WO2012015242A2 - Novel antimicrobial composition, and method for preparing same - Google Patents

Abstract

The present invention relates to an antimicrobial composition, containing a wheat germ extract or wheat bran extract having antimicrobial effects, which can replace preservatives or antibiotics added to food, cosmetics, animal feed, or the like. Wheat germ or wheat bran is a by-product of the mass production of flour, and thus can be obtained in an inexpensive and easy manner. The antimicrobial composition of the present invention using wheat germ or wheat bran can be widely used in food, beverages, cosmetics, or the like, and is also usable as an antimicrobial material which can replace antibiotics for animal feed, which has currently become a subject of great importance. Therefore, the antimicrobial composition of the present invention may prevent and treat pathogenic diseases mediated by food or the like, and may be used as a novel natural antimicrobial material for preventing an increase in antibiotics tolerance. The antimicrobial composition of the present invention may be applied to a wide range of fields including those of animal feed, food, beverages, cosmetics, food packaging, etc.

Description

New antimicrobial composition and preparation method thereof

The present invention relates to a novel antimicrobial composition and a method for producing the same, to an antimicrobial composition containing wheat germ extract or wheat bran extract having antimicrobial efficacy and a method for extracting antimicrobial components from wheat germ or wheat bran. More specifically, the present invention is an antimicrobial composition containing wheat germ extract or wheat bran extract as an active ingredient and an antimicrobial component from wheat germ or wheat bran as an active ingredient, which can replace preservatives or antibiotics added to food, cosmetics or animal feed, etc. It is about how to extract.

In particular, the present invention is a new natural antimicrobial material for preventing and treating pathogenic diseases caused by foods and preventing increased resistance to antibiotics, and is applicable to various fields such as food, beverages, cosmetics and food packaging. An antimicrobial composition containing wheat germ extract or wheat bran extract with possible antimicrobial efficacy and a method for extracting antimicrobial components from wheat germ or wheat bran.

Preservatives that can safely store food for a long time according to mass production and distribution of food, etc. are commonly used as additives in food or cosmetics. If preservatives are not used, food-borne pathogens may multiply in food or cosmetics and cause fatal harm to those who consume it or apply it to the skin. Therefore, preservatives that have been proven safe for humans have been widely used in foods and cosmetics. However, these preservatives have been reported as a factor that causes allergic diseases, atopic dermatitis, or even cancer by disturbing human immune function according to recent studies.

In addition, the development of animal husbandry and aquaculture has made it possible to raise large quantities of animals, and inexpensively and in large quantities to supply people with high-quality meat, fish, and processed products thereof, but raising large quantities of animals provides a poor environment for animals. When a virus or pathogenic disease occurs once a person's resistance is reduced, the damage spreads not only in the livestock farm or farm, but also in the region. In order to prevent a decrease in production due to these problems, a method of prophylactically injecting antibiotics into livestock or fish, or feeding them into animal feed is widely practiced. However, the abuse of antibiotics in the animal breeding process not only leads to the problem of antibiotic resistance but also affects human beings as end consumers of meat, fish and livestock products, causing certain antibiotics to become resistant to humans. It came to the following. In other words, the continuous use of antibiotics in the feed of human-eating animals increases the resistance of pathogens, resulting in side effects that can have a fatal effect on public health.

In light of these problems, the government has recently reduced the mix of antibiotics and antimicrobials in formulated feeds to produce safe livestock products, while adding common antibiotics for both humans and animals to feed for livestock and farmed fish. Since 2012, measures have been banned. Representative common antibiotics include penicillin, tetracycline-based antibiotics, dacitracin zinc, colistin sulfate, neomycin sulfate, and neomycin hydrochloride, and these acquired antibiotics will be banned in the future in the manufacture of animal feed.

However, despite the good policy aspects of national health protection, the prohibition of antibiotics is likely to lead to the spread of diseases caused by animal pathogen infections and the burden of livestock treatment on farms, and also to reduce the amount of livestock production in the future. Therefore, in order to prevent the reduction of livestock production resulting from the ban on the use of common antibiotics that affect humans, the necessity of suggesting a new animal husbandry industry which can prevent the infection of animal pathogens by replacing the existing antibiotics belongs to this invention. It can be said that in the technical field.

