JP6738626B2 - Bioreactor and bioreaction method using the bioreactor - Google Patents

Description

The present invention relates to a bioreactor for producing (fermenting/brewing) reaction products by microorganisms or cells (hereinafter referred to as “microorganisms”), or for proliferating or concentrating (culturing) microorganisms and the like, and a bioreactor for this. Regarding the biological reaction method using, the biological culture liquid extracted from the culture tank and supplied to the filter is referred to as a micro-nano bubble (hereinafter, “micro-nano bubble” is “MNB”, “micro bubble” is “MB”, “nano bubble”). Is sometimes referred to as “NB”), and a biological culture solution obtained by filtering the biological culture solution, collecting the filtrate, and removing the filtrate (hereinafter, “NB”). It is referred to as a "concentrated liquid of microorganisms") which is refluxed to a culture tank.

Unlike a chemical reaction, a biological reaction is a slow reaction, but since it does not use a lot of energy or many chemical substances, it is a mild and meaningful reaction for the environment.
However, the biological reaction has a problem that the reaction is generally mild and slow. That is, in many cases, a reaction within 1 hour is sufficient for a chemical reaction, whereas in the case of a biological reaction, a reaction time of several hours to several days in the case of a long reaction or several weeks or more in the case of a long reaction In some cases. Therefore, it is required to carry out biological reactions efficiently and economically.

As a method for carrying out a biological reaction (fermentation/brewing, culture) by microorganisms, (1) batch method (Batch method) and fed-batch method (Fed-Batch method) and (2) continuous method are generally used. Specifically, a continuous method as disclosed in Patent Document 1 is employed, which can stably maintain high yield and high productivity for a long time.

In Patent Document 1, as shown in FIG. 10, in a continuous fermentation method, a fermentation culture liquid in the apparatus is circulated between a fermentation reaction tank 101 and a membrane separation tank 112 by a fermentation culture liquid circulation pump 111 to separate a membrane. The element 102 filters the microorganisms and cultured cells with a separation membrane, collects the product from the filtrate, and at the same time, the filtered microorganisms and cultured cells are returned to the fermentation medium to increase the concentration of microorganisms and cultured cells in the fermentation medium. It is described that high material productivity can be obtained by maintaining.

Further, in Patent Document 2, in order to make a biological reaction in culture of a microorganism and the like efficient, a culture solution contains MNB or NB formed from air, thereby promoting activation of the microorganism and the like. It is disclosed that the reaction efficiency of the reaction, the reduction of the reaction time, and the like are aimed at.

Specifically, as shown in FIG. 11, the culture solution is extracted from the culture tank 207 and filtered with a bacterial cell filter 210 to obtain a filtrate, which is then stored in the MNB generation tank 215 and the MNB generator 216. It describes a method of generating and mixing MNB of air and refluxing it to the culture tank 207.

However, in the method described in Patent Document 1 described above, in which the biological culture solution is extracted from the culture tank and the concentrated liquid such as microorganisms is returned to the culture tank, the concentration of the microorganisms and the like in the biological culture solution in the culture tank is maintained at a high level. However, since a biological culture solution having a high concentration of microorganisms and the like is filtered, the filter membrane is likely to be clogged, and it is necessary to frequently wash or replace the filter membrane. Further, the clogging of the filtration membrane can be suppressed by increasing the circulation flow rate in the membrane, but if the circulation flow rate in the membrane is increased, stress and damage to microorganisms and the like will increase.

In addition, since a biological culture solution having a high concentration of microorganisms and the like is circulated outside the culture tank, oxygen necessary for respiration of the microorganisms and the like is not sufficiently supplied in this route, and the microorganisms and the like suffer stress and damage.

Further, in the method described in Patent Document 2 in which the biological culture solution is extracted from the culture tank and air MNB is contained in the filtrate, the biological culture solution is filtered in comparison with the amount of the biological culture solution extracted from the culture tank. Since the amount of the filtrate obtained by this method is quite small (the amount of the filtrate is usually about 1/10 to 1/100 of the amount of the biological culture solution extracted from the culture tank), the MNB containing the filtrate is used. However, it is difficult to maintain a high amount of MNB contained in the biological culture solution in the culture tank. Further, in order to increase the amount of the filtrate, if the amount of the biological culture solution to be extracted from the culture tank is increased or the filtration pressure is increased, stress and damage to the microorganisms and the like will be increased.

Further, although the technical field is different from the biological reaction such as fermentation, brewing, and culturing, in Patent Document 3, in order to suppress clogging of the membrane module in the water purification system, the water before being supplied to the membrane module is used. It is disclosed that ultrafine bubbles are generated and water containing the ultrafine bubbles is supplied to the membrane module.

Specifically, as shown in FIG. 12, raw water supplied from the raw water supply line 304 and/or concentrated circulating water from the membrane module 311 supplied from the concentrated circulating water line 317 is pressurized by the water supply pump 306. After that, a water purification method is described in which ultrafine bubbles are generated in water and water containing the ultrafine bubbles is supplied to the membrane module 311 to perform membrane filtration.

However, in the water purification method as described in Patent Document 3 above, a filtration membrane such as a membrane module simply removes suspended substances, bacteria, etc. from raw water such as river water, and filtered. Unlike the biological reaction in which microorganisms are returned to the culture tank and reused, it is not necessary to consider stress and damage to the microorganisms at all.

Furthermore, the "ultrafine bubble" used in Patent Document 3 has a bubble diameter of about 2 to 50 μm, which corresponds to MB and does not include NB which is an extremely fine bubble having a diameter of 100 nm or less. is there.

Japanese Patent No. 5092487 Japanese Patent No. 4146476 JP, 2011-83764, A

The subject of the bioreactor of the present invention and the bioreaction method using this bioreactor is to maintain a high amount of MNB contained in the bioculture liquid in the culture tank and suppress clogging of the filtration membrane. The purpose of the present invention is to reduce stress and damage to microorganisms and the like, thereby efficiently and economically performing biological reaction using microorganisms and the like and separation of microorganisms and the like.

In order to solve the above problems, in the bioreactor of the present invention and the bioreaction method using this bioreactor, the biological culture solution extracted from the culture tank and supplied to the filter, containing micro-nano bubbles. And a micro-nano bubble generator for

In addition, a means other than the above is also used as a means for containing MNB in the biological culture medium, MNB formed from air having an increased oxygen concentration is used as MNB, and a volume is used as a pump for conveying a liquid containing microorganisms and the like. The above problem can be further solved by using a formula pump and adding a pH adjuster to the filtrate that is refluxed in the culture tank.

In the present invention, the amount of MNB contained in the biological culture solution in the culture tank can be maintained at a high level, clogging of the filtration membrane can be suppressed, and further stress/damage that microorganisms and the like receive can be reduced. It is possible to efficiently and economically separate the used biological reaction and microorganisms.

It is a schematic diagram which shows the reference embodiment 1 of the biological reactor of this invention. It is a schematic diagram which shows the reference Embodiment 2 of the biological reactor of this invention. It is a schematic diagram which shows the reference embodiment 3 of the biological reaction apparatus of this invention. It is a schematic diagram which shows the reference embodiment 4 of the biological reaction apparatus of this invention. It is a schematic diagram which shows 1st Embodiment of the biological reaction apparatus of this invention. It is a schematic diagram which shows 2nd Embodiment of the biological reaction apparatus of this invention. It is a schematic diagram which shows 3rd Embodiment of the biological reaction apparatus of this invention. It is a schematic diagram which shows the reference embodiment 5 of the biological reaction apparatus of this invention. It is a schematic diagram which shows 4th Embodiment of the bioreactor of this invention. FIG. 1 of Patent Document 1 (Japanese Patent No. 5092487), which is a conventional example. FIG. 1 of Patent Document 2 (Japanese Patent No. 4146476), which is a conventional example. FIG. 1 of Patent Document 3 (Japanese Patent Laid-Open No. 2011-83764), which is a conventional example.

Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings, but the present invention is not limited thereto.

