阅读说明：本技术 一种惰性支撑体粘附式发酵方法及其应用 (Adhesive fermentation method for inert support and application thereof ) 是由 张笑然 曹振恒 万鹏 冯润毅 韩慧 黄兴雅 于 2021-08-23 设计创作，主要内容包括：本发明公开了一种惰性支撑体粘附式发酵方法。所公开的方法包括：将惰性支撑颗粒料和半固态培养基混合使得半固态培养基粘附于惰性支撑颗粒料表面；之后在灭菌后的混合料中接种进行发酵,发酵过程中惰性支撑颗粒料不发生降解；发酵完成后,分别回收发酵产物和惰性支撑颗粒料；所述半固态培养基由粘附保水剂、发酵营养物和水配制而成；所述惰性支撑颗粒料中的各惰性支撑颗粒包括颗粒体,该颗粒体上设有至少一个孔,且每个孔的径向尺寸和轴向尺寸均为毫米级；所述孔为通孔或盲孔。本发明的方法可以大幅度降低固态发酵生产中的物料消耗量,减少固体废弃物的产生量,同时可以改善料床的通透性,简化微生物固态培养物的分离过程。(The invention discloses an adhesion type fermentation method for an inert support body. The disclosed method comprises: mixing the inert support particle material with a semi-solid culture medium to enable the semi-solid culture medium to be adhered to the surface of the inert support particle material; then inoculating the sterilized mixture for fermentation, wherein the inert support particles are not degraded in the fermentation process; after fermentation is finished, respectively recovering a fermentation product and inert supporting granules; the semi-solid culture medium is prepared by adhering a water-retaining agent, a fermentation nutrient and water; each inert supporting particle in the inert supporting particle material comprises a particle body, at least one hole is formed in the particle body, and the radial dimension and the axial dimension of each hole are both millimeter-sized; the holes are through holes or blind holes. The method can greatly reduce the material consumption in the solid-state fermentation production, reduce the generation amount of solid wastes, improve the permeability of a material bed and simplify the separation process of the microorganism solid culture.)

1. An inert support adherent fermentation process, comprising:

mixing the inert support particle material with a semi-solid culture medium to enable the semi-solid culture medium to be adhered to the surface of the inert support particle material; then inoculating the sterilized mixture for fermentation, wherein the inert support particles are not degraded in the fermentation process; after fermentation is finished, respectively recovering a fermentation product and inert supporting granules;

the semi-solid culture medium is prepared by adhering a water-retaining agent, a fermentation nutrient and water; the adhesion water-retaining agent enables a semi-solid leavening agent to adhere to the inert support body and ensures that a semi-solid culture does not dry and knot in the fermentation process, and the adhesion water-retaining agent is not degraded by fermentation microorganisms or contains components which are not degraded by the fermentation microorganisms;

each inert supporting particle in the inert supporting particle material comprises a particle body, at least one hole is formed in the particle body, and the radial dimension and the axial dimension of each hole are both millimeter-sized; the holes are through holes or blind holes.

2. The inert support adhesive fermentation process of claim 1, wherein the inert support particles have a particle size of 2-10mm and the holes have a radial dimension of 1-9 mm.

3. The method of claim 1, wherein the inert support particles are folded or rolled from a mesh material into a cylindrical particle body with a hole in the middle.

4. The method according to claim 1, wherein the inert support particles are made of metal, ceramic or rigid polymer.

5. The method of claim 1, wherein 0.001-1 g of the semi-solid medium is attached to each cubic centimeter of the stack of inert support particles.

6. The inert support adhesive fermentation method according to claim 1, wherein the semi-solid medium comprises water 75-95 wt%, adhesive water-retaining agent 0.5-6 wt%, and fermentation nutrient in 100 wt%.

7. The inert support adherent fermentation method of claim 1 or 6, wherein the semi-solid medium has an adherence degree of 1 to 100 g-s.

8. The inert support adherent fermentation method of claim 1 or 6, wherein the semi-solid medium has a hardness of 50 to 300 g.

9. The inert support adhesive fermentation process of claim 1, wherein said adhesive water retention agent is selected from the group consisting of sodium alginate, sodium carboxymethylcellulose, sodium polyacrylate, xanthan gum, guar gum, curdlan, pectin, gellan gum, konjac gum, and locust bean gum.