As one of the solutions, the development of animal feed antimicrobial agents using phage has been attempted, but it is still in the early stages, and there is also a debate on the harmful effects on the human body. Therefore, the development of new natural antimicrobial agents to prevent and treat foodborne pathogenic diseases and to prevent increased resistance to antibiotics has recently attracted attention. That is, in the technical field to which the present invention belongs, as a new natural antimicrobial material for preventing and treating foodborne pathogenic diseases and preventing increased resistance to antibiotics, various fields such as food, beverage, cosmetics and food packaging There is an urgent need for the development of new natural antimicrobial agents having antimicrobial efficacy that can be applied to.

Wheat germ is a nutritious embryo of malt (wheat kernel) that is removed from the process of processing whole wheat into flour, which accounts for about 2-3% of the whole wheat grain. Wheat germ, which is a seed of wheat, is known to be rich in vitamin E (tocopherol), which is known as an antioxidant vitamin to date, and recent studies have shown that natural substances extracted from wheat germ have the effect of restoring the immune function of the human body.

In particular, Avemar, a water-soluble extract obtained by fermenting wheat germ with yeast at 30 ° C for 18 hours, has been reported to have an effect of halving fever or other complications caused by the use of anticancer products. In the case of chemo side effects have been reported to reduce by more than 50%. Looking at other research and patent applications using wheat germ, Korean Patent No. 10-0376538 discloses a hair shampoo composition for strengthening the hair internal fiber containing wheat germ protein as an additive, but as part of the shampoo composition It can be said that it stays at the level which utilizes the efficacy of the tocophenol which is the known component of wheat germ as mentioned above. Republic of Korea Patent No. 10-0970544 discloses a skin external preparation with excellent whitening and anti-aging effect, it discloses the extract of the main ingredient Sisters and plant extracts and a variety of plants added to it, as an additional additive extract wheat germ extract It is mentioned. However, the efficacy of the external skin preparation of the Republic of Korea Patent No. 10-0970544 mainly relates to the sister plant extract and wheat germ extract is referred to simply as an additional additive extract, and there is no disclosure about the exact efficacy of the wheat germ extract. Republic of Korea Patent Publication No. 10-2010-0038621 discloses a cosmetic composition for improving atopic dermatitis containing wheat germ oil and ceramide in the form of nano liposomes, strengthening the skin moisturizing power by reducing the amount of water loss by using wheat germ oil It only discloses the efficacy of improving atopic dermatitis, not only discloses or implies specific immune-improving efficacy, but does not suggest any antibacterial effect.

On the other hand, wheat bran (also known as "bran") is the remainder of the separation of flour and embryos from milled wheat, most of which are seed blood. In the milling process, three kinds of sheaths, sophies, and flakes come out but are mixed and shipped from the factory for feed. Wheat bran has a lot of fiber, but has a lot of protein, fat, minerals, etc., attracting attention as a feed.

Recent studies on the physiological activity of wheat bran has been reported that eating 50g of bran daily increases the amount of stool to prevent constipation and colon cancer. In particular, plant fiber of wheat bran is known to be effective for the prevention of colon cancer, it contains phytic acid and selenium and also contains antioxidants, attracting attention as a cancer prevention food. Looking at other research and patent applications using wheat bran, Republic of Korea Patent Publication No. 10-2005-0099141 is supplemented with nutrients such as vitamins and minerals in rice, such as rice snow and wheat bran to enable long-term preservation even after cooking Food additives containing supplementary nutrients have been disclosed, but this has been to provide supplementary nutrients using the nutrients in wheat bran and far from studying the useful bioactive function of wheat bran. Korean Patent Laid-Open Publication No. 10-1994-0013355 describes a method for preparing noodles with soft, chewy and savory flavor by adding wheat flour processed flour to wheat flour, but this also provides ingredients that give elasticity to noodles. It is only about improving the texture of the noodles using the physiological activity of wheat bran has not been disclosed at all.

As described above, many studies have been conducted on studies using tocophenols contained in wheat germ oil or wheat germ extract and suppressing side effects in anticancer treatment, but the efficacy and pharmacological mechanisms have not been fully demonstrated. Studies on the physiological activity and pharmacological efficacy of embryo extract or wheat bran extract are still poorly studied.

