KR100543242B1 - Precess of enzyme health food having high activity by microbial fermentation - Google Patents

Abstract

In the present invention, after soaking and increasing the grains of soybean, brown rice, white rice, barley, barley, black beans, etc., it is used in traditional fermented food and produces aspergillus duckase ( Aspergillus oryzae) , lyso Two strains of Rhizopus microsporus and Bacillus megaterium SMY-212 strains, which are protease-producing bacteria registered as patent strains, were sequentially cultured and fermented, followed by drying and addition of functional foods. It relates to a method for producing a high activity, high functional enzyme food through the molding process, and to the enzyme food.

 Aspergillus aurise, Rifupus microsporus, Bacillus megaterium, proteases, alpha-amylase, beta-amylase, chitosan oligosaccharides

Description

Preparation of enzyme activity food having high activity by microbial fermentation

1 is a photograph showing the blood clot dissolving ability of the functional enzyme food of the present invention.

Figure 2 is a simplified process diagram for producing a functional enzyme food of the present invention.

The present invention relates to a method for producing an enzyme food fermented by culturing a strain in cereals and a functional enzyme food thereof, and more particularly to a fungus Aspergillus duckling, which produces a large amount of proteases and amylases, and a pretreatment process of cereals. ( Aspergillus oryzae) , Rhizopus microsporus ( Rhizopus microsporus) and high-activity comprising the step of inoculating fermentation in cereals Bacillus megaterium SMY-212 registered as a patent strain A method for producing a functional enzyme food and a functional enzyme food.

In general, enzymes are organic compounds produced from various microorganisms that catalyze chemical reactions in living organisms. Cells of all living things, such as animals and plants, contain various kinds of enzymes. Life is sustained. In other words, enzymes are widely distributed in cells and are involved in life's chemical reactions.

 Foods designed to enhance the function of such enzymes are enzyme foods, which produce various biologically active substances and nutrients through fermentation and ripening of enzymes and microorganisms of grains, fruits, and vegetables themselves, and multiply beneficial bacteria to digest and absorb It is rich in a variety of trace elements and bioactive substances that help the human body (Huh, SH and Kim, MH The modern health and health food, Hongikjea press. Korea, 35-36 (1997)).

The enzyme food is largely classified into grain enzyme food, embryo enzyme food, fruit vegetable enzyme food and the like. Grain enzyme foods are processed foods containing more than 60% cereals, embryo enzyme foods are processed foods containing more than 40% of grain embryos, fruit vegetable enzyme foods are processed foods containing more than 60% fruit vegetables. In addition, there are other enzymatic foods processed by using ingredients other than cereal grain embryos and fruit vegetables as main raw materials (60% or more of vegetable raw materials). The present invention belongs to grain enzyme food of the above classification.

General manufacturing process of enzyme foods is to wash, select and increase the raw foods to remove the natural bacteria contained in the raw materials, and then to add the high starch or protease-producing enzymes for 36-48 hours under certain conditions. Incubate step. After the cultivation is completed through the step of drying under temperature conditions without the killing of enzymes and various useful microorganisms produced by the cultivation and the addition of micronutrients such as vitamins to the molding.

The efficacy of the enzyme food depends on the activity of the enzyme and its maintenance. Therefore, the conditions of the enzymatic process (fermentation and maturation) such as temperature, humidity or air control must be very demanding and strictly observed, and it is sensitive to hygienic conditions such as incorporation of various germs, so that the product can not Care should be taken, such as blocking contact. However, the current quality standard has only specified the compositional standards, properties, moisture and crude protein content and E. coli, and only the activities of alpha-amylase and protease are positive for the enzyme, the most important factor. There is a problem that defines only things (Ref. Eun-joo Lee, Chul-ho Lee, Korean Journal of Food Science, 461-468 (2001)).

Due to the lack of such enzyme-related regulations, the results of enzymatic activity measurements of commercially available enzyme foods show a wide variety of enzyme activities, and health foods with high enzyme activity were judged to be very rare.

In order to solve the above problems and to provide enzyme foods with superior enzyme activity than conventionally marketed enzyme foods, the pretreatment step of immersing and increasing the grains of main ingredients in various ways to show the optimum enzyme activity. , Highly active high functional enzymes, including the step of inoculating a variety of fungi that can be used as an edible fungus and edible bacteria ( Bacillus megatrium SMY-212) isolated from the traditional doenjang and fermentation, ripening step It provides a method for producing food and functional health food prepared by the above method.

