KR101836918B1 - Feed Composition for Mealworm, Breeding Method for Mealworm Using the Same, And Food Composition Obtained Therefrom - Google Patents

Abstract

The present invention relates to: a feed composition for mealworm; a breeding method using the same; a mealworm bred by the breeding method; and to a food composition comprising the same, and specifically, to a method for breeding the mealworm using the feed composition for the mealworm, which contains glutamic acid as an active ingredient to promote the growth of the mealworm and to enhance GABA and essential amino acid content in the body; to the mealworm bred by the breeding method; and to the food composition comprising the same.

Description

Technical Field [0001] The present invention relates to a feed composition for a wheat worm, a breeding method using the same, and an insect food composition obtained from the feed composition. [0002]

The present invention relates to a feed composition for wheat worms, a feeding method using the same and an insect food composition obtained therefrom, and more particularly to a feed composition for wheat worms containing glutamic acid as an active ingredient, a method for breeding wheat worms using the composition, (Gamma aminobutyric acid) content of the wheat worm.

In recent years, competition for securing biological resources through exploration and conservation of insect resources has become fierce, and their valuation and utilization research are being actively carried out. As part of this research, research on the development of food and medicine materials from insects has also been carried out domestically.

Tenebrionidae are insects in the beetle's neck that are common in warm, dry climates and live on mostly dry, decayed plants or animal tissues. The larvae of this species, Tenebrionidae, are about 25mm in length and cylindrical in shape, and are often called Mealworm, which is about 15 to 20 weeks old. The larval stage is relatively short, Is also very high and is a clean insect that feeds on grain.

Most of the larvae and larvae are used as food for pet birds, fish or hedgehogs. Among them, brown larvae (Tenebrio molitor, mealworm, TM) larvae are most commonly used. Brown ducks are also called brown rice gruel or soporaceae and are distributed all over the world including Korea. The brown duck contains abundant fat, protein, various kinds of amino acids, unsaturated fatty acids and minerals. Essential amino acid content is higher than pork, lamb and beans, and it is reported to be similar to beef and fish.

Brown ducks are fully transformed insects that pass through eggs, larvae, pupae, and adults. However, unlike other insects, they are relatively short in transformation period, adaptable, and easy to learn breeding techniques. Because of this short life cycle and strong survival rate, it is possible to breed with low feed costs, and it is eco-friendly because it can be used as a food source of straw and old cabbage leaves which are rarely used at the time of breeding.

The wheyworm is easy to mass-proliferate and has high nutritional value, so that there is a growing interest in the development of industrial applications such as feed and edible. Because of its high protein content, it is widely used for food in the Netherlands, Mexico, South America, Europe and China. Domestic animals have been widely used as feed for pet animals for more than 10 years and have begun breeding in insect lovers and zoos. In addition, in 2016, the KIRM has been officially registered as a general food ingredient in the KFDA, and farmers are growing in quantity for commercialization.

Γ-amino-n-butyric acid (GABA) is an inhibitory neurotransmitter of non-protein constituent amino acids composed of four carbons. In most people, it exists in the bone marrow of the brain, increases the neurotransmitter acetylcholine, and physiologically acts to promote brain function. Known effects of Gabba include prevention of stroke and dementia, strengthening concentration, improving memory, relieving insomnia and depression. In addition, neurostabilizing action, relieving hangover, relieving anxiety, lowering blood pressure, and increasing insulin effect have also been reported. In recent years, it has been shown that it has anticancer effect and it is attracting attention in the food industry. In abroad, supplements for the intake of GABA are available and marketed.

KR 101510260 B1 KR 1020170018509 A

 Holdiness M. R (1983) Chromatographic analysis of glutamic acid decarboxylase in biological samples. J. Chromatogr. 277, 1-24.  Rossetti V and Lombard A (1996) Determination of glutamate decarboxylase by high-performance liquid chromatography. Journal of Chromatography B, 681: 63-67

It is an object of the present invention to provide a feed composition for wheat worm which can improve the growth speed of the wheat worm and increase the body grain content.

The present invention aims at providing a feeding method for a wheat worm comprising feeding the wheat worm feed composition to the wheat worm.

It is another object of the present invention to provide a wheat worm which is raised according to the method for raising the wheat worm, thereby increasing the amount of garbage.

It is another object of the present invention to provide a food composition comprising the wheat worm or the extract thereof, wherein the gastric content is enhanced.

According to one aspect of the present invention, there is provided a wheat worm feed composition for promoting growth and gamma aminobutyric acid (GABA) synthesis comprising glutamic acid as an active ingredient.

In addition, the glutamic acid may be monosodium glutamate.

The glutamic acid may be contained in an amount of 5 to 50% by weight based on 100% by weight of the feed composition.

According to another aspect of the present invention, there is provided a method of raising a wheat worm, comprising feeding the feed composition to a wheat worm.

According to another aspect of the present invention, there is provided a wheat worm which is raised by the above-mentioned breeding method and has an increased amount of GABA.

According to another aspect of the present invention, there is provided a food composition containing the wheat worm or an extract thereof.

According to another aspect of the present invention, there is provided a feed additive containing the wheat worm or an extract thereof.

The feed composition for a wheat worm according to one aspect of the present invention can shorten the growth time by improving the growth speed of the wheat worm that ingests it. This makes it possible to improve the productivity of the wheat worm farm for the production of feed or food composition using the wheat worm.

