KR20050025138A - Feed for fry young fishes and method of producing hydrolyzate of low-phytin vegetable protein to be used therein - Google Patents

Abstract

It is intended to provide a feed which is appropriate for enhancing the survival rate of young fry fishes with underdeveloped digestion and absorption functions or promoting the growth of them. It is also intended to obtain a vegetable protein hydrolyzate to be used in this feed. To obtain a low-phytin vegetable protein hydrolyzate, a process for producing the low-phytin vegetable protein hydrolyzate which comprises (a) the step of digesting a protein with the use of a protease and (b) the step of digesting phytic acid with the use of an enzyme digesting phytic acid has been completed. In the case where the vegetable protein is soybean protein, for example, it is preferable that the low-phytin soybean protein hydrolyzate has an average molecular weight of from 200 to 10,000. It is also preferable that the phytic acid content is 0.05% by weight or less (based on dry solid matters). A feed for young fry fishes containing this low-phytin soybean protein hydrolyzate makes it possible to provide a feed for young fry fishes which promotes the growth of young fry fishes and enhances the survival rate.

Description

FEED FOR FRY YOUNG FISHES AND METHOD OF PRODUCING HYDROLYZATE OF LOW-PHYTIN VEGETABLE PROTEIN TO BE USED THEREIN}

The present invention provides a fine feed for fry using low-phytin vegetable protein hydrolyzate with reduced phytic acid. Moreover, it is related with the manufacturing method of the low phytin vegetable protein hydrolyzate which reduced this phytic acid.

Background Art Conventionally, soybean raw materials (soybean sesame seeds, soy milk, soybean protein, etc.) have been used as protein raw materials for fish feed. It is also known that the removal of phytic acid from soybeans increases feed efficiency.

As the invention for removing or decomposing phytic acid, for example, (a) feed (from Japanese Patent Application Laid-Open Nos. 11-000164 and 8-205785), (b) soybean derived from soybeans, bean curds and the like Feed material (Japanese Patent Laid-Open No. 9-140334), (c) soy protein (Japanese Patent Laid-Open No. 2000-300185 and Japanese Patent No. 4503002) or (d) Soymilk (Japanese Laid-Open Patent Publication 59- Japanese Patent Application Laid-Open No. 166049, Japanese Patent Application Laid-Open No. 2000-245340 by the applicant of the present application), and the like.

However, the use of low phytin protein hydrolyzate in fish farming has not been implemented.

On the other hand, the present applicant has studied the use of a protein hydrolyzate obtained by hydrolyzing soy protein raw material using an enzyme as a feed for fish farming. For example, the invention described in Japanese Patent Application Laid-Open No. 7-227223, the invention described in Japanese Patent Application Laid-Open No. 8-51937, etc. have been performed as a food for increasing the survival rate of small fry. And further, to improve the survival rate of small fry, soybean protein hydrolyzate was studied.

On the other hand, regarding soy protein hydrolyzate, the present applicant has disclosed the low phytin vegetable protein hydrolyzate from which phytic acid was removed using resin for patients with kidney disease (Japanese Patent Laid-Open No. 8-092123).

On the other hand, inoculum is inoculated with soybean protein and fermented to produce prophylactic and phytase-treated low-pitin vegetable protein hydrolysates (Japanese Patent Laid-Open No. 9-023822).

In addition, proteolytic enzymes and phytases work together when enzymes are used to enzymatically process soy protein raw materials, or some combinations of proteolytic and phytase treatments exist. There is neither disclosure nor teaching about the use of proteins for fish farming (Japanese Patent Laid-Open Nos. 51-125300 and 2002-51706).

The use of soy protein hydrolyzate in fish farming as described above has been studied by the present applicant, and low pitin soy protein hydrolyzate has been studied, but it is known to use them in fish farming feed, especially small fish. Not.

By the way, the method of removing phytic acid of soybean is (1) removing phytic acid in the water extraction process of soy protein using a salt (JP-A-8-173052 and 9-121780). Etc.), (2) a method of decomposing phytic acid with phytase, and (3) a method of adsorbing and removing a resin by adsorbing to a resin or the like (Japanese Patent Laid-Open No. 2001-163800), and the method of (1) and (3). On the other hand, the method of (2) is industrially advantageous because the process is not complicated.

(2) As for the method of decomposing phytic acid using phytase and the like, (a) soybeans and skim soybeans are widely known as feeds, and (b) soybean milk or separated soy protein. Although (c) vegetable protein hydrolyzate mixtures are not known.

For example, (a) feed (Japanese Patent Laid-Open Publication No. 11-000164 and Japanese Patent Laid-Open Publication No. 8-205785), and (b) soybean-derived feed materials such as skim soybean and bean curd (Japanese Patent Laid-Open Publication No. 9-140334). G), soybean protein (Japanese Patent Laid-Open No. 2000-300185 and Japanese Patent No. 4503002) or Soymilk (Japanese Patent Laid-Open No. 59-166049, Japanese Patent Laid-Open No. 2000-245340 by Applicant) Etc. are known.

However, the (c) low phytin vegetable protein hydrolyzate is not very well known. For example, Japanese Patent Laid-Open No. 8-092123 by the applicant of the present application discloses a method of using a resin.

In addition, Japanese Patent Laid-Open No. 9-023822 discloses a method of obtaining a peptide product having a low phytic acid content by inoculating fermented soybean protein with fermented yeast, followed by enzymatic digestion and phytase treatment.

However, a method for detecting phytic acid up to the detection limit is not known as in the present invention.

While the present inventors have progressed the research, in order to use the vegetable protein hydrolyzate for small fish or the young fish with insufficient development of the digestive absorption function, the vegetable protein hydrolyzate which is insufficient in the peptide product and has excellent digestive absorption ability It was found that extremely small amount of phytic acid is required as the decomposed product. Accordingly, an object of the present invention is to obtain a low phytic vegetable protein hydrolyzate having an extremely low phytic acid content, and a low phytic vegetable protein hydrolyzate having a phytic acid below a detection limit.

And it aimed at feed for fry using such a low phytin plant protein hydrolyzate.

As a result of intensive studies, the present inventors have found that by reducing the amount of phytic acid contained in the soy protein hydrolyzate used as feed, the survival rate of small fry and the growth of fry are made, and the present invention has been completed. .

