JP3030048B2 - Production method of veterinary drug - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to a method for producing a veterinary drug containing bacteria.

[Prior Art] Lactobacillus sali
varius) as a veterinary agent to promote livestock growth,
It is already known to prevent or treat intestinal diseases such as diarrhea and constipation (Japanese Patent Publication No. 59-46209). Enterococcus faecali
It is also known that s) has the same action as Lactobacillus salivarius.

[Content of the Invention] The present inventors have further studied the above effects of Enterococcus faecalis and Lactobacillus salivarius on domestic animals.

As a result, when at least one of alfalfa and brewer's yeast is added to at least one of the bacteria of Enterococcus faecalis and Lactobacillus salivarius, the growth of the above-mentioned bacteria is promoted, and the above-mentioned animal is caused by the bacteria. It has been found that the effects of promoting growth, preventing and treating intestinal diseases such as diarrhea and constipation are further improved, and it is also effective not only in domestic animals but also in other animals.

In addition, the present inventors have determined that Enterococcus faecalis or Lactobacillus salivarius has at least one of an alfalfa extract and a brewer's yeast extract.
It has been found that culturing using a medium containing seeds promotes the cultivation of the fungi, so that the fungi can be obtained at a high concentration and / or in a short time.

Accordingly, the present invention is a veterinary medicament containing (a) at least one of Enterococcus faecalis and Lactobacillus salivarius and (b) at least one of alfalfa and brewer's yeast.

Further, the present invention provides a method for culturing Enterococcus faecalis or Lactobacillus salivarius using a medium containing at least one of an alfalfa extract and a brewer's yeast extract. -Includes Salivarius culture method.

Both Enterococcus faecalis and Lactobacillus salivarius are bacteria which are abundant in the digestive tract of humans and animals and appear in feces. -Any type of Salivarius may be used, and its type, origin, culture method, preparation method and the like are not limited. However, the above-mentioned bacteria derived from a specific animal are preferably administered to animals of the same species. For example, when the bacteria are administered to a horse, it is preferable to prepare the agent of the present invention using bacteria derived from a horse.

In the medicament of the present invention, Enterococcus faecalis and Lactobacillus salivarius may be used alone or in combination.

In culturing these bacteria, any of a liquid medium, a semi-solid medium, and a fixed medium can be used. As the medium, basic inorganic components such as Na, K, Ca, Mg, P, and Cl, Mn,
Trace inorganic components such as Fe, carbon sources (sugars such as glucose, sucrose, maltose, soluble starch, etc.), nitrogen sources (meat extract,
(Peptone, tomato juice, etc.), amino acids, vitamins, hormones, etc. having a pH of about 4 to 8 can be used. Specific examples thereof include MBB, MRS (manufactured by OXOID) shown in Reference Example 1 below. , LSB (manufactured by BBL) and the like.

And, as described above, when at least one of an alfalfa extract and a brewer's yeast extract is further added to the medium containing the above components during the cultivation of the bacterium, the growth of the bacterium is promoted, High concentrations and / or can be obtained in a short time.

The culture may be performed by any culture method, for example, a stationary culture method, a shaking culture method, a submerged culture method, and a nitrogen gas aeration culture method, and a nitrogen gas aeration culture method is particularly preferable.

In the veterinary drug of the present invention, these bacteria may be in any form as long as they are viable, and include, for example, cultured cells, frozen products, freeze-dried products, spray-dried products, and the like.

The drug of the present invention contains at least one of alfalfa and brewer's yeast together with the above-mentioned bacteria.
In this case, alfalfa and brewer's yeast can be in any form, such as a dried product, a semi-dried product, and a paste-like product.Alfalfa and brewer's yeast are obtained by extracting water, alcohol, and other solvents with water. It may be used in the form of an extract (liquid).

The veterinary medicament of the present invention, the drug (dry basis) of at least one of Enterococcus faecalis and Lactobacillus salivarius per 10g is good to be included 10 ⁷ or more, also agents (dry basis) Alfalfa and / or brewer's yeast (on a dry matter basis) should make up about 40-95% of the weight.

The veterinary drug of the present invention has excellent effects on domestic animals such as pigs, cows, horses, sheep, and goats, among which pigs, horses and cattle, but also chickens, ducks, turkeys, It is also effective against poultry such as guinea fowl, pets such as dogs and cats, and other animals such as monkeys. Therefore, the livestock referred to in the present invention means animals including poultry and pets in addition to the above-mentioned livestock in a narrow sense. The drug of the present invention promotes growth and digestion and absorption of the administered livestock, prevents and treats diarrhea, constipation and other fire extinguisher diseases, improves feed efficiency, improves the shape and odor of feces, and the like.

