JPS58165773A - Apparatus for enzymic reaction - Google Patents

Abstract

PURPOSE:To improve remarkably the reaction efficiency, by means of a combination of a membranous module having membranes, flow path members for a stock solution and flow path members for a permeated solution with an enzyme. CONSTITUTION:A membranous module is of the spiral or pleated type. The pleated type has membranes 311 folded in the long direction in zigzags, flow path members 41 arranged to provide a flow path for a stock solution with one side of each membrane 311 and flow path members 321 arranged to provide a flow path for a permeated solution with the other side of each membrane 311. The membranes 311 and flow path members 41 and 321 are alternately arranged in plural rows through the membranes 311. An enzyme is immobilized in at least one of the membranes 311, flow path members 41 for the stock solution and the flow path members 321 for the permeated solution or an immobilized enzyme is introduced into any one of the gaps between the membranes 311 and the respective flow path members. Alternatively, a mixed state of both is taken to join the enzyme to the membranous module and improve the reaction efficiency.

Description

DETAILED DESCRIPTION OF THE INVENTION Misfire J relates to an enzyme reaction device, particularly an enzyme reaction device using a spiral type or pleated type membrane module.

Conventional enzyme reaction devices FET using immobilized enzymes are mainly flat membrane type (for example, Japanese Patent Application Laid-Open No. 58-10478
No. 5, Japanese Patent Publication No. 55-1023888) and tubular
type (for example, Science and Biology Vol. 16, page 685)
1978)) is known.

All of these conventional devices have low enzyme reaction efficiency per unit volume.

An object of the present invention is to provide an enzyme reaction device with excellent enzyme reaction efficiency per unit volume. Another object of the present invention is to provide an enzyme reaction device that can convert a two-reaction system into a composite enzyme system.

The enzyme reaction device of the present invention uses a spiral LV type membrane module or a pleated type membrane module, and an enzyme is immobilized on at least one of the membrane, the stock solution channel material, and the permeate channel material. . In another device of the present invention, an immobilized enzyme is interposed between the membrane of the membrane module and at least one of the raw solution channel material and between the membrane and the permeate channel material. In still another device of the present invention, an enzyme is immobilized on at least one of the membrane 9 undiluted solution channel material and the permeation channel material of the membrane module, and between the membrane and the undiluted solution channel material and between the membrane. An immobilized enzyme is interposed at least on one side between the permeate channel material and the permeate channel material. That is, the present invention includes a permeable membrane that partitions a first region that forms a flow path for the supplied stock solution and a second region that forms a flow path for the permeate; The present invention provides an enzyme reaction device in which an enzyme is combined with a membrane module provided with a hollow tube with a hole for leading out the enzyme.

The present invention will be explained below.

As shown in FIGS. 1(a) and 1(b), the spiral membrane module/I/1 used in the apparatus of the present invention basically consists of dl, (a) porous hollow tube 2, and (b) two membranes 31
(C) A permeate flow path material 32 placed inside the membrane envelope 3 to make the inside of the membrane envelope 3 a permeate flow path; (d) Membrane envelope 3 (e) A combination of the membrane envelope 3, the permeate channel material 32, and the concentrate channel material 4 is formed into a hollow tube. It is constructed by winding it in a spiral shape. There are two ways to supply stock solution to this type of membrane module. One is.

This is a method in which the stock solution is supplied from the side end of the wound body parallel to the axis of the hollow tube (Japanese Patent Publication No. 1421-1971).

In this case, the last end of the one-wound body is sealed liquid-tightly with an adhesive or the like. Another method is to supply the stock solution from the end of the wound body in a direction perpendicular to the hollow tube axis (
Special Publication No. 52-5431).

In this case, both ends of the one-wound body are sealed liquid-tightly with an adhesive or the like. For any type of membrane, -18 is undiluted solution, undiluted solution, botagasuke 1. Hello liquid.

The permeate then passes through the membrane 31 and passes through the permeate channel material 32.
1, the permeate flows out from the permeate flow path through the hollow tube 2 and out of the thread.

As shown in Fig. M2, the green type membrane module /I/11 used in the device of the present invention basically consists of (a) a membrane 311 folded in a zigzag shape in the longitudinal direction;

(b) An undiluted solution channel material 41 arranged on the membrane surface to make one surface of this membrane 311 a filtrate channel, and (C) to make the other surface of this membrane 311 a permeate channel. (d) A plurality of rows of these stock solution channels and permeate channels are formed alternately through the membrane 311. The membrane edge 310 of the permeate flow path is sealed liquid-tightly with adhesive or the like. The stock solution is supplied from the side end of the folded body, enters the stock solution channel via the stock solution channel material 41, and permeates through the membrane 311, and this membrane permeate passes through the permeated solution channel material 321.
The permeate is configured to flow out of the system from the permeate flow path (via the hollow tube 21).

