JPH04335892A - Continuous fermentation of ethanol by immobilized yeast - Google Patents

Abstract

PURPOSE:To stably obtain ethanol in high conversion efficiency over a long period by immobilizing yeast on the surface of a porous regenerated cellulose hollow fiber using repeated freezing and thawing process with PVA, packing the hollow fiber into a column and passing a glucose-containing culture liquid and air through the column. CONSTITUTION:An aqueous dispersion slurry of baker's yeast is added to a 7.5% aqueous solution of a polyvinyl alcohol and stirred at room temperature. A porous hollow fiber of regenerated cellulose is immersed in the obtained liquid mixture and cooled by spraying liquid nitrogen. The above operation is repeated thrice. The hollow fiber holding the yeast immobilized by the above repeated freezing and thawing process is cut to 5 cm long and packed in a jacket column. A culture liquid containing glucose as a substrate is introduced together with air into the column from the bottom while keeping the column temperature to 37 deg.C to effect the continuous fermentation. Ethanol can be stably produced in high conversion efficiency over a long period by this process.

Description

[Detailed description of the invention]

0001

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a continuous fermentation method for ethanol for consumption. More specifically, the present invention relates to a continuous fermentation method for producing ethanol continuously, highly efficiently, and stably over a long period of time in an ethanol fermentation process.

0002

BACKGROUND OF THE INVENTION At present, most of the ethanol for drinking and drinking is produced by the fermentation method. In this production, shortening the fermentation time and increasing the efficiency of the fermentation process by recovering and reusing yeast are important issues. Furthermore, in order to stabilize quality, it is desired to develop a continuous fermentation process under steady state conditions. In response to these demands, continuous fermentation processes using immobilized yeast have begun to attract attention.

[0003] Specifically, as a method for immobilizing yeast, a method has been adopted in which yeast is embedded in a gel using a polymer gelling agent such as sodium alginate or K-carrageenan. Although a carrier for immobilization is not particularly required in this case, granular or fibrous gel shapes have been proposed. In order for the ethanol fermentation process using immobilized yeast to be put into practical use on an industrial scale, at least (1) the conversion efficiency to ethanol must be at least 90%; (2)
The concentration of alcohol produced is 4% or more (
3) Has excellent yeast growth ability, (4) The performances of (1) to (3) above are stably maintained over a long period of time (for example, for 3 months or more), (5) Satisfies (1) above. The equivalent supply rate (hereinafter referred to as SV) (≡supply rate of culture solution containing substrate/column volume) under the conditions of The following conditions must be satisfied: (hereinafter referred to as STY) (≡ethanol production rate (kg/hr)/gel volume (m3 gel)) is large (for example, 20 kg/m2 ·hr or more).

[0004] With the current immobilization method using alginic acid, it is difficult to satisfy the conditions (1) to (6) above at the same time. Moreover, with a polymer gelling agent such as alginic acid, unless divalent metal ions such as calcium ions are constantly supplied, the polymer gel will dissolve and yeast will flow out. Contamination of calcium ions is not desirable, as it may result in contamination with the alcohol produced, or may impede satisfaction of the conditions (1) to (6) above.

[0005]

[Problems to be Solved by the Invention] The present invention relates to the above-mentioned SV
It is an object of the present invention to provide a method for continuous fermentation of ethanol for drinking and drinking that simultaneously satisfies the following conditions: a large value of STY and a large value of STY.

[0006]

[Means for solving the problem] The polymer gelling agent used for yeast immobilization becomes an obstacle to the diffusion of various components in the culture solution, including the substrate, into the yeast after immobilization, and the product It also becomes an obstacle to the diffusion of ethanol and carbon dioxide gas from the yeast into the culture solution. Therefore, (4) and (5) above
In order to satisfy the conditions (6), it is necessary to reduce the content of the polymer component in the gelling agent. A decrease in the content of the above-mentioned polymeric components increases the tendency of yeast to flow out. Therefore, there is an optimum concentration of the polymer component in the gelling agent.

However, the present inventors have discovered that these problems can be solved all at once by immobilizing yeast on the surface of a porous polymer having a special structure, leading to the present invention. That is, the present invention is a method of immobilizing yeast by repeated freeze-thaw method using polyvinyl alcohol. After immobilizing the yeast on the surface of porous regenerated cellulose hollow fibers, a column is filled, and glucose is injected from the bottom of the column. This is a continuous ethanol fermentation method characterized by flowing a solvent solution used as a substrate and air.

The most important feature of the method of the present invention is that yeast is immobilized on the surface of porous regenerated cellulose hollow fibers and then packed into a column. As a fixation method, a repeated freeze-thaw method using polyvinyl alcohol is preferred. By this method, yeast is immobilized on the outer wall surface or inner wall surface of the regenerated cellulose hollow fiber. It is thought that the yeast is immobilized thinly on the surface of the porous membrane, and because the membrane is a hollow fiber membrane, the rate of diffusion of substances into the yeast is significantly increased.

[0009] That is, by immobilizing it on the porous membrane surface, the STY value increases significantly. Further, by forming the fiber into a hollow fiber, the SV value increases, and at the same time, these values are stably maintained at a high level for a long period of time. The material for the porous hollow fibers used in the present invention is preferably a hydrophilic polymer, particularly regenerated cellulose, and more preferably ammonium ammonium regenerated cellulose. The reason for this is not clear, but since the long-term stability of ethanol conversion efficiency is particularly excellent, it is probably related to the fact that adsorption of yeast products is less likely to occur. The carbon dioxide produced can be easily removed from the system by forming into hollow fibers.