Grains used in the present invention include soybeans, black beans, brown rice, yulmu and rice barley.

Looking at the ingredients contained in the grains briefly, soybeans contain a large amount of functional ingredients such as dietary fiber, saponins, soybean peptides, isoflavones, lecithin, genistein, and the like. It is known that dietary fiber is effective in facilitating stool, removing cholesterol, and diabetes. Saponin is known to lower blood cholesterol and inhibit lipid peroxidation in the body. . Soybean peptides are effective in preventing hypertension by lowering blood pressure. Isoflavones are similar in structure to female hormones and are effective in preventing osteoporosis in women, and are also effective in diabetes and women's diseases. Lecithin is known to be effective in preventing dementia and obesity. . Zenithinin, which has a structure similar to isoflavone, has been reported to be effective in gastric cancer, colorectal cancer, and prostate cancer, including breast cancer.

Anthocyanin contained in black soybeans has antioxidant activity and is effective for night blindness.Brown rice is rich in free radicals, which releases toxins in the body, and it is good for nutrition because it is rich in vitamins and minerals. Effective in and diet.

The main characteristic of Yulmu is that it is excellent for diuretic action and also has abundant amino acids and is effective for diet. Barley is rich in vitamins, botanicals and phytins to promote digestion and improve the skin problems caused by intestinal dysfunction.

When fermented by edible microorganisms in the cereal grains, it is converted into nutrients that are easily absorbed, and high nutrients that are more physiologically active by adding high active enzymes and bioactive substances such as vitamins and minerals produced by the microorganisms. Contains nutritional foods.

All ingredients contained in the high-enzyme functional health food of the present invention are separated from high-quality cereals and traditional fermented foods produced in Korea and are functional additives used as food ingredients and food ingredients that have proven stability. It is completely harmless and its stability is excellent.

The grains are preferably in the ratio of 30% soybean, 15% black soybean, 15% barley, 25% brown rice, and 15% molybdenum. However, the present invention is not limited to the types and compounding ratios of the grains.

Method for producing a high-enzyme food of the present invention is a pretreatment step of immersing and increasing the grains of the main raw material; Culturing the seed; Inoculating the cultured spawn on the increased grain; Fermenting cereal seeded seed; Drying and powdering the fermented grains; It comprises the step of adding an additive to the powdered fermented grains and molding packaging the product.

The first pretreatment step is to remove various germs already contained in the grains, and consists of sterilizing the composition grains several times with natural groundwater and sterilizing them in a sterilizer at a constant temperature and pressure conditions. Preferred temperature and pressure ranges in the sterilization process is 100 to 130 ℃ and 1.2 to 1.7 atm.

The second stage spawn culture step is a step of culturing fungal and bacterial strains under certain conditions. First, in order to find a fungus showing optimal activity, it is a fungal strain that can be used for food, as a fungal strain (KCTC), which was distributed from Kukki ( Aspergillus oryzae 'KCTC-6373', Aspergillus oryzae 'KCTC-6292', Aspergillus awamori ' KCTC-6915 ') and two types of hair fungus ( Rhizopus microsporus 'KCTC-6969' and Rhizopus microsporus ' KCTC ' -1778). Activity results according to the above experiments were described in detail in Example 2. Since the fermented foods having similar enzymatic activity can be produced only by inoculating the same amount of fungi spawn on the increased grains, the present inventors obtained only the spores of the fungus among the cultured fungi cells and used them as spawns. When the spore formation time of H. pylori and fur fungus was examined, it was 4-7 days for H. pylori and 4-6 days for hair fungus. When spores were obtained and inoculated into cereals, it showed the fastest growth. In order to obtain the spores, the preformed fungi were plated on a PDA (potato dextrose agar) plate medium and incubated in a 28 ° C. incubator for 4-7 days. When spores were cultured and yellow spores were cultured on plate media for 4 days, the yellow spores showed green spores when cultured for 4 days or longer, and hairy fungi initially showed only white mycelium. When it came out black. After sterile distilled water was poured onto the mold plate medium showing the specific color, the spores were completely scratched with a smear of blot, and the spore suspension suspended in sterile distilled water was passed through a sterile gauze to remove the mycelium. Microscopic examination of the spore suspension passed through the gauze showed little mycelium. The spores thus separated were determined by a hemacytometer. The concentration of spores contained in the spore suspension was determined, and then diluted appropriately and used as a fungal spawn. In the present invention, in order to produce enzyme foods having a certain quality, only the spores obtained by the above method were used as spores obtained by the above method. Inoculating a certain amount of fungus can produce products with similar enzymatic activity.