As such, academic research and commercial use of wheat germ extract or wheat bran extract, which is a natural material, is still in its infancy. Especially, wheat germ extract or wheat bran extract has antimicrobial effects against food-borne pathogens. There was no study or report on this.

Therefore, continuous research on the bioactive function and pharmacological efficacy of the wheat germ extract or wheat bran extract is necessary, it is necessary to develop a new natural antimicrobial composition applicable to various fields such as food, cosmetics. Therefore, the present inventors are interested in the research on wheat germ extract or wheat bran extract, which is a natural material, and the wheat germ extract or wheat bran extract extracted by the extraction method developed by the present inventors has excellent antimicrobial efficacy against major food pathogens. It was confirmed that there is an excellent antiseptic effect also in cosmetics and came to complete the present invention.

The present invention provides a novel antimicrobial composition and a method for producing the same.

It is an object of the present invention to provide an antimicrobial composition containing wheat germ extract or wheat bran extract having an antimicrobial effect and a method for extracting the antimicrobial component from wheat germ or wheat bran.

More specifically, the present invention is an antimicrobial composition containing wheat germ extract or wheat bran extract as an active ingredient and an antimicrobial component from wheat germ or wheat bran as an active ingredient, which can replace preservatives or antibiotics added to food, cosmetics or animal feed, etc. The purpose is to provide a method for extracting.

In particular, the present invention is a new natural antimicrobial material for preventing and treating pathogenic diseases caused by foods and preventing increased resistance to antibiotics, and is applicable to various fields such as food, beverages, cosmetics and food packaging. It is an object of the present invention to provide an antimicrobial composition containing wheat germ extract or wheat bran extract having possible antimicrobial efficacy and a method for extracting the antimicrobial component from wheat germ or wheat bran.

In order to achieve the above object, the present invention provides an antimicrobial composition comprising wheat germ extract or wheat bran extract having an antimicrobial effect as an active ingredient.

In the antimicrobial composition of an embodiment of the present invention, the wheat germ extract or wheat bran extract is characterized in that extracted with at least one solvent selected from the group consisting of chloroform, methylene chloride, ethyl acetate and diethylene glycol monoethyl ether. . Preferably, the wheat germ extract or wheat bran extract is obtained by further filtration after extraction with the solvent.

In another antimicrobial composition of the present invention, the wheat germ extract or wheat bran extract is characterized in that extracted by the supercritical fluid extraction method. Preferably, the supercritical fluid used in the supercritical fluid extraction method may be carbon dioxide and the cosolvent may be ethyl alcohol. More preferably, the wheat germ extract or wheat bran extract is obtained by further filtration after mixing with the co-solvent.

In the antimicrobial composition of the present invention, the wheat germ extract or wheat bran extract is characterized in that it is produced in a dry powder form through a drying process.

The antimicrobial composition of the present invention may include 0.01 wt% to 100 wt% of the wheat germ extract or wheat bran extract based on the total weight of the composition.

The antimicrobial efficacy composition of the present invention may be used as an antimicrobial substance to replace an antimicrobial agent added to an animal feed or an antimicrobial agent to replace a preservative added to food, beverage or cosmetics.

Method for producing an antimicrobial composition of one embodiment of the present invention,

(a) grinding the prepared wheat germ or wheat bran and treating the ground wheat germ or wheat bran with at least one solvent selected from the group consisting of chloroform, methylene chloride, ethyl acetate and diethylene glycol monoethyl ether or wheat germ extract or Obtaining wheat bran extract,

(b) re-dissolving the wheat germ extract or wheat bran extract obtained in step (a) in the solvent and then filtering.

The method of preparing an antimicrobial composition of one embodiment of the present invention further comprises the step of evaporating the wheat germ extract or wheat bran extract in a vacuum evaporator to a maximum temperature of approximately 40 ° C. in the step (a).

In the method for preparing an antimicrobial composition of an embodiment of the present invention, the step (b) comprises the wheat germ extract or wheat bran extract obtained in step (a), or an evaporated concentrate of wheat germ extract or a wheat bran extract. After re-dissolving in the solvent, it is characterized in that the filtration through a PTFE (polytetrafluoroethylene) filter.