In addition, the following findings were made to complete the low phytin plant protein hydrolyzate. That is, the present applicant first completed the low phytic acid vegetable protein hydrolyzate (Japanese Patent Laid-Open No. 8-092123) by the resin treatment, but the resin treatment was complicated to make the phytic acid content below the detection limit.

On the other hand, the applicant has produced a low phytic soy protein by treating the soy protein with phytase (Japanese Patent Laid-Open No. 2000-300185), but the phytic acid content is below the detection limit even if the low phytic soy protein is simply enzymatically decomposed. It was difficult to obtain an extremely low soy protein hydrolyzate.

Therefore, as a result of more intensive studies, first, it was found that soy protein hydrolyzate having extremely low phytic acid (below the detection limit) was obtained by enzymatically digesting soy protein to a specific molecular weight range and then phytase treatment.

When soybean protein is enzymatically decomposed, soybean protein is extracted from soybean raw material, and then enzymatically decomposed without drying, phytase treatment is performed first, and even after enzymatic digestion, phytic acid can be removed to below the detection limit. It was found that it can.

More surprisingly, it was found that the extremely small soy protein hydrolyzate whose content of phytic acid is below the detection limit is superior in flavor to the soy protein hydrolyzate which is simply enzymatically digested.

The present invention has been completed based on these findings.

That is, the present invention is a fine fry feed characterized in that the feedstock contains low pitin vegetable protein hydrolyzate having a phytic acid content of 0.05% by weight or less (in dry solids).

Low phytin plant protein hydrolysates are suitable for plant protein hydrolysates having an average molecular weight of 200 to 10,000.

The present invention also provides a low phytic vegetable protein comprising (a) a step of decomposing a protein using a proteolytic enzyme and (b) a step of decomposing a phytic acid using an enzyme that decomposes phytic acid. It is a manufacturing method of a hydrolyzate.

It is preferable to decompose a protein using a proteolytic enzyme, and then decompose phytic acid using an enzyme that decomposes phytic acid.

It is preferable to decompose the undried protein using an enzyme that decomposes phytic acid, and then decompose the protein using a proteolytic enzyme.

The enzyme for decomposing phytic acid is preferably phytase.

The pH for phytase treatment is preferably 6-9.

The average molecular weight of the low phytic vegetable protein hydrolyzate is preferably 200 to 10,000.

It is preferable that the phytic acid content of the low phytic vegetable protein hydrolyzate is not detected by the vanado molybdate absorbance photometry (detection limit 5 mg / 100 g) per dry solid content.

Best Mode for Carrying Out the Invention

First, a description will be given of a small fry feed.

The low phytic vegetable protein hydrolyzate used in the petty feed of the present invention has a phytic acid content in the dry solids of 0.05% by weight or less, preferably 0.01% by weight or less, more preferably 0.004% by weight or less (less than the detection limit). Is suitable.

The smaller the amount of phytic acid contained in the plant protein hydrolyzate, the better the survival rate of the small fry. In addition, there is an effect of promoting the growth of the small fry to give the shit persistence to prevent the water from being clouded by the shit.

The average molecular weight of the low phytin plant protein hydrolyzate used in the petty feed of the present invention is preferably 200 to 10,000 (preferably 300 to 5,000). In the case of a large molecular weight, it is inferior to the effect of promoting the growth and the survival rate of the small fry, and when the molecular weight is lowered to the amino acid, the penetration pressure of the feed increases or becomes easier to dissolve.

If the feed for fish farming may be a soy protein hydrolyzate having a relatively high molecular weight, the smaller the fry of the fish, the lower the average molecular weight is preferable. In particular, the small molecular weight oligopeptide mixture is preferable for the small fish hatching from eggs.

It is suitable that the feed of the present invention contains 40 to 70% by weight of protein component, and preferably about 50 to 60% by weight as a composition.

It is suitable that the feed of the present invention contains 1 to 30% by weight, preferably 3 to 25% by weight, of the low phytin plant protein hydrolyzate. Typically, at least 3% by weight, preferably 10 to 80% by weight, of the protein components in the feed of the present invention is substituted with the low phytin protein hydrolyzate described above. Higher substitution rates make it difficult to granulate this feed.

When the low phytin protein protein hydrolyzate in the feed of the present invention is small, the effect of improving the survival rate and the growth rate of the small fry is small, and too large is not preferable because it causes growth inhibition. This is common to fish which are difficult to produce cultured seedlings such as red snapper and flounder, and is different from other fish farming.

Moreover, as protein components other than the low phytic vegetable protein hydrolyzate, krill, fish wheat, egg processed product, milk processed product, gelatin, fish meal, fish extract, yeast extract, and egg extract can be used together.

The feed of the present invention may contain phosphorus lipids such as carbohydrates, fats, vitamins, minerals, n-3 polyunsaturated fatty acids and soy lecithin, in addition to the low phytin protein protein hydrolysates and other proteins described above.

n-3 polyunsaturated fatty acids are essential fatty acids for saltwater fish such as halibut, and feed-based phosphorus lipids such as soybean lecithin can be included in the feed of the present invention because they are necessary for the culture of small fry. have.

The form of the feed of the present invention has a particle size, flotation, sedimentation rate, which is easy for fish to ingest, and moreover, it is preferable that nutrients are not eluted in water and digested and absorbed in the digestive tract, especially for small fish such as flounder or shrimp. As a microparticles feed, it is preferable to set it as microparticle feed.

As a method of microencapsulating the low phytin vegetable protein hydrolyzate of an average molecular weight of 200-10,000, the method of spray-drying the aqueous solution of a hydrolyzate, for example can be employ | adopted. In addition, in order to adjust elution, floatability, and dispersibility with respect to sea water, fats and oils can be added before spraying, and after spraying, it is prepared by adding a hardened fat and oil, stirring and coating, etc. after spraying. You may.

The feed of the present invention is fed in small amounts according to the age according to the number of days of fry in 30 minutes to 1 hour intervals in combination with a biodiuretic or alone. (給 餌) can be.

Next, one of the manufacturing methods of the low phytin vegetable protein hydrolyzate used for the above-mentioned small fry feed is described below.