In addition, by feeding the feed composition to the wheat worm, it is possible to remarkably increase the body burden of the wheat worm. Through this, it is possible to produce wheat worms with significantly increased body fat content.

In addition, a functional food composition containing a high content of GABA can be prepared using the wheat worm or the extract thereof, which has increased the GABA content.

FIG. 1 is a graph showing an activity and growth rate of a wheat worm fed with the feed composition of the present invention,
FIG. 2 is a graph showing the results of TLC analysis of the wheat-worm ethanol extract fed with the feed composition of the present invention,
FIG. 3 is a graph showing the results of TLC analysis of the wheat worm water extract fed with the feed composition of the present invention,
FIG. 4 is an HPLC chromatogram showing the concentration of GABA,
FIG. 5 is a graph showing HPLC chromatogram of a watery extract of wheat worm fed with the feed composition of the present invention.
FIG. 6 is a graph showing gene expression of cells injected with GABA cRNA,
FIG. 7 is a graph showing the concentration gradient experiment of GABA single component in cells injected with GABA cRNA,
FIG. 8 is a graph showing the results of concentration gradient experiments of wheat worm extracts fed with the feed composition of the present invention to cells injected with GABA cRNA.
FIG. 9 is a graph showing a concentration gradient experiment curve of a wheat worm extract fed with a feed composition of the present invention to cells injected with GABA cRNA,
FIG. 10 is a graph showing the results of voltage-dependency of the extract of the wheat worm obtained by feeding the feed composition of the present invention to cells injected with GABA cRNA.

The present invention relates to a wheat worm feed composition containing glutamic acid, a breeding method using the same, a wheat worm raised by the breeding method, and a food composition containing the wheat worm.

Hereinafter, the present invention will be described in more detail.

According to one aspect of the present invention, a feed composition for wheat worms is provided.

The feed composition for wheat worms of the present invention comprises glutamic acid. The glutamic acid may be monosodium glutamate (hereinafter referred to as MSG). Feeding the feed composition containing glutamic acid to the wheat worm promotes the growth and activity of the wheat worm. In addition, there is an effect of increasing GABA (gamma aminobutyric acid) and essential amino acid content. The glutamic acid may be contained in an amount of 5 to 50% by weight based on 100% by weight of the feed composition. If the content of glutamic acid is less than 5% by weight, it is difficult to exhibit the effect of addition. If the content of glutamic acid is more than 50% by weight, the production cost of the feed composition increases and the economical efficiency may decrease. Preferably, the glutamic acid is contained in an amount of 5 to 20% by weight based on 100% by weight of the feed composition in order to maximize the enhancing effect due to the content of glutamic acid while reducing the production cost of the feed composition. More preferably, it is contained in an amount of 10 to 15% by weight.

The feed composition may comprise at least one selected from the group consisting of cereals, vegetables and agricultural by-products. The cereals, vegetables, and agricultural by-products are not particularly limited as long as they are capable of eating wheat worms, and those normally added during the production of feed can be used. In order to lower the production cost of the feed composition and improve the economic efficiency, it is preferable that the feed composition includes agricultural by-products. When the feed composition of the present invention includes the agricultural by-products, the feed production cost can be reduced and the productivity can be increased. In addition, recycling of agricultural byproducts results in a natural circulation, which can be environmentally friendly. The agricultural by-products are products obtained by secondary production in agricultural production, for example, at least one selected from the group consisting of rice bran, wheat bran, cotton bran, cotton bran, jade bran, groundnut bran and beet pulp. In consideration of the eating symbol of the wheat worm, it is more preferable that wheat bran is selected. The cereals, vegetables and agricultural by-products may be contained in an amount of 50 to 95% by weight based on 100% by weight of the feed composition.

If desired, the feed composition may further comprise a feed additive. The additives may be those known in the art for the purpose of enhancing taste or functionality. More particularly, the present invention relates to a pharmaceutical composition for improving the condition, productivity and quality of a wheat worm, which comprises at least one selected from the group consisting of various antibiotics, probiotics, enzymes, organic acids, flavors, sweeteners, antioxidants, You can use one more.

According to another aspect of the present invention, there is provided a method of raising a wheat worm, comprising feeding the feed composition to a wheat worm.

There is no particular limitation on the time at which the feed composition for wheat worms of the present invention can be fed, and the feed can be fed from larva immediately after hatching to larva immediately before adult. Preferably, it is preferable to feed from immediately after hatching. In this case, the overall productivity can be increased by shortening the rearing period and reducing the effort required to deal with the wheat worm in supplying the food material using the same. In addition, when used as a food material, palatability and application range can be further improved.

The feeding period of the feed composition for wheat worms of the present invention is not particularly limited, and the feed can be fed for a period of one week to four months, preferably two to three months, in order to enhance the body burden of the worm.

By raising the wheat worm by the breeding method, a wheat worm having an increased body burden can be obtained.

The wheat worm raised by feeding the feed composition for wheat worm according to the present invention significantly improves the body burden. The gastric content in the body of the wheat worm may vary depending on an intake amount of the feed composition of the present invention, an amount of glutamate in the feed, and a feeding period of the feed composition. In addition to GABA, the above-mentioned wheat worm also increases essential amino acid content in the body.

The wheat worm having increased body burden of the present invention can be used as food itself, and can be prepared in various forms such as powder, capsule, ring, and extract.

