastix monocentric bacteria, multi-flagellated zoospores, Abundant filamentous rhizodiacs N.frontalis, N.patriciarum, N.hurleyensis, N.variabilis; Piromyces monocentric thalli, single-flagellate zoospore filamentous rhizoid system P.communis, P.maeP.dumbonicus P.rhizinflatus P.minutus P.spiralis Citronii; Orpinomyces multicentric thalli, multi-flagellate zoospore, filamentous rhizoid system O.joyonii, O.intercalaris, O.bovis; Anaeromyces multicentric thalli, single-flagellate zoospore, filamentous rhizoids A.elegans, A.mucronatus, in addition to being obtained by conventional technology, can also be obtained by mail from the China Industrial Microorganism Culture Collection Center.

[0007] The target product strains include anaerobic and facultative anaerobic bacteria such as ethanol-producing bacteria, butanol-producing bacteria, butyric acid-producing bacteria, and amino acid-producing bacteria.

[0008] The target microorganism is preferably an exogenous ethanol-producing bacteria, and the yeast that can utilize pentose and hexose is preferably Candida, ethanol-producing bacteria and mutant yeast that can utilize acetic acid. An ethanol yeast that can simultaneously utilize glucose and xylose is obtained from the China Industrial Microorganism Culture Collection Center: No. 1463 Genus name: tropicalis Genus name: Candida Isolation substrate: Chinese name: Tropical Candida; No. 1771 Species name: tannophilus Genus name: Pachysolen Isolation substrate: Chinese name: Tannophilic Pachysolen; No. 960 Species name: stipitis Genus name: Pichia Isolation substrate: Chinese name: Pichia stipitis; another is ethanol-producing bacteria; the third is a mutant yeast that can utilize acetic acid. In the later stage of amplification culture, rumen microbial culture metabolites are gradually added to acclimate to the rumen microbial culture conditions for co-cultivation, mainly with a culture temperature of 39°C, a weakly alkaline environment, and a low redox potential. The culture medium contains trace elements and nitrogen source elements required for the growth of rumen microorganisms. After startup, the nitrogen source for the growth process is mainly the protein in the straw and the protein produced by cell ablation. The target product strains also include anaerobic and facultative anaerobic bacteria such as butanol-producing bacteria, butyric acid-producing bacteria, and amino acid-producing bacteria.

[0009] The acclimated ethanol-producing bacteria are co-cultivated in a rumen anaerobic continuous culture system. The continuous culture system ensures that the products are separated by a membrane separation device, preventing antagonistic reactions. Rumen fungi, growing on the biomass, produce filamentous rhizomes that cause the biomass to float to the upper portion of the tank, while rumen microbial spores and unfermented straw remain in the lower portion. Due to the characteristics of the rumen fungi, continuous fermentation is achieved within a single tank.

[0010] As mentioned above, yeast is the preferred ethanol-producing bacteria. Years of research on cellulosic ethanol have enabled researchers to domesticate a variety of strains capable of utilizing both xylose and glucose. Yeast's adaptability has been demonstrated in numerous studies, and it can be domesticated according to the culture conditions of rumen fungi. Existing yeast strains cannot reproduce anaerobically, but anaerobic ethanol-producing bacteria do, resulting in equally high ethanol yields and the ability to utilize both xylose and glucose.

[0011] As mentioned above, rumen fungi can also coexist with a large number of competing rumen bacteria in the rumen. However, if acetyl-utilizing yeasts are introduced, the rumen fungi and these yeasts will form a synergistic symbiotic relationship, which is most conducive to the realization of co-cultivation. In addition to the adaptability of the microorganisms in the co-cultivated microecological environment, the continuous membrane separation device can eliminate inhibitory effects, mainly due to the culture temperature of 39°C, the weak alkaline environment, and the low redox potential of -350mV.

[0012] The co-cultivation is carried out as described above. During the co-cultivation cycle, the ethanol-producing bacteria need to be added again. The addition cycle is determined by measuring the increase in sugar concentration. Existing yeast cannot reproduce anaerobically, but anaerobic ethanol-producing bacteria do have the ability to reproduce anaerobically, and the ethanol yield is also high. They can also utilize both xylose and glucose.