(Vegetable protein raw material)

Although the vegetable protein used by this invention can use well-known vegetable protein, grains and a protein for a flow rate are easy to obtain, and since soy protein is produced industrially in large quantities, it is one of the preferable protein raw materials.

Hereinafter, an example of using a soy protein containing a small amount of phytic acid will be described. This method is also applicable to other vegetable proteins.

The soy protein raw material used in the preparation of the low phytic vegetable protein hydrolyzate of the present invention contains soy protein such as soy milk (including skim soy milk), concentrated soy milk, concentrated soy protein, isolated soy protein, skim soybean, and the like. If you can.

The soybean defatted soybean is preferably a soybean denatured soybean which is not accompanied by protein denaturation or has been subjected to light protein denaturation, and is not limited to varieties, producing regions and the like. In general, degreasing soybean treated with low temperature extraction with n-hexane as an extraction solvent is suitable as a raw material, and in particular, low-density degreasing soybean having a NSI (nitrogen solubility coefficient) of 60 or more, preferably 80 or more desirable.

(Enzymatic digestion of soy protein)

The method of enzymatic decomposition of soy protein can be obtained by hydrolyzing soy protein using an enzyme in an aqueous system (soy protein slurry or solution).

For example, when low-density soy protein is used, the concentration of the soy protein solution used for enzymatic treatment is 1% to 30% by weight, preferably 5 to 15% by weight, more preferably 8 to 12% by weight. It is suitable. Although this concentration is low, there is no problem in enzymatic degradation, but productivity is not preferable, which is not preferable.

The proteolytic enzyme (protease) used in the present invention may be used alone or in combination with exoprotease or endoprotease, and does not matter animal origin, plant origin or microbial origin. Specifically, serine proteases (trypsin derived from animals, chymotrypsin, subtilysin derived from microorganisms, carboxypeptidase, etc.), thiol proteases (Papain derived from plants, picin (ficin), bromelain and the like), carboxyprotease (Pepsin derived from an animal, etc.) can be used. In addition, specifically, "Protein FN" originated from Aspergillus aurise (Japanese topic), Streptomyces Griseus origin "Atinase" (Japanese topic), Bazil "Alcarase" (made by Novosa) derived from Ruth Lykeformis, and "Protein A" derived from Bacillus subtilus (a Japanese chemical agent). In addition, as an enzyme containing an endoprotease, "Protease S" made by Tennoga Co., Ltd. of Japan, and "Protein AC-10" made by Daiwa Chemical Co., Ltd. of Japan, and a bipolarase (Nagase of Japan) Biochemical Industry Co., Ltd.) etc. can be illustrated, and "protease M" by Tennog Co., Ltd. of Japan can be illustrated as a proteolytic enzyme containing exo and an endoprotease.

Although the conditions of the hydrolysis of the present invention vary somewhat depending on the type of protease used, it is generally preferable to use an amount sufficient to hydrolyze the soy protein at the pH range, the temperature range of action, and the optimum reaction time of the protease. desirable. When considering the use of a salt-limiting formula (eg, landscape nutrition, etc.) together with a lipid metabolism improving agent, if the pH is 5-10, preferably pH 6-9, formation of salts by neutralization It is preferable because it can reduce.

As for the degree of hydrolysis, the average molecular weight 200-10000, Preferably 300-5000 are suitable. The degree of hydrolysis can be adjusted according to the purpose or use. For example, in the case of diet or feed, even if the molecular weight is relatively large, there is no problem, but when used as a fish feed for small fish, a low molecular weight that is easy to digest is preferable, and an average molecular weight of 200 to 5000 is preferable. More preferably, about 200-2000 are suitable.

(Pyetic acid decomposition by phytic acid degrading enzyme)

Examples of enzymes for decomposing phytic acid used in the present invention include enzymes derived from plants such as wheat and potatoes, enzymes derived from animal organs such as intestinal tract, enzymes of microbial origin such as bacteria, yeast, mold, actinomycetes, etc. Enzymes, such as phytase and phosphatase which have acid-degrading activity, can be used.

As an enzyme for decomposing phytic acid, phytase and phosphatase are suitable, but phytase is more preferable. A phytase can use the thing derived from the strain which has various phytase production ability, such as the genus Aspergillus, the genus Rizopus, the Saccharomyces, the genus Mucor, and the genotricam. Preferably from the genus Aspergillus, more preferably from the genus Aspergillus: Phytase from Aspergillus ficuum, Aspergillus niger It can be selected from the group consisting of phytase derived from and phyta derived from Aspergillus terreus. In order to decompose phytic acid in soybean to inositol, it is necessary to cleave the ester group, and the enzyme that performs it is phytase.

It is also possible to use acidic phosphatase derived from fungi as acidic phosphatase. That is, acid phosphatase from Aspergillus ficuum, acid phosphatase from Aspergillus niger and acid phosphatase from Aspergillus terreus. You can choose from the group.

Since the decomposition reaction of phytic acid by the enzyme treatment can be performed under extremely mild conditions, the effect on the protein is extremely small. For example, the enzyme reaction of the present invention may be performed at 30 to 60 ° C. for 0.1 to 30 hours.

In this invention, pH at the time of a phytic acid decomposition reaction is especially important, It is good to carry out at pH 6-9, Preferably it is 6.2-8.5, More preferably, it is pH 6.2-7. Soy protein treated at a pH lower than 6.0 is not preferable because its solubility is lowered and its flavor is worsened. Moreover, even if pH exceeds 9.0, a flavor will worsen and it is unpreferable. By decomposing phytic acid within the pH range, it is possible to produce soy protein with reduced phytic acid.

Therefore, the enzyme suitably used in the present invention is preferably an enzyme capable of decomposing phytic acid and phytate in a neutral to alkaline pH range of pH 6 or higher, but the origin thereof is not particularly limited, and the enzyme described above may be used.

