JPH0441592B2 - - Google Patents

Description

[Detailed description of the invention]

(Industrial Application Field) The present invention relates to a bioreactor element formed by immobilizing microorganisms used in biochemical reactions. (Prior art) Biochemical reactions using this type of bioreactor emerent are used in a wide range of fields such as organic synthesis, food industry, and analytical chemistry. Various methods are known as means for immobilizing microorganisms (hereinafter referred to simply as microorganisms). By the way, in order to efficiently perform aerobic fermentation using such a bioreactor element, it is desirable to constantly supply air during the reaction process, and it is also desirable to constantly supply air during the reaction process.
Air supply is required during the growth process. (Problem to be solved by the invention) In these processes, it is necessary that the supplied air be applied evenly to the entire microorganism group,
Generally, a means is used to disperse the air supplied from one side using a diffuser plate. However, even with such means, it is difficult to uniformly disperse air, and air can only be applied locally to microorganism groups. Therefore, an object of the present invention is to immobilize microorganisms on an element substrate so as to reliably hold them, and to uniformly provide air to the immobilized microorganism group, thereby facilitating aerobic fermentation, microbial culture, and proliferation. The aim is to improve efficiency. (Means for Solving the Problems) The bioreactor element of the present invention has a first chamber that is partitioned by an inorganic porous element base having air permeability and is supplied with a reaction liquid, and a first chamber that is supplied with air. The device is characterized in that a microorganism that performs a biochemical reaction is immobilized at least on the first chamber side of the pores of the element substrate. Preferably, the element substrate is coated with a partition wall on which microorganisms that perform a biochemical reaction are immobilized at least on the first chamber side of the pore, and a microorganism that does not pass through the microorganism, covering the surface of the partition wall on the second chamber side. It is characterized by having a two-layer structure consisting of a thin film having holes. Furthermore, the bioreactor element of the present invention includes the above-mentioned bioreactor element,
The microorganism is a porous fermentation microorganism and air is supplied to the second chamber during the reaction process and the microbial culture and growth process, or the microorganism is an anaerobic fermentation microorganism and the microorganism is cultured and multiplication process. It is characterized in that air is supplied to the second chamber. Therefore, the element base constituting the bioreactor element according to the present invention is made of an inorganic material such as alumina, silica, silica alumina, zirconia, porous glass, carbon, etc., and has a honeycomb shape, monolith shape, pipe shape, or plate shape. It is formed into an appropriate shape in relation to the reaction vessel. The element substrate is porous and has at least air permeability, and has an average pore diameter of 0.1 to 300 μm, preferably 1 to 100 μm. Average pore diameter is 0.1μm
If it is less than 300 μm, the air supply pressure must be increased, and if it exceeds 300 μm, problems arise in terms of strength. The porosity of the element base is 30-80%,
Preferably it is 40-70%. If the porosity is less than 30%, air supply to the immobilized microorganisms will be uneven, and if it exceeds 80%, strength problems will occur. The thickness of the element base is 0.1~10mm,
Preferably it is 0.3 to 1.0 mm. If the thickness is less than 0.1 mm, strength problems will occur, and if it exceeds 10 mm, the air supply must be at high pressure. The microorganisms used in the present invention are not particularly limited to aerobic or anaerobic, but include bacteria, actinobacteria,
There are molds, yeasts, and fungi. The biochemical reactions carried out by these microorganisms include enzymatic reactions, and contributing enzymes include, for example, glucoamylase, aminoacidase, glucose isomerase, β-galactosidase, cellulase, invertase, asparaginase, aspartase, catalase, Examples include protease, reverse, lysine decarboxylase, hexokinase, tryptophan synthase, and glycerol dehydrogenase. These microorganisms can be immobilized on the element substrate, for example, by the following method. That is, a microorganism containing any of the enzymes described above is suspended in an aqueous solution containing nutrients to prepare a microorganism suspension, and the element substrate described above is immersed in the suspension. Next, in this state, the gas in the pores of the element base is replaced with a microorganism suspension by vacuum degassing, and then the microorganisms in the pores are cultured, grown, and immobilized in the pores. (Operations and Effects of the Invention) In the bioreactor element having such a configuration, the reaction liquid supplied to the first chamber reacts by the action of the enzyme contained in the microorganism immobilized on the element substrate. However, when air is supplied to the second chamber, the supplied air passes through the entire surface of the element base and flows into the first chamber, so the air is distributed evenly to the microorganisms immobilized on the element base. Granted. For this reason, for example, in aerobic fermentation, fermentation efficiency can be significantly improved, and the culture and proliferation of microorganisms can also be significantly improved. Since the microorganisms are immobilized in the pores of the element base, the retention of the microorganisms is ensured, and since the microorganisms are immobilized at least on the first chamber side of the pores, the microorganisms in the reaction liquid are In particular, if the microorganisms to be immobilized are unevenly distributed only in the first chamber side of the pore, it is difficult for the microorganisms to flow out to the second chamber side, and the second chamber is not contaminated by the microorganisms. This has the effect of making it less likely to be damaged. Example 1 Figures 1 to 3 show a reaction apparatus incorporating a bioreactor element according to the first example of the present invention, and Figure 4 shows a partially enlarged view of the bioreactor element. . The bioreactor Emerent 10A is formed of a cordierite honeycomb structure, has an outer diameter of 5 cm, a length of 20 cm, a cell opening length of 2 mm, a wall thickness of the element base 13 of 0.3 mm, an average pore diameter of 25 μm, and a porosity (porosity). 50%. In such an element 10A, as shown in black in FIG. 2, a large number of cells located at predetermined positions among the large number of cells have their bottom openings sealed, and are sealed with the cells 11 whose bottom openings are open. There are two types of cells 12. The former cell 11 (first cell) is the first cell of the present invention.
The reactant liquid is supplied into the cell 11 while it is incorporated in the reaction vessel 20A. The microorganisms are immobilized in the pores 13A of the element substrate 13 by the following method. First, acetic acid bacteria Acetobacter aceti were suspended in preculture medium A (10 g of glucose, 10 g of polypeptone, 10 g of yeast extract, 20 ml of ethanol, and 10 g of glacial acetic acid) at 10 ⁵ cells/ml. Element 10A is immersed in this suspension from the bottom side and left standing at 30° C. and pH 3.3 for 4 days to statically culture the acetic acid bacteria. Next, this element 1
0A into the reaction vessel 20A, each first cell 1
A medium similar to the preculture medium A was supplied into the second cell 1, and air was supplied into each second cell 12, and the cells were cultured for 36 hours while performing aeration. As a result, acetic acid bacteria A are immobilized on the first cell 11 side within the pores 13A of the element base 13 constituting the element 10A, as schematically shown in FIG. 4. The cylindrical body 21 of the reaction vessel 20A is a cylindrical body with a bottom, and the element 10A accommodated in the body 21
is received by the bottom of the main body. Cylindrical body 21
A supply pipe 22 for the liquid to be reacted is exposed at the bottom of the chamber. On the other hand, two chambers are formed in the upper part of the cylindrical body 21, and the first upper chamber _r1 contains the element 1.
Each of the second cells 12 of the element 10A is open, and an outflow pipe 23 for the reaction liquid faces, and each of the first cells 11 of the element 10A is open, and an air supply pipe is opened in the second upper chamber _r2. 24 is coming. In this example, an acetic acid fermentation (aerobic fermentation) test was conducted using the reaction apparatus. In this test, the above-mentioned preculture medium A was used at a temperature of 30°C and a pH of 3.3.
Supplying air at a supply rate of 40ml/hr and at the same time 200ml/hr of air.
It was fed at a rate of ml/min. The steady state concentration of acetic acid produced 10 days after the start of the test was measured and is shown in Table 1. For comparison, the above acetic acid bacteria were comprehensively immobilized on beads with a diameter of 3 mm using 3% by weight sodium alginate using the conventional method, and a bead-shaped bioreactor element was prepared and housed in the reaction vessel 20A at a height of about 20 cm. The same test as above was conducted using this reactor. However, air was supplied from the bottom of the reaction vessel 20A and dispersed using a diffuser plate.

