JPH0398552A - Liquid for processing edible meat - Google Patents

Abstract

PURPOSE:To increase glutamic acid in edible meat and remarkably improve taste and flavor of the meat by blending a glutaminase in a pickling liquid for processing the meat. CONSTITUTION:The above-mentioned liquid for processing edible meat is prepared by blending a glutaminase in a pickling liquid for processing the edible meat. Salt-tolerant glutaminse having the following physico-chemical properties is preferred as the aforementioned glutaminase. (a) action; hyrolyzing L- glutamine and producing L-glutamic acid and ammonia. (b) substrate specificity; 0.64mM Km value at 37 deg.C and pH6.0 (acetic acid buffer solution) for L- glutaminase, hydrolyzing L-glutamine and D-glutamine without hydrolyzing D- and L-asparagine, Z-glutamine, Dnp-Pro-Gln-Gly and Dnp-Gln-Ile-Ala-Gly-D- Arg. (c) optimum pH and stable pH range; optimum pH6 using L-glutamine as a substrate and stable pH range of 5-8. (d) salt tolerance; exhibiting relative activity of about >=90% in the presence of 18% (wt./vol.) common salt based on that in the absence thereof under conditions of 37 deg.C and pH5.5.

Description

[Detailed description of the invention] Industrial applications The present invention relates to an improved marinating solution for meat processing.

Conventional technology Pickling liquid for meat processing is also called salting liquid or bickle, and is used for the purpose of imparting storage properties to meat, adjusting the flavor and color of meat, improving meat quality, and fixing quality. It is indispensable for the production of processed meat products such as ham, sausage, and bacon. The liquid basically consists of salt and coloring agents such as nitrites, and may also contain coloring aids, binders, preservatives, spices, colorants, seasonings, binding enhancers, etc. depending on the purpose. Various ingredients can be mixed arbitrarily, and various studies have been conducted to improve the composition of the liquid and the meat processing process using the liquid (for example, Japanese Patent Application Laid-Open No. 118841/1984, (See Publication No. 62-29953, etc.).

On the other hand, as a way to effectively utilize livestock meat resources, for example, processing using enzymes such as papain is being carried out to soften meat.
There is currently no research linking such enzyme treatment and the above-mentioned pickling solution for meat processing.

This is due to the specificity of the enzyme itself. In other words, enzymes are substrate-specific and have limits to their working temperature and working pH.
Its activity is inhibited or deactivated by various substances, and its salt tolerance is weak.Therefore, it maintains sufficient activity in a solution like the above-mentioned pickling solution, which has a relatively high salt concentration and contains various ingredients. It is expected that it will be extremely difficult to exert the desired effect on meat.

Problems to be Solved by the Invention The present inventors aim not only to improve the seasoning technology using seasonings, spices, etc., but also to improve the quality of the raw meat itself with respect to the above-mentioned pickling liquid for meat processing. We have conducted extensive research with the aim of developing a new pickling liquid that can meet the trend of health preferences such as low calorie content, low chloride content, preference for natural products, and high quality products. In the process, the taste of meat and meat-like taste (Brolh7 taste)
The expression of 5'-inosinic acid (IMF
) and free amino acids (especially glutamic acid) are said to make a major contribution, and we believe that bringing out the inherent flavor and flavor of meat could be one way to improve meat quality. Based on this idea, we conducted further research. layered. Then, from a new perspective of allowing glutaminase to act on raw meat, we created a pickling liquid for meat processing containing the glutaminase, and salted meat with this liquid.It was truly surprising that in this processing step, Glutaminase in the liquid is very stable and exhibits excellent activity, and it specifically acts only on glutamine in raw meat, converting it to glutamic acid, and thus increasing the amount of glutamic acid, improving the taste and flavor of meat. We have discovered the fact that it can be significantly enhanced.

The present invention was completed based on this new knowledge.

Means for Solving the Problems, that is, the present invention relates to a meat processing pickling liquid characterized by containing glutaminase.