The enzyme can be used regardless of the form of powder or liquid, and is 0.01 to 10% by weight, preferably 0.05 to 2% by weight, more preferably 0.1 to 1% by weight, based on the weight of crude protein in soy protein. The enzyme titer is 0.1 to 100 U / g crude soy protein, preferably 0.5 to 20 U / g crude soy protein, more preferably 1 to 10 U / g crude soy protein. It is preferred that a degree of phytase be added. The enzyme activity was obtained by reacting a reaction solution containing 0.5 ml of 0.2 M Tris-HCl buffer (pH 6.5) containing 4 mM sodium phytate, 0.4 ml of distilled water, and 0.1 ml of enzyme solution for 30 minutes at 37 ° C. Oml is added to stop the reaction. The inorganic phosphoric acid content in this reaction solution was quantified by the Fiske-Subbarow method. Under the above conditions, the amount of enzyme which liberated 1 mol of inorganic phosphoric acid in 1 minute was 1 unit (U).

In the present invention, as long as it includes a step of decomposing a protein using a proteolytic enzyme and a step of decomposing a phytic acid using an enzyme that decomposes phytic acid, the order may be combined, and these steps may be combined. By this, the low phytinization, ie, the phytic acid content in the plant protein hydrolyzate can be 0.5% or less and 0.2% or less per dry solid content. Furthermore, in order to reduce the phytic acid content to 5 mg / 100 g or less, it is more preferable in the preparation of low phytic vegetable protein hydrolyzate to enzymatically decompose the protein and then phytase the protein. On the other hand, even if the protein is subjected to enzyme digestion after the phytase treatment, it is difficult to make the phytic acid content a detection limit of 5 mg / 100 g or less. However, in the case of using the unmodified soy protein as described above, the soy protein solution (not powder dried) is subjected to phytase as described above to decompose phytic acid, and then to decompose the protein as described above. It is possible to obtain a low phytin vegetable protein hydrolyzate having a target content of doptic acid below the detection limit.

As for the average molecular weight of the low phytin plant protein hydrolyzate obtained as mentioned above, 200-10000, Preferably 300-5000 are suitable.

In addition, the phytic acid content is 0.5% or less, preferably phytic acid is not detected by the vanadomolybdate absorption spectrophotometry (detection limit 5 mg / 100 g) in the dry solid of the low phytic vegetable protein hydrolyzate.

EMBODIMENT OF THE INVENTION Hereinafter, embodiment of invention is described by an Example.

First, the food will be described.

[Production Example 1] (Preparation of low phytic soy protein

7 times of water was added to 10 parts of degreased soybeans, extracted with stirring at 50 ° C. and pH 7 for 30 minutes, and the sebaceous acid and soy milk were separated in a centrifuge, the soy milk was adjusted to pH 4.5 with sulfuric acid, and then centrifuged to precipitate isoelectric point. After separating into protein and whey protein, 4 times of water was added to the isoelectric point precipitated protein, and then adjusted to pH 6.0 with NaOH to prepare 50 parts by weight of soy protein solution at 8% concentration.

This soy protein solution was heated to 50 degreeC, 0.04 weight part of phytase degrading enzyme "Sumichimu PHY" [manufactured by Nippon Chemical Co., Ltd.] was added, and it was made to react for 60 minutes. The reaction solution was sterilized at 150 ° C. for 7 seconds, cooled to 50 ° C., adjusted to pH 7.0 with NaOH, and reacted for 5 hours by adding 0.16 parts by weight of “protease M” (manufactured by Tenno Industries Co., Ltd.). . The mixture was adjusted to pH 6.5, sterilized at 150 ° C. for 7 seconds, and immediately dried by a spray dryer.

The content of phytic acid (mesonoinohexanoic acid) in this powder was measured by the vanadomolybdate absorption spectrophotometry, and it was not detected (detection limit 5 mg / 100 g). The average molecular weight was about 500 as measured by the electrophoresis method.

[Production Example 2] (Preparation of Soy Protein Hydrolyzate without Removing Phitic Acid)

Protein soybean protein [Fujipro R] manufactured by Japanese Soybean Co., Ltd., "Fujipro R" [100] Daiwa Chemical Co., Ltd. origin: Aspergillus genus] Using 1 part, enzymatic digestion at 50 ° C. for 5 hours, heating at 70 ° C. for 30 minutes to deactivate the enzyme, and then obtained by centrifugation after cooling. Supernatant was spray dried to prepare soy protein hydrolysate. And this was TCA (trichloroacetic acid) solubility rate (the value which multiplied the value of 15% TCA soluble nitrogen / total nitrogen by 100) was 100, and the average molecular weight was 676.

[Example 1 and Comparative Example 1]

Example 1

Thirty-eight days old (30 days old) flounder fry from 300 preparatory tanks were transferred to a 100-L experimental tank and 5 tanks were prepared as an experimental tank. The temperature of the seawater was maintained at 17 ° C. during the experiment.

As an experimental feed, 0, 2.5, 5.0, 10.0, and 20.0 parts by weight of low phytic soy protein hydrolyzate prepared in the same manner as in Production Example 1 was replaced with casein and Table 1 in the flounder cultured food of Table 1. It was added to obtain particulate feed by a conventional method. Table 1 shows the experimental feed composition. The unit is parts by weight. Group 5 is the control (control).

At the age of 39, the experiment was started, and feeding was given eight times from 9 am to 16:00 every hour, and at 17:00, Altemia was given as biological food, and fed nine times a day. Feeding amount was 0.31g ~ 0.50g / sashimi / fish, and Altemia was 55 ~ 80 / sigma / fish as the age of one day.

After the start of the experiment, when the age was 14 days and 52 days, about 50 body lengths, number of live tresses, and pigment abnormality were measured.

Comparative Example 1

As an experimental feed, the flounder was cultured in the same manner as in Example 1, using a soy protein hydrolyzate that was not removed with phytic acid, prepared in the same manner as in Production Example 2.

TABLE 1

   ingredient     Group 1      Group 2      Group 3       4th group      5 groups Flesh White Fish Wheat     30.0     30.0      30.0      30.0     30.0 Casein     11.3     20.6      25.3      27.7     30.0 Soy Protein Hydrolysates     20.0     10.0       5.0       2.5      0.0 dextrin      7.0      7.0       7.0       7.0      7.0 Vitamin Mix      5.3      5.3       5.3       5.3      5.3 Mineral mixtures      5.0      5.0       5.0       5.0      5.0 Pollack cod liver oil      8.0      8.0       8.0       8.0      8.0 n-3 polyunsaturated fatty acids      0.5      0.5       0.5       0.5      0.5 Soy Lecithin      4.0      4.0       4.0       4.0      4.0 cellulose      0.9      1.6       1.9       2.0      2.2 Casein      8.0      8.0       8.0       8.0      8.0       system      100      100       100       100      100

The results of Example 1 and Comparative Example 1 were as shown in Table 2.