10. The method of claim 1, wherein the fermentation product and the inert support particles are separated by washing with water and filtering after the fermentation is completed, and then recovered separately.

11. Preparing a filamentous fungus, yeast or bacterium by the method of claim 1.

Technical Field

The invention relates to a microbial fermentation technology, in particular to a method for carrying out solid state fermentation by adhering microbial cells and a fermentation substrate to the surface of an inert support body which can be recycled repeatedly.

Background

In the field of industrial fermentation, the history of solid state fermentation is long, and products mainly comprise traditional fermented food, wine, feed, microbial thalli, enzyme preparations and partial microbial metabolites. Compared with liquid submerged fermentation, solid state fermentation has the advantages of water saving, energy saving, low cost and the like, and has good development prospect. However, in the traditional solid-state fermentation, granular materials such as bran, rice hulls, bagasse, sorghum, bean cakes and the like are generally used as culture mediums, the heat transfer efficiency and the mass transfer efficiency of the solid materials are low, and the material bed is easy to shrink and harden along with the mass degradation and consumption of the solid mediums by microorganisms and the loss of water, so that the spatial distribution of temperature and water content is not uniform. In addition, in addition to some products that facilitate distillation (e.g., white spirits) or washing (e.g., vinegar), separation of the solid substrate from the general solid fermentation product is difficult, and the large amount of solid waste remaining after extraction of the product is liable to cause environmental stress. The above problems become major technical problems limiting the development of solid state fermentation.

In order to solve the above problems in the conventional solid state fermentation field, an adsorption carrier solid state fermentation technology has emerged. The technology uses natural or artificial porous granular inert or semi-inert material as a carrier, uses developed pores inside the carrier to adsorb water and soluble culture medium components, and contains microorganisms to grow and metabolize in the internal pores and on the surfaces of the granules, and the used adsorption carrier is generally porous granules such as bagasse, corncob granules, polyurethane foam blocks, vermiculite and the like. Compared with the traditional solid fermentation, the adsorption carrier solid fermentation technology has the advantage of difficult hardening, and the heat transfer efficiency and the mass transfer efficiency are improved. However, such an adsorption carrier achieves high water binding property by virtue of high porosity, and the carrier such as a polyurethane foam block therein has an inner surface of its pores as a place for growth and metabolism of microorganisms, and thus has the following problems:

(1) in the process of product elution and separation, thalli and metabolite carriers adsorbed in carrier pores are difficult to completely separate due to a capillary phenomenon, so that product loss is caused.

(2) Porous supports are generally not mechanically strong and are susceptible to damage during batch-to-batch cleaning, limiting the useful life of the support particles. Meanwhile, due to the accumulation of residual thalli and metabolites in pores, the porous carrier is difficult to realize long-term recycling, and the waste carrier still causes environmental protection pressure.

(3) Although the porous inert carrier can relieve the problem of hardening of the fermentation material bed, the heat and mass transfer resistance in the pores inside the carrier particles is still large.

Disclosure of Invention

Aiming at the defects or shortcomings of the prior art, the invention provides an inert support adhesion type fermentation method.

To this end, the method provided by the invention comprises the following steps:

mixing the inert support particle material with a semi-solid culture medium to enable the semi-solid culture medium to be adhered to the surface of the inert support particle material; then inoculating the sterilized mixture for fermentation, wherein the inert support particles are not degraded in the fermentation process; after fermentation is finished, respectively recovering a fermentation product and inert supporting granules;

the semi-solid culture medium is prepared by adhering a water-retaining agent, a fermentation nutrient and water; the adhesion water-retaining agent enables the semi-solid fermentation agent to adhere to the inert support body and ensures that the semi-solid fermentation agent does not dry and knot in the fermentation process, and the adhesion water-retaining agent is not degraded by fermentation microorganisms or contains components which are not degraded by the fermentation microorganisms;

each inert supporting particle in the inert supporting particle material comprises a particle body, at least one hole is formed in the particle body, and the radial dimension and the axial dimension of each hole are both millimeter-sized (namely, the dimension can be accurate to millimeter unit); the holes are through holes or blind holes.

Optionally, the inert support particles have a particle size of from 2 to 10mm, preferably from 3 to 8mm, and the radial dimension of the holes is less than the particle size of the granules and is from 1 to 9 mm.

Optionally, the inert support particles are folded or rolled from a mesh material into a cylindrical particle body with a hole in the middle.