First, general items of the biological reaction apparatus and biological reaction method of the present invention will be described.
INDUSTRIAL APPLICABILITY The bioreactor of the present invention can be used for both a biological reaction (fermentation/brewing) for the purpose of producing a reaction product by a microorganism and a biological reaction for the purpose of growing or concentrating (culturing) the microorganism. it can. Further, the bioreactor of the present invention can be used in any of the batch method (Batch method), the fed-batch method (Fed-Batch method), and the continuous method. It can be preferably used in a continuous method capable of maintaining the property.

In a biological reaction for the purpose of producing a reaction product by a microorganism or the like, the reaction product is continuously collected together with a filtrate, and in a biological reaction for the purpose of proliferating or concentrating the microorganism, etc. The microorganisms and the like are collected by growing and concentrating.

The biological reaction of the present invention includes not only the production of foods, drugs, chemicals, etc. by brewing, fermentation, etc., the production of reaction products by microorganisms such as the production of bioethanol using biomass, but also the growth or concentration of microorganisms etc. It is also a useful thing that can be applied to.

In the biological reaction device of the present invention and the biological reaction method using this biological reaction device, a filter is used to separate the biological culture liquid extracted from the culture tank into a filtered liquid and a concentrated liquid such as a microorganism. Cross-flow filtration using a hollow fiber membrane module can be preferably used.

The “micro/nano bubble” of the present invention means “micro bubble” and/or “nano bubble”. "Normal bubbles" rise rapidly in water and burst and disappear at the surface, whereas micro bubbles with a diameter of 50 μm or less called "micro bubbles" shrink and disappear in water. With the free radicals, "nano bubbles", which are extremely minute bubbles having a diameter of 100 nm or less, are generated, and these "nano bubbles" remain in water for a certain period of time.

In the present invention, bubbles having a number average diameter of 100 μm or less are called “micro bubbles”, and bubbles having a number average diameter of 1 μm or less are called “nano bubbles”. As a method for measuring the bubble diameter of microbubbles, an image analysis method, a laser diffraction scattering method, an electrical detection zone method, a resonance mass measurement method, an optical fiber probe method, etc. are generally used, and a method for measuring the bubble diameter of nanobubbles is used. As the method, a dynamic light scattering method, a Brownian motion tracking method, an electric detection zone method, a resonance type mass measurement method and the like are generally used.

"Nano bubbles", which are extremely small bubbles, are also called "ultra fine bubbles". In addition, at present, ISO (International Organization for Standardization) is considering making an international standard for fine bubble technology, and if the international standard is made, the commonly used name “nano bubble” is now used. There is a possibility that it will be unified into "Ultra Fine Bubble".

As the micro-nano bubble generator, a known or commercially available device can be used. For example, after dissolving a sufficient amount of gas in water at a certain high pressure, the pressure is released to dissolve the dissolved gas. "Pressurized dissolution type micro bubble generator" that creates supersaturation condition, using a phenomenon that a vortex is generated by causing a water flow, a large bubble is entrained in the vortex, and when the vortex is collapsed, the bubble is fragmented into pieces The “gas-liquid two-phase flow swirl type micro bubble generator” described above can be used.

In addition, examples of the nanobubble generator include those disclosed in JP2007-321690A, JP2006-289183A, JP2005-245817A, JP2007-136255A, and JP2009-39600A. What was described etc. can be used.

When a micro-nano bubble generator that is driven by a water flow (nozzle method) is used, a large amount of MNB can be economically generated, stress and damage given to microorganisms can be reduced, and clogging can be suppressed. preferable.

The biological reaction of the present invention is to cause a reaction product to be produced in a microorganism or the like, or to proliferate or concentrate the microorganism or the like in a biological culture solution containing the microorganism or the like contained in a culture tank.
As a nutrient source in the biological culture solution, one containing a saccharide and a nitrogen source is used. As the saccharide, usually, saccharides such as maltose, sucrose, glucose, fructose and a mixture thereof are used, and the concentration of the saccharide in the biological culture solution is not particularly limited, but is set to 0.1 to 10 w/v%. Is preferred. As the nitrogen source, ammonium chloride, ammonium sulfate, corn steep liquor, yeast extract, meat extract, peptone or the like is used, and it is preferably set to 0.1 to 10 w/v %. Furthermore, in addition to sugars and nitrogen sources, vitamins, inorganic salts, etc. are preferably added to the biological culture solution, if necessary.

As the microorganism in the present invention, brewing, conventionally used in the technical field such as fermentation, koji molds such as Aspergillus, natto, acetic acid bacteria, yeasts, aerobic and facultative anaerobic microorganisms such as lactic acid bacteria Various aerobic and facultative anaerobic microorganisms created by gene recombination technology can be used. Examples of the cells include animal cells for producing physiologically active peptides or proteins used as antibody drugs, especially genetically modified animal cells.

The concentration of the microorganism or cell added to the biological culture solution is not particularly limited, but is preferably 0.5 to 10.0 g/L, more preferably 3.0 to 6.0 g/L.

Next, the features of the biological reaction apparatus and biological reaction method of the present invention will be described.
The first feature of the present invention is, as described above, that the biological culture solution extracted from the culture tank and supplied to the filter is provided with the MNB generator.

In the biological reaction in which the reaction product is produced by a microorganism or the like (fermentation/brewing), or the microorganism or the like is proliferated or concentrated (cultured),
(1) The biological culture solution extracted from a culture tank containing a biological culture solution containing a medium or reaction raw materials, microorganisms, etc. and supplied to a filter is assumed to contain MNB,
(2) The biological culture solution containing this MNB is separated into a filtered solution and a concentrated solution of microorganisms by a filter,
(3) By collecting the filtrate and refluxing the concentrated liquid of microorganisms and the like into the culture tank,
Surprisingly, the amount of MNB contained in the biological culture solution in the culture tank can be maintained at a high level, clogging of the filtration membrane can be suppressed, and the stress and damage to the microorganisms can be reduced. The present invention was accomplished by finding that a biological reaction using can be carried out efficiently and economically.

As described above, in the continuous fermentation method described in Patent Document 1, in which the biological culture solution is extracted from the culture tank and the concentrated solution of microorganisms is returned to the culture tank, the biological culture solution having a high concentration of microorganisms and the like is used. Since it is circulated outside the culture tank for filtration, clogging of the filtration membrane is likely to occur, and there is a problem that oxygen is not sufficiently supplied to microorganisms and the like in the circulation route, but like the present invention, it is extracted from the culture tank. The biological culture solution supplied to the filter is assumed to contain MNB, and the biological culture solution containing this MNB is supplied to the filter to separate the filtrate and the concentrated liquid such as microorganisms and collect the filtrate. At the same time, by refluxing the concentrated liquid such as microorganisms to the culture tank, such a problem can be solved.

That is, in the present invention, by filtering the biological culture solution containing MNB with a filter, MNB is present between the filtration membrane and the substances (microorganisms, suspended substances, etc.) contained in the biological culture solution. Since the substance acts to prevent the substance from adhering to the filtration membrane, it is considered that clogging of the filtration membrane can be suppressed.

Thus, for example, when performing cross-flow filtration using a hollow fiber membrane, in order to perform filtration while stripping substances adhering to the surface of the hollow fiber membrane from the membrane surface, the filtration flow rate is usually relative to the membrane cross-sectional area. However, in the case of filtering a biological culture solution containing MNB as in the present invention, even if the filtration flow rate is 0.5 m/s or less, clogging of the hollow fiber membrane is caused. Since it is possible to carry out filtration while suppressing the above, it is possible to reduce stress and damage received by microorganisms and the like in the filtration step.

Furthermore, since the filtration efficiency is improved by this, the filtration device can be downsized, and since the supply amount of the biological culture solution to the filtration device can be reduced, the facility cost and operating cost of the biological reaction device can be reduced. In addition, it is possible to reduce stress/damage to the microorganisms.

Furthermore, in the present invention, the biological culture solution extracted from the culture tank containing the biological culture solution and supplied to the filter contains MNB, so that the biological culture solution circulated through the circulation route Since oxygen can be sufficiently supplied to microorganisms and the like, stress and damage to the microorganisms and the like can be reduced.