TABLE 2

   Example 1: With phytic acid removed    Comparative Example 1: No Petri Acid Removal    group    N    Length average (mm)  Fresh water    group     N     Length average (mm)  Fresh water    1 2 3 4 5   48 54 50 54 52      18.11 17.89 17.65 17.36 17.15   252 271 244 230 189     1 2 3 4 5    52 49 49 54 53       17.91 18.18 17.43 17.31 17.01   194 215 200 195 194

From the results of Example 1, all of the groups 1 to 4 to which the low phytic acid soy protein hydrolyzate was added were promoted growth compared to the control group 5, and especially in the groups 1 to 3, the significant difference was significant. appear. In terms of fresh water, good results were found in the 1-4 groups compared to the 5 groups. In addition, the pigment | dye abnormality rate did not differ in each group, and the abnormality did not appear also in the group which gave soy protein hydrolyzate.

In addition, the soybean protein hydrolyzate (oligopeptide mixture) having a low phytic acid content also significantly increased the viability compared to Comparative Example 1 below.

[Manufacture example 3]

100 parts by weight of isolated soy protein [New Fujipro R, manufactured by Daiichi Co., Ltd.] was dissolved in 900 parts of water, and 2 parts of proteolytic enzyme [Protein, manufactured by Daiwa Chemical Co., Ltd. After incubation at 50 ° C. for 5 hours, centrifugation (5000 rpm × 30 minutes) was used to remove insoluble matters, and then heated at 80 ° C. for 30 minutes to inactivate and sterilize enzymes. Got it.

An acrylic weakly basic anion exchange resin (KA890, manufactured by Nippon Chemical Co., Ltd.) was charged to a column of 1.4 cm in diameter up to 15 cm in height (resin volume of 23 cm ² ), 50 ml of 5% caustic soda solution and ion exchanged water. 500 ml was passed through to wash.

On the other hand, the above enzymatic digest (SH) was dissolved in ion exchanged water, the pH was adjusted to 4.5 with 10% hydrochloric acid, and adjusted with ion exchanged water so that the final protein concentration was 10%.

The preparation was passed through 51 ml / hr at the top of a column filled with KA890 and washed, and a treatment solution eluted from the bottom of the column was fractionated.

Enzyme digestion liquid was passed through, and the enzyme digestion liquid until the amount became 1219 ml (121.9 g as protein amount, 5.3 g per 1 ml of resin), that is, the eluate in which phytic acid was completely adsorbed and removed from the resin, was collected and freeze-dried. A low phosphorus enzymatic digest (SHR) was obtained. The phytic acid content was measured by the method of Mohamed et al. (Cereal Chem. 63, 475, 1986). As a result, phytic acid was not detected (detection limit 0.005% by weight).

[Example 2 and Comparative Example 2]

Using low phytic soybean protein hydrolyzate obtained in Preparation Example 3 and soy protein hydrolyzate from which phytic acid obtained in Preparation Example 2 was not removed, fertilized eggs obtained from mother shrimp were hatched and pre-cultured to Zoea stage 1 Breeding experiments were conducted.

As for the breeding conditions, 100 Zoea 1 was accommodated in the 1-liter beaker as shown in Table 3, and it was breeding at room temperature using particulate feed. The composition of the test feed is shown in Table 4.

As the test feed, 10.0 parts by weight of each soy protein hydrolyzate prepared in the same manner as in Production Example 3 and Production Example 2 was replaced with casein (control), and the particulate feed was prepared by a conventional method. It was set as.

In addition, the effect of addition was determined by the growth index of the larvae (growth stage reached) and the survival rate. The results are shown in Table 5.

[Table 3] Breeding Conditions of Prawn Larvae

         Condition             Contents Larval (at the beginning of breeding) Breeding time Breeding Density Water temperature Breeding Feeding Feeding Frequency Zoea 1 stage 100 rats / tank (11) until reaching post-larvae Room temperature (25 ~ 27 ℃) pH 8.4 ± 0.2100% / day 2 times / day Particle size of feed Zoea stage 1 Mysis stage Post-larvae stage 53 μm 125 μm 250 μm Feed Intake (mg Feed / Larva / day) Zoea Stage 1 Mysis Stage Post-larvae Stage 0.160.200.24

Table 4 Test Feed Composition

   Ingredient / Group      Control    Preparation Example 3    Preparation Example 2 Casein       42.8      32.8      32.8 L-arginine HCl        2.4       2.4       2.4 Low Phytin Soy Protein Hydrolyzate        0.0      10.0      10.0 Glucose        5.5       5.5       5.5 Sucrose       10.0      10.0      10.0 α-starch        4.0       4.0       4.0 Cod liver oil        4.0       4.0       4.0 ω3-HUFA * 1        0.5       0.5       0.5 Soy Lecithin        3.0       3.0       3.0 cholesterol        1.0       1.0       1.0 Mineral mixtures        8.6       8.6       8.6 Vitamin mixtures        3.0       3.0       3.0 Glucosamine HCl        0.8       0.8       0.8 Sodium citrate        0.3       0.3       0.3 Sodium succinate        0.3       0.3       0.3 Carrageenan        5.0       5.0       5.0 Cellulose        8.8       8.8       8.8      system      100.0     100.0     100.0

    ※ 1 (20: 5ω3): (22: 6ω3) = 3: 2

[Table 5] Results of breeding experiments of prawn larvae

     group Length 11th day (mm)   Survival rate (%)  Growth Index (Day 11)   Control    4.56 ± 0.27    79.5      6.9   Preparation Example 3    4.68 ± 0.25    91.2      7.7   Preparation Example 2    4.66 ± 0.26    83.5      7.4

 * Growth rate = (1n1 + 2n2 + 3n3 + 4n4 + 5n5 + 6n6 + 7n7) / N

   (Where N = n1 + n2 + ----- + n7)

 * n1, n2, n3, n4, n5, n6 and n7 represent the number of digits of Zoea 1, Zoea 2, Zoea 3, Mysis 1, Mysis 2, Mysis 3 and Post larvae 1 larvae.