If necessary, the wheat worm of the present invention can be used in the form of an extract obtained by extracting it. The method for separating the extract of the mildew worm is not particularly limited, but a method commonly used in the art can be used for producing the extract. For example, a method using an extraction device such as hot water extraction, immersion extraction, reflux cooling extraction, supercritical extraction, subcritical extraction, high temperature extraction, high pressure extraction and ultrasonic extraction, or a method using an adsorption resin containing XAD and HP-20 Method, but the present invention is not limited thereto. Preferably, it can be obtained by treating with an extraction solvent, extracting it with hot water, and concentrating it under reduced pressure. As the extraction solvent, for example, at least one selected from the group consisting of water, ethanol, butylene glycol, propylene glycol, dipropylene glycol and glycerin can be used. Preferably, water is used.

The wheat worm or its extract of the present invention contains a promoted content of GABA, so that it can be usefully used as an edible material for GABA ingestion or replenishment. In addition, the wheat worm or the extract thereof of the present invention can be easily produced and mass-produced, so that high-quality edible materials containing high-quality can be produced at low cost.

The wheat worm or extract thereof of the present invention can be produced with a food composition. Examples of the food composition include drinks, meats, sausages, bread, biscuits, rice cakes, chocolate, candy, snacks, confectionery, pizza, noodles, gum, ice cream, soups, drinks, tea, But is not limited thereto. In addition, it includes all functional foods in a conventional sense. The functional food refers to a food prepared by adding a wheat-worm extract to a food material, or a food prepared by preparing a wheat-worm extract in tablets, capsules, powders, suspensions or the like, and has a usual meaning Of healthy foods. The form of the food may be in the form of a powder, a granule, a tablet, a capsule, a liquid or a drink.

The wheat worm or extract thereof of the present invention can be added directly to the food or can be used together with other food or food ingredients, and can be suitably used according to a conventional method. The mixed content of the active ingredient may be appropriately determined depending on the purpose of use thereof. Generally, the content of the wheat worm or its extract in the food composition may be 0.0001 to 99.9% by weight of the total food weight.

If desired, the food composition of the present invention may further comprise ingredients that are conventionally added in food preparation. For example, there may be mentioned nutrients, vitamins, minerals (electrolytes), flavors, coloring agents, thickening agents, pectic acid, pectate salts, alginic acid, alginates, organic acids, protective colloid thickening agents, pH adjusting agents, stabilizers, preservatives, , Carbonating agent, pulp, and the like, but is not limited thereto.

The ingestion of the food composition of the present invention can exhibit the effects of improving nervous stability, relieving stress, improving memory, lowering blood pressure, relieving depression, preventing stroke and dementia, insomnia, obesity, menopausal disorder and diabetes. In addition, wheat worm is a high-protein food containing a large amount of unsaturated fatty acids. It can take various essential or non-essential amino acids through its ingestion, so that it can maintain the healthy body by exercising the metabolism or promoting function of the human body. There is a health improvement effect.

The wheat worm or its extract of the present invention can be produced as a feed or feed additive. The feed additive is added to the feed of livestock and can be added to the livestock for specific purposes so as to exert various physiological functions in vivo. Examples of the feed additives include, but are not limited to, probiotics, enzymes, minerals, antibiotics, and vitamins. The feed or feed additive may be added to animal feed fed to cattle, pigs, chickens or the like or aquaculture feed fed to aquatic organisms.

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in more detail with reference to the following examples. It will be apparent to those skilled in the art that these embodiments are illustrative of the present invention and are not intended to limit the scope of the appended claims and that various changes and modifications may be made to the embodiments within the scope and spirit of the invention , And it is natural that such variations and modifications fall within the scope of the appended claims.

Example

<Confirmation of growth promoting effect of MSG supplemented feed>

Feeds of the present invention containing glutamic acid were prepared at the blending ratios shown in Table 1 below. MSG was used as added glutamic acid.

bran MSG Comparative Example 100% 0% Example 1 95% 5% Example 2 90% 10% Example 3 85% 15%

The brown duck, which is a subject of rearing, was sieved using adult sperm and larvae, and only four eggs were prepared in four breeding boxes. The prepared diets were added to each breeding box and breeding was carried out for 60 to 90 days while maintaining the temperature at 25 to 29 캜 and the humidity of 50 to 70%.

As a result, the wheat worms belonging to the feed group of the MSG supplemented groups exhibited better activity and growth rate, as compared to the comparative feed ward group to which no MSG was added. The results of feeding the diets of the above Comparative Examples and Example 2 are shown in Fig.

<Nutritional analysis of wheat worms fed MSG diet>

The general components of wheat worms were analyzed by the AACC method. As for the samples, the larvae of the feed group of the comparative example were used, and the basis weight was determined by using atmospheric pressure heating drying method, semi micro-kjeldahl method for moisture, Soxhlet method for fat content, and ash content at 550 ° C. for 6 hours. And the results are shown in Table 2 below. In Table 2, the unit of each component is g / 100g.

Content by ingredient moisture 4.82 Ash 3.36 Crude fat 35.38 Crude protein 55.96 carbohydrate 1.48

As a result of the general composition analysis, the crude fat content of the wheat worm was 35.38 g / 100 g. Considering the result of the fatty acid measurement described below, it is considered that crude lipid was also high because of the high unsaturated fatty acid content of the wheat worm.