[0013] The state of the biomass corresponds to the co-cultivation conditions. When rumen fungi and yeast are co-cultivated, the biomass particles are selected to be larger, ranging from 1 to 10 mm, preferably 2 to 5 mm. When rumen fungi and ethanol-producing bacteria are co-cultivated, the biomass particles are also larger. During the co-cultivation of rumen fungi, rumen bacteria and ethanol-producing fungi, the straw particles are smaller, ranging from 1 to 3 mm. Since the symbiosis inhibits the production of other miscellaneous bacteria, the biomass does not need to be sterilized, which will significantly reduce the energy consumption of ethanol production.

[0014] The fermentation system uses 3 sets, with ethanol yield as the target, and fermentation is carried out in parallel. After 24 hours, the straw that has completed fermentation in the upper part of the tank is removed from the tank, and then continuous fermentation is carried out by adding feed. The 3 tanks are started in different time periods. When a problem occurs in one tank, it can be introduced through other tanks.

[0015] The biomass mentioned above refers to corn straw, rice straw, wheat straw, cotton straw, and hemp stalks. Preferably, the pith and skin of corn straw are fermented separately, and the fermentation residue of the pith is subsequently used as a biological feed raw material, and the fermentation residue of the straw skin is subsequently used as a biological fiber raw material.

[0016] The beneficial effects of the invention are as follows: the present invention provides a method for preparing biomass monomers by co-culturing and fermenting microorganisms, in which the microorganisms can propagate on their own, without using enzyme preparations, and adopting a continuous fermentation device to separate products in situ, thereby improving efficiency. Microorganisms are co-cultured, and the metabolites inhibit the harmful bacteria of the corresponding co-cultured microorganisms. Sterilization and pure culture are not required, thereby reducing energy consumption. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] The present invention will be further described below in conjunction with the accompanying drawings:

[0018] Figure 1 A schematic diagram of a system for co-cultivating microorganisms to produce biomass monomers. DETAILED DESCRIPTION

[0019] In order to better understand the present invention, the technical solutions of the present invention are described in detail below with specific examples, but the present invention is not limited thereto.

[0020] Example 1

[0021] Ruminant goats were selected for biomass utilization, using straw-separated pith as a carbon source. The animals were fed a gradually reduced diet of concentrated feed and fed a co-culture of ethanol-producing microorganisms, Candida species, which can utilize pentoses and hexoses. The animals were fed this diet for over one month. Rumen contents were aspirated using a negative pressure gastric tube, and anaerobic tubes were used to purify the rumen microorganisms. The culture medium was then cultured in an anaerobic incubator. After a period of incubation, the ethanol-producing microorganism metabolites were gradually added to the culture medium to acclimate the co-culture. The culture medium was then expanded using a small-scale continuous culture and membrane separation device. The purified rumen microorganisms, including Caecomyes monocentric thallus, single-flagellated zoospores, and C. communis, which has a spherical, branching rhizoid system, were cultured. Rumen microorganisms were added to the medium-scale anaerobic continuous culture and membrane separation device at a concentration of 3-5% by weight of the biomass. After 8 hours of incubation, the rumen microorganisms were expanded. For the introduction of exogenous ethanol-producing bacteria, yeasts that can utilize pentose and hexose are selected, with Candida being the first choice. An ethanol yeast that can utilize both glucose and xylose was obtained from the China Industrial Microorganism Culture Collection: No. 1463, Genus name: tropicalis, Genus name: Candida, Isolation substrate: Chinese name: Candida tropicalis. In the later stage of amplification culture, rumen microbial culture metabolites are gradually added to acclimate the bacteria towards co-cultivation conditions under rumen microbial culture conditions, mainly with a culture temperature of 38°C, a weakly alkaline environment, and a low redox potential. Acclimated ethanol-producing bacteria were added to the rumen anaerobic continuous culture system at 6% biomass for co-cultivation. The culture medium contained trace elements and nitrogen sources required for rumen microbial growth. After startup, protein from straw and protein produced by cell ablation served as the primary nitrogen source during the growth process. The continuous culture system ensured that metabolic products, such as ethanol, were separated by a membrane separation device without antagonistic interactions. Samples were collected at different time intervals for observation. As the rumen fungi grew on the straw, they produced filamentous rhizomes that caused the straw to float to the upper part of the tank, where they were expelled by the tank mechanism. Rumen microbial spores and unfermented straw remained in the lower part of the tank. A mixture of straw pith and culture medium was continuously added at a constant solid-to-liquid ratio, with a biomass-to-liquid ratio of 1:15. Due to the characteristics of rumen fungi, continuous fermentation was achieved within a single tank. During the co-cultivation cycle, ethanol-producing bacteria were added again. The addition period was determined by measuring the sugar concentration to see if it increased again. In this example, the addition was made after 20 hours. Rumen fungi and yeast are co-cultured, with biomass particles ranging from 3 to 5 mm in diameter. Because the symbiotic growth inhibits the growth of other bacteria, sterilization of the biomass is unnecessary, significantly reducing energy consumption for ethanol production. Three or more fermentation systems are operated in parallel, targeting ethanol yield. After 24 hours, the fermented straw in the upper tank is removed from the tank, and then fed back into the system for continuous fermentation. The three tanks are started in phases, allowing for the introduction of seed from the others if a problem arises in one tank.The fermentation residues of corn stalks and pith are subsequently utilized as biological feed raw materials.