The drug of the present invention can be directly administered to an animal as it is in any dosage form such as a powder, a granule, a tablet, a pellet, a capsule, a liquid and a paste, or can be administered together with feed or drinking water. .

The dose varies depending on the animal type, age, dosage form, administration method, etc., but when given orally, it is usually 10 to 200 mg / body weight 1 kg / day, calculated as a dry drug. 3 days ~
Preferably, it is administered for one month.

The agent of the present invention may further comprise, if necessary, a palatability-enhancing component (for example, one or two of spices, seasonings, etc.) in addition to the above-mentioned bacteria and alfalfa and / or brewer's yeast.
Species or more).

The drug of the present invention is highly safe since it is based on alfalfa and / or brewer's yeast, which are naturally added to the animal's digestive tract and fungi which are naturally present in the animal's digestive tract, and can be used continuously. It does not cause harm such as appearance of resistant bacteria. Therefore, the agent of the present invention can be continuously administered to an animal, in which case the function of the digestive tract of the animal is improved as compared to the case where Enterococcus faecalis or Lactobacillus salivarius is administered alone. As a result, the appetite is further increased, the intake of the feed is increased, and the utilization rate of the ingested feed in the animal is also improved, whereby the growth of the animal can be greatly promoted. Such an effect is particularly remarkable in a young animal before it has completely grown.

In addition, the drug of the present invention is more effective for the prevention and treatment of transient digestive diseases such as diarrhea and constipation, in which case it should be administered only during the period that requires prevention or treatment. Can be. In addition, since digestive diseases such as diarrhea and constipation often occur at the time of weaning, training, transportation, etc., the agent of the present invention is administered to an animal in a weaning period, an animal before, during, and during training or transportation. It has excellent effects when administered.

In addition, due to the above-mentioned property of being safe, the agent of the present invention is not only safe for the animal to which it is administered, but also safe for humans who eat the meat, milk, eggs, etc. of the animal to which it is administered. It is.

Further, as described above, the present invention also includes a culture method of culturing Enterococcus faecalis or Lactobacillus salivarius using a medium containing at least one of an alfalfa extract and a brewer's yeast extract. As the alfalfa extract and brewer's yeast extract in this case, an extract obtained by extracting alfalfa and brewer's yeast with water, alcohol, or the like is preferable. Such an extract can be obtained by adding an extraction medium such as water or alcohol about 3 to 10 times the weight of alfalfa and / or brewer's yeast, performing an extraction treatment, and then removing solids. In culturing the fungi, it is preferable to add about 0.1 to 5 parts by volume of the alfalfa and / or brewer's yeast extract (liquid) thus prepared per 10 parts by volume of the culture medium.

When such a culture method of the present invention is employed, Enterococcus faecalis and Lactobacillus salivarius can be cultured at a high concentration and / or in a short time. As a result, the drug of the present invention and the above-mentioned JP-B-59-46209 can be used. The availability of Lactobacillus salivarius or Enterococcus faecalis for use in the No. drug is greatly facilitated.

Next, the present invention will be specifically described with reference to examples, but the present invention is not limited thereto. In the following examples,% indicates% by weight.

Example 1 (Bacterial culture) 20 g of alfalfa (water content 7%) and 100 m of distilled water
, And the mixture was stirred well, and then subjected to a heat extraction treatment at 100 ° C. for 60 minutes. After cooling, the precipitate was removed by centrifugation and filtration, and the residual liquid (50 m) was recovered as an aqueous alfalfa extract.

In addition, water extraction treatment was performed in the same manner as described above using 20 g of brewer's yeast powder (water content: 5.5%) instead of alfalfa to obtain 70 m of a brewer's yeast water extract.

MBB medium having the following composition (pH 6.8)
It was used.

MBB medium composition (content per medium) Baker's yeast extract 6 g Polypeptone 15 g Dextrose 20 g Soluble starch 0.5 g Sodium glutamate 2 g Cysteine hydrochloride hydrate 0.2 g Ascorbic acid 3.4 g Dipotassium phosphate 4 g Sodium acetate Hydrate 10 g Sodium carbonate 1.25 g Sodium chloride 5 g Polysorbate 1 g Tomato juice 400 m To the above MBB medium 10 m, enterococcus faecalis derived from horse intestinal tract was added at a ratio of 1.0 × 10 ⁵ / m medium. To this was added 1.0 m of the alfalfa aqueous extract obtained above or 0.5 m of the aqueous extract of brewer's yeast, and the mixture was cultured at 37 ° C for 20 hours.

Absorbance (Abs ₆₀₀ ) at a wavelength of 600 nm of the solution at the start of culture and after culture for 20 hours was measured using Bioscreen (manufactured by Labsystem Japan).

In addition, the above-mentioned Enterococcus faecalis was added to the medium without adding the aqueous extract of alfalfa or the aqueous extract of brewer's yeast, and cultivation was carried out in the same manner as above, and the absorbance (Abs ₆₀₀ ) was measured.