[Table] Example 2 Figures 5 to 7 show a reaction apparatus incorporating a bioreactor element according to a second example of the present invention, and Figure 8 shows a partially enlarged view of the bioreactor element. It shows. The bioreactor emerent 10B is formed of a cordierite honeycomb structure, and the emerent base 18 constituting the bioreactor element 10B has the second chamber side surface of the partition wall 14 coated with an α-alumina thin film 15. It has a two-layer structure. Thin film 1 of this emerent substrate 18
The partition walls 14 except for the element membrane 15 of the first embodiment are the same as the element base 13 of the first embodiment, and have pores 14A.
The thickness of the thin film 15 is 50 μm, and the pores 15
The average pore diameter of A is 1 μm. In this element 10B, the cell 16 (first cell) to which the thin film 15 is not exposed constitutes the first chamber of the present invention, and has an open bottom opening and is connected to the reaction vessel 20.
A reactant liquid is supplied into the cell 16 while it is incorporated in the cell B. Further, the cell 17 (second cell) in which the thin film 15 is exposed constitutes the second chamber of the present invention, and the bottom opening of the second cell 17 is shown in black in FIG. Air is supplied to the cell 17 when the microorganisms are cultured and multiplied while being assembled in the reaction container 20B. Immobilization of microorganisms into the pores 14A of the partition wall 14 of the element base 18 is carried out using the element 10B in the reaction vessel 20.
The reaction vessel 20B is incorporated into the vessel 20B and carried out in the following manner.
The container is substantially the same as the container of the device shown in Publication No. 198383. When immobilizing microorganisms, first, the alcohol-fermented yeast Satucharomyces cerevisiae was added to a culture solution (yeast extract 0.15%, NH ₄ Cl 0.25%, K ₂ PO ₄ 0.55%,
_MgSO4・_7H2O0.025 %, NaCl0.1%, _CaCl2 0.001
%, citric acid 0.3%...all weight%) at PH5.4 10 ⁵
Suspend at 1/ml. The first cell 1 of the element 10B in which such a yeast suspension was incorporated into the reaction vessel 20B
6 and fill it up, and multiply yeast B while supplying air into the second cell 17 for 3 days.
Fix it on the side. In this example, an ethanol production test (anaerobic fermentation) was conducted using the reaction apparatus. In this test, a glucose solution (containing 20% glucose in the above culture medium) was supplied to each first cell 16 from the bottom supply pipe 25 of the apparatus at a rate of 40 ml/hr at pH 5.4.
and reflux through passage 26.
During this time, a portion of the glucose solution flowing through the first cell 16 passes through the partition wall 14 and the thin film 15, flows through each second cell 17 as a reaction liquid, and flows out from the outflow pipe 27 at the top of the apparatus. The yeast immobilized on the partition wall 14 contributes to alcohol fermentation, and the thin film 15 restricts the outflow of free yeast to the second cell 17. After performing this fermentation for 20 consecutive days, a cycle of supplying air into the second cell 17 for 2 days to multiply yeast was performed three times, and the ethanol production concentration and free microorganism concentration in the reaction solution during this cycle were measured. It was measured. The obtained results are shown in Table 2, and as a comparative example, yeast was
The results of using a bioreactor element that is comprehensively immobilized on mm beads are also shown.