Optionally, the inert support particles are made of metal, ceramic or a high molecular polymer which can be made into a rigid body.

Optionally, 0.001-1 g of semi-solid medium is adhered to each cubic centimeter of the stack of inert support particulate material.

Optionally, the weight percentage of water in the semi-solid culture medium is 75% -95%, the weight percentage of the adhering water-retaining agent is 0.5% -6%, and the balance is fermentation nutrients, wherein the weight percentage is calculated by 100%.

Optionally, the semi-solid medium has an adherence of 1 to 100 g.s.

Optionally, the semi-solid medium has a hardness of 50-300 g.

Optionally, the adhesive water-retaining agent is selected from one or more of sodium alginate, sodium carboxymethylcellulose, sodium polyacrylate, xanthan gum, guar gum, curdlan, pectin, gellan gum, konjac gum and locust bean gum.

Optionally, after fermentation, the fermentation product and the inert support particles are separated by washing with water and filtering, and are recovered respectively.

The method of the invention and the use for the preparation of filamentous fungi, yeasts or bacteria. The method of the invention has the following advantages:

(1) the invention utilizes the characteristic that the inert support body is not easy to deform when being extruded, so that the material bed can keep good permeability in the whole fermentation process; the larger surface area and the porosity of the inert support body are utilized to provide sufficient space for the growth and the fermentation of microorganisms; furthermore, by utilizing the characteristic that the inert support body and the culture medium and the culture substance adhered to the surface of the inert support body are easy to separate through water washing, water is added for washing after the fermentation is finished, so that all the inert support bodies are recycled, the repeated cyclic utilization of the inert support bodies is realized, the material consumption in the fermentation process is reduced, the generation amount of solid wastes is reduced, and the extraction difficulty of the fermentation product is also reduced.

(2) The inert support body of the invention does not depend on capillary phenomenon to keep water and nutrient substances, but realizes the solid state fermentation process of microorganisms by supporting the culture medium layer adhered to the surface of the support body, and the support body can be separated from the microorganisms, metabolites and residual culture medium by simple washing after the fermentation is finished, thereby simplifying the separation and extraction process of solid state fermentation.

(3) After the fermentation of the process is finished, the inert support body is easy to separate from the culture medium layer through washing, so that the inert support body can be repeatedly recycled for a long time, the consumption of the fermented materials is reduced, and the production of solid wastes is greatly reduced.

(4) The fermentation material bed has good permeability, and the material bed can not shrink and harden along with the fermentation and the consumption of the culture medium, thereby relieving the problem of uneven macroscopic distribution of the temperature and the water in the material bed.

Drawings

FIG. 1 is a graph showing the effect of example 2 adhesion of water retention agent on water transpiration rate of solid fermentation material;

FIG. 2 is the apparent state of the solid-state fermentation material adhered on the inert support in example 3;

FIG. 3 shows the temperature trend at the center of the material bed in three different Aspergillus niger culture methods of example 6.

Detailed Description

Unless otherwise indicated, the terms or methods herein are understood or implemented using known methods as would be recognized by one of ordinary skill in the relevant art.

The term "inert" as used herein is understood to mean that it does not chemically react with the adhering water retention agent, fermentation nutrients and other additives, does not affect the growth and metabolism of the microorganisms, and can be recycled in the fermentation process. For example, 022Cr17Ni12Mo2 stainless steel is an inert material and has been widely used in the manufacture of fermentors, pharmaceutical equipment, and surgical instruments. The term "support" as used herein is understood to mean a support that supports the semi-solid adherent substance adhered to its surface so that the semi-solid adherent substance also maintains a stable shape without aggregating to form a large mass.

The difference between the support granular material used in the fermentation process of the invention and the carrier used in the existing solid state fermentation process is as follows: on the one hand, in the existing traditional solid state fermentation process, solid medium particles not only play a role of bearing fermenting microorganisms, but also serve as fermentation nutrients, such as but not limited to flaky bran; the material of the support body of the invention is inert. On the other hand, in the existing carrier adsorption solid-state fermentation process, a liquid culture medium is adsorbed on the surface and inside of a carrier through capillary action, and the fermentation mechanism is that a capillary channel on the carrier is utilized to adsorb the liquid culture medium, and meanwhile, a space can be provided for the growth of microorganisms; unlike the prior art, the fermentation mechanism of the present invention is to support the culture substrate adhered to the surface of the support to realize the fermentation process of the microorganism, the size and structure of the support are required to meet the requirement of semi-solid culture medium adhesion or mounting, and from another perspective, the "adhesion or mounting" of the present invention can be understood as "non-capillary adhesion". In an embodiment, the particle size of the single particle body of the support particle material suitable for the present invention is in the millimeter level, and the surface of the single particle body is provided with at least one hole, the radial dimension and the axial dimension of the hole are both in the millimeter level, and the structure of the hole is a through hole or/and a blind hole.