The amino acid content of the wheat worm was analyzed using an automatic analyzer. Specifically, 20 ml of anhydrous alcohol was added to 2 g of the wheat-worm sample for each of Comparative Examples and Examples, homogenized, and centrifuged at 3000 rpm for 30 minutes using a centrifugal separator to separate the supernatant (A) (B), which was obtained by homogenization and centrifugation again in the same manner as above, was mixed with the supernatant A, and the supernatant A was mixed with the supernatant A (WB-2001, Heidolph , Kelheim, Germany) to remove the alcohol in the supernatant. 8 ml of HPLC water was added to the concentrated solution, and 0.2 g of sulfosalicylic acid reagent was dissolved. The mixture was allowed to stand at 4 ° C for 1 hour, centrifuged again at 3000 rpm for 30 minutes, and the supernatant was collected, adjusted to 10 ml with water for HPLC, and filtered through a 0.2 μm membrane filter (Whatman, Maidstone, UK) Followed by filtration to prepare a sample solution for analysis. The free amino acid analyzer (S430, SYKAM, Eresing, Germany) was used for the analysis of free amino acids. The buffer used was analyzed for two hours per sample using lithium buffer solution and ninhydrin reagent. . The analysis results of the free amino acids are shown in Table 3 below. In Table 3, the unit of each component is g / 100g.

Comparative Example Example 1 Example 2 Example 3 Phosphoserine N.D. N.D. N.D. N.D. Taurine N.D. N.D. N.D. N.D. Phosphoethanolamine N.D. N.D. N.D. N.D. Urea N.D. N.D. N.D. N.D. Aspartic acid 2,177.67 2,740.50 2,720.86 2,823.69 Hydroxyproline N.D. N.D. N.D. N.D. Threonine 892.53 1,670.70 1,114.31 1,209.37 serine 1,816.45 1,964.47 2,306.88 2,256.01 Asparagine N.D. N.D. N.D. N.D. Glutamic acid 3,598.14 4,680.99 4,545.63 4,765.58 Sarcocine 27.26 77.56 41.68 51.05 alpha -aminoadipic acid N.D. N.D. N.D. N.D. Proline 2,728.42 3,737.12 3,451.11 3,296.31 Glycine 1,642.52 2,209.28 2,089.31 2,083.62 Alanine 2,664.58 3,419.53 3,346.74 3,285.08 Citrulline N.D. N.D. N.D. N.D. alpha-aminobutyric acid N.D. N.D. N.D. N.D. Valine 2,022.74 2,673.34 2,554.77 2,605.48 Cystine 216.92 206.95 352.49 244.69 Methionine 431.99 568.80 531.05 570.86 Cysthathionine N.D. N.D. N.D. N.D. Isoleucine 1,367.19 1,794.55 1,737.30 1,785.81 Leucine 2,372.90 3,097.30 2,997.32 3,064.55 Tyrosine 2,667.50 3,524.47 3,165.65 3,469.39 피 phenylalanine 1,345.26 1,727.16 1,637.70 1,729.14 β-domain 4.10 4.43 5.65 4.85 β-aminoisobutyric acid 15.27 N.D. N.D. N.D. Histidine 1,045.15 1,372.55 1,309.80 1,350.75 1-methylhistidine 5.12 8.81 6.15 6.67 3-methylhistidine N.D. N.D. N.D. N.D. Carnosine N.D. N.D. N.D. N.D. Anserine N.D. N.D. N.D. N.D. Tryptopan N.D. N.D. N.D. N.D. Hydroxylysine 3.875 8.380 5.472 6.818 Ornitine 15.904 29.461 18.006 23.299 Lysine 2,063.89 2,714.06 2,518.36 2,793.01 Ethanolamine 86.34 121.19 114.62 112.36 Arginine 1,635.73 2,115.78 2,124.18 2,190.57 Total 30,864.25 40,497.66 38,717.28 39,751.49

Table 3 shows that the content of various essential amino acids was increased in the worm body fed with the feed composition of the present invention. Glutamic acid was the most abundant amino acid in food, followed by Aspartic acid, Proline, Alanine, Valine, Leucine and Tyrosine. Aspartic acid has been reported to be able to be produced in MSG, and it is considered that the content of Aspartic acid is higher in the Examples than the Comparative Example which is the control group due to the feeding effect of the feed of the present invention including MSG.

In order to analyze the fatty acid composition of the wheat worm, 2 g of the wheat worm sample is extracted with chloroform-methanol, filtered, and 100 mg of the fat concentrated under reduced pressure is taken in an eggplant-shaped flask and mixed with 3 ml of 1N-KOH ethanol solution And then 3 ml of 14% (v / v) BF3-Methanol was added thereto. The mixture was heated to 80 ° C for 5 minutes with a reflux condenser and methylesterized. To this solution was added 3 ml of a saturated solution of NaCl, and 1 ml of hexane was added. The mixture was shaken and transferred to a test tube. The upper layer was separated and washed with anhydrous Na ₂ SO ₄ , And 0.5 ml was collected in a vial and analyzed by GC (Shimadzu GC-17A, Shimadzu co., Japan). The analysis results of the fatty acids are shown in Table 4 below. In Table 4, the unit of each component is g / 100 g of TFA (total fatty acids).