Such a method of containing MNB is not limited to the cross-flow filtration of a biological culture solution, but can be applied to general cross-flow filtration used for purification of water, concentration of foods, purification of abrasive particles, etc. , The same excellent effect is exhibited. That is, even in general cross-flow filtration, by including MNB in the liquid to be filtered, it is possible to perform filtration while suppressing clogging of the hollow fiber membranes due to the solid substance contained in the liquid. Equipment costs and operating costs can be reduced.

Further, as described above, in the method of adding MNB to the filtrate of the biological culture solution extracted from the culture tank as described in Patent Document 2, compared with the amount of the biological culture solution extracted from the culture tank, In the culture tank, the amount of the filtrate obtained by filtering the biological culture solution is quite small (the amount of the filtrate is usually about 1/10 to 1/100 of the amount of the biological culture solution extracted from the culture tank). There is a problem that it is difficult to maintain a high amount of MNB contained in the biological culture solution, and in order to solve this problem, if the amount of the biological culture solution extracted from the culture tank is increased or the filtration pressure is increased. However, in the present invention, the biological culture solution extracted from the culture tank and supplied to the filter contains micro-nano bubbles. Since the generator is provided, such a problem can be solved.

That is, in the present invention, the biological culture solution extracted from the culture tank contains MNB, and the biological culture solution containing the MNB is supplied to the filter, thereby preventing extra stress and damage to microorganisms and the like. The MNB can be sufficiently contained in the biological culture solution circulated through the circulation route, and the amount of MNB contained in the biological culture solution in the culture tank can be maintained high.

As a means for containing the MNB in the biological culture solution extracted from the culture tank and supplied to the filter, an MNB generating device is provided in the path for extracting the biological culture solution from the culture tank and supplying it to the filter. It is preferable to use a means for containing MNB in the culture solution (hereinafter referred to as “first means”). When the first means is used, MNB can be sufficiently contained in the biological culture solution that is supplied to the filter and circulates in the circulation path. Therefore, as described above, suppression of clogging of the filtration membrane, microorganisms, etc. It is possible to sufficiently exert the effect of reducing stress and damage received by the person.

Further, instead of the first means, an MNB generator is provided in the path for supplying the culture medium or the reaction raw material to the culture tank, and means for containing MNB in the biological culture solution (hereinafter referred to as "second means"), culture tank. An MNB generator is installed in the biological culture medium, and a means for containing the MNB in the biological culture medium (hereinafter, referred to as “third means”) and a MNB generator in the conduit for returning the concentrated liquid such as microorganisms from the filter to the culture tank. The use of one or a plurality of means (hereinafter referred to as “fourth means”) for causing MNB to be contained in the concentrated liquid such as microorganisms that is refluxed in the culture tank also prevents clogging of the filtration membrane, microorganisms, etc. It is possible to exert effects such as reduction of stress and damage received.

Further, the first means and one or more means of the second means to the fourth means can be used together. For example, when the first means is used alone, it takes time to bring the MNB content of the biological culture solution in the culture tank to an appropriate value, and when it is necessary to shorten this time, It is effective to use one or more of the second to fourth means in combination. In particular, the second means is preferable as the means used in combination with the first means because the microorganisms and the like are not stressed or damaged by the blowing of MNB.

The second feature of the present invention is to use MNB having an increased oxygen concentration as MNB contained in the biological culture solution.
As a result, even if the amount of the biological culture solution extracted from the culture tank is reduced, and even if the amount of MNB contained in the biological culture solution is reduced, the high concentration of easily absorbed oxygen in the MNB state, such as microorganisms in the culture tank, is reduced. The activity of microorganisms can be maintained. Furthermore, by reducing the amount of the biological culture solution extracted from the culture tank, it is possible to reduce stress and damage to microorganisms and the like, and it is possible to reduce energy required for circulating the biological culture solution. Furthermore, the energy required to drive the MNB generator can be reduced by reducing the amount of MNB contained in the biological culture solution.

In order to obtain air having an increased oxygen concentration, known oxygen enrichment means such as PSA method using adsorbent, VSA method, water electrolysis method, cryogenic separation method, membrane separation method, chemisorption method, etc. However, from an economical point of view, it is preferable to use an oxygen-enriched membrane and obtain air having an increased oxygen concentration by passing air through the oxygen-enriched membrane. When the PSA method, VSA method, and chemisorption method are used, it is preferable to obtain oxygen-enriched air by mixing oxygen produced by these methods and air with a line mixer or the like.

The oxygen concentration of the oxygen-enriched MNB is preferably 25 to 40%, more preferably 35 to 40%. When the oxygen concentration is 25% or more, the respiratory action of microorganisms and the like can be promoted and the activity of microorganisms and the like can be enhanced. When the oxygen concentration is 40% or less, microorganisms and the like are less likely to be damaged by oxidation.

A third feature of the present invention is a pump for delivering a biological culture solution containing microorganisms such as a pump for extracting the biological culture solution from the culture tank, a diaphragm pump, a tube pump, a screw pump, a rotary pump, etc. The positive displacement pump is used.

By using such a positive displacement pump to convey a biological culture solution containing microorganisms and the like, stress and damage to the microorganisms and the like can be further reduced.

The fourth feature of the present invention is to provide means for adding a pH adjusting agent to the conduit for returning the filtrate to the culture tank.
Microorganisms generate by-products such as organic acids in the biological reaction in the culture tank, but if this causes a change in the pH of the biological culture solution in the culture tank, adjust this to a range suitable for the microorganisms, etc. Need to be adjusted.

As a general pH adjusting means, a pH adjusting agent such as an acid or an alkali is directly added to a biological culture solution in a culture tank. However, with this means, the acid/alkali concentration is locally increased and microorganisms etc. It causes stress and damage.

In the present invention, since the pH adjusting agent such as acid or alkali is added to the filtrate returned to the culture tank, compared with the case where the pH adjusting agent is directly added to the biological culture solution, the microorganisms etc. in the culture tank are The pH of the biological culture solution in the culture tank can be adjusted without giving stress and damage due to the difference in concentration.

<Reference Embodiment 1 (FIG. 1)>
First, a reference embodiment 1 of the present invention will be described with reference to FIG.
The biological reaction device of Reference Embodiment 1 uses the first means as a means for containing the MNB in the biological culture solution extracted from the culture tank and supplied to the filter.

The bioreactor of Reference Embodiment 1 is used for any of bioreactions (fermentation/brewing) for producing reaction products by microorganisms or the like, and bioreactions for growth or concentration (culture) of microorganisms or the like. be able to. When the reaction product is intended to be produced, the reaction product is continuously collected together with the filtrate, and when the purpose is to grow or concentrate the microorganism or the like, after the microorganism or the like is grown, The biological culture solution in the culture tank is concentrated to collect microorganisms and the like.

First, a case where a biological reaction for the purpose of producing a reaction product by a microorganism or the like is performed will be described.
a) While agitating with the culture tank agitator 7, the biological reaction is carried out in the biological culture solution 3 contained in the culture tank 2 and containing the medium or the reaction raw material 1 and the microorganisms.
b) While carrying out the reaction of the above-mentioned a), the pump 8 is driven to continuously withdraw the reacted biological culture solution 3 from the culture tank 2 and supply it to the MNB generator 6a to contain the MNB of the air A. ..
c) The biological culture solution containing the MNB of the air A is supplied to the filter 4 and separated into a filtered solution B containing a reaction product and a concentrated solution C such as microorganisms.
e) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

Next, a case where a biological reaction for the purpose of growing or concentrating microorganisms or the like is performed will be described.
As the first step, until the microorganisms and the like grow to an appropriate amount, operations are carried out in the same steps as the biological reaction for the purpose of producing reaction products by the microorganisms and the like. By separating and recovering the filtrate B, by-products such as organic acids produced by biological reactions of microorganisms and the like can be recovered together with the filtrate B.

As a second step, after the microorganisms and the like grow to an appropriate amount, the supply of the medium or the reaction raw material 1 to the culture tank 2 is stopped, and the microorganisms and the like are concentrated and recovered by the following steps.
a) While proliferating microorganisms and the like, the pump 8 is driven to continuously withdraw the reacted biological culture liquid 3 from the culture tank 2 and supply it to the MNB generator 6a to contain the MNB of the air A.
b) The biological culture solution containing MNB of the air A is supplied to the filter 4 and separated into the filtered solution B and the concentrated solution C such as microorganisms.
c) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

As described above, in the second stage, the biological culture solution in which the concentration of microorganisms or the like is increased is circulated outside the culture tank 2 through the MNB generator 6a and the filter 4, but in the present invention, the culture is performed. Since the biological culture medium 3 extracted from the tank 2 contains MNB, clogging of the filtration membrane can be suppressed, and oxygen can be sufficiently supplied to the microorganisms in the circulation path, which in turn causes stress and stress on the microorganisms. The damage can be reduced.