Method for producing an antimicrobial composition of another embodiment of the present invention,

(a) pulverizing and drying the prepared wheat germ or wheat bran and supercritical fluid extraction method using carbon dioxide as a supercritical fluid and ethyl alcohol as a co-solvent for milled or dried wheat germ or wheat bran to extract wheat germ or Obtaining wheat bran extract,

(b) mixing the wheat germ extract or wheat bran extract obtained in step (a) with the co-solvent and filtering the mixture.

In the method for preparing an antimicrobial composition of another embodiment of the present invention, the step (b) is a wheat germ extract or wheat bran extract obtained in the step (a) after mixing with the co-solvent through a PTFE (polytetrafluoroethylene) filter It is characterized in that the filtration treatment.

The antimicrobial composition containing wheat germ extract or wheat bran extract having the antimicrobial efficacy of the present invention may replace preservatives or antibiotics added to food, cosmetics or animal feed. In particular, wheat germ and wheat bran can be obtained in large quantities inexpensively and easily as a by-product of the flour production process, so the antimicrobial composition containing wheat germ extract or wheat bran extract of the present invention can be widely used in food, beverages, cosmetics, etc. In addition, there is an advantage that can be used as an antimicrobial substance that can replace the animal feed antibiotics that are currently the topic.

Therefore, the antimicrobial composition containing wheat germ extract or wheat bran extract of the present invention can be used as a novel natural antimicrobial material for preventing and treating pathogenic diseases such as food-borne media and preventing increased resistance to antibiotics, As well as animal feed, there is an advantage that can be applied to various fields such as food, beverage, cosmetics and food packaging.

The above and other technical problems and features of the present invention will be apparent to those skilled in the art through the description of the embodiments of the present invention made with reference to the accompanying drawings.

1 is a graph showing a strong growth inhibitory effect against Staphylococcus aureus which is a food-mediated pathogen, the antimicrobial composition containing wheat germ extract of the present invention as an example of food Staphylococcus aureus in soy milk It is a graph showing the antimicrobial effect against.

Figure 2 is a graph showing a strong growth inhibitory effect against staphylococcus aureus, the main pathogen through which the antimicrobial composition containing wheat germ extract of the present invention is a food-borne pathogen to Staphylococcus aureus in milk as an example of food. It is a graph showing the antimicrobial effect against.

Figure 3 is a graph showing the results of the Staphylococcus aureus killing rate for the effect of killing the Staphylococcus aureus antibacterial composition containing the bran extract extracted by the method of extracting bran of one embodiment of the present invention.

Figure 4 is a graph showing a strong growth inhibitory effect against staphylococcus aureus, the main pathogen through which the antimicrobial composition containing wheat bran extract of the present invention as a food-borne pathogen to Staphylococcus aureus in soy milk as an example of food. It is a graph showing the antimicrobial effect against.

5 is a graph showing a strong growth inhibitory effect against Staphylococcus aureus, which is a food-mediated pathogen, is an antimicrobial composition containing wheat bran extract of the present invention for Staphylococcus aureus in milk, which is an example of food. It is a graph showing the antimicrobial effect.

Hereinafter, the present invention will be described in more detail based on examples. The following examples of the present invention are not intended to limit or limit the scope of the present invention only to embody the present invention. From the detailed description and examples of the present invention, those skilled in the art to which the present invention pertains can easily be interpreted as belonging to the scope of the present invention. References cited in the present invention are incorporated herein by reference.

Example 1 Materials Used in Embodiments of the Invention

Wheat germ or wheat bran was purchased from Dong-A Won Co., Ltd. (Seoul, Korea) and used, and pathogens that are the antimicrobial and antiseptic test of wheat germ extract or wheat bran extract of the present invention (in the present invention, human or animal) As a term encompassing microorganisms such as viruses, bacteria, and fungi that cause disease, Salmonella typhimurium , Escherichia coli , Listeria monocytogenes , Staphylococcus aureus , and Bacillus Bacillus cereus , Pseudonomas aeruginosa , Candida albicans and Aspergillus niger are known microorganisms in the art, such as the Korean Bank of Biotechnology for Type Cultures, KCTC) (Daejeon Metropolitan City, Korea).

In addition, nutritional medium for assaying antimicrobial activity (containing 0.03% beef extract and 0.05% peptone) was obtained from the BBL Microbiology System (Cockeysville, MD, USA.), And beef extract and peptone were obtained from Difco Laboratories (Detroit, MI, USA). Purchased from Other materials were purchased from Sigma Co., U.S.A., USA, unless otherwise noted.