[0022] Example 2

[0023] Ruminant dairy cows were selected for biomass utilization, and straw-separated pith was used as a carbon source. The animals were fed a gradually reduced amount of concentrate feed. Candida species, which can utilize pentoses and hexoses, were also co-cultured with the ethanol-producing microorganisms, for at least one month. Rumen contents were extracted using a negative pressure gastric tube, and anaerobic tubes were used to purify the rumen microorganisms. Ruminococcus bacteria were then cultured in an anaerobic incubator. After a period of incubation, the culture medium was gradually supplemented with metabolites of the ethanol-producing microorganisms to acclimate the culture toward the co-culture direction. The rumen microorganisms were then scaled up using a small-scale continuous culture and membrane separation device to produce the purified Ruminococcus bacteria. Rumen microorganisms are placed in a medium-sized anaerobic continuous culture and membrane separation device at 3-5% of the biomass weight for rumen microorganism amplification culture. For the introduction of exogenous ethanol-producing bacteria, yeasts that can utilize pentose and hexose are selected, with Candida being the first choice. An ethanol yeast that can utilize both glucose and xylose is obtained from the China Industrial Microorganism Culture Collection: No. 1463, Genus name: tropicalis, Genus name: Candida, Isolation substrate: Chinese name: Candida tropicalis. In the later stage of amplification culture, rumen microorganism culture metabolites are gradually added to acclimate to the rumen microorganism culture conditions for co-cultivation, mainly with a culture temperature of 39°C, a weakly alkaline environment, and a low redox potential. Acclimated ethanol-producing bacteria are co-cultivated in a rumen anaerobic continuous culture system at 5% biomass. The culture medium contains trace elements and nitrogen sources necessary for rumen microbial growth. After startup, the primary nitrogen source for growth is protein from straw and protein produced by cell ablation. The continuous culture system ensures that metabolic products such as ethanol are separated by a membrane separation device without antagonistic interactions. Samples are collected at different time intervals for observation. As yeast grow on the straw, the filamentous rhizomes they produce cause the straw to float to the top of the tank, where they are expelled by the tank mechanism. Rumen microbes remain on the unfermented straw leaves in the lower part of the tank. A mixture of straw leaves and culture medium is continuously added at a constant solid-to-liquid ratio, with a biomass-to-culture medium ratio of 1:15. Due to the characteristics of rumen fungi, continuous fermentation within a single tank is possible. Existing yeast strains are not capable of anaerobically propagating. Although rumen fungi can coexist with numerous competing rumen bacteria within the rumen, the introduction of acetic acid-utilizing yeast creates a synergistic symbiotic relationship between the rumen fungi and these yeasts, maximizing the potential for co-cultivation. In addition to the inherent adaptability of microorganisms within the microbiome, continuous membrane separation can eliminate inhibitory effects. During co-cultivation, the existing yeast cannot reproduce anaerobically, necessitating the addition of ethanol-producing bacteria during the co-cultivation cycle. This addition period is determined by measuring the increase in sugar concentration; in this example, it is added again after 20 hours.The biomass's state corresponds to the co-cultivation conditions. During the co-cultivation of rumen bacteria and ethanol-producing fungi, the straw and pith particles are relatively small, ranging in size from 1 to 3 mm. Because the symbiotic relationship suppresses the growth of other bacteria, the biomass does not require sterilization, significantly reducing energy consumption for ethanol production. Three or more fermentation systems are operated in parallel, targeting ethanol yield. After 24 hours, the fermented straw in the upper tank is removed from the tank, and then fed back into the system for continuous fermentation. The three tanks are started in phases, allowing for the introduction of seed from the others if a problem arises in one tank. The residual corn straw and pith fermentation residue, which contains microbial ablation proteins, can be used as biofeed or fertilizer.