Comparative Example Example 1 Example 2 Example 3 Saturated Butyric acid (C4: 0) 0.00 0.00 0.00 0.00 Caproic acid (C6: 0) 0.00 0.00 0.00 0.00 Caprylic acid (C8: 0) 0.00 0.00 0.00 0.00 Capric acid (C10: 0) 0.01 0.01 0.02 0.00 Undecanoic aicd (C11: 0) 0.00 0.00 0.00 0.00 Luric acid (C12: 0) 0.37 0.39 0.42 0.37 Tridecanoic acid (C13: 0) 0.07 0.08 0.14 0.05 Myristic acid (C14: 0) 3.32 3.60 3.60 3.41 Pentadecanoic acid (C15: 0) 0.33 0.36 0.33 0.00 Palmitic acid (C16: 0) 8.96 9.50 9.66 9.59 Heptadecanoic acid (C17: 0) 5.01 4.94 4.54 6.35 Stearic acid (C18: 0) 0.00 0.00 0.00 0.00 Arachidic acid (C20: 0) 0.00 0.00 0.00 0.00 Heneicosanoic acid (C21: 0) 0.00 0.00 0.00 0.00 Behenic acid (C22: 0) 0.00 0.00 0.00 0.00 Tricosanoic acid (C23: 0) 0.18 0.16 0.18 0.25 Lignoceric acid (C24: 0) 0.00 0.00 0.00 0.00 sub Total 18.25 19.04 18.89 20.02 Monounsaturated Myristoleic acid (C14: 1) 0.07 0.07 0.08 0.06 cis-10-Pentadecenoic acid (C15: 1) 0.00 0.00 0.00 0.00 Palmitoleic acid (C16: 1) 1.49 1.70 1.61 1.36 cis-10-Heptadecenoic acid (C17: 1) 0.17 0.24 0.17 0.14 Elaidic acid (C18: 1n9t) 0.00 0.00 0.00 0.00 Oleic acid (C18: 1n9c) 24.36 22.24 25.27 6.50 cis-11-Eicosenoic acid (C20: 1) 0.00 0.00 0.00 0.00 Erucic acid (C22: 1n9) 0.00 0.00 0.00 0.00 Nervonic acid (C24: 1) 0.46 0.52 0.23 0.23 sub Total 26.55 24.77 27.36 8.29 Polyunsaturated Linolelaidic acid (C18: 2n6t) 45.51 44.89 42.65 61.05 Linoleic acid (C18: 2n6c) 9.62 11.06 11.00 10.57 cis-11,14-Eicosadienoic acid (C20: 2) 0.00 0.03 0.00 0.00 cis-13,16-Docosadienoic acid (C22: 2) 0.00 0.00 0.00 0.00 aicd (C18: 3n6) 0.00 0.00 0.00 0.00 Linolenic aicd (C18: 3n3) 0.00 0.00 0.00 0.00 cis-8, 11,14-Eicosatrienoic AICd (C20: 3n6) 0.00 0.00 0.00 0.00 cis-11,14,17-Eicosatienoic acid (C20: 3n3) 0.00 0.00 0.00 0.00 Arachidonic acid (C20: 4n6) 0.07 0.08 0.10 0.07 cis-5,8,11,14,17-Eicosapentaenoic acid (C20: 5n3) 0.00 0.13 0.00 0.00 cis-4,7,10,13,16,19-Docosahexaenoic acid (C22: 6n3) 0.00 0.00 0.00 0.00 sub Total 55.20 56.19 53.75 71.69

The content of oleic acid and linolelaidic acid was higher in the unsaturated fatty acid of mullum. Oleic acid is known to have the effect of improving the health of the human body such as memory enhancement and prevention of dementia. Since the maintenance ingredient of the wheat worm contains a high content of high-functional unsaturated fatty acid, .

<Standardized manufacturing process of functional food materials using MSG-fed wheatgrass>

The manufacturing process for standardization as food material was divided into the steps of raw material weighing step, raw material washing step, freeze drying step and raw materializing step.

The feeds were prepared by mixing wheat bran and MSG in a weight ratio of 9: 1. 100kg of wheat worms fed with the prepared feeds were used as 3 raw materials. After washing for 10 minutes with warm water at 30 ° C, it was lyophilized at -70 ° C and 5 mTorr. The yield of the insect powder was 42 ㎏ and the average content of GABA was 13.96 ㎎ / g.

Experiment result
(GABA, unit mg / g) Average
(mg / g) 1 Lot 2 Lots 3 lots Milworm 13.92 14.92 13.05 13.96

<Functional food material property test using MSG-fed wheat worm>

The sugar content and pH were measured by homogenizing the insect powder and distilled water at a ratio of 1:10, homogenizing the mixture using a homogenizer (IKA T 25 digital ULTRA-TURRAX, IKA Ltd., Staufen, Germany) Were measured with a pH meter (SevenEasy 520, Mettler-Toledo Ltd., Schwerzenbach, Switzerland) and a pH meter (Pal-1, Atago Co., Ltd., Tokyo, Japan). Lightness, redness, and yellowness of the insect powder were measured using a colorimeter (CR-300, Minolta Co, Osaka, Japan). The average value of the measurement was repeated three times for each sample and the results are shown in Table 6 below.

pH Sugar content
(Brix) Chromaticity L * (yellow degree) a * (redness degree) b * (brightness) 6.66 3.75 45.35 2.31 10.50

The texture of the insect powder was measured with a texture analyzer (TX-XT2, Stable Micro System, Haslemere, Surrey, UK) using a P / 50R cylinder probe. After loading 4 g of sample into a cylinder of diameter and height of 3.5 cm, the pre-test speed, test speed, posttest speed was 2.0 mm / s, distance was 10.0 mm, trigger force was 5.0 g and compression mode. Shown in Table 7 below.