As a result, the postlarvae length (measured on the 11th day) showed no difference between the low phytin soy protein hydrolyzate added in Preparation Example 3 and the soy protein hydrolyzate added in Preparation Example 2, Low phytin soy protein hydrolyzate showed good tendency in growth index.

EXAMPLE 3

Seventy flounder fry of 70 to 80 days after birth were transferred to a 20L experimental water tank and three tanks were prepared for breeding experiments. The temperature of the breeding water was maintained at 18 ℃ during the experiment.

As experimental feed, a commercial flounder feed (EP feed manufactured by Higashimaru Co., Ltd. S-6 for seedlings) was prepared as a base. That is, the commercial feed is warmed, coated with the low phytin soy protein hydrolyzate prepared in Preparation Example 1 and the phytase untreated soy protein hydrolyzate prepared in Preparation Example 2, and then dried and tested. Feed was used. Table 6 shows the composition of the commercial feed and the composition of the experimental feed added with soy protein hydrolyzate.

TABLE 6

(Commercial feed composition) 75.5% fish meal, krill wheat 12.7% wheat flour, starch, corn flour 11.8% feed yeast, refined fish oil, soy lecithin, yeast extract, calcium dihydrogen phosphate, trace elements, vitamins 67.5% Fish Meal, Krill Wheat 10% Soy Protein Hydrolyzate 11.7% Wheat Flour, Starch, Corn Flour 10.8% Feed Yeast, Refined Fish Oil, Soy Lecithin, Yeast Extract, Calcium Dihydrogen Phosphate, Trace Elements, vitamins 67.5% Fish Meal, Krill Wheat 10% Low Phytin Soy Protein Hydrolyzate 11.7% Wheat Flour, Starch, Corn Flour 10.8% Feed Yeast, Refined Fish Oil, Soy Lecithin, Yeast Extract, Phosphoric Acid 2 Calcium hydrogen, trace elements, vitamins

The experiment was started using 80.3 mm overall length, fish weight 4.4 g, and body weight / total length = 55.5 mg / mm and fed four times a day from 9 am to 6 hours. The body length of 60 small fry and fish body weights after 10 and 20 days after the start of the experiment were measured, and the weight / total length was calculated from the measurement results.

TABLE 7 Day 0 of measurement start (60 fish)

 Control sphere Soy Protein Hydrolysates Low Phytin Soy Protein Hydrolyzate Average length (mm) Max Min        81.4 90 73     79.0 88 72         80.4 90 71 Average weight (g) max min        4.49 6.5 2.9     4.26 6.2 3.1         4.54 6.0 3.1 Average weight / length (mg / mm) max min        55.2 72.2 39.7  53.9 (-1.3) 70.5 (-1.7) 43.1 (+3.4)      56.5 (+1.3) 66.7 (-5.5) 43.7 (+4.0)

Note: () inside and outside the control

[Table 8] Day 10 of measurement start (60 test fish, 60 fresh water)

 Control sphere Soy Protein Hydrolysates Low Phytin Soy Protein Hydrolyzate Average length (mm) Max Min     86.1 95 73     82.5 95 74       86.4 101 71 Average weight (g) max min     4.99 6.8 2.7     4.57 7.3 4.1       5.31 8.2 3.2 Average weight / length (mg / mm) max min     57.9 71.6 37.0   55.4 (-2.5) 76.8 (+5.2) 43.2 (+6.2)    61.5 (+3.6) 81.2 (+9.6) 53.9 (+16.9)

Note: () inside and outside the control

Table 9 Measurement start 20th day (60 test fish, 60 live fish)

 Control sphere Soy Protein Hydrolysates Low Phytin Soy Protein Hydrolyzate Average length (mm) Max Min      94.0 106 76     89.3 106 78         93.4 101 81 Average weight (g) max min      7.02 10.6 3.3     6.05 10.9 3.9         6.97 11.8 5.3 Average weight / length (mg / mm) max min      74.7 100.0 43.4  67.7 (-10.0) 102.8 (+2.8) 50.0 (+6.6)      74.6 (-0.1) 106.3 (+6.3) 65.4 (+22.0)

Note: () inside and outside the control

As a result, compared with the control and soy protein hydrolyzate addition, the low phytin soy protein hydrolyzate addition group showed higher elongation at the average body weight / length (mg / mm) at 10 days and showed a growth promoting effect. Moreover, on day 20, maximum solids were also observed in body weight / length (mg / mm). The minimum value was higher than the control and soy protein hydrolyzate additions, but the solids were due to the concentration of food in large fish, and the variation between the solids was large, and the average did not change much with the control.

(Breeding progress)

The water temperature, food supply, and food intake during the breeding period are shown below.

In the control group, the stool was in the form of diarrhea after feeding, and the water in the tank was clouded. In the soy protein hydrolyzate and low phytin soy protein hydrolyzate, the feces remained morphological. The water in the tank was clean as it was. In addition, low phytic soy protein hydrolyzate tended to have persistent fish poop and a large amount of poop.