[0024] Example 3

[0025] Ruminant cattle were selected for biomass utilization, using straw husks from separated straw as a carbon source. The animals were fed a gradually reduced diet of concentrated feed and fed with ethanol-producing microorganisms that could utilize pentoses and hexoses for co-culture for at least one month. Rumen contents were extracted using a negative pressure gastric tube, and anaerobic tube feeding was performed to purify the rumen microorganisms. The rumen microorganisms were then cultured in an anaerobic incubator. After a period of incubation, the culture medium was gradually supplemented with ethanol-producing microbial metabolites to acclimate the culture toward the co-culture. The rumen microorganisms were then scaled up using a small-scale continuous culture and membrane separation device. The purified rumen microorganisms contained one or more of the following: Neocallimastix monocentric thallus, multiflagellated zoospores, and abundant rhizobacteria: N. frontalis, N. patriciarum, N. hurleyensis, and N. variabilis. Rumen microorganisms are placed in a medium-sized anaerobic continuous culture and membrane separation device at 3-5% of the biomass weight for rumen microorganism amplification culture. For the introduction of exogenous ethanol-producing bacteria, ethanol-producing bacteria that can utilize pentose and hexose are selected. The strains with relevant characteristics can be mailed from local strain collection centers. In the later stage of amplification culture, rumen microorganism culture metabolites are gradually added to acclimate to the rumen microorganism culture conditions for co-cultivation, mainly with a culture temperature of 36-39°C, a weak alkaline environment, and a low redox potential. Acclimated ethanol-producing bacteria were added to the rumen at a biomass ratio of 5% for co-cultivation in an anaerobic continuous culture system. The culture medium contained trace elements and nitrogen sources necessary for rumen microbial growth. During the initial growth phase, protein from straw and protein produced by cell ablation served as the primary nitrogen source. The continuous culture system ensured that metabolic products, such as ethanol, were separated by a membrane separation device without antagonistic interactions. Samples were collected and observed at different time intervals. As the rumen fungi grew on the straw, they produced filamentous rhizomes that caused the straw to float to the top of the tank, where they were expelled by the tank mechanism. Rumen fungal spores and unfermented straw remained in the lower part of the tank. A mixture of straw peels and culture medium with a constant solid-to-liquid ratio was continuously added, with a biomass:culture medium ratio of 1:15. Due to the characteristics of rumen fungi, continuous fermentation was achieved within a single tank. Anaerobic ethanol-producing bacteria possess the ability to reproduce anaerobically, resulting in high ethanol yields and the ability to utilize both xylose and glucose. Rumen fungi and these ethanol-producing bacteria form a synergistic symbiotic relationship, optimal for co-cultivation. In addition to the inherent adaptability of microorganisms within the microbiome, continuous membrane separation devices can eliminate inhibitory effects. Co-cultivation eliminates the need for re-introducing ethanol-producing bacteria, or reduces the amount of ethanol-producing bacteria added during the co-cultivation cycle. The biomass's state corresponds to the co-cultivation conditions, with rumen fungi and ethanol-producing bacteria co-cultivated, using biomass particles with a diameter of 3-5 mm. Because the symbiotic relationship inhibits the growth of other bacteria, the biomass does not require sterilization, significantly reducing energy consumption for ethanol production.During operation, three or more fermentation systems are operated in parallel, targeting ethanol yield. After 24 hours, the fermented straw in the upper tank is removed from the tank, and then fed back into the tank for continuous fermentation. The three tanks are started in phases, so if one tank experiences a problem, the others can be used to introduce new seeds. The remaining corn straw husk fermentation residue is then used as a biofiber feedstock.