The inert support particles used in the invention are made of stainless steel, titanium alloy, ceramics or high molecular polymers (including but not limited to polyethylene, polypropylene, polycarbonate and polytetrafluoroethylene) which can be made into a rigid body, the surface of the inert support particles can be directly exposed, and a high molecular material coating can be coated to protect the substrate of the support body and improve the suspension capacity of the semi-solid culture medium. The inert support body is granular and made of stainless steel meshes or stainless steel sheets, and has enough rigidity, so that the inert support body cannot be extruded and deformed when being stacked into a material bed, thereby maintaining the permeability of the material bed and being beneficial to relieving the uneven distribution phenomenon of temperature and water content in the material bed.

The semi-solid culture medium is prepared from an adhesive water-retaining agent, a fermentation nutrient and water; the used adhesive water-retaining agent can be one or a combination of more of sodium alginate, sodium carboxymethylcellulose, sodium polyacrylate, xanthan gum, guar gum, curdlan, pectin, gellan gum, konjac glucomannan, locust bean gum, carrageenan and the like, the adhesive water-retaining agent has the functions of increasing the hanging amount of a culture medium on the surface of a support body and delaying the drying of the culture medium in the culture process, and in order to avoid drying in the fermentation process, the selected component of the adhesive water-retaining agent does not participate in fermentation, namely is not degraded by fermentation microorganisms or contains the component which is not degraded by the fermentation microorganisms, so that the adhesive and water-retaining effects are continuously exerted in the fermentation process; the fermentation nutrient is determined according to the specific fermenting microorganism. The fermentation process of the present invention is applicable to all microorganisms that can be grown or produced by solid state fermentation, such as filamentous fungi, yeasts and bacteria. Calculated by 100 percent, the water content of the semi-solid culture medium is about 75 to 95 percent (the water content of the solid culture medium in the prior art is 30 to 75 percent), the weight content of the adhesive water-retaining agent is 0.5 to 6 percent, and the balance is fermentation nutrient.

The semi-solid culture medium of the present invention has different properties in terms of viscosity (or adhesiveness), or/and hardness, in addition to having different components from those of a general liquid culture medium. The viscosity (or apparent viscosity) of a typical liquid medium measured with a rotational viscometer should be 5 to 1000000 mPas, usually 100 to 1000 mPas, and the liquid medium has no hardness; the semisolid culture medium of the present invention has an adhesion degree of 1 to 100 g.s, preferably 5 to 50 g.s, as measured by a food physical analyzer (such as a food physical analyzer of type ta.xt.plus, manufactured by TA corporation, uk); a hardness of 50 to 300g, preferably 100 to 200 g; the adhesion degree unit reflects the adhesive capacity of the semi-solid medium, and the hardness can reflect the compression resistance and deformation resistance of the semi-solid medium. The adhesion and hardness in the following examples were measured by a food physical property analyzer of ta.xt.plus, manufactured by TA corporation, uk.

When the semi-solid culture medium is prepared, the adhesion water-retaining agent, the fermentation nutrient components and water are mixed to form the semi-solid culture medium with higher viscosity, and then the semi-solid culture medium is mixed and stirred with the support body, so that the high-viscosity mixture of the adhesion water-retaining agent and the nutrient components of the culture medium is hung on the inner surface and the outer surface of the support body. In the implementation process of the invention, the adhesion can be carried out firstly and then the sterilization is carried out or the sterilization is carried out firstly and then the adhesion is carried out.