In the present invention, in order to ferment grains, in addition to the fungus, inoculated bacteria showing the optimal activity. Bacillus subtilis KCTC-1028, Bacillus megateum, Bacillus subtilis, Bacillus megatilium SMY-212, which was isolated from traditional doenjang and deposited as a patent strain in order to culture the bacteria showing optimal activity Leeum ( Bacillus megaterium) KCTC-217 is used as a spawn. Activity results for the bacteria are described in detail in Example 3. The bacteria were plated on Luria-Bertani (LB) plate medium and incubated overnight at 30 ° C. to form colonies. Then, one colony was inoculated into 5 ml LB liquid medium (20 ml test tube) and overnight at 37 ° C. shaking incubator. Incubated. The culture medium thus cultured was inoculated into 500 ml LB liquid medium contained in a 2-liter Erlenmeyer flask. After inoculation, the cells were shaken at 200 rpm for 20 hours at 37 ° C., and the culture solution was aseptically placed in a sterile centrifuge bottle to recover the bacterial cells, and then centrifuged at 6,000 rpm for 20 minutes. The precipitated cells were suspended in sterile distilled water and washed again by centrifugation. The recovered cells were suspended in sterile distilled water and used as seed grains of cereals. In the present invention, when cultured and washed in the same manner as in the above method, the degree of fermentation is appropriate, less off-flavor, and the highest enzyme activity can be obtained.

The third step is inoculating seed spawned in the second step to the grains increased in the first step.

The fourth step is a step of fermenting cereal seed seeded inoculation, which is described in detail in Example 4. The fifth step is drying and powdering the fermented grains, and the sixth step is to add an additive such as chitosan oligosaccharides to about 5% to the powdered fermented grains, and then add water appropriately and shape the granulator. Enzyme activity that is killed in the drying and molding step of the present invention is less than 10%, it can be seen that the enzyme activity is maintained stable.

Preferred embodiments of the present invention and significant effects obtained in comparison with existing enzyme products are described in detail below. However, this is only a description to help understand the present invention, but the scope of the present invention is not limited thereto.

Example 1

Enzyme productivity according to culture conditions

1-1. Alpha-amylase activity measurement

10 g of the sample was accurately measured, dissolved in a pH 5.2 buffer solution to a final 100 ml, and filtered to use as a crude enzyme solution. Alpha-amylase activity measurement using coenzyme was used by modifying the method of measuring the blue value (blue value). After dissolving soluble starch in 1% concentration in pH 5.2 buffer, 1 ml of crude enzyme solution was added and the reaction was carried out at 40 ° C. for 30 minutes. After the reaction was stopped with 1N-acetic acid, 10 ml of 0.005% KI + I ₂ solution was developed, the absorbance was measured at 660 nm, the test difference was subtracted, and the starch digested per minute per gram of the sample was expressed as ug. Amylase activity was shown and the starch content was calculated as a standard curve by coloration of the same iodine (I ₂ ) solution.

1-2. Beta-amylase activity measurement

10 g of the sample was accurately measured, dissolved in a pH 5.2 buffer solution to a final 100 ml, filtered and used as a crude enzyme solution. After adding 1 ml of 0.5% soluble starch (pH 4.8) and 1 ml of crude enzyme solution, the mixture was reacted at 30 ° C. for 30 minutes. 3 ml of dinitrosalicylic acid (DNS reagent) was taken to stop the reaction, and then heated and developed for 5 minutes. The color development was measured at absorbance of 550 nm. After subtracting the absorbance of the blank, the maltose decomposed maltose per gram of sample per minute and reaction time was expressed as ug, indicating beta-amylase activity. After preparing a standard curve of maltose by the same method as above, the content of maltose was calculated.