Hardness (g) Springiness Gumminess (g) Chewiness (g) Cohesiveness 2100 0.17 139.40 24.07 0.07

Protein, fat, saturated fat, trans fat, cholesterol, and sodium content of the insect powder were analyzed according to the method described in the Food and Drug Administration, and the results are shown in Table 8 below.

content Unit (Daily nutritional intake%) calorie 327.7 kcal / 100g salt 110.5 mg / 100 g (6) Carbohydrate Mall 7.0 g / 100 g (2)   sugars 1.6 g / 100g  Fat 33.3 g / 100 g (65)  Saturated fat 8.1 g / 100 g (54)  trans fat Non-detection g / 100g cholesterol 44.0 mg / 100 g (15) protein 53.0 g / 100 g (96)

Antioxidant activity was determined by measuring the radical scavenging activity. DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging activity was obtained by mixing 2 g of sample with 20 ml of 80% methanol and homogenizing for 3 minutes using homogenizer. The supernatant was separated by centrifugation, and the methanol extract was filtered using a filter paper. The DPPH solution was added to 150 μl of the sample, reacted in the dark room for 30 minutes, and absorbance was measured at 517 nm. As a result, the antimicrobial activity of the wheat worm was shown to be DPPH radical (Trolox (uM)) 7.57.

As a result of the test, it can be confirmed that the larva of the wheat worms fed with the feed according to the present invention has characteristics suitable for food and can be used for the ingestion of GABA. In addition, protein content, which is an important nutrient of insects, is so high that it can be absorbed in a small amount to a large amount of protein intake.

<Confirmation of production of gabas and physiologically active substances in the body of MSM-fed wheat worms>

Experiments were conducted to determine the amount of GABA production in wheat worms according to MSG concentration and drying method. The larvae were cultured for 90 days in a medium containing various concentrations of MSG. Extract samples of wheat worms were prepared by different drying methods and extraction solvents.

division Drying method Extraction solvent MSG concentration in feed
[% (w / v)] Production Example 1 Freeze-dried water
0 Production Example 2 5 Production Example 3 10 Production Example 4 15 Production Example 5 ethanol 0 Production Example 6 5 Production Example 7 10 Production Example 8 15 Production Example 9 hot air dry water 0 Production Example 10 5 Production Example 11 10 Production Example 12 15 Production Example 13 ethanol 0 Production Example 14 5 Production Example 15 10 Production Example 16 15

Each sample was pulverized and extracted with a extraction solvent at a rate of 1.5 g / 10 ml at 30 ° C for 24 hours. After extraction, 1 ml of each sample was taken and centrifuged (13,000 rpm, 3 minutes) to remove impurities. The physiologically active substance was purified and purified.

GABA qualitative analysis was performed using Thin-layer Chromatography (TLC). Holdiness M. R (1983) Chromatographic analysis of glutamic acid decarboxylase in biological samples. J. Chromatogr. 277, 1-24. Reference is made to the literature. 0.5 μl of each sample was loaded onto a silica gel TLC plate and then developed using a developing solvent of n-butanol-acetic acid-water (5: 3: 2, v / v / v). After the second development, it was heated and dried at 110 ° C for 10 minutes using 0.2% (w / v) ninhydrin (in ethanol) color developing reagent. The results are shown in Figures 2 and 3. 2 and 3, 1% (w / v) of GABA was used as a control.

2 and 3, GABA was not confirmed in the extract using ethanol as an extraction solvent, and GABA was extracted when it was extracted with water. GABA content was significantly increased when the feed MSG concentration was 10 ~ 15% (w / v).

Quantitative analysis of GABA was performed by PITC (phenylisothiocyanate) derivatization. Rossetti V and Lombard A (1996) Determination of glutamate decarboxylase by high-performance liquid chromatography. Journal of Chromatography B, 681: 63-67. Add 10 μl of each sample to the tube, remove the solvent through a vacuum concentrator, add 20 μl of ethanol-water-triethylamine (TEA) (2: 2: 1, v / v / v) And the solvent was removed again through a vacuum concentrator. 50 μl of ethanol-water-TEA-PITC (7: 1: 1: 1, v / v / v / v) solvent was added to the residue and the PITC derivatization was carried out at room temperature for 20 minutes. The reagent was removed through a vacuum concentrator and dissolved in 600 μl of solvent A, followed by HPLC analysis. The results of HPLC measurement according to GABA concentration are shown in FIG. 4, and the results of HPLC measurement of each of the water extracts of wheat worm are shown in FIG. The results of GABA quantitative analysis are shown in Table 10.

division GABA concentration
(mg / mL) Extract volume
(mL) Total GABA
content GABA content
(mg / g) Production Example 1 0.009 10 0.09 0.06 Production Example 2 0.301 3.01 2.01 Production Example 3 2.238 22.38 14.92 Production Example 4 2.423 24.23 16.15 Production Example 9 0.000 0.000 0.00 Production Example 10 0.135 1.35 0.90 Production Example 11 0.174 1.74 1.16 Production Example 12 0.355 3.55 2.37

As shown in Table 10, the freeze-dried samples contained 0.009, 0.301, 2.238 and 2.423 mg / mL of GABA in the MSG-free sample, 5%, 10% and 15% MSG, respectively.