Example 2: Antimicrobial efficacy measurement method used in this example

In the following examples the antimicrobial activity was detected by modified disk diffusion method (Bauer, AW, WM Kirby, JC Sherris, and M. Turck. 1966. Antibiotic susceptibility testing by a standardized single disk method.Am J. Clin. Pathol. 45: 493-496), to facilitate the diffusion of antimicrobial material and to analyze the antimicrobial activity of wheat germ extract or wheat bran extract of the present invention using agar well diffusion assay well known in the art.

First, Salmonella typhimurium , Escherichia coli , Listeria monocytogenes , Staphylococcus aureus , and Bacillus cereus , respectively, prepared in Example 1, were passaged on a medium. The cells were cultured and incubated at 37 ° C. (but Bacillus cereus bacteria were 30 ° C.) for 18 hours. Then, each pathogen cell was suspended in physiological saline according to the McFarland protocol to make a suspension of about 10 ⁵ CFU / ml.

0.5 ml of each pathogen suspension was mixed with 150 ml of nutrient agar at 40 ° C. and poured into agar plates (23 × 23 cm) in a laminar flow cabinet. Then, agar well plates were prepared by cutting agar wells from agar medium sprayed with pathogens using a 7 mm diameter hollow tube, and applying a slight negative pressure to remove agar plugs.

On the other hand, the sensitivity of each pathogen to the test substance was determined by the formation of an inhibition zone after incubating the agar well plate for 18 hours at 37 ° C. The experiments were conducted in triplicates and the results were expressed as mean values for the three measurements.

In addition, the minimum inhibitory concentration is Christophet al.Christoph, M.S., P. Klaus, W. Annette, S. Erwin, and J. C. Simon. 1999. Antibacterial activity of hyperforin from St. John's wort, against multiresistantStaphylococcus aureusand gram-positive bacteria.The Lancet.353: 2129-2131) and macrodilution test. Serial 2-fold dilutions of the test substance were prepared in dimethylsulfoxide, and 0.5 ml of dilution of test substance was added to 9.0 ml of nutrient medium. Salmonella cultured overnight in the diluted solution of each test substance thus obtained (Salmonella typhimurium), E. coli (Escherichia coli),Listeria monocytogenesListeria monocytogenes), Staphylococcus aureus (Staphylococcus aureus), Or Bacillus cereus bacteria (Bacillus cereus0.5 ml of the culture medium) was injected. After 48 hours of incubation at 37 ° C. (but Bacillus cereus bacterium at 30 ° C.), a minimum inhibitory concentration was determined, which is the minimum concentration of test substance that results in no visible growth of pathogens (see Table 1 below).

Table 1: Measurement result of minimum inhibitory concentration of wheat bran extract of the present invention

According to Table 1, the wheat bran extract of the present invention has a minimum inhibitory concentration of <1.25 mg / ml, which is the minimum concentration of a test substance which does not show visible growth of Staphylococcus aureus , and the wheat bran extract is higher than wheat germ extract. It is confirmed that about 5 times lower, the antibacterial activity is about 5 times better.

On the other hand, 150 μl of a solution in which a test substance was dissolved in a solvent was added to each agar well plate containing the pathogen prepared as described above, and the antimicrobial activity by the well diffusion method was measured. At this time, an agar well plate containing 100 ppm, 500 ppm or 1000 ppm of chloramphenicol was used as a positive control (indicated by "CMP" in Tables 2 to 9), and chloroform, methylene chloride, ethyl acetate, or diethylene glycol mono Agar well plates containing ethyl ether (carbitol) were used as negative controls. The antimicrobial activity of wheat germ extract or wheat bran extract in Tables 2 to 9 of the following examples was measured by growth inhibition zone test compared to the positive control and the negative control, and the results are expressed in mm. . The results shown in Tables 2 to 9 represent the mean value ± standard deviation for the measured values obtained from a total of three experiments, and all the data in Tables 2 to 9 were statistical analysis programs [Statistical Package for Social Science (SPSS; SPSS Inc., Chicago, IL, USA).