In the reference embodiment 1, the MNB generator 6a is provided in the pipe line connecting the pump 8 and the filter 4, but the MNB generator 6a is provided in the pipe line connecting the culture tank 2 and the pump 8. However, this aspect is also included in the reference embodiment 1. This also applies to the following first to fourth embodiments and reference embodiments 2 to 5.

The filter 4 includes a filtration membrane and a container that houses the filtration membrane. The filtration membrane may be an organic membrane or an inorganic membrane. The shape of the filtration membrane may be any shape such as a flat membrane, a hollow fiber membrane, and a spiral type, but among them, a hollow fiber membrane module is preferable, and if the hollow fiber membrane module is, an external pressure type, an internal type Any type of pressure type can be adopted.

As a filtration method, cross-flow filtration using a hollow fiber membrane module is preferable. In this filtration method, a culture solution containing reaction products, microorganisms, etc. is supplied to the inside of the hollow fiber membranes to be filtered, and the filtrate is taken out from the outside. Microorganisms deposited inside the hollow fiber membranes. Since membrane stains such as the above are scraped off by the shearing force of the parallel flow of the culture solution, a stable filtration state can be maintained for a long period of time.

When performing cross-flow filtration using a hollow fiber membrane module, in order to scrape the membrane fouling, it is necessary to flow the liquid to be filtered into the hollow fiber membrane at a flow rate above a certain level. Since the biological culture solution containing microorganisms, which is the object of filtration, contains MNB, the membrane dirt can be sufficiently scraped off even if it is flowed at a lower flow rate than usual, and stress or damage given to microorganisms etc. Can be significantly reduced.

Specifically, in a general cross-flow filtration, the circulation flow rate is about 1 to 2 m/s when an organic membrane is used, and about 1 to 3 m/s when a ceramic membrane is used, the steady operation is performed. However, by including MNB in the biological culture medium, the membrane fouling can be reduced and the filtration resistance can be kept small. Therefore, the circulation necessary to obtain the same flux (membrane filtration water amount per unit time/unit membrane area). The flow velocity can be reduced to about 0.2 to 1.5 m/s. When operating at the same circulation flow rate, the flux can be increased by 1.2 to 2.0 times.

As the filtration membrane, an organic polymer compound can be preferably used from the viewpoint of separation performance, water permeability performance, and stain resistance. For example, polyethylene resin, polypropylene resin, polyvinyl chloride resin, polyvinylidene fluoride resin, polysulfone resin, polyethersulfone resin, polyacrylonitrile resin, cellulose resin and cellulose triacetate resin, and the like, It may be a mixture of resins containing these resins as main components. Polyvinyl chloride resin, polyvinylidene fluoride resin, polysulfone resin, polyether sulfone resin, and polyacrylonitrile resin, which are easy to form a film by solution and have excellent physical durability and chemical resistance, are preferable. A vinylidene chloride-based resin or a resin containing the vinylidene-based resin as a main component is more preferably used because it has the characteristics of having both chemical strength (particularly chemical resistance) and physical strength.

Here, as the polyvinylidene fluoride resin, a vinylidene fluoride homopolymer is preferably used. Further, as the polyvinylidene fluoride-based resin, a copolymer of vinylidene fluoride and a vinyl-based monomer copolymerizable with each other may be used. Examples of vinyl monomers copolymerizable with vinylidene fluoride include tetrafluoroethylene, hexafluoropropylene, and trifluorochloroethylene.

The average pore diameter of the filtration membrane can be appropriately determined depending on the purpose and situation of use, but it is preferably small to some extent, and is usually 0.01 μm or more and 1 μm or less. If the average pore diameter of the hollow fiber membrane is less than 0.01 μm, components such as microorganisms such as sugars and proteins and aggregates of the membrane such as aggregates block the pores and stable operation cannot be performed. In consideration of the balance with water permeability, the thickness is preferably 0.02 μm or more, more preferably 0.03 μm or more. On the other hand, when it exceeds 1 μm, the smoothness of the membrane surface and the shearing force due to the flow of the membrane surface and the peeling of the dirt component from the pores due to physical washing such as backwashing and air scrubbing become insufficient, and stable operation can be performed. Disappear.

Further, when the average pore size approaches the size of microorganisms, etc., these may directly block the pores. Further, some of the microorganisms may be killed to produce crushed products, and in order to avoid clogging of the pores with these crushed products, the average pore diameter is preferably 0.4 μm or less, and 0.2 μm or less. The following are preferred:

Here, the average pore diameter of the filtration membrane can be obtained by measuring and averaging the diameters of a plurality of pores observed by scanning electron microscope observation at a magnification of 10,000 times or more. It is preferable to randomly select 10 or more pores, preferably 20 or more pores, measure the diameters of the pores, and average them to obtain a number. If the pores are not circular, it is also preferable to use a method such as an image processing device to obtain a circle having an area equal to that of the pores, that is, an equivalent circle, and obtain the equivalent circle diameter as the diameter of the pore. it can.

<Reference Embodiment 2 (FIG. 2)>
A second embodiment of the present invention will be described with reference to FIG.
The bioreactor of Reference Embodiment 2 is the same as the bioreactor of Reference Embodiment 1 except that a pipe line equipped with a valve for refluxing the filtrate to the culture tank is provided. By adjusting the amount of the filtrate flowing back to the culture tank through this conduit, the concentration of the medium or the reaction raw material, the microorganism, etc. in the culture tank can be adjusted.

The bioreactor of Reference Embodiment 2 is used for any of bioreactions (fermentation/brewing) for the purpose of producing reaction products by microorganisms or the like, and bioreactions for the purpose of growing or concentrating (culturing) microorganisms or the like. be able to. When the reaction product is intended to be produced, the reaction product is continuously collected together with the filtrate, and when the purpose is to grow or concentrate the microorganism or the like, after the microorganism or the like is grown, The biological culture solution in the culture tank is concentrated to collect microorganisms and the like.

First, a case where a biological reaction for the purpose of producing a reaction product by a microorganism or the like is performed will be described.
Normally, the valve 10 is closed and the valve 11 is opened, and the biological reaction is performed by the following steps.
a) While agitating with the culture tank agitator 7, the biological reaction is carried out in the biological culture solution 3 contained in the culture tank 2 and containing the medium or the reaction raw material 1 and the microorganisms.
b) While carrying out the reaction of the above a), the pump 8 is driven to continuously withdraw the reacted biological culture solution 3 from the culture tank 2 and supply it to the MNB generator 6a to contain the MNB of the air A. ..
c) The biological culture solution containing the MNB of the air A is supplied to the filter 4 and separated into a filtered solution B containing a reaction product and a concentrated solution C such as microorganisms.
e) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

Further, in order to keep the amount or concentration of the medium or the reaction raw material and the microorganisms in the culture tank 2 constant, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B refluxed to the culture tank 2. Alternatively, an operation such as newly supplying the medium, the reaction raw material 1, the microorganisms or the like to the culture tank 2 can be appropriately performed.

Next, a case where a biological reaction for the purpose of growing or concentrating microorganisms or the like is performed will be described.
As the first step, until the microorganisms and the like grow to an appropriate amount, operations are carried out in the same steps as the biological reaction for the purpose of producing reaction products by the microorganisms and the like. By separating and recovering the filtrate B, by-products such as organic acids produced by biological reactions of microorganisms and the like can be recovered together with the filtrate B. In this first step, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2 in the same manner as the biological reaction for producing the reaction product by the microorganisms and the like. be able to.