1-3. Determination of Protease Activity

1 ml of coenzyme solution was added using 2% casein solution (pH 7.0) as a substrate and reacted at 30 ° C. for 20 minutes. After stopping the reaction with 0.4M Trichloroacetic acid (TCA), add 0.4M sodium carbonate solution and foline solution for 20 minutes to develop tyrosine per gram of sample. ug indicates protease activity. A tyrosine standard curve was prepared in the same manner as above and then the tyrosine amount was calculated.

Example 2

Investigation of Alpha-amylase and Protease Productivity in Mold Cultures

In order to investigate the most suitable fungi to be used in the present invention, the fungi were previously cultured with fungi alone and used as edible microorganisms, and then alpha-amylase and protease productivity were investigated. In this example, the fungus was not inoculated into cereals, and was cultured in PDB (Potato dextrose broth) medium, and then the enzyme productivity was examined. Two types of hair fungi ( Rhizopus microsporus KCTC-6969, Rhizopus microsporus KCTC - 1778) and three types of Rhesus bacteria ( Aspergillus oryzae KCTC-6373, Aspergillus oryzae KCTC-6292) reported to be edible and have high productivity in amylase and proteases. Aspergillus awamori KCTC-6915) was plated on a PDA plate medium and incubated in a 28 ° C. incubator for 4 days to form spores. The spores were suspended in sterile distilled water, the mycelium was removed by passing through sterile gauze, and the number of spores was determined using a hemocytometer. The spore solution thus prepared was diluted with sterile distilled water and inoculated in a 250 ml PDB medium so that the concentration of spores was 10 ⁵ / ml. After inoculation, the culture medium was taken at time of shaking at 28 ° C. and 200rpm, and the alpha-amylase and protease activities were measured. Aspergillus awamori KCTC-6915 alone resulted in the highest alpha-amylase activity (3.99 unit) at 48 hours (Table 1) Rhizopus microsporus KCTC-1778 The highest protease activity (4.60 unit) at 120 hours of incubation (Table 2).

Investigation of Alpha-amylase Activity According to Incubation Time of Fungi                       Incubation Time Mold 48 hours 72 hours 96 hours 120 hours Refupus microsporus KCTC-6969 Refupus microsporus KCTC-1778 Aspergillus Orize KCTC-6373 Aspergillus Orize KCTC-6292 Aspergillus Awamori KCTC-6915 0.77 1.39 1.44 1.11 3.99 1.92 2.87 1.31 1.32 2.16 1.46 1.65 1.26 1.16 1.92 1.01 1.16 1.14 1.06 1.78

 Investigation of Protease Activity According to Incubation Time of Fungi                         Incubation Time Mold  48 hours 72 hours 96 hours 120 hours  Refupus Microsporus KCTC-6969 Refupus Microsporus KCTC-1778 Aspergillus Auris KCTC-6373 Aspergillus Auris KCTC-6292 Aspergillus Awamori KCTC-6915 1.14 1.58 0.77 0.83 1.84 2.90 2.65 1.65 1.71 2.72 3.47 3.53 1.46 1.96 1.96 4.10 4.60 1.02 2.09 1.90

Example 3

Investigation of Alpha-amylase and Protease Productivity According to Bacterial Culture

In order to investigate the most appropriate bacteria to be used in the present invention, it was intended to investigate the productivity of alpha-amylase and protease after culturing alone bacteria which can be used as edible microorganisms. In this example, bacteria were not inoculated into the grains, and were simply cultured in LB liquid medium, and then the enzyme productivity was examined. Bacillus megaterium SMY-212, already registered as a patent strain, Bacillus subtilis KCTC-1028 and Bacillus megaterium KCTC-217 received from Gene Bank were inoculated with platinum in a 5 ml LB liquid medium and shaken at 37 ° C. Incubated overnight at 200 rpm conditions. This was used as a seed culture solution and inoculated to 250% LB liquid medium to 1% and shaken in the same manner. After inoculation of the seed culture solution, the culture supernatant obtained after 8 hours and 18 hours of centrifugation was used as coenzyme. Table 3 and Table 4 show the results of measuring the enzyme activity using the crude enzyme by the method of the above example. The activity of alpha-amylase showed similar productivity in all three bacteria, but it was found that Bacillus megaterium SMY-212 was produced relatively high and stable. Protease productivity was 2.96 and 3.53 units for Bacillus megaterium SMY-212, respectively, which showed higher productivity than other bacteria.