【table】

【table】 [Brief explanation of drawings]

FIG. 1 is a partially cutaway perspective view of a reaction device incorporating a bioreactor element according to a first embodiment of the present invention, FIG. 2 is a cross-sectional plan view of the same device in the arrow-line direction, and FIG. A cross-sectional plan view in the arrow-line direction, FIG. 4 is a partially enlarged sectional view of the same element,
FIG. 5 is a partially cutaway perspective view of a reaction device incorporating a bioreactor element according to the second embodiment, and FIG. 6 is a cross-sectional plan view of the same device in the arrow-line direction.
FIG. 7 is a cross-sectional plan view of the same device in the direction of the arrow line, and FIG. 8 is a partially enlarged sectional view of the same element. Explanation of symbols, 10A, 10B... Bioreactor element, 11, 16... First cell (first
(chamber), 12, 17...Second cell (second chamber), 1
3, 18... Element base, 14... Partition wall, 1
5... Thin film, 20A, 20B... Reaction container.

Claims (1)

[Scope of Claims] 1. A first element that is partitioned by an inorganic porous element base having air permeability and to which a reactant liquid is supplied.
1. A pie reactor element comprising a chamber and a second chamber to which air is supplied, and a microorganism that performs a biochemical reaction is immobilized on at least the first chamber side of the pores of the element base. 2. The element substrate has a partition wall in which microorganisms that perform a biochemical reaction are immobilized on at least the first chamber side of the pores, and a thin film that covers the surface of the partition wall on the second chamber side and has micropores that do not pass through the microorganisms. The bioreactor element according to claim 1, which has a two-layer structure consisting of.