 The above results are shown in Table 1 below.

From the results in Table 1 above, when culturing was performed using a medium to which alfalfa or brewer's yeast extract was added, the value of Abs ₆₀₀ after 20 hours of culturing was compared to the case where a medium without the extract was used. Is larger, indicating that the growth of Enterococcus faecalis is promoted. In addition, it can be understood from this that alfalfa and brewer's yeast can increase the activity of Enterococcus faecalis in animal drugs containing Enterococcus faecalis and improve its activity.

Example 2 (Culture of bacteria) Culture was performed in the same manner as in Example 1 except that Lactobacillus salivarius IIb derived from a horse intestine was used instead of Enterococcus faecalis, and Abs ₆₀₀ of the culture solution was measured.

 The results are shown in Table 2 below.

From the results in Table 2 above, when culturing was performed using a medium to which alfalfa or brewer's yeast extract was added, the value of Abs ₆₀₀ after 20 hours of culturing was lower than when a medium without the extract was used. , Which indicates that the growth of Lactobacillus salivarius is promoted. In addition, it can be understood from this that alfalfa and brewer's yeast can enhance the activity of Lactobacillus salivarius in animal drugs including Lactobacillus salivarius and enhance its activity.

Example 3 In the same MBB medium 50 used in Example 1 above, 6.0 × 1 of Enterococcus faecalis derived from horse intestine was used.
The cells were added at a ratio of ⁰⁹ cells / medium and cultured at 35 ° C. for 8 hours to obtain a culture solution having about 1.3 × 10 ¹² cells / cell. This culture was centrifuged to collect the Enterococcus faecalis solids, dried at a temperature of 25 ° C. and a pressure of 10 ⁻³ mmHg, and dried to obtain 85 g of a dry powder of Enterococcus faecalis (water content: 2.8%).
I got Sterilized and purified corn starch was added thereto to dilute the bacterial count to about 2 × 10 ⁷ cells / g, and this was used as Enterococcus faecalis powder.

Enterococcus faecalis powder 2k obtained above
g, 2 kg of dried brewer's yeast powder (water content 5.5%) and 6 kg of alfalfa (water content 3.4%) were mixed to obtain a horse dust.

This horse powder was administered to 7 thoroughbreds (2 years old: 6 females and 1 male) at a rate of 20 g / day / head with a normal feed for 16 days, and the palatability was good and the ingestion was continued. The feces of all horses were slightly dry at the start of the test, but on the 5th day after the start of the administration, the feces of 5 horses became moderately moist, and on the 7th day after the start of the administration. Showed that all horse feces had moderate wettability.

Example 4 6.5 Lactobacillus salivarius IIb derived from horse intestine was added to the same MBB medium 50 used in Example 1 above.
The cells were added at a rate of × 10 ⁹ cells / medium and cultured at 39 ° C. for 10 hours to obtain a culture solution having about 1.2 × 10 ¹² cells / cell. This culture is centrifuged to obtain Lactobacillus salivarius II b
Was recovered as a solid, and was freeze-vacuum dried at a temperature of 25 ° C. and a pressure of 10 ⁻³ mmHg to obtain 82 g of a dry powder of Lactobacillus salivarius IIb (water content: 2.8%). Sterilized and purified corn starch was added thereto, and diluted so that the number of bacteria was about 2 × 10 ⁷ cells / g. This was used as Lactobacillus salivarius IIb bacterial powder.

Lactobacillus salivarius II b bacterial powder 2k obtained above
g, 2 kg of dried brewer's yeast powder (water content 5.5%) and 6 kg of alfalfa (water content 3.4%) were mixed to obtain a horse dust.

This horse powder was administered to three 1-year-old male Thoroughbreds and 3 2-year-old female Thoroughbreds at a rate of 25 g / day / head mixed with a normal diet, and the feeding by horses was reduced. It was taken without leaving it well, and its physical condition and fecal condition were also good.

Continuation of the front page (72) Inventor Hideo Oishi 2-4-240 Nakano-cho, Shizunai-cho, Shizunai-gun, Hokkaido (56) References JP-A-62-104552 (JP, A) JP-B-59-46209 (JP, B2) ) JP-B-59-46208 (JP, B2) JP-B-59-9146 (JP, B2) (58) Fields investigated (Int. Cl. ⁷ , DB name) A61K 35/72 A61K 35/74 A61K 35 / 78 C12N 1/20

Claims (1)

(57) [Claims] 1. An Enterococcus faecalis or Lactobacillus salivarius obtained by culturing Enterococcus faecalis or Lactobacillus salivarius in a medium containing at least one of an alfalfa extract and a brewer's yeast extract. Production method for veterinary medicine.