Tables 9 and 10 show that MSG fed in the breeding of wheat worms plays an important role in the generation of GABA, which is an indirect result of the presence of microorganisms growing in the worms. As a result of feeding MSG concentration differently, GABA production did not increase linearly according to MSG concentration, and it was confirmed that high concentration of MSG did not produce high concentration of GABA even when added to feed. The production of useful substances by microorganisms seems to be different from that of enzymatic reactions and that the production of metabolites by adaptation to microorganisms and environment and the inherent characteristics of microorganisms do not seem to produce linear GABA according to MSG concentration . Therefore, considering the economical efficiency, it was confirmed that it is necessary to examine the breeding condition for effectively producing GABA by adding low concentration of MSG.

<EVALUATION OF EMERGENCY OF GABA INGREDIENTS IN THE MILS WRONGED WITH MSG>

After GABA receptor expression, GABA potency was evaluated. RNA transcription was performed for receptor expression. Plasmid DNA linearization was performed by adding DW to 1.2 μl of DNA, 1.2 μl of 10 × buffer, 1 μl of restriction enzyme and 1 μl of restriction enzyme, adding DW at 37 ° C for 2 hours, adding 70 μl after 2 hours to 100 μl, (100 μl) of phenol (pH 8) was added, and phenol extraction was carried out after centrifuging at 12,000 rpm for 5 minutes. The supernatant was transferred to a new ET tube and stored at -20 ° C for 30 minutes at a volume of 0.1 × NaOAC and twice the volume of 100% EtOH. The supernatant was then centrifuged for 20 minutes at 13,000 to 14,000 rpm and 4 ° C, did. 400 μl of 70% (v / v) EtOH was added, and the supernatant was again removed by centrifugation at 13,000 rpm for 5 minutes at 4 ° C. At this time, once again centrifuged for 1 minute, the EtOH was removed as much as possible and dried for 10 minutes. After drying, the pellet was dissolved with 6 μl DEPC.

For in vitro transcription, dissolve the pellet, add 2 μl of 2X NTP, 2 μl of 10 × buffer, and 2 μl of RNA polymerase, and incubate at 37 ° C for 2 hours. Add 1 μl of DNase and incubate at 37 ° C for 15 minutes . After incubation, 115 μl of DEPC and 15 μl of NaOAC were added, 150 μl of phenol was added, and the supernatant was transferred to a new ET tube by centrifugation at 12,000 rpm for 5 minutes.

Two-fold volume of isopropanol was added to the transferred tube, stored at -20 ° C for 30 min, centrifuged at 13,000 ~ 14,000 rpm and 4 ° C for 25 min, and the supernatant was removed. At this time, once again centrifuged for 1 minute, the EtOH was removed as much as possible and dried for 10 minutes. After dissolving the pellet with 11 μl of DEPC and confirming it by electrophoresis, it was divided into several pieces and stored at -80 ° C deep freezer to synthesize GABA gene cRNA.

Then, oocytes were injected to inject synthetic cRNA. Xenopus oocytes were removed from the culture after sufficient hibernation, and oocytes removed were placed in OR2 buffer (5 mM HEPES at pH 7.6 and 18 ° C, 82.5 mM NaCl, 2.5 mM KCl, 1 mM MgCl ₂ ) Treated with 1 unit of collagenase (2 mg) and shaked on shaker for 1 hour. After 1 hour, Xenopus oocytes were first washed with OR2 buffer and then with ND96 buffer (5 mM HEPES, 96 mM NaCl, 2 mM KCl, 1 mM MgCl ₂ , 1.8 mM CaCl ₂ ) healthy oocytes with a clear border between vegetable poles and an animal pole as a whole were selected.

Use a Narishige's PC-10 micropipette puller to make injection needles for the injection tube. PC-10 micropipette puller can adjust the sharpness of the needle by changing the temperature. Needles were made by heating to 89.9 ℃ for 1step and 50.3 ℃ for 2 steps to fit the oocyte pipette equipped in the laboratory. The needle was slanted or flattened to 4μm using Narishige MF-900 microforge. The bruised needle was filled with mineral oil, inserted into an oocyte pipette, and cRNA was injected into Xenopus oocytes at 40ng. After injection, the cells were placed in ND96 buffer and stored in a shaker at 18 ° C. The RNA injected into the oocytes was expressed 1-2 days later, and the buffer was changed twice a day, morning and evening to maintain the state of oocytes.