[0026] Example 4

[0027] Ruminant goats were selected for biomass utilization, using straw-separated pith as a carbon source. The animals were fed a gradually reduced amount of concentrated feed and co-cultured with ethanol-producing yeasts that utilize acetic acid, a metabolite of rumen microorganisms, for at least one month. Rumen contents were extracted using a negative pressure gastric tube, and anaerobic tubes were used to purify the rumen microorganisms. The microorganisms were then cultured in an anaerobic incubator. After a period of incubation, the culture medium was gradually supplemented with metabolites of the ethanol-producing microorganisms to acclimate the culture toward the co-culture. The microorganisms were then cultured in a small-scale continuous culture and membrane separation device for scale-up. Purified rumen microorganisms included one or more of the following: Anaeromyces multicentrosomes, zoospores with single flagella, A. elegans rhizobium, and A. mucronatus. Rumen microorganisms were placed in a medium-sized anaerobic continuous culture and membrane separation device at 3-5% of the biomass weight for rumen microorganism amplification culture. Exogenous ethanol-producing bacteria were introduced, and mutant yeasts that could utilize acetic acid were selected. In the later stage of amplification culture, rumen microorganism culture metabolites were gradually added to acclimate to the rumen microorganism culture conditions for co-cultivation, mainly with a culture temperature of 39°C, a weakly alkaline environment, and a low redox potential. Acclimated acetic acid-utilizing mutant yeast was added to a rumen anaerobic continuous culture system at 6% biomass for co-cultivation. The culture medium contained trace elements and nitrogen sources required for rumen microbial growth. During the initial growth phase, protein from straw and protein produced by cell ablation served as the primary nitrogen source. The continuous culture system ensured that metabolic products such as ethanol were separated by a membrane separation device, preventing antagonistic interactions. Samples were collected at different time intervals for observation. Rumen fungi, growing on the straw, produced filamentous rhizomes that caused the straw to float to the upper portion of the tank, where they were expelled by the tank mechanism. Rumen microbial spores and unfermented straw remained in the lower portion of the tank. A mixture of straw pith and culture medium with a constant solid-to-liquid ratio was continuously added, with a biomass-to-culture ratio of 1:10. Due to the characteristics of rumen fungi, continuous fermentation was achieved within a single tank. The introduction of acetic acid-utilizing yeast creates a synergistic symbiotic relationship between the rumen fungi and these yeasts, maximizing the potential for co-cultivation. In addition to the inherent adaptability of the microorganisms within the microecology, the continuous membrane separation device eliminates inhibitory effects. During the co-cultivation cycle, ethanol-producing bacteria need to be reintroduced. The reintroduction period is determined by measuring the increase in acetic acid concentration. The existing yeast cannot reproduce anaerobically. The state of the biomass corresponds to the co-cultivation situation. During the co-cultivation of rumen fungi and ethanol-producing fungi, straw and leaf particles are small, ranging from 1 to 3 mm. Because the symbiosis inhibits the production of other bacteria, the biomass does not need to be sterilized, which will significantly reduce the energy consumption of ethanol production.During operation, three or more fermentation systems are operated in parallel, targeting ethanol yield. After 24 hours, the fermented straw in the upper tank is removed from the tank, and then fed back into the tank for continuous fermentation. The three tanks are started in phases. If a problem occurs in one tank, the other tanks can be used to introduce new seeds. The remaining fermentation products of the corn straw and pith are then used as a biofeed raw material.