As a second step, after the microorganisms and the like have grown to an appropriate amount, the supply of the medium or the reaction raw material 1 to the culture tank 2 is stopped, and the valve 10 is closed so that the filtrate B is refluxed to the culture tank 2. Stop and concentrate and collect microorganisms by the following steps.
a) While proliferating microorganisms and the like, the pump 8 is driven to continuously withdraw the reacted biological culture liquid 3 from the culture tank 2 and supply it to the MNB generator 6a to contain the MNB of the air A.
b) The biological culture solution containing MNB of the air A is supplied to the filter 4 and separated into the filtered solution B and the concentrated solution C such as microorganisms.
c) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

As described above, in the second stage, the biological culture solution in which the concentration of microorganisms or the like is increased is circulated outside the culture tank 2 through the MNB generator 6a and the filter 4, but in the present invention, the culture is performed. Since the biological culture medium 3 extracted from the tank 2 contains MNB, clogging of the filtration membrane can be suppressed, and oxygen can be sufficiently supplied to the microorganisms in the circulation path, which in turn causes stress and stress on the microorganisms. The damage can be reduced.

<Reference Embodiment 3 (FIG. 3)>
A third embodiment of the present invention will be described with reference to FIG.
The biological reaction apparatus of Reference Embodiment 3 is a combination of the first means and the second means as means for containing the MNB in the biological culture solution extracted from the culture tank and supplied to the filter. is there. Similar to Reference Embodiment 1 and Reference Embodiment 2, Reference Embodiment 3 also aims at a biological reaction (fermentation/brewing) for the purpose of producing a reaction product by a microorganism or the like, and a growth or concentration (culture) of the microorganism or the like. Can be used for any of the biological reactions mentioned above.

First, a case where a biological reaction for the purpose of producing a reaction product by a microorganism or the like is performed will be described.
Normally, the valve 10 is closed and the valve 11 is opened, and the biological reaction is performed by the following steps.
a) The medium or the reaction raw material 1 is supplied to the MNB generator 6b to contain the MNB of the air A, and the medium or the reaction raw material D containing the MNB of the air A is supplied to the culture tank 2.
b) The biological reaction is carried out in the biological culture solution 3 containing the medium or the reaction raw material 1 and the microorganisms and the like, which is stored in the culture tank 2 while being stirred by the culture tank agitator 7.
c) While carrying out the reaction of b) above, the pump 8 is driven to continuously withdraw the reacted biological culture liquid 3 from the culture tank 2 and supply it to the MNB generator 6a to contain the MNB of the air A. ..
d) The biological culture solution containing the MNB of the air A is supplied to the filter 4 and separated into a filtered solution B containing a reaction product and a concentrated solution C such as microorganisms.
e) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

Further, in order to keep the amount or concentration of the culture solution and the microorganisms in the culture tank 2 constant, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2, It is possible to appropriately perform operations such as newly supplying the medium or the reaction raw material 1, the microorganisms and the like to the culture tank 2.

Next, a case where a biological reaction for the purpose of growing or concentrating microorganisms or the like is performed will be described.
As the first step, until the microorganisms and the like grow to an appropriate amount, operations are performed in the same steps as the biological reaction for the purpose of producing reaction products by the microorganisms and the like. By separating and recovering the filtrate B, by-products such as organic acids produced by biological reactions of microorganisms and the like can be recovered together with the filtrate B. In this first step, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2 in the same manner as the biological reaction for producing the reaction product by the microorganisms and the like. be able to.

As a second step, after the microorganisms and the like have grown to an appropriate amount, the supply of the medium or the reaction raw material 1 to the culture tank 2 is stopped, and the valve 10 is closed so that the filtrate B is refluxed to the culture tank 2. Stop and concentrate and collect microorganisms by the following steps.
a) While proliferating microorganisms and the like, the pump 8 is driven to continuously withdraw the reacted biological culture liquid 3 from the culture tank 2 and supply it to the MNB generator 6a to contain the MNB of the air A.
b) The biological culture solution containing MNB of the air A is supplied to the filter 4 and separated into the filtered solution B and the concentrated solution C such as microorganisms.
c) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

As described above, in the second stage, the biological culture solution in which the concentration of microorganisms or the like is increased is circulated outside the culture tank 2 through the MNB generator 6a and the filter 4, but in the present invention, the culture is performed. Since the biological culture medium 3 extracted from the tank 2 contains MNB, clogging of the filtration membrane can be suppressed, and oxygen can be sufficiently supplied to the microorganisms in the circulation path, which in turn causes stress and stress on the microorganisms. The damage can be reduced.

<Reference Embodiment 4 (FIG. 4)>
A reference embodiment 4 of the present invention will be described with reference to FIG.
The biological reaction apparatus of Reference Embodiment 4 uses the second means and the third means in combination with the first means as means for containing the MNB in the biological culture solution extracted from the culture tank and supplied to the filter. It was done.

Similar to Reference Embodiments 1 to 3, Reference Embodiment 4 also aims at a biological reaction (fermentation/brewing) for the purpose of producing a reaction product by a microorganism or the like, or a growth or concentration (culture) of the microorganism or the like. Can be used for any of the biological reactions mentioned above.

First, a case where a biological reaction for the purpose of producing a reaction product by a microorganism or the like is performed will be described.
Normally, the valve 10 is closed and the valve 11 is opened, and the biological reaction is performed by the following steps.
a) The medium or the reaction raw material 1 is supplied to the MNB generator 6b to contain the MNB of the air A, and the medium or the reaction raw material D containing the MNB of the air A is supplied to the culture tank 2.
b) The medium or the reaction raw material 1 stored in the culture tank 2 while being stirred by the culture tank agitator 7 and supplying the biological culture solution E containing the MNB of the air A by the MNB generator 6c to the culture tank 2. A biological reaction is performed in the biological culture solution 3 containing microorganisms and the like. The biological culture solution 3 is supplied to the MNB generator 6c by extracting the biological culture solution 3 from the culture tank 2 by the pump 9.
c) While carrying out the reaction of b) above, the pump 8 is driven to continuously withdraw the reacted biological culture solution 3 from the culture tank 2 and supply it to the MNB generator 6a to contain the MNB of air A. ..
d) The biological culture solution containing the MNB of the air A is supplied to the filter 4 and separated into a filtered solution B containing a reaction product and a concentrated solution C such as microorganisms.
e) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

Further, in order to keep the amount or concentration of the culture solution and the microorganisms in the culture tank 2 constant, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2, It is possible to appropriately perform operations such as newly supplying the medium or the reaction raw material 1, the microorganisms and the like to the culture tank 2.

Next, a case where a biological reaction for the purpose of growing or concentrating microorganisms or the like is performed will be described.
As the first step, until the microorganisms and the like grow to an appropriate amount, operations are carried out in the same steps as the biological reaction for the purpose of producing reaction products by the microorganisms and the like. By separating and recovering the filtrate B, by-products such as organic acids produced by biological reactions of microorganisms and the like can be recovered together with the filtrate B.

In the second step, after the microorganisms and the like grow to an appropriate amount, the valve 10 is closed to stop the reflux of the filtrate B to the culture tank 2, and the microorganisms and the like are concentrated and recovered by the following steps.
a) While performing the growth, the pump 8 is driven to continuously extract the biological culture solution 3 from the culture tank 2, supply it to the MNB generator 6a to contain the MNB of the air A, and then to the filter 4. Supply.
b) In the filter 4, the biological culture liquid is separated into a filtered liquid B and a concentrated liquid C such as microorganisms.
c) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

As described above, in the second stage, the biological culture solution having a high concentration of microorganisms and the like is circulated to the outside of the culture tank 2 via the filter 4, but in the present invention, the biological culture extracted from the culture tank 2 is used. Since the liquid 3 contains MNB, clogging of the filtration membrane can be suppressed, oxygen can be sufficiently supplied to the microorganisms and the like in the circulation route, and stress and damage to the microorganisms and the like can be reduced.

<First embodiment (FIG. 5)>
A first embodiment of the present invention will be described with reference to FIG.
The biological reaction apparatus of the first embodiment uses the second means alone as a means for containing the MNB in the biological culture solution extracted from the culture tank and supplied to the filter.

Similarly to the first to fourth embodiments, the first embodiment also aims at a biological reaction (fermentation/brewing) for the purpose of producing a reaction product by a microorganism or the like, and a growth or concentration (culture) of the microorganism or the like. Can be used for any of the biological reactions mentioned above.