The pickling liquid for meat processing of the present invention can have various conventionally known compositions, except that it contains glutaminase. Typical examples of the liquid include those containing at least one selected from coloring aids, binders, preservatives, seasonings, spices, and binding reinforcing agents in addition to common salt and coloring agents. In the above, the coloring agent is, for example, a nitrite such as sodium nitrite or potassium nitrite, and the coloring aid is, for example, L-ascorbic acid or a salt such as its sodium salt, erythorbic acid or a salt such as its sodium salt, or nicotinic acid amide. etc., as a binder, for example, polymerized phosphates such as sodium pyrophosphate, sodium tripolyphosphate, sodium metaphosphate, etc., as a preservative, for example, salts such as sorbic acid or its potassium salt, etc., as a seasoning. Nucleic acid-based taste substances such as sodium succinate, sodium 5'-inosinate, and sodium 5'-ribonucleotide, saccharides such as sugar, sugar alcohols such as sorbitol, saccharides such as dextrin, etc., and spices such as pepper, Coriander, cucumber, allspice, nutmeg, chili pepper, fennel, kozuishi, sage, thyme, bay leaf, hinge, ginger, onion, garlic, cinnamon bark, etc., and binding agents such as egg white protein, milk, etc. Protein, caseins, gelatin,
Examples include animal and plant proteins such as wheat protein and soybean protein, their partially decomposed products, and starch. Furthermore, it is also possible to blend coloring agents such as tar-based synthetic pigments, vegetable natural pigments, Food Red No. 3 and Food Red No. 4, and edible oils and fats into the above-mentioned meat processing liquid, if necessary.

The amount of each of the above ingredients is not particularly limited,
It is determined as appropriate depending on the type and purpose of the raw meat, but for normal common salt it is about 5 to 20% (wt% to the raw meat, the same applies hereinafter), preferably around 10%, and NaN as a coloring agent.
About 0.01-0.025% for O2, about 0.1-0.2% for NaNO3, about 0.02-0.07% for L-ascorbic acid as a coloring aid, and about 0.02-0.07% for its sodium salt. 0.05-0.07%, about 0 for erythorbic acid
．． 02-0.07%, about 0.05 for its sodium salt
~0.07%, and approximately 0.03-0.07% for nicotinamide.
0.05%, and about 0.05% for the binder. 1-0.5%, usually about 0
．． Approximately 3% or less, preservatives approximately 0. It is appropriate to select from a range of 2% or less. In addition, other ingredients can be added and blended as appropriate; for example, the binding reinforcing agent is about 3
%, spices approx. Suitably, amounts of up to 3%, amino acids as seasonings up to about 0.5%, and sugars up to about 1.0% are used. The use of each of the above components should be determined with consideration given to improving the quality of the raw meat. In particular, in order to bring out the flavor and aroma of the meat itself, the use of the above nucleic acid-based taste substances and spices should be determined. The use of glutaminase should be kept to the minimum possible level, and even without the addition of glutaminase, the present invention enhances glutamic acid in meat through the unique action of glutaminase, which is an essential ingredient, and improves the color and taste of meat. , flavor etc. can be significantly improved, and unpleasant odor of meat can also be reduced.

Glutaminase, which is an essential component of the meat processing liquid of the present invention, is selected from those having the ability to convert L-glutamine into L-glutamic acid, and is particularly stable in the meat processing pickling liquid prepared by blending it. Preferably, those that can maintain and exhibit excellent enzyme activity. The enzymes include, for example, various enzymes originating from microorganisms belonging to the genus Bacillus, Escherichia, and the like. A specific example of the above-mentioned enzyme is the salt-tolerant glutaminase (
An example is Japanese Patent Application No. 1-82444.

The salt-tolerant glutaminase originates from the genus Bacillus, can be obtained, for example, from a commercially available enzyme preparation by the method shown in Examples below, and has the following physical and chemical properties.

a) Action: Hydrolyzes L-glutamine to produce L-glutamic acid and ammonia.

b) Substrate specificity: Km value for L-glutamine is 37°C, pI{6.0
(acetate buffer), and the L-glutamine and D-glutamine are degraded, but D- and L-asparagine, Z-glutamine, Dnp-Pro-Gln-
Gl7 and Dnp-Gln-11e-Ala-Gly
-D1^rg does not decompose.

C) Optimal pH and stable pH range: The optimal pH is 6 using L-glutamine as a substrate, and the stable pH range is 5 to 8.

d) Salt tolerance: 18% (W/V) at 37°C and pH 5.5
In the presence of sodium chloride, it exhibits a relative activity of about 90% or more of that in the absence.

By utilizing the salt-tolerant glutaminase related to the applicant's previous application in the present invention, it is possible to provide a meat processing liquid that can exhibit a particularly excellent effect of improving meat quality.