Example 3: Antimicrobial efficacy test by wheat germ or wheat bran extract preparation and disk diffusion method

Example 3-1: Preparation of wheat germ or wheat bran extract and antibacterial efficacy test using chloroform

First, the prepared wheat germ was pulverized and 2.5 L of secondary distilled water was added to 100 g of the ground wheat germ, stirred for 1 hour, and then 1 L of chloroform (CHCl ₃ ) solution was added to the solution for 30 minutes. The solvent layer was recovered by stirring and centrifugation. At this time, the chloroform layer was removed after centrifugation, and the remaining filtrate layer was subjected to chloroform extraction three times in the same manner. The chloroform layers were combined, washed with distilled water and concentrated at a maximum temperature of 40 ° C. using a vacuum evaporator. The concentrated material (6.2 g) was finally redissolved in chloroform and filtered through a 0.45-μm PTFE filter. Salmonella (Salmonella typhimurium), as described in Example 2 by using the wheat germ extract obtained is filtered, treated according to the process as a sample, E. coli (Escherichia coli), Listeria monocytogenes Zenith (Listeria monocytogenes), Staphylococcus aureus (Staphylococcus aureus ) or 150 μl of the sample was put into each agar well plate in which Bacillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 2).

In addition, the prepared wheat bran was pulverized and 2.5 L of secondary distilled water was added to 100 g of the ground wheat bran, followed by stirring for 1 hour. Then, 1 L of chloroform (CHCl ₃ ) solution was added again to the solution and re-stirred for 30 minutes. The solvent layer was recovered by centrifugation. At this time, the chloroform layer was removed after centrifugation, and the remaining filtrate layer was subjected to chloroform extraction three times in the same manner. The chloroform layers were combined, washed with distilled water and concentrated at a maximum temperature of 40 ° C. using a vacuum evaporator. The concentrated material (5.8 g) was finally redissolved in chloroform and filtered through a 0.45-μm PTFE filter. Using the bran extract obtained by filtration according to the above process as a sample, as described in Example 2 Salmonella typhimurium , Escherichia coli , Listeria monocytogenes , Staphylococcus aureus ) Or 150 μl of the sample was put into each agar well plate in which Bacillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 3).

Looking at the results shown in Table 2, the wheat germ extract (1.5g fresh weight (FW; 150μl)) of this Example 3-1 was analyzed by the antimicrobial activity of five food and cosmetic harmful bacteria, It can be seen that it exhibits even antimicrobial activity against Gram-positive and Gram-negative bacteria, especially Staphylococcus aureus (approximately 27.22 mm growth inhibition zone) and Bacillus cereus (approximately 12.99 mm growth). It can be seen that it has a strong growth inhibitory effect on the zone of inhibition).

TABLE 2

In addition, looking at the results shown in Table 3, the wheat bran extract (1.5g fresh weight (FW; / 150μl) of this Example 3-1) of the antimicrobial activity analysis results for five harmful bacteria and food products , It can be seen that it shows even antimicrobial activity against Gram-positive and Gram-negative bacteria, in particular Staphylococcus aureus (approximately 15.91mm growth inhibition zone) and Bacillus cereus (approximately 12.94mm It can be seen that it has a strong growth inhibitory effect on the growth inhibition zone).

TABLE 3

These results strongly suggest that wheat germ extract or wheat bran extract of the present invention can be used as a useful antimicrobial material for food poisoning prevention. That is, the wheat germ extract or wheat bran extract of the present invention can be confirmed that the material is applicable as an antiseptic for the safe production of food or drink and can be used as an antibacterial and antibiotic.

Example 3-2: Preparation of wheat germ or wheat bran extract and antibacterial efficacy test using methylene chloride

First, the prepared wheat germ was pulverized, and 2.5 L of secondary distilled water was added to 100 g of the ground wheat germ, followed by stirring for 1 hour. Then, 1 L of methylene chloride (CH ₂ Cl ₂ ) solution was further added to the solution. The solvent layer was recovered by restirring for minutes and centrifuging. At this time, the methylene chloride layer was removed after centrifugation, and the remaining filtrate layer was subjected to methylene chloride extraction three times in the same manner. The methylene chloride layers were combined, washed with distilled water and concentrated at a maximum temperature of 40 ° C. using a vacuum evaporator. The concentrated material (5.9 g) was finally redissolved in methylene chloride and filtered through a 0.45-μm PTFE filter. Salmonella (Salmonella typhimurium), as described in Example 2 by using the wheat germ extract obtained is filtered, treated according to the process as a sample, E. coli (Escherichia coli), Listeria monocytogenes Zenith (Listeria monocytogenes), Staphylococcus aureus (Staphylococcus aureus ) or 150 μl of the sample was put into each agar well plate in which Bacillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 2 above).