TABLE 10

Month day Days Water temperature Target sphere Soy Protein Hydrolysates Low Phytin Soy Protein Hydrolyzate Feed Intake Status Fresh water Feed Intake Status Fresh water Feed Intake Status Fresh water 4/2 One 17.7 0 20 0 20 0 20 4/3 2 18.0 8 Inactive, but ingested 20 8 Inactive, but ingested 20 8 Inactive, but ingested 20 4/4 3 18.8 4 There is cloudy of water and good intake 20 4 There is no cloud, good intake 20 4 There is no cloud, good intake 20 4/5 4 18.3 4 There is cloudy of water and good intake 20 4 There is no cloud, good intake 20 4 There is no cloud, good intake 20 4/6 5 18.3 4 There is cloudy of water and good intake 20 4 There is no cloud, good intake 20 4 There is no cloud, good intake 20 4/7 6 18.4 4 There is cloudy of water and good intake 20 4 There is no cloud, good intake 20 4 There is no cloud, good intake 20 4/8 7 18.4 4 There is cloudy of water and good intake 20 4 There is no cloud, good intake 20 4 There is no cloud, good intake 20 4/9 8 18.4 4 There is cloudy of water and good intake 20 4 There is no cloud, good intake 20 4 There is no cloud, good intake 20 4/10 9 17.8 4 There is cloudy of water and good intake 20 4 There is no cloud, good intake 20 4 There is no cloud, good intake 20 4/11 10 18.3 5 There is cloudy of water and good intake 20 5 There is no cloud, good intake 20 5 There is no cloud, good intake 20 4/12 11 18.5 5 There is cloudy of water and good intake 20 5 There is no cloud, good intake 20 5 There is no cloud, good intake 20 4/13 12 18.5 5 There is cloudy of water and good intake 20 5 There is no cloud, good intake 20 5 There is no cloud, good intake 20 4/14 13 18.6 5 There is cloudy of water and good intake 20 5 There is no cloud, good intake 20 5 There is no cloud, good intake 20 4/15 14 18.8 5 There is cloudy of water and good intake 20 5 There is no cloud, good intake 20 5 There is no cloud, good intake 20 4/16 15 18.9 8 There is cloudy of water and good intake 20 8 There is no cloud, good intake 20 8 There is no cloud, good intake 20 4/17 16 19.4 6 There is cloudy of water and good intake 20 6 There is no cloud, good intake 20 6 There is no cloud, good intake 20 4/18 17 18.4 6 There is cloudy of water and good intake 20 6 There is no cloud, good intake 20 6 There is no cloud, good intake 20 4/19 18 18.4 6 There is cloudy of water and good intake 20 6 There is no cloud, good intake 20 6 There is no cloud, good intake 20 4/20 19 18.4 6 There is cloudy of water and good intake 20 6 There is no cloud, good intake 20 6 There is no cloud, good intake 20 4/21 20 18.4 8 There is cloudy of water and good intake 20 8 There is no cloud, good intake 20 8 There is no cloud, good intake 20

Next, examples of low phytin soy protein hydrolysates will be described.

EXAMPLE 4

Seven times of water was added to 10 kg of defatted soybeans, extracted with stirring at 50 ° C. and pH 7 for 30 minutes. After separating the soy and soy milk in a centrifuge, the soymilk was adjusted to pH 4.5 with sulfuric acid, followed by centrifugation to precipitate isoelectric point. Protein and Whey protein were separated, 4 times of water was added to the isoelectric point precipitated protein, and then adjusted to pH 7.0 with NaOH to adjust 50 liters of the protein solution at 8% concentration.

160g of proteolytic enzymes "protease S" and "protease M" (manufactured by Tennog Co., Ltd., Japan) were added to the protein solution, and the mixture was hydrolyzed by reaction at 50 ° C for 5 hours (15% TCA solubility rate). 85%). Subsequently, pH was adjusted to 6.0, 40 g of phytic acid decomposing enzyme "Sumichimu PHY" [manufactured by Nippon Chemical Co., Ltd.] was added thereto, and reacted at 50 ° C for 60 minutes, and then adjusted to pH 6.5 with NaOH, 150 After sterilization for 7 seconds at < RTI ID = 0.0 >

The content of phytic acid (mesoinocit hexaphosphate) in the powder was not detected as a result of measurement by vanadomolybdate absorption spectrophotometry (detection limit 5 mg / 100 g), and the phytic acid was extremely reduced.

[Example 5]

Seven times of water was added to 10 kg of skim soybeans, extracted with stirring at 50 ° C. and pH 7 for 30 minutes, and the sebaceous acid and soy milk were separated by a centrifuge. After separating into whey protein, 4 times of water was added to the isoelectric point precipitated protein, and then adjusted to pH 6.0 with NaOH to adjust 50 liters of the protein solution at 8% concentration.

The protein solution was warmed to 50 ° C, and 40 g of phytase degrading enzyme "Sumichimu PHY" (manufactured by Nippon Chemical Co., Ltd.) was added thereto and reacted for 60 minutes. After that, the pH was adjusted to pH 7.0 with NaOH. 160 g of M "(made by Tennoga Co., Ltd. of Japan) was added, and it was made to react for 5 hours. The mixture was adjusted to pH 6.5, sterilized at 150 ° C. for 7 seconds, and immediately dried by a spray dryer.

The content of phytic acid (mesoinocit hexaphosphate) in the powder was not detected as a result of measurement by vanadomolybdate absorption spectrophotometry (detection limit 5 mg / 100 g), and the phytic acid was extremely reduced.

EXAMPLE 6

Seven times of water was added to 10 kg of skim soybeans, extracted with stirring at 50 ° C. and pH 7 for 30 minutes, and the sebaceous acid and soy milk were separated by a centrifuge. After separating into whey protein, 4 times of water was added to the isoelectric point precipitated protein, and then adjusted to pH 6.0 with NaOH to adjust 50 liters of the protein solution at 8% concentration, and immediately powder dried with a spray dryer.

The protein solution was adjusted to an 8% solution, warmed to 50 ° C, 40 g of phytase-degrading enzyme "Sumichimu PHY" (manufactured by Nippon Chemical Co., Ltd.) was added and reacted for 60 minutes, followed by pH 7.0 with NaOH. It adjusted to and added 160 g of "protease M" (made by Tenno Industries Co., Ltd. of Japan), and made it react for 5 hours. The mixture was adjusted to pH 6.5, sterilized at 150 ° C. for 7 seconds, and immediately dried by a spray dryer.

As a result of measuring the phytic acid (mesinoinohexanoic acid) content in this powder by the vanado molybdate absorption spectrophotometry, the phytic acid was reduced to 0.5% (detection limit 5 mg / 100 g).

EXAMPLE 7

Seven times of water was added to 10 kg of skim soybeans, extracted with stirring at 50 ° C. and pH 7 for 30 minutes, the sebaceous milk and soy milk were separated in a centrifuge. After separating into whey protein, 4 times of water was added to the isoelectric point precipitated protein, and then adjusted to pH 6.0 with NaOH to adjust 50 liters of the protein solution at 8% concentration.