The pressure of this reactor is determined by the molecular size of the substrate during operation and the degree of separation by the membrane. The membrane is not limited to a reverse osmosis membrane, and any membrane having fractionation performance such as an ultrap-permeation membrane or a precision p-permeation membrane can be used. The material may be any material such as cellulose conductor, saponified ethylene-vinyl acetate copolymer, polyamide, polyimide, etc. The selection can be made depending on whether or not to immobilize the enzyme on the membrane, whether to perform fractionation using the membrane or not, the reaction solvent for the enzyme 2 to be immobilized, the substrate, etc. Ionic bonding is the method of immobilizing the membrane.

Any method such as covalent bonding, inclusion, etc. can be used.

For example, when decomposing starch by immobilizing α-amylase on a membrane, a saponified ethylene-vinyl acetate copolymer having performance as an ultrap-filter membrane is used as the membrane.

Covalent bonding using glutaraldehyde may be employed for immobilization. At this time.

In order to separate the butt/substrate and the reaction product using a membrane, a membrane with a molecular cutoff of 10,000 or less is selected. When only reaction is performed without separation, the 1-fraction molecule source can be freely selected.

As an undiluted solution channel material, it can hold the undiluted solution channel.

Any material may be used as long as it can reduce the concentration polarization of the stock solution. Examples include mensches made of polypropylene, nylon, rayon, vinylon, and the like. In the case of immobilizing enzymes, nylon, rayon, vinylon, etc. are suitable. To immobilize enzymes on rayon 2 For example,
This is done by treating rayon with cyanuric chloride and reacting it with an enzyme.

As the permeate channel material, any material that can maintain the channel even under pressure may be used, and in general.

Cotton, wool felt, nylon, polyester.

Rayon, rayon viscose, acrylic/L/fiber, glass cloth, or polyester mesh treated with a resin such as melamine or epoxide is used. To immobilize an enzyme thereon, for example, a polyester mensch is partially hydrolyzed in an aqueous solution such as sodium hydroxide, and an enzyme is immobilized via the hydroxyl or carboxyl groups formed. In this case, the hydroxyl group is glutaraldehyde.

Terephthalaldehyde, cyanuric chloride, etc. are used.

For carboxyl groups, glutaraldehyde.

Terephthalaldehyde, water-soluble carbodiimide, etc. are used.

For the Spiral A/ffl membrane module, the substrate solution can be supplied to the undiluted solution channel material in parallel to the central axis of the spiral, or from the end of the spiral winding to the undiluted solution channel material. methods can be used.

Also, the immobilized enzyme that is sandwiched between the two pieces is in the form of a piece.

It may be in any form such as w4 fibrous form. The immobilization method may be any method such as ionic bonding or covalent bonding.

As described above, the enzyme reaction device of the present invention has a large contact area between the enzyme and the substrate, so the efficiency of the two reactions is high. moreover,
Since the enzyme is immobilized on each channel material and the immobilized fermentation agent is sandwiched between the membrane and the channel material, the efficiency of one reaction is significantly increased. Furthermore, by immobilizing different types of enzymes, multi-step reactions become possible.

An example of non-explosion will be described below. The present invention is not limited to this.

Example 1 1. Creation of immobilized enzyme membrane A saponified ethylene-vinyl acetate copolymer membrane (80μ film made by casting method from an aqueous solution of Kuraray PVA-H) was cross-linked with glutaraldehyde. ) with 0.1M phosphate buffer to pH 6,
The sample was immersed in a 5% by weight aqueous glutaraldehyde solution at room temperature for 1.5 hours to react. Then, change this to PH
Thoroughly washed with 7.0 phosphate buffer. After washing, this film-like carrier was immersed in a 10% glucose isomerase phosphate buffer solution (PH 7.0) at room temperature for 20 hours to immobilize glucose isomerase.・PH6,
An immobilized glucose isomerase membrane was obtained by thoroughly washing with the phosphate buffer solution described in step 5. Its specific activity was 20%. This film was used as a film for a membrane module for Spiral A/
Incorporated into a type membrane module. Its membrane area is Q, 3gm
It was 2. Add 1% glucose solution to this module.
When supplied at a flow rate of z/min, the fructose concentration in the permeate was 0.01%.

Example 2 An immobilized enzyme (specific activity: 80%) was placed between the membrane of a Spira/L/type membrane module and the stock solution channel material.

This enzyme was prepared by immobilizing siglucose isomerase on a weakly basic ion exchange resin by ionic bonding. As a result, the concentration of 7 lactis in the transmitted wave is 0.
It became 0.5%.

Example 3 A stock solution channel material made of rayon mesh was prepared at pH 6.0.
The sample was immersed in a 5% by weight aqueous glutaraldehyde solution at room temperature for 6 hours to react. this.