[0010] That is, in the case of hollow fibers, gaseous carbon dioxide gas continues in the axial direction of the fibers and escapes from the system. On the other hand, when the carbon dioxide gas is immobilized in the form of granules, the carbon dioxide gas bubbles are retained between the granules and cannot be removed from the system. Therefore, in the long term, the carbon dioxide concentration in the culture solution increases and the rate of alcohol production decreases.

It is particularly preferable that the hollow fibers have an inner diameter of 200 μm or more. If the inner diameter becomes extremely small, the results will be similar to those obtained when yeast is immobilized on the fiber surface,
In order for the characteristics of the hollow fiber shape to be exhibited in the fermentation reaction, the inner diameter is preferably 50 μm or more, more preferably 200 μm or more. The rate at which the substrate or active ingredient in the culture solution diffuses into the yeast, or the rate at which the produced carbon dioxide and ethanol are diffused out of the system, is determined by the rate at which the substrate or active ingredient in the culture solution diffuses out of the system. is strongly influenced by the rate of diffusion.

[0012] Therefore, it is preferable that the average pore diameter and porosity of the hollow fibers be relatively large. Experimentally, when the porosity of the hollow fiber is 30% or more and the average pore diameter of the hollow fiber calculated from the water filtration rate is 0.01 μm or more, it is characterized as a porous hollow fiber (condition (4) above). (to be satisfied) is demonstrated and desirable. Another feature of the method of the present invention is that
As a method for supplying the culture solution and air as a substrate to the immobilized yeast, it is preferable to forcefully flow into the column from the bottom of the column using a pump or the like. By introducing air from the bottom, it is easy to supply oxygen to the yeast and remove carbon dioxide from the production system.

[0013] At this time, if granules or the like having a relatively non-anisotropic shape are used as the immobilization carrier, air bubbles will remain between these granules, making it impossible to efficiently convert oxygen and carbon dioxide gas. . Therefore, the ethanol conversion efficiency and the alcohol concentration produced decrease. On the other hand, by flowing the culture solution from the bottom, the rate of bubble retention is reduced and the ethanol concentration in the collected solution is increased. It is necessary to control the pH of the culture fluid flowing into the column to around 4.0 to prevent bacterial contamination.

However, if a copper ammonium regenerated cellulose membrane with virus removal performance is installed at the inlet and only the filtrate is allowed to flow in, the pH will be optimal for yeast growth and conditions suitable for alcohol production. It becomes possible to do so. At this time, it is necessary that the incoming air be filtered in advance through the copper ammonium regenerated cellulose membrane. Here, the copper ammonium method regenerated cellulose membrane has an average pore diameter of 20-100 nm, a porosity of 30-60%, an inner diameter of 200-500 μm, and a wall thickness of 25-100 nm.
A 50 μm hollow fiber membrane is preferred.

[0015]

[Example] The present invention will be explained in detail below using examples.

[0016]

[Example 1] 7.5% of polyvinyl alcohol (hereinafter referred to as PVA, manufactured by Kuraray Co., Ltd., trade name Opal 117)
An aqueous dispersion slurry of baker's yeast (baker's yeast concentration: 30 g of baker's yeast/50 g of water) was added to 220 g of the aqueous solution and mixed with stirring at room temperature. The resulting mixed solution had an average pore size of 35 nm and an inner diameter of 3.
00 μm, wall thickness 35 μm copper ammonium regenerated cellulose hollow fibers (manufactured by Asahi Kasei Corporation, trade name BMM35) were immersed.
-5 at a cooling rate of 0.255℃/min by spraying liquid nitrogen
Cooled to 0°C. This operation (freeze-thaw operation) was repeated three times.

[0017] The thus obtained hollow fiber with immobilized yeast was cut into a length of approximately 5 cm, and this was cut into a 25φ×500
The mixture was packed into a L jacketed column. Column temperature 37℃
It was set to 10% glucose from the bottom of the column,
Yeast extract 60ppm, malt extract 60ppm
m, a solvent solution containing 100 ppm of peptone (pH = 4.0
) and an air filter, and the air was allowed to flow out from the bottom at a rate of 1 ml/min.

The concentration of the produced alcohol was measured using a gas chromatograph (manufactured by Shimadzu Corporation, trade name: Gas Chromatograph G).
C-PA, column packing material Polaropak QS).

[0019]

[Table 1]

[0020]

[Comparative Examples 1 to 4] As a comparative example, when PVA was used as a gelling agent and cotton thread was used as an immobilization carrier, (Comparative Example 1)
The same operation as in Example 1 was carried out for the case where the gel produced by the gelling agent was granulated without using a carrier (Comparative Example 2) and the case where Ca-alginic acid was used as the gelling agent (Comparative Examples 3 and 4). Alcohol was produced under certain conditions. Table 1 shows the produced ethanol concentration, ethanol conversion efficiency,
Actual values of SV value, STY value, and steady state duration time are shown.

[0021]

[Effects of the Invention] According to the continuous ethanol fermentation method of the present invention, the SV value and STY value are improved. Furthermore, superior results can be obtained compared to conventional methods in terms of the concentration of ethanol produced, the efficiency of ethanol conversion, and the duration of steady state. Especially the value of SV, STY
The effect of improving the value of , the duration of steady state is very large. Therefore, the method of the present invention enables continuous fermentation of ethanol with high efficiency.

Claims (1)

[Claims] Claim 1: In a method of immobilizing yeast using polyvinyl alcohol by a repeated freeze-thaw method, the yeast is immobilized on the surface of a porous regenerated cellulose hollow fiber and then packed into a column, and glycose is added to the substrate from the bottom of the column. A continuous ethanol fermentation method characterized by flowing a culture solution and air.