JPS6279741A - Feed for raising fish - Google Patents

Abstract

PURPOSE:A feed for raising fish, obtained by uniformly dispersing a growth hormonal substance of fishes in a water-insoluble carrier and capable of effectively incorporating the growth hormone into the fish body and promoting the growth of the fishes. CONSTITUTION:A feed obtained by uniformly dispersing a growth hormonal substance of fishes, e.g. substance collected from pituitary gland of dog salmon or eel or substance produced by using a recombinant DNA technique, in a water-insoluble carrier, e.g. casein or glutelin.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of Industrial Application The present invention relates to a new fish feed, and is expected to be used in the fish farming industry.

Prior Art 1 Problem and Solution It has been known that fish growth hormone exists in the pituitary gland of fish. It is difficult to extract growth hormone from the pituitary gland, especially since it has very similar molecular weight and properties to the related hormone prolactin, making it difficult to separate and have not succeeded in pure isolation.

The applicant isolated fish growth hormone from the pituitary glands of chum salmon and eel, and also succeeded in producing fish growth hormone from Escherichia coli using genetic recombination techniques, and filed a patent application. 59-68670
, 60-151847. 60-50096゜6
0-161429).

When cultivating fish, even if they are kept in the same aquarium under certain conditions, their growth varies greatly among individuals. For example, for rainbow trout, which is a typical freshwater fish, even if you start from fertilized eggs obtained by fertilizing the eggs from two young fish with the sperm of one male and culture them under the same environment, After 6 months, the difference in weight between the maximum and minimum growth points is about three times.

In red sea bream, which is a marine fish, when floating eggs from male and female pairs that are fertilized on the same day and hatched on the same day are cultured, the maximum weight after 3 months is 21.2 kg and the minimum weight is 3.5 kg. The difference is about 6 times.

In this way, even when genetically identical fish are cultured under the same environment, significant individual differences occur in the fish weight, resulting in differences in the swimming ability of the young fish. These individual differences cause differences in feeding ability and competitiveness, and the growth of small individuals is further suppressed, so during aquaculture, cultivation must be repeated according to individual differences. These things cause labor and over-current of facilities, and furthermore, damage the fish bodies during cultivation and cause diseases.

It is expected that the above-mentioned problems in aquaculture can be solved by using the fish growth hormone.

However, like the conventional pituitary gland extract, the fish growth hormone is poorly water-soluble and difficult to incorporate into the fish body during aquaculture, and it is desired to develop a method for efficiently incorporating it into the fish body.

The present inventor conducted research on a method for effectively incorporating fish growth hormone into the fish body, and found that if the hormone was uniformly dispersed in water-insoluble protein,
The present invention was completed by discovering that the protein can be orally taken into the fish body together with the protein and used effectively. The present invention will be explained in detail below.

The present invention provides a fish feed containing a fish growth hormone-like substance mixed with a water-insoluble carrier.

Chum salmon is a growth hormone-like substance for fish.

DNA collected from the pituitary gland of eels etc. or recombinant DNA
It is possible to use the one described in the above-mentioned patent application, which is manufactured using the technique. Regarding the growth hormone used in the examples, the manufacturing method thereof is specifically shown in the reference example.

Examples of water-insoluble carriers include casein and glutelin.

Proteins such as zein are used. These proteins can be dissolved in a solvent depending on the conditions; for example, casein becomes water-soluble when converted to sodium salt, glutelin is easily soluble in dilute acids and dilute alkalis, and zein is soluble in alcohol.

Therefore, each of these proteins is made into a solution under appropriate conditions, growth hormone is added to it, and the protein is dissolved or dispersed.
Thereafter, by returning the protein to water-insoluble state, the growth hormone can be uniformly dispersed and coated in the water-insoluble protein. An isolated protein may be used, or a protein contained in minced fish or the like may be used as it is.

The growth hormone contained in the water-insoluble carrier is used in an amount of 10 to 10 g per kg of the carrier.

During aquaculture, a water-insoluble carrier containing growth hormone is mixed with aquaculture feed and administered orally to fish as a mixed feed. Growth hormone in compound feed is 1-1000 mg
/kg, and the dosage per fish body varies depending on the species, but it may be administered in the range of 0.1 to 100 μg/g/7 days of body weight.