The mixing effect of the inert support particles and the semi-solid medium can be controlled and evaluated in the fermentation process of the invention through the 'hanging amount' defined as the total mass (wet weight) of the semi-solid medium adhered to the surfaces of the inert support particles per unit stacking volume in g cm to ensure the fermentation productivity^-3. In an example, the value range of the carrying capacity is 0.001-1 g-cm^-3. The detection method of the hanging amount used in the following examples is as follows: filtering the mixture of the inert support particle material and the semi-solid culture medium by using a 20-mesh stainless steel net, weighing the mass of the filtrate, namely the mass of the mixture which can not be carried, wherein the difference between the mass of the semi-solid culture medium and the mass of the filtrate is the mass of the carried semi-solid culture medium, and dividing the mass by the stacking volume of the inert support particle material to obtain the carrying capacity:

on the basis of the scheme of the invention, aiming at the microorganism or bacteria to be produced, the technical personnel in the field adopt a conventional experimental method to select the components of a proper semi-solid culture medium, the addition amount of the components, the proper adhesion degree and hardness, and the proper material and size of an inert support body, the carrying capacity and the conventional fermentation process parameters (temperature, pH, nutrient selection and the like) to obtain the ideal yield.

The following are specific embodiments of the present invention to explain the features, aspects and effects of the present invention in detail.

Example 1:

the fermentation nutrient in the embodiment is selected according to aspergillus niger, the specific components are bran, tapioca starch and glucose, the adhesion water retention agent respectively selects carrageenan, sodium polyacrylate and xanthan gum, a plurality of groups of semi-solid fermentation agents are prepared, the adhesion degree of each group of semi-solid fermentation agents and the hanging amount of the semi-solid fermentation agents on the west tower ring packing are tested, and the results are shown in table 1.

TABLE 1

The mixture of the adhering water retention agent, the fermentation nutrient and the water with the formula amount of each group shown in the table 1 is fully mixed with the theta-ring carrier (the diameter of the theta-ring filler particles used in the embodiment is 5mm, and the height of the theta-ring filler particles is 5mm), so that the semi-solid culture medium is adhered to the whole inner and outer surfaces of the support body particles and is filled with the small holes on the metal net;

respectively measuring the hanging amount of the sterilized (high-temperature steam sterilization condition is 121 ℃, and the time is 20min) and the non-sterilized mixed materials of each group of mixed materials;

the result shows that whether the mixture is sterilized by high-temperature steam or not, the mixture can be ensured to be hung on the surface of an inert support after the water-retaining agent is adhered, and the mixture can be lost to different degrees without the water-retaining agent.

Example 2:

this example sets up two sets of semi-solid media and measures the rate of water transpiration during the culture.

A first group: bran powder 1.25g, glucose monohydrate 1.0g, water 25.0g, Sita ring 5mm particle size 23.0 g;

second group: bran powder 1.25g, monohydrate dextrose 1.0g, water 25.0g, Sita ring 23.0g with particle size of 5mm, carrageenan 0.375g, sodium polyacrylate 0.25g, xanthan gum 0.167 g.

Placing the two groups in a triangular flask, sealing with cotton cloth, sterilizing at 121 deg.C for 35min, cooling, placing the flask mouth upward in a 35.0 deg.C incubator, standing, and weighing the total mass of the rest materials at regular time.

The results are shown in fig. 1, when the adhesive water retention agent comprising carrageenan, sodium polyacrylate and xanthan gum is added, the water evaporation rate is obviously lower than that of the group without the adhesive water retention agent, and the adhesive water retention agent can be shown to play the roles of retaining water and reducing the evaporation rate.

Example 3:

this example is an Aspergillus niger M288 (available from Shanghai institute, science and technology Co., Ltd.) produced by the process of the present invention, and sets up multiple sets of fermentation examples, as shown in Table 2.

The specific fermentation process comprises the following steps:

the formula amounts of the adhesion water-retaining agent (wherein carrageenan and sodium polyacrylate are not degraded by aspergillus niger and continuously exert adhesion and water-retaining effects in the fermentation process), the mixture of the fermentation nutrient and water and the theta-ring carrier (the diameter of the theta-ring filler particles used in the embodiment is 5mm, the height of the theta-ring filler particles is 5mm) of each fermentation group shown in table 2 are respectively filled in corresponding containers and fully mixed, so that the semi-solid culture medium is adhered to the whole inner and outer surfaces of the support body particles and is filled with small holes on a metal net;

then, bundling and sealing each fermentation group container with four layers of pure cotton cloth, sterilizing for 35min by saturated steam at 121 ℃, cooling, inoculating into bacterial suspension, and starting fermentation after fully mixing;

the fermentation temperature is 35 ℃, the mixture is kept still and fermented in a constant temperature incubator, the fermentation materials are respectively shaken and mixed in 24h and 48h in the fermentation process, and after the mixture is continuously cultured for 7 days, the aspergillus niger thallus grows on a culture medium layer adhered to the west tower ring carrier in a large quantity, as shown in figure 2. Spore yields were detected and calculated by the following method (same as in the following examples):