The electrophysiological experiments were performed by two microcrocrode voltage clamp method. Place one of the oocyte in ^{plexiglass recording chamber 1.8 mM Ca 2 +} frog Ringer's solution ND96 ( composition components mM: 115 NaCl, 2 KCl, 1.3 Na 2 HPO 4, 1.8 CaCl 2; pH 7.4) with a flow rate of 3-7 per minute ㎖ . The resistance of the two glass electrodes was adjusted to within 1-3 MΩ by filling 3 M KCl solution. Oocyte clamps (OC 725C, Warner Instruments, Hamden, CT, USA) were used and the oocyte with a basal current of 0.2 μA or more was not used in the experiment. To measure the maximum current induced by GABA through the GABA receptor, 100 uM GABA in the previous study was used and the current flowed in the ND96 solution for 2 min was measured beforehand. GABA was administered for about 10 to 30 seconds until the peak current was reached. It was confirmed that the baseline values were restored before and after each experimental current, and it was confirmed that the reference value recovered more than 90% in order to exclude the residual effect due to the previous drug administration. It took 5 ~ 30 minutes of recovery period between experiments. GABA was administered via a miniature solenoid valve (LFAA series, Westbrook, CT, USA) under computer control using Clampex v 5.7 (Axon Instruments, Burlingame, CA, USA) The solution exchange time of this valve was 0.5 seconds.

In order to confirm the expression of the GABA receptor gene after culturing for 2 days after the injection of the GABA receptor gene into the cells, the GABA receptor monoclonal antibody, which was purchased from Sigma and having a purity of 99% or more, was treated to confirm the cell expression of the GABA receptor. This is shown in Fig.

100 μM of the pure GABA single component was treated to confirm the expression. However, in order to confirm the treatment of the experimental group, the experiment was conducted in w / w unit. Therefore, since the molecular weight of GABA is 101.12, the mass of 100 uM of pure GABA single substance is 10.312 ug / ml. Therefore, the wormworm extract of about 10 ug / ml was isolated and treated in ND96 buffer used for electrophysiological experiments. Ten milligrams of Millworm extract was prepared by pulverizing 10 ml of water, centrifuging after O / N incubation, separating supernatant and mixing with ND96 buffer. After confirming the expression level, a concentration gradient dependence test was conducted and the results are shown in Fig. GABA-expressing cells were confirmed to have cell activity expressed by pure single substance GABA, and it was confirmed that they exhibited activity in a concentration-dependent manner, and their activity was regulated by reversible action.

Thus, the control of the activity was confirmed by using the water extract of the wheat worms fed with MSG. After dissolving in the solvent of extracts of Millworm, the supernatant was separated by centrifugation, and diluted in ND96 buffer for electrophysiological experiments. The first GABA single component was treated with GABA receptor expressing cells to confirm the receptor activity and then the concentration of the honey extract was increased to confirm the action. The results are shown in FIG.

Referring to FIG. 8, it was confirmed that the wheat worm extract activates the GABA receptor, and this action acts in a concentration-dependent manner and acts reversibly. In order to confirm the concentration-dependent properties, the concentration-based effects were analyzed and generalized, and the concentration dependence was confirmed by Hill equation analysis showing the effect of the concentration-dependent effect. The results are shown in FIG. 9 and Table 11. The equation and the analytical values are shown in Table 12.

Wheat worm extract concentration
(ug / ml) Average error One 1.2 0.1 3 5.1 2.1 10 10.7 8.5 30 31.2 15.4 100 51.2 22.4 300 82.5 15.52 1000 100.1 6.1 3000 98.2 5.2

Equation y = A1 + (A2-A1) / (1 + 10 ^ ((LOGx0-x) * p))) Adj. R-Square 0.99429 Value Standard Error mealworm A1 -1066.10776 16664.49643 A2 98.18555 2.87081 LOGx0 -219.50798 1649.57729 p 0.00472 0.00475 EC50 3.10E-220

Referring to FIG. 9 and Table 11, smooth concentration-dependent efficacy can be confirmed through a curve.

In addition, a voltage dependency study was performed. The purpose of this study is to determine whether the action of GABA is a physiological, natural, and specific action, in order to confirm whether the Wormworm extract can physically and safely control the activity of the GABA receptor. If the wheat-worm extract acts physically and irregularly on the activity of the GABA receptor, for example, it does not act in a voltage-dependent manner but acts independently, which may result in physical destruction and non-specific action on the GABA receptor rather than biological action.

As a control, the reverse potential voltage was checked before treatment with GABA alone and the GABA single substance was treated. After the reverse potential voltage was changed or shifted when treated with the wormwood extract, it was not a physiological effect but a physical nonspecific action . For example, the reverse potential voltage will change if it acts on carbohydrates outside the cell membrane as well as the action of specific substances on the cell membrane or nonspecific action on the membrane protein.

Referring to FIG. 10, it can be seen that the treatment of the extract of the wheat worm exhibits the action of the receptor while maintaining the change of the reverse potential voltage. This shows the biological correct action of GABA contained in the wormworm extract of the present invention.

It can be confirmed that the enhanced GABA in the worm body fed with the feed composition containing glutamic acid according to the present invention acts biologically accurately on the GABA receptor as in the conventional GABA. As a result, it can be confirmed that the wheat worms fed with the glutamate feed composition of the present invention are safe as a food material for GABA supplementation and ingestion.

Claims (7)

A composition for enhancing growth and gamma aminobutyric acid (GABA) incorporating monosodium glutamate as an active ingredient.
delete The method according to claim 1,
Wherein the monosodium glutamate is contained in an amount of 5 to 50% by weight based on 100% by weight of the feed composition.
A method for breeding wheat worms, comprising feeding the feed composition of claim 1 or 3 to a wheat worm.
The wheat worm according to claim 4, wherein the wheat worm is raised by the breeding method to increase the content of GABA.
A food composition comprising the wheat worm of claim 5.
A feed additive comprising the wheat worm of claim 5.

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