[0028] Example 5

[0029] Ruminant goats were selected for biomass utilization, using straw-separated pith as a carbon source. The animals were fed a gradually reduced amount of concentrated feed and co-cultured with butyrate-producing bacteria, specifically those that utilize acetic acid, a metabolite of rumen microorganisms. The animals were fed this diet for at least one month. Rumen contents were extracted using a negative pressure gastric tube, and anaerobic tube feeding was performed to purify the rumen microorganisms. The microorganisms were then cultured in an anaerobic incubator. After a period of incubation, the culture medium was gradually supplemented with metabolites of the butyrate-producing microorganisms to acclimate the culture toward the co-culture. The microorganisms were then cultured in a small-scale continuous culture and membrane separation device for scale-up. The purified rumen microorganisms included one or more of the following: Anaeromyces multicentroides, single-flagellated zoospores, A. elegans rhizobacteria, and A. mucronatus. Rumen microorganisms were placed in a medium-sized anaerobic continuous culture and membrane separation device at 3-5% of the biomass weight for rumen microorganism amplification culture. Exogenous butyrate-producing bacteria were introduced and butyrate-producing bacteria were selected. In the later stage of amplification culture, rumen microorganism culture metabolites were gradually added to acclimate to the rumen microorganism culture conditions for co-cultivation, mainly with a culture temperature of 39°C, a weak alkaline environment, and a low redox potential. Acclimated butyrate-producing bacteria were added to the rumen at 6% biomass for co-cultivation in an anaerobic continuous culture system. The culture medium contained trace elements and nitrogen sources necessary for rumen microbial growth. After startup, protein from straw and protein produced by cell ablation served as the primary nitrogen source for growth. The continuous culture system ensured that metabolites, such as butyrate production, were separated by a membrane separation device, preventing antagonistic interactions. Samples were collected at different time intervals for observation. As the rumen fungi grew on the straw, they produced filamentous rhizomes that caused the straw to float to the upper part of the tank, where they were expelled by the tank mechanism. Rumen microbial spores and unfermented straw remained in the lower part of the tank. A mixture of straw pith and culture medium with a constant solid-to-liquid ratio was continuously added, with a biomass-to-culture ratio of 1:10. Due to the characteristics of the rumen fungi, continuous fermentation was achieved within a single tank. The introduction of butyrate-producing bacteria, a registered strain from a national collection, established a symbiotic relationship between the rumen fungi and butyrate-producing bacteria, maximizing the potential for co-cultivation. In addition to the inherent adaptability of the microorganisms in the microbiome, continuous membrane separation ensures the elimination of inhibitory effects. During the co-cultivation cycle, butyrate-producing bacteria need to be reintroduced. The reintroduction period is determined by measuring the increase in butyrate production. The biomass state corresponds to the co-cultivation status. During the co-cultivation of rumen fungi and butyrate-producing bacteria, straw and leaf particles are small, ranging from 1 to 3 mm. Because the symbiosis inhibits the production of other bacteria, the biomass does not require sterilization, significantly reducing the energy consumption of butyrate production. During operation, three or more fermentation systems are operated in parallel, targeting butyrate yield. After 24 hours, the fermented straw in the upper tank is removed from the tank, and then the fermentation is continued with additional feed. The three tanks are started in phases, and if a problem occurs in one tank, the biomass can be reintroduced from the others.The fermentation residues of corn stalks and pith are subsequently utilized as biological feed raw materials.

[0030] Example 6

[0031] Ruminant goats were selected for biomass utilization, using straw-separated pith as a carbon source. The animals were fed a gradually reduced amount of concentrated feed and co-cultured butanol-producing bacteria, a species preserved in a collection center, for at least one month. Rumen contents were extracted using a negative pressure gastric tube, and anaerobic tubes were used to purify the rumen microorganisms. The rumen microorganisms were then cultured in an anaerobic incubator. After a period of incubation, butanol-producing microbial metabolites were gradually added to the culture medium to acclimate the culture toward the co-culture. The rumen microorganisms were then cultured in a small-scale continuous culture and membrane separation device for scale-up. Purified rumen microorganisms included one or more of the following: Anaeromyces multicentrophores, single-flagellated zoospores, A. elegans rhizobacteria, and A. mucronatus. Rumen microorganisms were placed in a medium-sized anaerobic continuous culture and membrane separation device at 3-5% of the biomass weight for rumen microorganism amplification culture. Exogenous butanol-producing bacteria were introduced, and in the later stage of amplification culture, rumen microorganism culture metabolites were gradually added to acclimate to the rumen microorganism culture conditions for co-cultivation, mainly with a culture temperature of 39°C, a weakly alkaline environment, and a low redox potential. Acclimated butanol bacteria were added to the rumen at 6% biomass for co-cultivation in an anaerobic continuous culture system. The culture medium contained trace elements and nitrogen sources necessary for rumen microbial growth. During the initial growth phase, protein from straw and protein produced by cell ablation served as the primary nitrogen source. The continuous culture system ensured that metabolic products such as butanol were separated by a membrane separation device, preventing antagonistic interactions. Samples were collected at different time intervals for observation. Rumen fungi, growing on the straw, produced filamentous rhizomes that caused the straw to float to the upper portion of the tank, where they were expelled by the tank mechanism. Rumen microbial spores and unfermented straw remained in the lower portion of the tank. A mixture of straw leaves and culture medium with a constant solid-to-liquid ratio was continuously added, with a biomass-to-culture ratio of 1:10. Due to the characteristics of rumen fungi, continuous fermentation was achieved within a single tank. The introduction of butanol bacteria created a synergistic symbiotic relationship between the rumen fungi and these yeasts, maximizing the potential for co-cultivation. In addition to the inherent adaptability of the microorganisms within the microecology, the continuous membrane separation device eliminated inhibitory effects. During the co-cultivation cycle, butanol-producing bacteria need to be reintroduced. The reintroduction period is determined by measuring the increase in butanol concentration. The existing butanol bacteria are not yet capable of anaerobic reproduction. The state of the biomass corresponds to the co-cultivation status. During the co-cultivation of rumen fungi and butanol bacteria, the straw and pith particles are small, ranging from 1 to 3 mm. Because the symbiotic growth inhibits the growth of other bacteria, the biomass does not require sterilization, significantly reducing the energy consumption of butanol production. During operation, three or more fermentation systems are operated in parallel, with ethanol yield as the target. After 24 hours, the fermented straw in the upper tank is removed from the tank, and then fed back into the tank for continuous fermentation. The three tanks are started in phases. If a problem occurs in one tank, the other tanks can be used to reintroduce the bacteria. The residual fermentation products of corn straw and pith are subsequently used as a biofeed raw material.