(1) adding a culture medium containing 2 g.L^-1Tween-80 and 12 g.L^-1Washing with 100mL of citric acid water by shaking; 5mL of spore suspension obtained by washing was aspirated and washed with a solution containing 2 g.L^-1Tween-80 and 12 g.L^-1Diluting the citric acid solution to 25mL, and detecting the spore concentration by using a blood counting chamber;

(2) calculating total spore yieldYield, total spore yield ═ volume of wash water (100mL) × spore concentration (mL)^-1)；

(3) Spore yield ═ total spore yield/dry weight of total fermentation nutrients in the medium (adhering water retention agents not included), in g^-1。

TABLE 2

The 1-11 fermentation groups shown in table 2 had higher spore yields than the aspergillus niger fermentation process shown in table 2, group 12; it is demonstrated that while the culture medium is adhered to the surface of the support body by using the adhesive water-retaining agent, the amount of the suspended spore can be increased by using a culture medium component having a certain adhesive force, thereby further increasing the spore yield.

Example 4:

this example is an Aspergillus niger produced by the process of the present invention, and sets up a plurality of fermentation examples shown in Table 3, in each of which the theta ring packing used in this example is 5mm in diameter and 5mm in height. The specific fermentation process was the same as in example 3.

TABLE 3

The 1-8 fermentation groups shown in Table 3 had higher spore yields compared to the Aspergillus niger fermentation process shown in group 9 of Table 3, indicating that the presence of an adherent water retention agent is beneficial for improving spore yields.

Example 5:

this example is for Aspergillus niger produced by the process of the present invention, and sets up multiple sets of fermentation examples shown in Table 4, each set using different inert support particles. The specific fermentation conditions were the same as in example 3.

The spore recovery rate of water washing extraction and the solid waste production amount of the whole process of the culture-extraction process refer to that:

washing with 300mL of deionized water for 2 times, 100mL for 1 time and 200mL for 2 times, and removing free water by suction filtration with 20-mesh filter cloth as a filter medium after each washing; then, the cleaned theta-ring is moved away from the filter cloth, a little residue is deposited on the surface of the filter cloth, and the filter cloth is dried to constant weight at 103 ℃ to obtain dry solid residue;

and (4) carrying out suction filtration on the total amount of the spores obtained after the first washing to obtain the percentage of the total amount of the spores obtained by the batch of cultivation, namely the recovery rate of the spores extracted by the first washing.

The spore suspension obtained is washed by water, and the solid residue obtained by filtering the spore suspension by using a 20-mesh filter cloth is the solid waste yield.

The proportion of the decrease in the packed bed volume at the end of fermentation is the amount of decrease in the bed volume at the end of fermentation as a percentage of the bed volume at the beginning of fermentation.

As shown in Table 4, the spore yield was significantly affected by the size of the support body, with the same amount of the water retention agent and the same amount of the culture medium, and the same volume of the inert support body. In addition, besides the theta ring, a ceramic raschig ring, a ceramic cross ring, and a stainless steel rolled ring can also be used as an inert support for spore culture.

As the corncob particles are used as the adsorption carrier, the principle is that the nutrient substances in the suspension state are adsorbed by utilizing the small holes in the particles, so that the nutrient substances in the suspension state can be ensured to be attached to the inner part and the surface of the corncob particles without adding an adhesive water-retaining agent. Compared with the corncob particle carrier culture method in the prior art, the inert support culture method basically avoids the generation of solid waste in the process of washing and extracting spores, because the inert support of the theta ring can be completely recycled and reused, and the corncob particles are difficult to recycle and reuse; moreover, the inert support body is adhered to the surface of the support body, the internal porosity of the corncob particles is relatively high, and the spores which are eluted are easily adsorbed when the spores are extracted by water washing, so that part of the spores are difficult to separate from the corncob particle carrier. Therefore, the inert support is also more efficient in extracting spores after completion of cultivation by water washing than the corncob particles.