That is, compared to other glutaminases, this enzyme exhibits excellent resistance to all components of pickling liquids for meat processing, exhibits extremely high salt tolerance, and is specific only to L,D-glutamine. Therefore, it is possible to impart excellent taste, flavor, etc. to processed meat, thereby significantly improving its quality, and has the advantage that it does not have any adverse effect on the texture of the meat.

The amount of various glutaminase, including the above-mentioned salt-tolerant glutaminase, added to the meat processing liquid of the present invention is usually about 0.001 units/g1, preferably about 0.000 units/g, based on the raw meat.
It is desirable to select from the range of 0.005 to 0.05 units/g.

The meat processing liquid of the present invention may be simply a predetermined amount of the above-mentioned glutaminase added and dissolved in water, and is usually used in combination with various conventionally known pickling liquids for meat processing. Available for This combination need not be done at the same time, and it is also possible to apply one of them to the meat first and then the other, or to apply both of them sequentially to the meat. The processing liquid of the present invention is preferably used for meat processing by adding and mixing a predetermined amount of glutaminase into water together with predetermined amounts of various pickling liquid components to form a single agent.

The pH of the meat processing liquid of the present invention prepared in this way does not need to be particularly adjusted, and even if it contains various ingredients, it is usually in the range of about 4.5 to 8, and it can be applied to meat as it is. The action of glutaminase in the liquid can be fully expected. Of course, the pH may be adjusted using a general pH adjusting agent.

A particularly preferred pH is in the range of about 5-6.

The meat processing liquid of the present invention can be applied to raw meat in the same manner as conventional pickling liquids (salting liquids or pickles) of this kind. The meat used as a raw material here may be any of the various meats that can be processed into regular ham, sausage, bacon, etc., such as pork, beef, chicken, horsemeat, mutton, etc. The processing liquid of the present invention can be applied to various forms such as cuts and steaks. A typical example of its application is the so-called wet salting method, in which raw meat is immersed in the above pickle.
There are two methods: pickle cutting) and so-called pickle pumping, in which the pickle is injected into the raw meat using an injector, and either method can be employed in the present invention. The operations and conditions in each of these methods are not particularly different from those in general.

Effects of the Invention According to the use of the meat processing liquid of the present invention, the taste of meat can be significantly improved by enhancing glutamic acid, resulting in meat quality with good flavor. In particular, the processing liquid of the present invention can be used as a salting process for meat, such as beef, whose coloration and taste are completed earlier than the completion of ripening. It can be converted into meat quality with the above-mentioned good flavor, which greatly contributes to the naturalization and high-quality production of processed meat products. In addition, the meat processing liquid of the present invention can also be used to reduce the unpleasant odor of meat.

EXAMPLES Examples will be given below to explain the present invention in more detail.

The glutaminase activity in each example was measured by the following method.

<Glutaminase activity measurement method> 1% (w/v%) L-glutamine 0.5 zll, LM acetate buffer (pH 6.0) 0. lz/, test enzyme solution 0.1
After reacting a liquid consisting of zl and water Q and 3 tll at 37°C for a predetermined time, the reaction was stopped by heat treatment at 100°C for 5 minutes, and after cooling the reaction solution in ice water, a certain amount of L produced in the reaction mixture was reacted. - Glutamic acid is quantified using F-kit L-glutamic acid (manufactured by Boehringer Mannheim Yamanouchi).

One unit (U) of this enzyme is the amount of enzyme that produces 1 μmol of glutamic acid per minute at 37°C.

Example 1 Salt 4.5g, sodium nitrite 0. 0 1 8
g, sodium tripolyphosphate 0.68 g, sodium L-ascorbate 0.09 g, and potassium sorbate 0.09 g. A predetermined glutaminase is added to 451 liters of an aqueous solution (salting solution) and mixed thoroughly to produce the meat processed according to the present invention. A solution was prepared.

The above-prepared meat processing liquid was added to 100 g of minced beef, mixed, and then stored at 4°C. After a certain period of time, add the same amount of water as the meat, process at 100℃ for 15 minutes, and then cool with water.
Centrifugation was performed at 3500 rpm for 10 minutes, and the amount of L-glutamic acid in the resulting supernatant was measured.

In addition, as the glutaminase, the following A-C were used,
The salting solutions prepared by adding each glutaminase are designated by the same symbols.