[0032] Example 7

[0033] Ruminant goats were selected for biomass utilization, using straw-separated pith as a carbon source. The animals were fed a diet supplemented with concentrated feed, gradually reducing the amount of concentrate added. The animals were then fed a co-culture of amino acid bacteria for at least one month. Rumen contents were extracted using a negative pressure gastric tube, and anaerobic tube feeding was performed to purify the goat rumen microorganisms. The microorganisms were then cultured in an anaerobic incubator. After a period of incubation, ethanol-producing microbial metabolites were gradually added to the culture medium to acclimate the culture toward the co-culture. The microorganisms were then cultured in a small-scale continuous culture and membrane separation device for scale-up. The purified rumen microorganisms included one or more of the following: Anaeromyces multicentrum, single-flagellated zoospores, A. elegans rhizobium, and A. mucronatus. Rumen microorganisms were placed in a medium-sized anaerobic continuous culture and membrane separation device at 3-5% of the biomass weight for rumen microorganism amplification culture. Exogenous amino acid-producing bacteria were introduced, and in the later stage of amplification culture, rumen microorganism culture metabolites were gradually added to acclimate to the rumen microorganism culture conditions for co-cultivation, mainly with a culture temperature of 39°C, a weakly alkaline environment, and a low redox potential. Acclimated amino acid-producing bacteria were added to the rumen at 6% biomass for co-cultivation in an anaerobic continuous culture system. The culture medium contained trace elements and nitrogen sources necessary for rumen microbial growth. During the initial growth phase, protein from straw and protein produced by cell ablation served as the primary nitrogen source. The continuous culture system ensured that metabolic products such as ethanol were separated by a membrane separation device, preventing antagonistic interactions. Samples were collected at different time intervals for observation. As the rumen fungi grew on the straw, they produced filamentous rhizomes that caused the straw to float to the upper part of the tank, where they were expelled by the tank mechanism. Rumen microbial spores and unfermented straw remained in the lower part of the tank. A mixture of straw pith and culture medium with a constant solid-to-liquid ratio was continuously added, with a biomass-to-culture ratio of 1:10. Due to the characteristics of rumen fungi, continuous fermentation was achieved within a single tank. The introduction of amino acid-producing bacteria created a synergistic symbiotic relationship between the rumen fungi and the amino acid-producing bacteria, maximizing the potential for co-cultivation. In addition to the inherent adaptability of the microorganisms within the microecology, the continuous membrane separation device eliminated inhibitory effects. During the co-cultivation cycle, amino acid-producing bacteria need to be reintroduced. The reintroduction period is determined by measuring the increase in amino acid concentration. The existing amino acid-producing bacteria are not yet capable of anaerobically reproducing. The biomass state corresponds to the co-cultivation status. During the co-cultivation of rumen fungi and amino acid-producing bacteria, the straw and pith particles are small, ranging from 1 to 3 mm. Because the symbiosis inhibits the growth of other bacteria, the biomass does not require sterilization, significantly reducing the energy consumption of ethanol production. During operation, three or more fermentation systems are operated in parallel, targeting amino acid yield. After 24 hours, the fermented straw in the upper tank is removed from the tank, and then fed back into the tank for continuous fermentation. The three tanks are started in phases. If a problem occurs in one tank, the other tanks can be reintroduced. The residual fermentation products of corn straw and pith are subsequently used as a biofeed raw material.