The material bed of the traditional bran koji culture method is easy to agglomerate, the phenomenon of internal overtemperature is more serious, and therefore the traditional bran koji culture method is not suitable for supplementing high-efficiency culture media such as glucose, starch and the like.

Example 6:

in this example, the temperature variation trend of the center point of the material bed in the three methods of traditional bran koji culture, corncob particle carrier culture and inert support adhesion culture is compared by comparing the excess temperature range of the material bed in the aspergillus niger fermentation process of the present invention with that of the aspergillus niger spore culture process of the prior art:

500mL triangular flasks are used as culture containers, and the thickness of the culture materials is 5cm at the beginning of fermentation; detecting the central temperature of the material bed at 24h, 48h and 72h after the fermentation is started respectively, wherein the detection point is positioned at the position on the axis of the material bed and away from the top 1/2 in thickness;

the formula (g) of each group of culture materials is as follows:

traditional bran koji culture: flaky wheat bran 54, water 70;

culturing a corncob carrier: 50 parts of corncob particles, 1.6 parts of cassava starch, 6.0 parts of bran powder and 100 parts of water;

the invention discloses an inert support adhesion culture method, which comprises the following steps: the raw materials comprise, by weight, 92 parts of citalopram, 1.5 parts of carrageenan, 1.0 part of sodium polyacrylate, 1.6 parts of xanthan gum, 1.6 parts of cassava starch, 6.0 parts of bran powder and 100 parts of water.

Referring to the results shown in fig. 3, the phenomenon of overheating of the inner temperature of the material bed in the inert support cultivation method according to the present invention is significantly reduced, compared to the conventional bran koji cultivation method and the prior art corncob particle carrier adsorption cultivation method. The reason is that the inert support particles with enough rigidity are easy to maintain the porosity of the material bed not to be reduced along with the growth of microorganisms in the culture process, thereby keeping the permeability of the material bed and ensuring that the interior of the material bed has higher convective heat transfer strength; secondly, the inert support body is made of stainless steel, the heat conductivity coefficient of the inert support body is far higher than that of soaked biomass particles (corncob particles, bran and the like), and high heat conduction strength in the material bed is ensured.

Example 7: the process of the present invention is used in preparing saccharomycete

The embodiment is an example of producing the yeast by adopting the fermentation process, the yeast strain is Saccharomyces cerevisiae CICC1049, and the specific process is as follows:

0.375g of carrageenan, 0.25g of sodium polyacrylate, 0.25g of yeast powder, 0.5g of monohydrate glucose, 0.5g of peptone and 25mL of water are added into a 250mL conical flask and stirred uniformly. Then adding the mixture with the particle size of 5mm and the total stacking volume of 62.5cm³The theta ring packing of (1);

sterilizing, cooling to room temperature, inoculating Saccharomyces cerevisiae, mixing, culturing at 30 deg.C for 3 days to obtain yeast cell with cell yield of 9.05 × 10⁹g^-1(the cell productivity was measured and calculated in the same manner as in the spore productivity in the above examples).

Example 8: preparation of bacteria Using the Process of the invention

This example is an example of producing bacteria using the fermentation process of the present invention, the Escherichia coli used is Escherichia coli CMCC44102, and the specific process is:

0.375g of carrageenan, 0.25g of sodium polyacrylate, 0.125g of yeast powder, 0.125g of NaCl, 0.25g of peptone and 25mL of water are added into a 250mL conical flask and stirred uniformly. Then adding the mixture with the particle size of 5mm and the total stacking volume of 62.5cm³The theta ring packing of (1). Sterilizing, cooling to room temperature, inoculating Bacillus subtilis, mixing, culturing at 35 deg.C for 3 days to obtain Bacillus subtilis cell yield of 4.66 × 10¹⁰g^-1(the cell productivity was measured and calculated in the same manner as in the spore productivity in the above examples).

The above examples adopt specific aspergillus niger, yeast and bacillus subtilis for fermentation, but the present invention is not limited to producing the thallus of these microorganisms, and based on the principle and concept of the present invention, the technical scheme of the present invention can be used for fermentation production of other microorganisms, and the above process parameters can be optimized by combining the growth characteristics of the corresponding microorganisms to obtain the ideal effect, and the technical schemes optimized by the technical scheme of the present invention by adopting routine experiments within the scope of the present invention are within the protection scope of the present invention.