When the feed of the present invention is orally administered, it is taken up by fish and exhibits its effects. Oral administration methods include the general method of administering growth hormone mixed with other feed ingredients, the method of injecting it using a stomach tube, and the method of directly administering it into the stomach using a gelatin capsule or the like. The general method is preferred in the present invention.

Examples of the present invention will be shown below.

Example 1 A group of 50 juvenile sweetfish (average body length: 63 mm, average weight: 1.6 g), 130 days old, were cultured for 30 days.

During the cultivation period, 0.05 g/g of the feed prepared in Reference Example 1 (containing chum salmon growth hormone (SGH) 1.10.100.1100 h/g) for 7 days was orally administered using a standard method. . The converted body weight was measured 30 days after the start of culture.

The results are shown in Table 1.

Table 1 Feed 30-day mouth weight gain (%) (SG) I
Content mg/kg) Average weight (g) 1
6.8±0.4 32510 8,
5±0.3 431100 8, 1±
0.4 4061000 7, 2±0
．． 3 350 control 5.5±0.
6 244 Example 2 Rainbow trout (70 days old, average weight 0.5 g) Flounder (55 days old, average weight 0.2 g) Red sea bream (46 days old, average weight 0.3 g) Eel ( Glass eel, average weight 0.2 g) per group 5
0 fish were cultured for 40 days. During the cultivation period, feed prepared in Reference Example 2 (including chum salmon growth hormone 200 fish/g)
0.05 g/g body weight twice a week.

A total of 8 orifices were administered.

The converted body weight was measured 40 days after the start of culture. The eel was fed as a paste feed, and the chum salmon growth hormone was dissolved and mixed immediately before feeding and administered orally using a conventional method. The results are shown in Table 2.

Table 2 Rainbow trout 15 87 61 87±13
61 19 flounder 20 128 110
128±11 110±21 red sea bream 20 1
40 123 140±10 123±23 Eel 25 145 120 145±12 1
20±30 Example 3 Using the eel growth hormone obtained by the method of Reference Example 5 and the feed produced in Reference Example 3, red sea bream, flounder, and eel were cultured according to the method of Example 2.

The amount of eel growth hormone added is 100■ per kg of feed.
It is. The eel was fed as a paste feed, and the eel growth hormone was dissolved and mixed immediately before feeding and administered orally using a conventional method. The results are shown in Table 3.

Table 3 Red sea bream 550 420 Flounder
720 510 eel
430 215 Reference Example 1 10.100°1 in 100g of casein sodium salt
000, 10000mg of chum salmon growth hormone (SG)
) l) was added with 400mM water to suspend and disperse, and 50g of this suspension was mixed with 50g of Beiyo Fish Meal (manufactured by Nippon Suisan Co., Ltd.).

Brewer's yeast (HOg manufactured by Kirin Brewery, 5 g of Fitsch Soluble (manufactured by Nippon Kagaku Feed Co., Ltd.), 5 g of vitamin mixture (manufactured by Kawai Pharmaceutical Co., Ltd.), mineral mixture (Naiver)
(manufactured by Kawai Pharmaceutical Co., Ltd.) 5 g, refined cod liver oil (manufactured by Nippon Kagaku Feed Co., Ltd.) 3 g, synthetic binder CMC (manufactured by Daicel Chemical Co., Ltd.) 2 g
, Refined soybean oil cake (manufactured by Showa Sangyo Co., Ltd.) 10g, total 100g
A mixed feed was made. As a control, a sample containing the same amount of casein without growth hormone was prepared.

Reference Example 2 5 g of glycerin was added and dissolved in 50 ml of water at pH 2.0. After adding and dissolving 20 mg of SGH into this solution, 50 g of northern sea fish meal and 15 g of brewer's yeast were added.

Ficin soluble 5g, vitamin mixture 5g, mineral mixture (Halver) 5g, refined cod liver oil 3g
.

A compound feed containing 2 g of synthetic binder and 10 g of refined soybean oil meal was prepared, containing a total of 100 g. As a control, a sample containing no SGH was prepared in the same manner.

Reference Example 3 5 g of zein was added and dissolved in 3 Q ml of 60% ethanol. Add 10 mg of eel growth hormone (
eGH) was suspended and dispersed. In this suspension, 5
0g, brewer's yeast 15g, fishini soluble 5g, vitamin mixture 5g, mineral mixture (lalver)
5 g of refined cod liver oil, 2 g of synthetic binder, and 10 g of refined soybean oil meal, a total of 100 g of mixed feed was prepared. As a control, a sample containing no eGH was prepared using the same method.