Then, it was thoroughly washed with a phosphate buffer solution of pH 7.0. After washing, this mensch-like carrier was immersed in a 10% glucose isomerase phosphate buffer solution (PH 7.0) at room temperature for one day to immobilize glucose isomerase. Its specific activity was 30%.

This was combined with the glucose isomerase fixed membrane of Example 1 to form a module, and a 1% glucose solution was added to it. It was supplied at a flow rate of /min. The 7-lactose bs in the obtained permeate was 0.05%.

Furthermore, when the immobilized enzyme obtained in Example 2 was placed between these membranes and the stock solution channel material, the concentration of 7-lactose in the permeation was 0.08%.

Example 4 After immersing a permeate channel material made of polyester mensch in an IN aqueous sodium hydroxide solution for 1 hour.

Washed thoroughly with water. The obtained partially hydrolyzed polyester mesh was immersed in a 5% by weight aqueous gulguraldehyde solution adjusted to pH 6.0 at room temperature for one day to react, and then thoroughly washed with a phosphate buffer solution of pH 7.0. Next, this mensch-like carrier was mixed with 10% of glucose isomerase.
Glucose isomerase was fixed by immersing it in a phosphate buffer solution (PH7.0) at room temperature for one day. Its specific activity is 2
It was 0%.

Glucoamylase was immobilized on the membrane in the same manner as in Example 1 above by using a 10% phosphate buffer solution (PH 7.0) of glucoamylase on top of the glucose isomerase solution (specific activity 10%). ). α for undiluted solution channel material
- fixed amylase specific activity (40%). A module was created by combining these membranes and stock solution channel material with this permeate channel material on which glucose isomerase was immobilized. The membrane area is 0.38m! Met.

Add 1% starch aqueous solution to this module as a stock solution.
The reaction solution obtained as the permeate through the L roller, which was flowed at a flow rate of //min, contained 0.03% of 7-lactose.

Example 5 In addition to the combination of Example 4 above, glucoamylase immobilized on an ion exchange resin was placed between the membrane and the stock solution channel material, and Ll glucose isomerase immobilized on the ion exchange resin was placed between the membrane and the ion exchange resin. When this was done between the permeate channel materials, the concentration of 7-lactose in the obtained permeate was 0.05%.

Example 6 In addition to the combination of Example 5 above, an immobilized enzyme in which α-amylase was immobilized by ionic bonding to an ion exchange resin was inserted between the membrane and the permeate channel material. .

As a result, the concentration of 7-lactose in the permeate became even higher, O,OS%.

Example 7 An immobilized glucose isomerase membrane obtained in the same manner as in Example 1 was folded in the longitudinal direction, and a stock solution channel material was placed on the front side.
A permeate channel material was placed on the back side to create the module shown in Figure 2. The membrane area was 0.5 m2. When a 1% glucose solution was supplied at a liquid volume of lj'/min,
The concentration of 7-lactose in the permeate was 0.01%. Furthermore, when glucose isomerase was immobilized on the stock solution channel material, the concentration of lactose in the permeate was 0.03%. Furthermore, when an immobilized enzyme in which glucose isomerase was immobilized was placed between the stock solution channel material and the membrane, the concentration of 7-lactose in the permeate was 0.05%.

[Brief explanation of the drawing]

FIG. 1(a) is a partially cutaway perspective view showing an example of a membrane module used in the device of the present invention, FIG. 1(b) is an exploded perspective view thereof, and FIG. 2 is a semi-illustrated diagram showing another example of the membrane module. FIG. 1... Spira/l/type membrane module, 2.21...
・Hollow tube, 31. 311...membrane, 4.41...
- Raw solution channel material, 32° 321... Permeated solution channel material. Agent: Patent Attorney Hidetaka Yamaki

Claims (1)

[Scope of Claims] 1. So that the stock solution entering the stock solution channel through the stock solution channel material permeates the membrane, and the membrane permeate flows out of the system from the permeate channel via the permeate channel material. An enzyme reaction device comprising a membrane module configured such that an enzyme is immobilized on at least one of the membrane, the stock solution channel material, and the permeate channel material. 2. In a membrane module configured such that the stock solution entering the stock solution channel via the stock solution channel material permeates the membrane, and the membrane permeate flows out of the system from the permeate channel via the permeate channel material. , between the membrane and the stock solution channel material. and an enzyme reaction device comprising an immobilized enzyme interposed between at least one of the membrane and the permeate channel material. 3. In a membrane module configured such that the stock solution entering the stock solution channel via the stock solution channel material permeates the membrane, and the membrane permeate flows out of the system from the permeate channel via the permeate channel material. , an enzyme is immobilized on at least one of the membrane, the concentrate channel material, and the permeate channel material; An enzyme reaction device comprising an immobilized enzyme interposed between at least one of the path materials.