JPH0822229B2 - Stabilized enzyme solution - Google Patents

Description

TECHNICAL FIELD OF THE INVENTION The present invention relates to a stabilized aqueous enzyme solution, more specifically,
The present invention relates to an aqueous solution of α-amylase which can be stored for a long period of time in the form of an aqueous solution, and also has little deterioration in activity and improved stability.

Conventional technology Enzyme solution products have the advantages over powder products that they do not have to be redissolved during handling, do not have the problem of dust scattering, and can be injected with a metering pump, saving the labor of metering and addition. Therefore, it is widely used in the industry that uses large amounts of enzymes. In general, however, such an enzyme product in the form of an aqueous solution is remarkably unstable and has poor storage stability as compared with a powder product, and particularly when the liquid temperature rises in the summer and the like, it is easily deactivated during transportation and storage and cannot be used. There's a problem.

Conventionally, in order to enhance the storage stability of the enzyme aqueous solution as described above, it is known to add an antiseptic agent such as salt, but the antiseptic agent originally prevents the spoilage of the enzyme and loses it. It suppresses the activity and cannot improve the stability itself of the enzyme. Further, it has been empirically known that polyhydric alcohols such as sucrose and glycerol have an effect of stabilizing the enzyme, but
The stabilizing effect is not fully satisfactory. Further, a stabilizer which is a combination of the above-mentioned polyhydric alcohols and gelatin and / or casein and ethyl alcohol has been proposed (see Japanese Patent Publication No. 41-152). However, the proposed stabilizer usually requires a total amount of about 20% by weight or more to be added to the aqueous enzyme solution in order to achieve the desired stabilizing effect, which is of course economically disadvantageous. However, it is unavoidable that a large amount of the stabilizer component is mixed into various products obtained by using the enzyme aqueous solution in which it is present, which is not preferable in this respect.

Problems to be Solved by the Invention An object of the present invention is to eliminate the drawbacks found in the above-mentioned conventional techniques for improving the stabilization of enzyme aqueous solutions, and in particular, to achieve excellent stability-improving effects even when used in small amounts. It is intended to provide a new stabilized α-amylase aqueous solution product to which an agent is added and blended.

Means for Solving the Problems According to the present invention, there is provided a stabilized enzyme aqueous solution characterized by containing 0.01% by weight to a saturated concentration of methionine in the α-amylase aqueous solution.

Since the enzyme aqueous solution of the present invention contains a predetermined amount of methionine as described above, its storage stability is remarkably improved. In fact, for example, as shown in Examples below, the methionine-added α-amylase-derived Bacillus subtilis culture solution of the present invention shows a residual activity of 74 to 90% after being left at 40 ° C. for 3 months. It retains, and it has less deactivation about twice as much as methionine-free (residual activity 46%) and has very excellent storage stability. However, according to the studies conducted by the present inventors, the stabilizing effect is peculiar to methionine, and histidine, glycine, lysine, sodium glutamate, etc., which are amino acids similar to methionine, are shown in Comparative Examples below. As described above, it has been found that the stabilizing effect of the aqueous enzyme solution is not substantially recognized.

Since the enzyme aqueous solution of the present invention has excellent stability as described above, it can be stored for a long period of time without requiring storage in a cold place even under high temperature conditions such as summer, The activity of the initial production can be continuously expressed without being inactivated even by the above.

It is essential that the aqueous enzyme solution of the present invention contains a predetermined concentration of methionine. Here, as methionine, D
Type, L type and DL type (racemic mixture) may be used, and no substantial change is observed in the stability improving effect on the aqueous enzyme solution among them. The amount of methionine added to the enzyme aqueous solution is selected from the range of 0.01 wt% to its saturation concentration (which varies depending on the enzyme aqueous solution and the liquid temperature, but is usually about 3 wt% for DL-methionine). The higher the concentration is, the more the stabilizing effect due to the addition thereof is improved, but the concentration and the effect are not in proportion to each other, and the improvement tendency of the effect becomes less remarkable as the concentration becomes higher. Therefore, it is more practical to usually select the amount of methionine added from the range of about 0.5 to 2.0% by weight.