The protein solution was heated to 50 ° C, 40 g of phytase degrading enzyme "Sumichimu PHY" (manufactured by Nippon Chemical Co., Ltd.) was added thereto and allowed to react for 60 minutes, followed by powder drying with a spray dryer immediately.

The solution was adjusted to 8%, adjusted to pH 7.0 with NaOH, and 160 g of "Protease M" (manufactured by Tenno Industries Co., Ltd.) was added thereto to react for 5 hours, and sterilized at 150 ° C for 7 seconds, immediately. The powder was dried with a spray dryer.

As a result of measuring the phytic acid (mesonoinohexanoic acid) content in this powder by the vanado molybdate absorbance photometric method, the phytic acid was reduced to 0.2% (detection limit 5 mg / 100 g).

EXAMPLE 8

Seven times of water was added to 10 kg of skim soybeans, extracted with stirring at 50 ° C. and pH 7 for 30 minutes, and the sebaceous acid and soy milk were separated by a centrifuge. After separating into whey protein, 4 times of water was added to the isoelectric point precipitated protein, and then adjusted to pH 6.0 with NaOH to adjust 50 liters of the protein solution at 8% concentration.

The protein solution was heated to 50 ° C, 40 g of phytase degrading enzyme "Sumichimu PHY" (manufactured by Nippon Chemical Co., Ltd.) was added thereto and allowed to react for 60 minutes, followed by powder drying with a spray dryer immediately.

The solution was adjusted to 8%, adjusted to pH 7.0 with NaOH, and 160 g of "Protease M" (manufactured by Tenno Industries Co., Ltd.) was added thereto to react for 5 hours (pH 6.2 at the time of reaction). 40 g of phytase degrading enzyme "Sumichimu PHY" (manufactured by Nippon Chemical Co., Ltd.) was added thereto, followed by 60 minutes of reaction, followed by sterilization at 150 ° C. for 7 seconds, followed by powder drying with a spray dryer.

As a result of measuring the phytic acid (mesonoinohexanoic acid) content in this powder by the vanado molybdate absorbance photometric method, the phytic acid was reduced to 0.2% (detection limit 5 mg / 100 g).

EXAMPLE 9

30 kg of isolated soybean protein [New Fuji pro R, made by Nippon Kogyo Co., Ltd., once made powder] 10% aqueous solution of pH 7.0, protease S [protease S] of Japan Agent] 1.2 kg and 0.3 kg of "protease M" [manufactured by Tenno Pharmaceutical Co., Ltd.], were made to work and hydrolyzed at 50 ° C for 5 hours (15% TCA solubility 85%), and then the pH was adjusted to 6.0. It was adjusted, 0.6 kg of phytase degrading enzyme "Sumichimu PHY" (manufactured by Nippon Chemical Co., Ltd.) was added, and hydrolysis was performed at 45 ° C for 2 hours.

This digestion liquid was adjusted to the feed rate of 100 liters / hour in the high speed centrifuge which can process continuously, and the settling component produced | generated was isolate | separated and removed. The obtained centrifugal supernatant (70% yield of solid content) was adjusted to pH 6.5, sterilized at 150 degreeC for 7 seconds, and then powder-dried immediately by the spray dryer.

The content of phytic acid (mesoinocit hexaphosphate) in the powder was not detected as a result of measurement by vanadomolybdate absorption spectrophotometry (detection limit 5 mg / 100 g), and the phytic acid was extremely reduced.

By feeding the feed of the present invention as a food, it is possible to promote the growth of small fry and greatly increase the survival rate.

In particular, it is possible to increase the survival rate of the small fish which becomes difficult to fish, and it is possible to farm fish conventionally difficult to fish.

In addition, it is possible to promote the growth of the fry which has grown to some extent, and to prevent turbidity of the water by keeping the poo sticky, thereby improving the growth environment.

In addition, the low phytin protein protein hydrolyzate suitable for such breeding is enabled.

That is, according to the present invention, the phytic acid content is extremely low, and the low phytin vegetable protein hydrolyzate is less than the limit of detection by the vanadomolybdate absorption spectrophotometry. In addition, the low phytin protein protein hydrolyzate by the method of the present invention is superior to the low phytic soy protein hydrolyzate by the resin adsorption method (presumably due to the decomposition products of phytic acid).

In addition, the low phytic vegetable protein hydrolyzate of the present invention has a low molecular weight and extremely low phytic acid, so that the development of the digestive absorption function is insufficient, and when used in feed for the born animals, the digestive absorption is excellent, and the trace metals such as calcium It is extremely effective by promoting absorption.

Claims (10)

A small pomaceous feed comprising a plant protein hydrolyzate having a phytic acid content of 0.05% by weight or less (in dry solids) as a feedstock. The petty fry of Claim 1, wherein the plant protein hydrolyzate is a plant protein hydrolyzate having an average molecular weight of 200 to 10,000. 3. The particulate feed of claim 1 or 2, wherein the form of the feed has a particle size, suspension, and settling velocity that are easy to ingest in small fry, and further microencapsulated so that nutrients do not elute in water and are digested and absorbed in the digestive tract. Good pet food. A method for producing a low phytin plant protein hydrolyzate, comprising the steps of: (a) degrading a protein using a proteolytic enzyme; and (b) decomposing a phytic acid using an enzyme that decomposes phytic acid. The manufacturing method of Claim 4 which decomposes a soy protein using a proteolytic enzyme, and then decomposes a phytic acid using the enzyme which decomposes phytic acid. The manufacturing method of Claim 4 which decomposes an undried soy protein using the enzyme which decomposes a phytic acid, and then uses a proteolytic enzyme. The production method according to any one of claims 4 to 6, wherein the enzyme for decomposing phytic acid is phytase. The production method according to any one of claims 4 to 7, wherein the phytase-treated pH is 6-9. The production method according to any one of claims 4 to 8, wherein the average molecular weight of the low phytic vegetable protein hydrolyzate is 200 to 10,000. The production method according to any one of claims 4 to 9, wherein the content of phytic acid in the low phytic vegetable protein hydrolyzate is not detected by vanadic molybdate absorbance photometry (detection limit 5 mg / 100 g) per dry solid content. .