Production of Alpha-amylase According to Bacterial Incubation Time       Strain culture time Bacillus Megaterium SMY-212 Bacillus subtilis KCTC-1028 Bacillus megaterium KCTC-217 8 hours 0.36 .0.58 .0.13 18 hours 0.62 .0.27 .0.67

Production of proteases according to the incubation time of bacteria       Strain culture time Bacillus Megaterium SMY-212 Bacillus subtilis KCTC-1028 Bacillus megaterium KCTC-217 8 hours 2.96 1.70 1.33 18 hours 3.53 1.58 1.39

According to the above results, the Bacillus megaterium SMY-212 strain had higher alpha-amylase and protease productivity than the other strains. Therefore, the Bacillus megaterium SMY-212 strain was used for the fermentation of cereals. It was estimated to be suitable for making products.

Example 4

Investigation of Enzyme Productivity According to Spawn During Cereal Fermentation

The above examples were the contents of the enzyme production after culturing each spawn (bacteria, fungus) in a basic liquid medium. Using these spawns, the inventors insist on inoculating and fermenting spawns with grains to make fermented foods containing high enzymes, so inoculating spawns on cereals and cooked cereals in various ways and investigating enzyme productivity. We investigated whether we can produce high-enzyme foods. Example Preprocesses 8 species of cereals according to the method of Example 2 after 1 and Bacillus mega prepared by the method in the same manner as in Te 3 Solarium (Bacillus megaterium) SMY-212, Bacillus MEGATHERIUM (Bacillus megaterium) KCTC-217, Bacillus subtilis KCTC-1028, Rhizopus microsporus KCTC-6969, Rhizopus microsporus KCTC-1778, Aspergillus orize ) Inoculated with KCTC-6373, Aspergillus oryzae KCTC-6292, Aspergillus awamori KCTC-6915 and then fermented at 28 ° C for 2 days for bacteria and for 4 days for fungi. . After fermentation, the grains were completely dried in a drying chamber at 30 ° C. for 2 days and then powdered. Chitosan oligosaccharides (95% purity chitosan oligosaccharides purchased from Amicogen) were added thereto at a 5% concentration, and then distilled water was added and molded into granulators. The products thus prepared were suspended in buffer again to investigate the activity of alpha-amylase and proteases, and the enzyme activity per 1 g of product was investigated. As a result, the strain of Bacillus megaterium SMY-212 showed the highest protease activity in cereals at 352 U / g, and the alpha-amylase activity, Rhizopus microsporus KCTC-1778 and Aspergillus oryzae KCTC-6373 were high at 90 U / g and 105 U / g, respectively. Bacillus bacteria with high protease activity had relatively low amylase activity, and amylase activity was higher in the fungus group, but in this case, the protease activity was relatively low. Therefore, if these strains, which are reported to be isolated from traditional fermented foods, are reported to produce high levels of both amylase and protease activity when mixed with grain fermentation, these strains reported as H. pylori, fur fungus, and natto-producing bacteria will be produced. The experiment was inoculated at the same time to fermentation. In the case of bacteria, growth was faster than mold, and when the fungus and bacteria were mixed and fermented, mold was inoculated first, incubated for 1 day, and bacteria were further inoculated to incubate for 2 days. As a result, when Bacillus megaterium SMY-212 and Rhizopus microsporus KCTC-1778 were sequentially cultured, the activity of alpha-amylase and protease was 1760 U / g and 250 U, respectively. / g and Bacillus megaterium SMY-212 and Aspergillus oryzae KCTC-6373 were sequentially cultured at 1,830 U / g and 247 U / g. In the case of sequential culture, it was found that the activities of alpha-amylase and protease were higher than those of single culture. In particular, when Bacillus megaterium SMY-212, Rhizopus microsporus KCTC-1778, Aspergillus oryzae KCTC-6373 are inoculated and fermented The activities of alpha-amylase and protease were 2,590 U / g and 250 U / g, respectively. Therefore, we tried to make a product using the method of sequentially inoculating three kinds of microorganisms (Table 5).