A... Salt-tolerant glutaminase derived from the genus Bacillus B... Glutaminase derived from the genus Bacillus (derived from Proleather) C-... Escbetichia
Bacillus-derived salt-tolerant glutaminase of Doki A was obtained by the following method.

That is, 1300 g of the commercially available enzyme preparation "Protin M3
．． 5), thoroughly dialyzed against the same buffer, and centrifuged (12,000 rpm x 15 minutes) to remove insoluble matter.
DEAE-cellulose force ram (H7.5) equilibrated with
5.6 x 27 cm, manufactured by Wako Pure Chemical Industries), and 20 m
M}Lis-HCl buffer (pH 7.5, O, IM NaC
After washing with 20mM Tris-HCl buffer (pH
7.5, containing 0.3M NaCA') to elute glutaminase. Next, ammonium sulfate was added to the obtained 0.3M NaCA' elution fraction containing glutaminase at a concentration of 0.8 saturation, left overnight at 4°C, and then centrifuged (12,
OOOrpm x 15 minutes) to collect the glutaminase active fraction.

The ammonium sulfate precipitate obtained was again diluted with 20mM phosphate buffer (pH
7.4) and dialyzed against the same buffer, then hydroxyl apatite (2.8x
20cm, Naca!ai tesq
UE), 100-350 mesh), washed with the same buffer, and then 20mM phosphate buffer (pH 7.4).
400mM phosphate buffer (pH 7.4) while continuously increasing the buffer concentration, and the buffer concentration was 350mM.
The glutaminase active fraction eluted in
DEAE-Toyopearl (dialyzed against Tris-HCl buffer (pH 7.5) and equilibrated with the same buffer)
1.6 x 7.5 cm (manufactured by Tosoh), washed with the same buffer, and then eluted stepwise with 10mM Lis-HCl buffer (pH 7.5) containing 0.05-0.25M NaCA'. The elution fraction of 0.15M NaCA' was further added for 20m
M}Lis-HCl buffer (pH 7.5, 0.2M NaC
/ containing) on a Sephacryl S-200 column (2.8 x 4 1 Cm, manufactured by Pharmacia),
2 yl portions were collected at a rate of 12'yl/hour.

Finally, the active fractions were collected and dialyzed against 10 mM phosphate buffer (pH 6.0) containing 25% ammonium sulfate.
Phenirucef 70-SCL pre-equilibrated with the same buffer
-4B (1.6 x 5 cm, manufactured by Pharmacia), and after washing with the same buffer, the ammonium sulfate concentration was reduced to 25
% to 15% (W/V) while increasing the ethylene glycol from O to 20% (V/V) to elute the glutaminase to obtain the desired salt-tolerant glutaminase.

As a result of disk electrophoresis using 7% gel at pH 8.0, it was revealed that this substance was single, and its specific activity was l
It was 7 units/mg.

The above glutaminase B was prepared using DEAE Toyopearl (Tosoh Co., Ltd.), which was obtained by equilibrating a commercially available Proleather (Amano Pharmaceutical Co., Ltd.) solution that had been thoroughly dialyzed against 10 mM Tris-HCl buffer (pH 7.2) with the same buffer. After washing and removing impure proteins with the same buffer, the glutaminase was eluted with the same buffer containing 0.3M NaCl, and the active fraction was collected at 0.3M NaCl. 8
A gel was obtained by salting out and recovering with saturated ammonium sulfate, and then applying the gel to a Sephacryl S-200 column (manufactured by Pharmacia) equilibrated with 10 mM phosphate buffer (pH 7.2). According to the above procedure, a sample that does not contain protease and has a specific activity increased by 274 times can be obtained.

Furthermore, as a control, the same test was conducted using the above-mentioned salt pickling solution containing no added glutaminase.

The results of each of the above tests repeated twice are shown in Table 1 below.

Chapter 1 Table Table l (continued) 1 indicates the amount of enzyme added to 1 g of meat.

From Table 1 above, it is clear that the amount of L-glutamic acid increases significantly when the meat processing liquid containing glutaminase of the present invention is used.

This effect of increasing the amount of L-glutamic acid is particularly remarkable in the case of the glutaminase derived from the genus Bacillus shown as A, and this enzyme exhibits the desired enzyme activity very stably in the meat processing liquid of the present invention. I know what I can get.