Claims

1. A system for producing biomass monomers by co-cultivation of microorganisms, comprising an anaerobic incubator, a small-scale continuous culture and membrane separation device, a rumen anaerobic continuous culture system, and a membrane separation device. The preparation steps are as follows: (a) Before collecting rumen microorganisms, conduct in vivo intensive culture, select ruminants, select the biomass to be utilized as a carbon source, feed the experimental animals, gradually reduce the addition of concentrated feed, feed the microorganisms to be co-cultured to prepare monomers, and feed the experimental animals for at least one month; (b) using a negative pressure gastric tube method to aspirate rumen contents, anaerobic tube rolling, purifying rumen microorganisms, constructing a small-scale continuous culture and membrane separation device in an anaerobic incubator to amplify rumen fungi culture, and after a period of culture, gradually adding target product microbial metabolites to the culture medium for co-cultivation acclimation culture, and the purified rumen microorganisms are fiber-decomposing bacteria, and the fiber-decomposing bacteria are selected from the following fungi: Caecomyes monocentric thallus, uniflagellated zoospores, spherical, branched rhizoid system C.communis , C.equi; Cyllamyces multicentric thallus, multiflagellate zoospores, spherical, branched sporangium Cy.Aberensis; Neocallimastix has a monocentric thallus, multi-flagellated zoospores, and abundant filamentous rhizoids, such as N.frontalis, N.patriciarum, N.hurleyensis, and N.variabilis. Piromyces has a monocentric thallus, single-flagellated zoospores, and a filamentous rhizoid system, such as P.communis and P.mae, P.dumbonicus, P.rhizinflatus, P.minutus, and P.spiralis Citronii. Orpinomyces multicentric thallus, multiflagellate zoospores, filamentous rhizoid system O. joyonii, O. intercalaris, O. bovis; Anaeromyces multicentric bacteria, single flagellated zoospores, and thread-like rhizodendrocytes A.elegans, A. mucronatus; the rumen microorganism used in the co-culture was the isolated single cellulolytic bacterium described above; (c) introduction of an exogenous target product bacteria, wherein the target product bacteria is Candida, which is an ethanol yeast that can utilize both glucose and xylose and is obtained from the China Industrial Microorganism Culture Collection Center: No. 1463, Species Name: tropicalis, Genus Name: Candida, Isolate Substance: Chinese Name: Candida tropicalis; (d) Gradually add rumen microbial culture metabolites in the late stage of amplification culture to acclimate to the rumen microbial culture conditions for co-cultivation. The main conditions are a culture temperature of 39°C, a weakly alkaline environment, and a low redox potential. The culture medium contains trace elements and nitrogen sources required for rumen microbial growth. After startup, the main nitrogen sources for the growth process are protein in straw and protein produced by cell ablation. (e) adding the acclimated target product bacteria to the rumen anaerobic continuous culture system for co-cultivation. The continuous culture system ensures that the rumen microbial metabolites and the target microbial products are separated by the membrane separation device without antagonism, thereby achieving high-efficiency straw degradation by the rumen microbial extracellular enzymes. The equipment releases monosaccharides or disaccharides produced by the extracellular enzymes during biomass degradation that have not yet been utilized by the rumen microorganisms for utilization by the target product microorganisms. (f) Rumen fungi grow on the straw and produce filamentous rhizomes, which cause the biomass to float to the upper part of the tank and be removed from the system by the mechanism within the fermentation equipment. Rumen microbial spores and newly added unfermented straw remain in the lower part of the tank. Due to the characteristics of rumen fungi, continuous fermentation and symbiotic culture are achieved in one tank. (g) Three fermentation systems are used, with the target product as the target, and fermentation is carried out in parallel. After 24 hours, the straw that has been fermented in the upper part of the tank is removed from the tank, and then the feed is added for continuous fermentation. The three tanks are started in different time periods. If a problem occurs in one tank, the seeds are introduced through the other tanks.

2. The system for preparing biomass monomers by co-cultivation of microorganisms according to claim 1, characterized in that: The state of the biomass corresponds to the co-cultivation situation. Rumen fungi and yeast are co-cultivated, and the biomass particles are larger, 2 to 10 mm. Rumen fungi have hyphae that enter the biomass, and the biomass does not need to be ground into powder without consuming a lot of energy.

3. The system for preparing biomass monomers by co-cultivation of microorganisms according to claim 1, characterized in that: The biomass used is a variety of agricultural residues, namely corn straw, rice straw, wheat straw, cotton straw, and hemp straw.

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