First, a case where a biological reaction for the purpose of producing a reaction product by a microorganism or the like is performed will be described.
Normally, the valve 10 is closed and the valve 11 is opened, and the biological reaction is performed by the following steps.
a) The medium or the reaction raw material 1 is supplied to the MNB generator 6b to contain the MNB of the air A, and the medium or the reaction raw material D containing the MNB of the air A is supplied to the culture tank 2.
b) The biological reaction is carried out in the biological culture solution 3 containing the medium or the reaction raw material 1 and the microorganisms and the like, which is stored in the culture tank 2 while being stirred by the culture tank agitator 7.
c) While carrying out the reaction of the above b), the pump 8 is driven to continuously withdraw the reacted biological culture solution 3 from the culture tank 2, and the biological culture solution containing the MNB of the air A is transferred to the filter 4. It is supplied and separated into a filtrate B containing a reaction product and a concentrated liquid C such as microorganisms.
e) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

Further, in order to keep the amount or concentration of the culture solution and the microorganisms in the culture tank 2 constant, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2, It is possible to appropriately perform operations such as newly supplying the medium or the reaction raw material 1, the microorganisms and the like to the culture tank 2.

Next, a case where a biological reaction for the purpose of growing or concentrating microorganisms or the like is performed will be described.
As the first step, until the microorganisms and the like grow to an appropriate amount, operations are carried out in the same steps as the biological reaction for the purpose of producing reaction products by the microorganisms and the like. By separating and recovering the filtrate B, by-products such as organic acids produced by biological reactions of microorganisms and the like can be recovered together with the filtrate B.

In the second step, after the microorganisms and the like grow to an appropriate amount, the valve 10 is closed to stop the reflux of the filtrate B to the culture tank 2, and the microorganisms and the like are concentrated and recovered by the following steps.
a) While performing the growth, the pump 8 is driven to continuously withdraw the biological culture solution 3 from the culture tank 2 and supply it to the filter 4.
b) In the filter 4, the biological culture liquid is separated into a filtered liquid B and a concentrated liquid C such as microorganisms.
c) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

As described above, in the second stage, the biological culture solution having a high concentration of microorganisms and the like is circulated to the outside of the culture tank 2 via the filter 4, but in the present invention, the biological culture extracted from the culture tank 2 is used. Since the liquid 3 contains MNB, clogging of the filtration membrane can be suppressed, oxygen can be sufficiently supplied to the microorganisms in the circulation path, and stress and damage to the microorganisms can be reduced. ..

<Second Embodiment (FIG. 6)>
A second embodiment of the present invention will be described with reference to FIG.
The bioreactor of the second embodiment is a combination of the second means and the third means as means for containing the MNB in the biological culture liquid extracted from the culture tank and supplied to the filter. ..

In the second embodiment, as in the first to fourth embodiments and the first embodiment, a biological reaction (fermentation/brewing) for the purpose of producing a reaction product by a microorganism or the like, a growth or concentration of the microorganism or the like. It can be used for any biological reaction for the purpose of (culturing).

First, a case where a biological reaction for the purpose of producing a reaction product by a microorganism or the like is performed will be described.
Normally, the valve 10 is closed and the valve 11 is opened, and the biological reaction is performed by the following steps.
a) The medium or the reaction raw material 1 is supplied to the MNB generator 6b to contain the MNB of the air A, and the medium or the reaction raw material D containing the MNB of the air A is supplied to the culture tank 2.
b) The medium or the reaction raw material 1 stored in the culture tank 2 while being stirred by the culture tank agitator 7 and supplying the biological culture solution E containing the MNB of the air A by the MNB generator 6c to the culture tank 2. A biological reaction is performed in the biological culture solution 3 containing microorganisms and the like. The biological culture solution 3 is supplied to the MNB generator 6c by extracting the biological culture solution 3 from the culture tank 2 by the pump 9.
c) While carrying out the reaction of the above b), the pump 8 is driven to continuously withdraw the reacted biological culture solution 3 from the culture tank 2, and the biological culture solution containing the MNB of the air A is transferred to the filter 4. It is supplied and separated into a filtrate B containing a reaction product and a concentrated liquid C such as microorganisms.
e) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

Further, in order to keep the amount or concentration of the culture solution and the microorganisms in the culture tank 2 constant, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2, It is possible to appropriately perform operations such as newly supplying the medium or the reaction raw material 1, the microorganisms and the like to the culture tank 2.

Next, a case where a biological reaction for the purpose of growing or concentrating microorganisms or the like is performed will be described.
As the first step, until the microorganisms and the like grow to an appropriate amount, operations are carried out in the same steps as the biological reaction for the purpose of producing reaction products by the microorganisms and the like. By separating and recovering the filtrate B, by-products such as organic acids produced by biological reactions of microorganisms and the like can be recovered together with the filtrate B.

In the second step, after the microorganisms and the like grow to an appropriate amount, the valve 10 is closed to stop the reflux of the filtrate B to the culture tank 2, and the microorganisms and the like are concentrated and recovered by the following steps.
a) While performing the growth, the pump 8 is driven to continuously withdraw the biological culture solution 3 from the culture tank 2 and supply it to the filter 4. At this time, the MNB generator 6c causes the biological culture solution 3 in the culture tank 2 to contain MNB of air A.
b) In the filter 4, the biological culture liquid is separated into a filtered liquid B and a concentrated liquid C such as microorganisms.
c) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution C of microorganisms containing MNB is refluxed to the culture tank 2.

As described above, in the second stage, the biological culture solution having a high concentration of microorganisms and the like is circulated to the outside of the culture tank 2 via the filter 4, but in the present invention, the biological culture extracted from the culture tank 2 is used. Since the liquid 3 contains MNB, clogging of the filtration membrane can be suppressed, oxygen can be sufficiently supplied to the microorganisms in the circulation path, and stress and damage to the microorganisms can be reduced. ..

<Third Embodiment (FIG. 7)>
A third embodiment of the present invention will be described with reference to FIG.
The biological reaction apparatus of the third embodiment uses the fourth means as a means for containing the MNB in the biological culture solution extracted from the culture tank and supplied to the filter.

Similarly to the first to fourth embodiments and the first to second embodiments, the third embodiment also includes a biological reaction (fermentation/brewing) for the purpose of producing a reaction product by a microorganism or the like, a microorganism. Can be used for any biological reaction for the purpose of proliferation or concentration (culturing).

First, a case where a biological reaction for the purpose of producing a reaction product by a microorganism or the like is performed will be described.
Normally, the valve 10 is closed and the valve 11 is opened, and the biological reaction is performed by the following steps.
a) While agitating with the culture tank agitator 7, the biological reaction is carried out in the biological culture solution 3 contained in the culture tank 2 and containing the medium or the reaction raw material 1 and the microorganisms.
b) While carrying out the reaction of a) above, the pump 8 is driven to continuously withdraw the reacted biological culture solution 3 from the culture tank 2 and supply it to the filter 4 to obtain a filtrate containing the reaction product. B and a concentrated solution C such as microorganisms are separated.
c) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution F of microorganisms containing MNB of the air A is refluxed to the culture tank 2 by the MNB generator 6d.

Further, in order to keep the amount or concentration of the culture solution and the microorganisms in the culture tank 2 constant, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2, It is possible to appropriately perform operations such as newly supplying the medium or the reaction raw material 1, the microorganisms and the like to the culture tank 2.

Next, a case where a biological reaction for the purpose of growing or concentrating microorganisms or the like is performed will be described.
As the first step, until the microorganisms and the like grow to an appropriate amount, operations are performed in the same steps as the biological reaction for the purpose of producing reaction products by the microorganisms and the like. By separating and recovering the filtrate B, by-products such as organic acids produced by biological reactions of microorganisms and the like can be recovered together with the filtrate B. In this first step, the opening and closing of the valve 10 and the valve 11 are adjusted to adjust the amount of the filtrate B flowing back to the culture tank 2 in the same manner as the biological reaction for producing the reaction product by the microorganisms and the like. be able to.