Example 2 Salt 5% (w/v%, same hereinafter), sodium nitrite 0604%, sodium tripolyphosphate 1%, sodium L-ascorbate 0.05%, and potassium sorbate 0.2% based on meat weight The meat processing liquid of the present invention was prepared by mixing 0.004 U/g meat of the salt-resistant glutaminase derived from the genus Bacillus shown as A in Example 1 into the salting liquid consisting of %.

Add 0.7 g of the meat processing liquid prepared above to 5 g of beef block.
5zA' was injected into the meat using a syringe and left at 4°C for 5 days. Next, add the same amount of water to the processed meat and boil it to 100
After heat treatment at ℃ for 15 minutes, water cooling and 3500 rpm
Centrifugation was performed for 10 minutes, and the amount of L-glutamic acid in the resulting supernatant was measured.

In addition, as a control, the same test was conducted using the above-mentioned salt pickling solution that did not contain glutaminase.

The results obtained by repeating the above test three times for each liquid are as follows:
The results are shown in Table 2 in the same manner as in Table 1.

Table 2 That is clear.

Example 3 20g each of thinly sliced pork, chicken, and beef, 33g of water
21 l and 32xl of 27 mM NaN were added, and glutaminase (enzyme A described in Example 1) was added in an amount of 0.01 U per 1 g of meat, and the mixture was allowed to stand at 4°C for 5 days. The processed meat liquid was then heat treated at 100° C. for 15 minutes, cooled with water, centrifuged at 3500 rpm for 15 minutes, and the amount of L-glutamic acid in the resulting supernatant was measured.

Further, as a control, the same treatment as above was performed except that glutaminase was not used.

The results of the above test are shown in Table 3.

As shown in Table 2 above, according to the present invention, the amount of L-glutamic acid increases by about 4 times compared to the control. is recognized.

Example 4 Salt 15% (w/v%, same below), sodium nitrite 0.04%, sodium L-ascorbate 0.15%
, sodium polyphosphate 0.25%, sodium pyrophosphate 0.1%, sodium acid pyrophosphate 0.06%,
Potassium sorbate 0.3%, sodium caseinate 1%
, egg white powder 1%, sugar 1%, white pepper 0.4%
IU of glutaminase derived from the genus Bacillus shown as A in Example 1 to 50 zl of pickle having a composition consisting of
A meat processing liquid of the present invention was prepared by mixing the following.

5021 of the pickle prepared above to 100g of pork thigh
After injecting (glutaminase O.OIU/g meat), tumbling was performed and salting treatment was performed at 4°C for 5 days.

Next, the same amount of water as the processed meat was added, heat treated at 100°C for 30 minutes, cooled with water, and the soup portion was heated to 350°C.
After centrifugation at rpm for 10 minutes, the amount of L-glutamic acid in the resulting supernatant was measured.

In addition, as a control, the same treatment was performed using the same set of pickles as above, except that glutaminase was added.

Table 4 shows the results obtained by repeating the above test twice.

Table 4 From Table 4 above, it is clear that the use of the pickle of the present invention can significantly increase the amount of glutamic acid.

(that's all)

Claims (3)

[Claims] (1) A pickling liquid for meat processing, characterized in that it contains glutaminase. (2) The meat processing liquid according to claim (1), wherein the glutaminase is a salt-tolerant glutaminase having the following physical and chemical properties. a) Action: Hydrolyzes L-glutamine to produce L-glutamic acid and ammonia. b) Substrate specificity: The Km value for L-glutamine is 0.64 mM at 37°C and pH 6.0 (acetate buffer);
- Glutamine and D-glutamine are degraded, but D- and L-asparagine, Z-glutamine, Dnp-Pro
-Gin-Gly and Dnp-Gln-lle-Ala
-Gly-D-Arg does not decompose. c) Optimal pH and stable pH range: The optimal pH is 6 using L-glutamine as a substrate, and the stable pH range is 5 to 8. d) Salt tolerance: 18% (w/v) at 37°C and pH 5.5
In the presence of sodium chloride, it exhibits a relative activity of about 90% or more of that in the absence. (3) Claim (1) wherein the pickling liquid for meat processing contains at least one selected from coloring aids, binding agents, preservatives, seasonings, spices, and binding reinforcing agents in addition to salt and coloring agents. ) or the meat processing liquid described in (2).