Reference example 4. Production of chum salmon growth hormone Escher
ichiacoli ESGHIB2 (FERMBP
-612) to the method of Birnboim et al.
boim) et al.: Nucleic Acid Research (N
ucl'eic Acicls Res,) Yu, 151
3 (1979)) using the recombinant plasmid psGHIB2 isolated according to the conventional method [Proceedings of the National Academy of Sciences (1979)].
Proc, Natl, Acad.

Sci, ), USA 69.2110 (1972
)) was used to transform Escherichia coli W311ONα49 strain. The resulting ampicillin-resistant colonies were diluted with 3mM M
CG medium [0.6% NaaHPOa, 0.3% K
H, P○,.

0.5% NaCC0,1% NH,Cj! , 0.5%
Glucose, 0.5% Casamino Acids, 1mM Mg5O
+.

4u/ml vitamin 7B, pH 7-2),
The cells were cultured at 30°C for 18 hours. The obtained culture solution was 8.00
Orpm.

The cells were collected by centrifugation for 10 minutes. Laem this bacterial body
After suspending in sample buffer of mli, 5DS-polyacrylamide gel electrophoresis was performed, and Coomasieve IJ
Stained with IJ Ant Blue, molecular weight approximately 25,000
A polypeptide band was detected at the site. This band was not present when E. coli not containing the plasmid was used. As a result, it was found that Escherichia coli harboring psGHIB2 produces a large amount of chum salmon growth hormone polypeptide.

The cells were treated with 3QmMTri containing 30mM Na (l).
It was washed twice with s-HCβ buffer (pH 7,5). The washed bacterial cells were suspended in Qml of the above buffer solution I and disrupted by ultrasonication at 0°C [Branson Sonic Power Company (B
ranson 5onic power company
y) Company Sonifier Cell Dislab Coo (So
cell disruptor) 2
Q Q, output control
control) 2, 10 minute treatment]. This was centrifuged at 15,000 rpm for 30 minutes to obtain bacterial cell residue. From this bacterial cell residue, the method of Marston et al.
on) et al.: Biotech/D.
ogy) 2, 800 (1984)], the salmon growth hormone polypeptide was extracted and purified.

Reference example 5. Production of eel growth hormone Escheri
chiacoli EUPAI (FERM BP
-825) to the method of Birnboim et al.
Nucleic Acids Res, 15
13 (1979):] using the recombinant plasmid pUPA 1 isolated according to the standard method [Procedures of the National Academy of Sciences (Proc. Natl., Acad.

Sci.), USA 69.2110 (1972)
) was used to transform Escherichia coli W311ONα49 strain.

The resulting ampicillin-resistant colonies were transferred to 3+nl of M
CG medium [0.6% Na2HP O<,
0.3% K82PO,.

0.5% NaCβ, 0.1% NH, Cj! , 0.5
% glucose, 0.5% casamino acids, 1mM MgS
○、.

4 g/ml vitamin B1, pH 7, 2) and 3
The cells were cultured at 0°C for 18 hours. The obtained culture solution was heated to 8.00O
The cells were collected by centrifugation at rpm and IQ for minutes. After suspending the bacterial cells in Laemmli sample buffer, S
DS ~ Performed polyacrylamide gel electrophoresis and stained with Coomassie brilliant blue, molecular weight approximately 23.00.
A polypeptide band was detected at the 0 site. This band was not present when E. coli not containing the plasmid was used. As a result, it was found that E. coli harboring pL]PA1 produced a large amount of eel growth hormone polypeptide.

This bacterial cell was treated with 30mM NaCj! 30mMTr containing
is-HCj! The cells were washed twice with a buffer solution (pH 7.5), and the washed bacterial cells were treated in the same manner as in Reference Example 4 to extract and purify the eel growth hormone polypeptide.

Patent Applicant (102) Kyowa Hakko Kogyo Co., Ltd. Voluntary author of procedural amendments) October 30, 1985

Claims (1)

[Claims] A fish feed containing a fish growth hormone-like substance uniformly dispersed in a water-insoluble carrier.