As a second step, after the microorganisms and the like have grown to an appropriate amount, the supply of the medium or the reaction raw material 1 to the culture tank 2 is stopped, and the valve 10 is closed so that the filtrate B is refluxed to the culture tank 2. Stop and concentrate and collect microorganisms by the following steps.
a) While propagating microorganisms and the like, the pump 8 is driven to continuously withdraw the reacted biological culture liquid 3 from the culture tank 2 and supply it to the filter 4, and the filtrate B and the concentrated liquid C such as microorganisms C And separate.
b) The filtrate B is collected in the filtrate storage tank 5, and the concentrated solution F of microorganisms containing MNB of the air A is refluxed to the culture tank 2 by the MNB generator 6d.

As described above, in the second stage, the biological culture solution in which the concentration of microorganisms or the like is increased is circulated to the outside of the culture tank 2 via the filter 4 and the MNB generator 6d, but in the present invention, Since the microorganism-concentrated liquid F separated from the filter 4 by the MNB generating device 6d is refluxed to the culture tank 2 by the MNB generator 6d, clogging of the filtration membrane can be suppressed. In addition, oxygen can be sufficiently supplied to the microorganisms and the like in the circulation route, and consequently stress and damage to the microorganisms and the like can be reduced.

<Reference Embodiment 5 (FIG. 8)>
A fifth embodiment of the present invention will be described with reference to FIG.
The bioreactor of Reference Embodiment 5 is the same as that of Reference Embodiment 2 of the present invention, except that the pH adjusting agent 12 is added to the filtrate B that is refluxed in the culture tank 2. By adjusting the addition amount of the pH adjuster 12, the pH of the biological culture solution 3 in the culture tank 2 can be adjusted to a range suitable for microorganisms and the like.

Similarly to the second embodiment, the fifth embodiment also includes a biological reaction (fermentation/brewing) for producing a reaction product by a microorganism, a biological reaction for growing or concentrating (culturing) the microorganism. It can be used for both.

In the reference embodiment 5, normally, the valve 10 is closed and the valve 11 is opened to perform the biological reaction, but in order to keep the amount or concentration of the biological culture solution 3 and the microorganisms in the culture tank 2 constant, The valve 10 is opened and a part or all of the filtrate B separated by the filter 4 is refluxed to the culture tank 2.

At that time, in Reference Embodiment 5, the pH adjuster 12 is added by the following steps.
a) From the state where the biological reaction is performed by closing the valve 10 and opening the valve 11, the valve 10 is opened and a part or all of the filtrate B separated by the filter 4 is mixed with the pH adjusting agent mixing tank 13 Supply to.
b) In the filtrate B in the pH adjusting agent mixing tank 13, an amount of the pH adjusting agent 12 (acid, alkali) necessary for adjusting the pH of the biological culture solution 3 in the culture tank 2 to a range suitable for microorganisms and the like. Etc.) is added.
c) The filtrate B to which the pH adjuster 12 has been added is refluxed to the culture tank 2.

When a pH adjusting agent such as an acid or an alkali is directly added to the biological culture liquid 3 in the culture tank 2, the acid/alkali concentration locally increases, which causes stress or damage to microorganisms or the like. However, by adding the pH adjusting agent as described above, the pH of the biological culture solution 3 in the culture tank 2 is adjusted without giving stress and damage due to the concentration difference to the microorganisms and the like in the culture tank 2. be able to.

<Fourth Embodiment (FIG. 9)>
A fourth embodiment of the present invention will be described with reference to FIG.
The bioreactor of the fourth embodiment is the same as that of the second embodiment of the present invention, but is provided with a means for adding the pH adjuster 12 to the filtrate B refluxed in the culture tank 2. By adjusting the addition amount of the pH adjuster 12, the pH of the biological culture solution 3 in the culture tank 2 can be adjusted to a range suitable for microorganisms and the like.

Similarly to the second embodiment, the fourth embodiment also includes a biological reaction for the purpose of producing a reaction product by a microorganism (fermentation/brewing) and a biological reaction for the purpose of growing or concentrating (culturing) a microorganism. It can be used for both.

In the fourth embodiment, normally, the valve 10 is closed and the valve 11 is opened to perform the biological reaction, but in order to keep the amount or concentration of the biological culture solution 3 and the microorganisms in the culture tank 2 constant, The valve 10 is opened and a part or all of the filtrate B separated by the filter 4 is refluxed to the culture tank 2.

At that time, in the fourth embodiment, the pH adjuster 12 is added by the following steps.
a) From the state where the biological reaction is performed by closing the valve 10 and opening the valve 11, the valve 10 is opened and a part or all of the filtrate B separated by the filter 4 is mixed with the pH adjusting agent mixing tank 13 Supply to.
b) In the filtrate B in the pH adjusting agent mixing tank 13, an amount of the pH adjusting agent 12 (acid, alkali) necessary for adjusting the pH of the biological culture solution 3 in the culture tank 2 to a range suitable for microorganisms and the like. Etc.) is added.
c) The filtrate B to which the pH adjuster 12 has been added is refluxed to the culture tank 2.

When a pH adjusting agent such as an acid or an alkali is directly added to the biological culture liquid 3 in the culture tank 2, the acid/alkali concentration locally increases, which causes stress or damage to microorganisms or the like. However, by adding the pH adjusting agent as described above, the pH of the biological culture solution 3 in the culture tank 2 is adjusted without giving stress and damage due to the concentration difference to the microorganisms and the like in the culture tank 2. be able to.

As described above, in the bioreactor of the present invention and the bioreaction method using this bioreactor, the biological culture solution containing MNB is filtered by a filter to reduce stress and damage to microorganisms and the like. The amount of MNB contained in the biological culture solution in the culture tank can be maintained at a high level, and the biological culture solution can be appropriately filtered and circulated, thereby efficiently performing biological reactions using microorganisms and the like. It is an excellent thing that can be done economically and economically.

Further, in the present invention, as a means for containing MNB in the biological culture medium, by using the first means together with one or a plurality of means for the second means to the fourth means, it is possible to use the medium in a culture tank in a short time. The content of MNB in the biological culture can be set to an appropriate value.

Further, in the present invention, as the MNB contained in the biological culture medium, MNB formed from air having an increased oxygen concentration is used, whereby the amount of the biological culture medium extracted from the culture tank is reduced and the biological culture medium contains it. Even if the amount of MNB is reduced, a high concentration of easily absorbed oxygen in the state of MNB can be supplied to the microorganisms in the culture tank, so that the stress/damage received by the microorganisms can be reduced and the activity of the microorganisms can be maintained. In addition, the energy required for circulating the biological culture solution and the energy required for driving the MNB generator can be reduced.

Further, in the present invention, 1) a pump for extracting the biological culture solution from the culture tank, 2) a pump for supplying the biological culture solution containing MNB from the MNB generator to the filter, and 3) a culture tank. As a pump that conveys a liquid containing microorganisms such as a pump for refluxing the biological culture liquid excluding the filtrate, a diaphragm pump, a tube pump, a screw pump, a rotary pump, etc. that causes relatively little stress and damage to the microorganisms By using the positive displacement pump, it is possible to further reduce the stress and damage that the microorganisms receive and further maintain the activity of the microorganisms.

Further, in the present invention, a conduit for refluxing the filtrate to the culture tank is provided with a means for adding a pH adjuster, and by adding a pH adjuster such as acid or alkali to the filtrate refluxing to the culture tank. The pH of the biological culture solution in the culture tank can be adjusted without giving stress or damage to the microorganisms in the culture tank due to the difference in concentration.

Next, the action and effect such as clogging of the filtration membrane, which is exhibited by the feature of the present invention that MNB is contained in the biological culture solution extracted from the culture tank and supplied to the filter Will be described with reference to experimental examples and comparative experimental examples, but the present invention is not limited to these experimental examples and comparative experimental examples.

<Experimental Examples 1-5 and Comparative Experimental Examples 1-5>
As a filter, a filter (manufactured by Asahi Kasei Co., Ltd., trade name “Microza MF USP-343”) in which a large number of hollow fiber membranes are bundled and housed in a cylindrical cartridge is used. The standard strain of coryneform bacteria (Corynebacterium glutamicum) is contained at a turbidity (value of OD660): 60], and the pressure and flow rate of the culture solution supplied to this filter are adjusted to obtain each hollow. The average pressure difference between the inside and outside of the fiber membrane (hereinafter referred to as “transmembrane pressure difference”) and the average flow velocity of the culture solution flowing inside each hollow fiber membrane (hereinafter referred to as “circulation flow velocity”) are set to predetermined values. The value was adjusted and the filtration rate by the filter was measured.