Further, in the present invention, α to which the above methionine is added
-The amylase aqueous solution is not particularly limited, but a sterilized α-amylase aqueous solution is preferable. The aqueous solution may be a crude enzyme solution prepared according to a conventional method such as a bacterial culture solution or an enzyme extract, and a conventional method from the crude enzyme solution, for example, ion exchange chromatography,
It may be a purified liquid purified by a method such as gel filtration or adsorption. A particularly preferred sterilized α-amylase aqueous solution is Bacillus subtilis.
Examples of the culture solution, its purified solution, and the like. The aqueous enzyme solution may contain an antiseptic such as salt, etc., as it is usually commercially available. Further, not only the aqueous solution form, but also a paste or cream containing the aqueous enzyme solution. It may have a shape such as a shape.

In using the enzyme aqueous solution of the present invention, it is of course desirable to adjust the enzyme aqueous solution to a stable pH range of the enzyme.

Examples Hereinafter, examples and comparative examples will be given in order to explain the present invention in more detail. Unless otherwise specified as methionine
DL-methionine was used.

Example 1 As an aqueous enzyme solution, a culture solution of α-amylase derived from Bacillus subtilis containing 20% of sodium chloride (6100 units (L _J
/ g), pH 6.6) 3 methionine 0.05%, 0.1%, 0.5%,
1.0% and 2.0% (both by weight, the same applies below) were added and mixed to obtain the stabilized enzyme aqueous solution of the present invention.

A stability test was conducted on each of the aqueous enzyme solutions of the present invention (Samples 1 to 5) obtained above. That is, each of them is 40 ℃
Were stored for 1 month to 3 months, the enzyme activity after that was determined, and the residual activity (%) was calculated.

The results are shown in Table 1 below. In addition, Table 1 also shows the results for the enzyme aqueous solution without methionine as a control.

Comparative Examples 1 to 16 In Example 1, histidine 0.5 was used instead of methionine.
%, Glycine 0.5%, lysine 0.5% and sodium glutamate 0.5% were added to the same enzyme aqueous solution and mixed to obtain comparative enzyme aqueous solutions (Comparative Samples 1 to 4).

The same stability test as in Example 1 was performed on these comparative aqueous enzyme solutions. The results of the residual activity after storage at 40 ° C. for 1 month are shown in Table 2 below.

Example 2 As an enzyme aqueous solution, a purified solution of Bacillus subtilis-derived α-amylase containing 20% of sodium chloride (a solution obtained by elution and purification after starch adsorption, 11000 units (AU / g), initial pH 5.7) was used. Methionine 0.1%, 0.25%, 1.0% and 2.0%
Were added and mixed to obtain stabilized enzyme aqueous solutions of the present invention (Samples 1 to 4).

Table 3 shows the results of a stability test conducted on each of the samples obtained above in the same manner as in Example 1.

Example 3 8% salt and 2.5% calcium chloride as an enzyme aqueous solution
And a culture solution of α-amylase derived from Bacillus subtilis containing 0.25% cresol (3300 units (AU / g), pH
6.1), 0.1%, 0.25%, 0.5%, 1.0 of L-methionine
% And 2.0% were respectively added and mixed to obtain the stabilized enzyme aqueous solution of the present invention.

The stability test results for each of the aqueous enzyme solutions (inventive samples 1 to 5) obtained above are shown in Table 4 below.

Example 4 In Example 3, an aqueous solution of the enzyme as a starting material was added to a culture solution of Bacillus subtilis-derived thermostable α-amylase containing 1000% (L _J / L /) containing 8% sodium chloride, 2.0% calcium chloride and 0.2% cresol. g), pH 6.7), except that the stabilized enzyme aqueous solution of the present invention was obtained in the same manner.

The results of the same stability test conducted on each of these are shown in Table 5 below.

Claims (1)

[Claims] 1. A stabilized enzyme aqueous solution comprising 0.01% by weight to a saturated concentration of methionine in an α-amylase aqueous solution.