 Investigation of Enzyme Activity Produced in Pretreated Cereals                        Enzyme-activated strain Alpha-amylase (ug / ming) Proteases (ug / ming) Culture method Bacillus megaterium SMY-212 150 352 Exclusive Bacillus megaterium KCTC-217 180 213 Exclusive Bacillus subtilis KCTC-1028 200 151 Exclusive R hizopus microsporus KCTC-6969 1,200 80 Exclusive Rhizopus microsporus KCTC-1778 2,340 90 Exclusive Aspergillus oryzae KCTC-6373 2,870 105 Exclusive Aspergillus oryzae KCTC-6292 1,620 62 Exclusive Aspergillus awamori KCTC-6915 1,350 54 Exclusive Bacillus megaterium SMY-212 R hizopus microsporus KCTC-1778 1,760 251 mix Bacillus megaterium SMY-212 Aspergillus oryzae KCTC-6373 1,830 247 mix Bacillus megaterium SMY-212 R hizopus microsporus KCTC-1778 Aspergillus oryzae KCTC-6373 2,590 295 mix

Example 5

Investigation of Enzyme Productivity According to Inoculation Volume of Fungal Spawn

According to the above examples it was possible to select the seed to be used in the present invention. The selected seed was Bacillus megaterium SMY-212 with high protease activity, Rhizopus microsporus KCTC-1778 with high alpha-amylase activity, and Aspergillus oryzae KCTC. -6373. These three kinds of bacteria are, if sikyeoteul fermentation was inoculated in sequence was used as a sequential culturing method since this enzyme productivity was higher, 10 per was pre-treated with soaking and steaming grain 1g In order to examine the productivity of the enzyme according to the inoculum size of the fungal inoculum ^4, 5x10 ⁴ and 10 ⁵ spores were inoculated and incubated at 28 ° C. for 2 days. After incubation, the moldy grains were evenly mixed and inoculated with the same amount of bacteria (100 ml culture per kg grain), followed by further incubation at 28 ° C for 2 days. The fermented grains were dried and made through a molding process to measure the enzyme activity per 1g of product. As a result, when the fungus spores were inoculated at a concentration of 10 ⁵ per gram of grains, the activities of alpha-amylase and protease were the highest at 2,960 U / g and 321 U / g, respectively. However, 5x10 ⁴ , which is half of this concentration, was not significantly different from the enzyme activity. Therefore, when producing this product, 5x10 ⁴ concentration per g was considered to be the most suitable, and 5x10 ⁴ spores were inoculated in future experiments and production. (Table 6).

Investigation of Enzyme Productivity According to Inoculation Volume of Fungal Spawn           Enzyme Activity Inoculation Alpha-amylase (ug / ming) Proteases (ug / ming) 10 ⁴ 1,985 240 5 x 10 ⁴ 2,870 310 10 ⁵ 2,960 321

Example 6

Product production according to optimized culture method and quality comparison with other products

According to the method of the above embodiments, the product of the present invention was made by using the fermentation method optimized, and the final produced product conforms to the quality standard of food industry, and the products and enzymes of other companies that are produced and sold in the past The activity of enzyme production was investigated by comparing the activities. The current Food Code permits only alpha-amylases and proteases to be positive, so it is difficult to know the exact enzymatic activity of other products. , 461-468 (2001)) to compare the enzyme activity of the present invention. Since the enzyme activity measurement method used in this example was performed in the same manner as the reference, it is determined that the enzyme activity can be compared. As a result, when fermented to cereals under the optimum conditions of the present invention, the average activity of alpha-amylase, beta-amylase, and protease was 3,033 U / g, 12,150 U / g, and 332 U / g, respectively. Seemed. As a result of measuring repeated cultures, the error of activity was about 20%, so it was possible to produce a product of constant quality as well as high activity. The result of comparing the result with the reference data is shown in Table 7. The results show that cereal enzyme products generally have protease activity of 0.24-27.57 U / g, higher than 0.41-2.68 U / g of vegetables and other enzyme foods, and amylase activity of embryos, vegetables, and other enzyme foods. Could know. However, no products with high levels of amylase and protease activity were found. Compared with the activity of the present invention, alpha-amylase showed a maximum activity of 1.5-19 times higher than all the products, beta-amylase 89-1,100 times higher, and protease 12-1,844 times higher. Therefore, the high enzyme activity enzyme food produced by the present invention has a high enzymatic activity of up to 2,000 even when comparing only one enzyme, and all the enzyme activities of alpha-, beta-amylase, protease are superior to the existing products. Could know. When these enzymes are determined to act simultaneously in vivo, it can be estimated that they will be thousands of times more effective than existing products in enzyme activity. As a result of examining the items of the food regulations according to the measurement of the food code, the finished product was found to be 8-9% of moisture, 15-18% of crude protein, and negative for E. coli, which can be used as enzyme food.