In Experimental Examples 1 to 3 and Comparative Experimental Examples 1 to 3, the transmembrane pressure difference was set to 0.1 MPa, the circulation flow rate was changed, and the filtration rate was measured. In addition, in Experimental Examples 4 to 5 and Comparative Experimental Examples 4 to 5, the transmembrane pressure difference was set to 0.2 MPa, the circulation flow rate was changed, and the filtration rate was measured.

The filtration rate gradually decreases at the beginning of filtration due to the action of coryneform bacteria clogging the pores of the hollow fiber membrane, but eventually this clogging action and the action of removing the clogging due to the flow of the culture solution are in equilibrium. Thus, the filtration rate becomes stable (hereinafter, this state is referred to as “first stable state”, and the filtration rate in this state is referred to as “no MNB filtration rate”). Next, in the first stable state, when MNB was blown into the culture solution supplied to the filter using the MNB generator (trade name: OKE-25L, manufactured by OK Engineering Co., Ltd.), the filtration rate gradually increased. After that, the above two actions are again in an equilibrium state, and the filtration rate becomes stable again (hereinafter, this state is referred to as "second stable state", and the filtration rate in this state is referred to as "MNB-containing filtration rate"). ..

The MNB generators used in Experimental Examples 1 to 5 and Comparative Experimental Examples 1 to 5 are of the nozzle type, and the flow rate of the culture solution supplied to the filter and the concentration of MNB contained in the culture solution. Has a positive correlation.

<Experimental Example 1>
When the culture solution was supplied to the filter so that the transmembrane pressure difference was 0.1 Mpa and the circulation flow rate was 0.19 m/s, the MNB-containing filtration rate was 50.3 L/m ² /h.

<Comparative Experimental Example 1>
When the culture solution was supplied to the filter at the same transmembrane pressure difference and circulating flow rate as in Example 1, the MNB non-filtration rate was 27.4 L/m ² /h.

<Experimental example 2>
When the culture solution was supplied to the filter so that the transmembrane pressure difference was 0.1 MPa and the circulation flow rate was 0.33 m/s, the MNB-containing filtration rate was 59.2 L/m ² /h.

<Comparative Experimental Example 2>
When the culture solution was supplied to the filter at the same transmembrane pressure difference and circulation flow rate as in Example 2, the MNB non-filtration rate was 43.0 L/m ² /h.

<Experimental example 3>
When the culture solution was supplied to the filter so that the transmembrane pressure was 0.1 MPa and the circulation flow rate was 0.51 m/s, the MNB-containing filtration rate was 73.6 L/m ² /h.

<Comparative Experimental Example 3>
When the culture solution was supplied to the filter at the same transmembrane pressure difference and circulating flow rate as in Example 3, the MNB non-filtration rate was 67.0 L/m ² /h.

The results of Experimental Examples 1 to 3 and Comparative Experimental Examples 1 to 3 are summarized in Table 1.

<Experimental example 4>
When the culture solution was supplied to the filter so that the transmembrane pressure difference was 0.2 MPa and the circulation flow rate was 0.33 m/s, the MNB-containing filtration rate was 79.5 L/m ² /h.

<Comparative Experimental Example 4>
When the culture solution was supplied to the filter at the same transmembrane pressure difference and circulation flow rate as in Example 4, the MNB non-filtration rate was 43.6 L/m ² /h.

<Experimental example 5>
When the culture solution was supplied to the filter so that the transmembrane pressure difference was 0.2 MPa and the circulation flow rate was 0.51 m/s, the MNB-containing filtration rate was 102.8 L/m ² /h.

<Comparative Experimental Example 5>
When the culture solution was supplied to the filter at the same transmembrane pressure difference and circulating flow rate as in Example 5, the MNB non-filtration rate was 77.7 L/m ² /h.

The results of Experimental Examples 4 to 5 and Comparative Experimental Examples 4 to 5 are summarized in Table 2.

As is clear from the results of Experimental Examples 1 to 5 and Comparative Experimental Examples 1 to 5, by including MNB in the culture solution supplied to the filter, clogging of the hollow fiber membranes can be suppressed and the filtration rate can be maintained high. I know that I can do it.

In addition, Table 3 plots the results of Experimental Examples 1 to 3 and Comparative Experimental Examples 1 to 3 with the circulation flow rate (m/s) as the horizontal axis and the filtration rate (L/m ² /h) as the vertical axis. Is. As shown by the upper line (Experimental Example 1→2→3) and the lower line (Comparative Experimental Example 1→2→3), it was demonstrated by adding MNB to the culture solution supplied to the filter. It can be seen that the effect of suppressing clogging of the filtration membrane is more remarkable as the circulation flow rate is lower (the stress and damage to the microorganisms are less).

1 medium or reaction raw material 2 culture tank 3 biological culture solution 4 filter 5 filtrate storage tank 6a to 6d MNB generator 7 culture tank agitator 8, 9 pump 10, 11 valve 12 pH adjuster 13 pH adjuster mixing tank A air B Filtrate C Concentrated liquid of microorganisms D D Medium or reaction raw material containing MNB of air A E Biological culture liquid containing MNB of air A F Concentrated liquid of microorganism B containing MNB of air A 101 Fermentation reaction tank 102 Separation Membrane Element 111 Fermentation Broth Circulation Pump 112 Membrane Separation Tank 207 Culture Tank 210 Bacterial Cell Filter 215 MNB Generation Tank 216 MNB Generator 304 Raw Water Supply Line 306 Water Supply Pump 311 Membrane Module 317 Raw Water and/or Concentrated Circulating Water Line

Claims (11)

A culture tank containing a culture medium or a biological culture solution containing a reaction raw material and a microorganism or a cell, a filter for separating the biological culture solution extracted from the culture vessel into a filtrate and a biological culture solution excluding the filtrate, the filtration A bioreactor comprising a pipe for collecting a liquid, and a pipe for refluxing the biological culture liquid excluding the filtrate to the culture tank,
The reaction by microorganisms or cells, characterized in that the biological culture solution supplied to the filter is provided with a micro-nano bubble generator that contains micro-nano bubbles , and the micro-nano bubble generator is provided in the culture tank. A bioreactor for producing a product, or for growing or concentrating a microorganism or cell. The bioreactor according to claim 1, characterized in that the bioreactor further comprises a conduit for returning the filtrate to the culture tank. The micro-nano bubble generator is provided in at least one place of a path for supplying the culture medium or the reaction raw material to the culture tank, the culture tank, and a conduit for returning the biological culture solution excluding the filtrate to the culture tank. The bioreactor according to claim 1 or 2, which is characterized by being provided. The bioreactor according to any one of claims 1 to 3, wherein the micro-nano bubbles are micro-nano bubbles formed from air having an increased oxygen concentration. The bioreactor according to claim 4, wherein the oxygen content of the oxygen-enriched air is 25 to 40%. The bioreactor according to claim 4 or 5, wherein the oxygen-enriched air is obtained by passing air through an oxygen-enriched membrane. The air having an increased oxygen concentration is obtained by mixing oxygen produced by any one of the PSA method, the VSA method and the chemisorption method with air by a line mixer or the like. The bioreactor according to claim 4 or 5. As a pump for delivering a biological culture solution containing microorganisms or cells such as a pump for extracting the biological culture solution from the culture tank, a diaphragm pump, a screw pump, a positive displacement pump such as a rotary pump is used, The biological reaction device according to claim 1. Bioreactor according to claim 8, characterized in that the positive displacement pump is a tube pump. The bioreactor according to any one of claims 2 to 9, wherein a means for adding a pH adjusting agent is provided in a pipe line for returning the filtrate to the culture tank. A bioreaction method, comprising the step of producing a reaction product by a microorganism or a cell, or proliferating or concentrating a microorganism or a cell, by the bioreactor according to any one of claims 1 to 10.

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