Comparison of Enzyme Activity between Existing Products and Inventive Products Enzyme food type (company) Alpha-amylase (ug / ming) Beta-amylase (ug / ming) Proteases (ug / ming) Products of the Invention Grain Enzyme Food (A) Grain Enzyme Food (B) Grain Enzyme Food (C) Grain Enzyme Food (I) (D) Grain Enzyme Food (II) (D) Embryo Enzyme Food (E) Fruit Vegetable Enzyme Food (F) Other Enzyme Food (G) Other Enzyme Food (H) Other Enzyme Food (I) Other Enzyme Food (F) Other Enzyme Food (G) 3,033 339 1,410 821 159 451 1,455 1,630 1,563 1,973 1,593 1,430 1,611 12,150 171 39 40 136 108 32 11 109 29 11 21 40 332.00 0.56 0.24 21.44 23.26 27.57 7.68 0.41 1.55 2.68 0.41 0.18 0.26

Lee, Eun-Ju and Chul-Ho Lee: Korean Society of Food Science and Technology Vol.

* Enzyme activity was measured and compared with the same method as the paper presented above.

Example 7

Investigation of thrombi lysis ability of the present invention

In the present invention, because the microorganisms are separated from the traditional fermented food and known to have excellent blood clot dissolving ability, the present invention was to investigate whether the product has a blood clot dissolving ability. Fibrin plate method based on fibrin according to the method of Astrup (Astrup, T. and Mullertz, S.,: Arch. Biochem. 40: 346, 1952), etc. to investigate the thrombolytic activity Was used. 10 ml of 0.06% fibrinogen solution was added to a petri dish and 20 units of thrombin solution (40 unit / ml) dissolved in borate saline buffer (pH 7.8) were added. Add uniformly and harden. This plate was dropped by concentration of the fermentation broth and reacted at 37 ° C to investigate the ring formed. After diluting the fermented product 10 times, the extract was dropped 5ul, 10ul, 20ul, respectively, and reacted for 20 minutes at 37 ° C. As shown in FIG. Therefore, although not quantitative, it was judged to be excellent in thrombus resolution.

 As described above, the enzyme food produced according to the method of pretreatment, seed inoculation, fermentation and aging of cereals according to the present invention shows an enzymatic activity that is thousands of times higher than that of conventional products, and is an optimized process. As it can produce enzyme food of the same quality, it can be usefully used in the production of functional enzyme food of high enzyme activity.

Claims (8)

In the method for producing a high activity enzyme food, A pretreatment step of immersing and increasing soybean, black soybean, rice barley, brown rice, and pearl barley; mold Incubating Aspergillus oryzae KCTC-6373, Rhizopus microsporus KCTC-1778 and Bacterial Bacillus megaterium SMY-212; Inoculating the cultured spawn on the cooked cereals; Fermenting the grains inoculated with the spawn; Drying and powdering the fermented grains; And A method of producing a high activity enzyme food comprising the step of adding an additive to the powdered fermented grains. The method of claim 1, wherein the composition of the cereal grains in the pretreatment step is 30% soybean, 15% black soybean, 15% barley, 25% brown rice, and 15% molybdenum. delete The method of claim 1, wherein the fungus is inoculated at the start of inoculation, followed by incubation for 1 day, and then inoculated with bacteria for 2 days of culturing. The method of claim 1, wherein the fermentation step is a bacterium for 2 days, the fungus is fermented at 28 ° C for 4 days. The method according to claim 1, wherein the additive is chitosan oligosaccharide. The method of claim 3, wherein the Bacillus megaterium SMY-212, Rhizopus microsporus KCTC-1778 and Aspergillus oryzae KCTC-6373 are sequentially inoculated and fermented. Method for producing a highly active enzyme food, characterized in that. A high activity enzyme food prepared by the method of any one of claims 1 to 7.

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