In addition, the prepared wheat bran was pulverized and stirred for 1 hour by adding 2.5 L of secondary distilled water to 100 g of the crushed wheat bran, and then 1 L of methylene chloride (CH ₂ Cl ₂ ) solution was added to the solution for 30 minutes. The solvent layer was recovered by restirring and centrifugation. At this time, the methylene chloride layer was removed after centrifugation, and the remaining filtrate layer was subjected to methylene chloride extraction three times in the same manner. The methylene chloride layers were combined, washed with distilled water and concentrated at a maximum temperature of 40 ° C. using a vacuum evaporator. The concentrated material (5.4 g) was finally redissolved in methylene chloride and filtered through a 0.45-μm PTFE filter. Using the bran extract obtained by filtration according to the above process as a sample, as described in Example 2 Salmonella typhimurium , Escherichia coli , Listeria monocytogenes , Staphylococcus aureus ), Or 150 μl of each sample was put into an agar well plate in which Bacillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 3 above).

Looking at the results shown in Table 2, the wheat germ extract (1.5g fresh weight (FW; 150μL)) of this Example 3-2 was analyzed by the antimicrobial activity of five harmful food and cosmetic bacteria, It can be seen that it exhibits even antimicrobial activity against Gram-positive and Gram-negative bacteria, in particular Staphylococcus aureus (approximately 29.59 mm growth inhibition zone) and Bacillus cereus (approximately 14.49 mm It can be seen that it has a strong growth inhibitory effect on the growth inhibition zone).

In addition, looking at the results shown in Table 3, the wheat bran extract (1.5g fresh weight (FW; 150μL) of this Example 3-2) of the antimicrobial activity analysis results for five harmful bacteria and food products , It can be seen that it shows even antimicrobial activity against Gram-positive and Gram-negative bacteria, especially Staphylococcus aureus (approximately 16.13 mm growth inhibition zone) and Bacillus cereus (approximately 13.81 mm It can be seen that it has a strong growth inhibitory effect on the growth inhibition zone).

These results strongly suggest that wheat germ extract or wheat bran extract of the present invention can be used as a useful antimicrobial material for food poisoning prevention. That is, the wheat germ extract or wheat bran extract of the present invention can be confirmed that the material is applicable as an antiseptic for the safe production of food or drink and can be used as an antibacterial and antibiotic.

Example 3-3 Preparation of Wheat Germ or Wheat Bran Extract Using Ethyl Acetate and Antimicrobial Efficacy Test

First, the prepared wheat germ was pulverized, and 2.5 L of secondary distilled water was added to 100 g of the ground wheat germ, followed by stirring for 1 hour, and then 1 L of ethyl acetate (CH ₃ COOC ₂ H ₅ ) solution was added to the solution. The mixture was restirred for 30 minutes and the solvent layer was recovered by centrifugation. At this time, the ethyl acetate layer was removed after centrifugation, and the remaining filtrate layer was extracted three times with the same method. The ethyl acetate layers were combined, washed with distilled water and concentrated at a maximum temperature of 40 ° C. using a vacuum evaporator. The concentrated material (5.5 g) was finally redissolved in ethyl acetate and filtered through a 0.45-μm PTFE filter. Salmonella (Salmonella typhimurium), as described in Example 2 by using the wheat germ extract obtained is filtered, treated according to the process as a sample, E. coli (Escherichia coli), Listeria monocytogenes Zenith (Listeria monocytogenes), Staphylococcus aureus (Staphylococcus aureus ) or 150 μl of samples were put into each agar well plate in which Ba cillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 4).

In addition, the prepared wheat bran was pulverized and 2.5 L of secondary distilled water was added to 100 g of the crushed wheat bran, followed by stirring for 1 hour. Then, 1 L of ethyl acetate (CH ₃ COOC ₂ H ₅ ) was added to the solution. The solvent layer was recovered by restirring for minutes and centrifuging. At this time, the ethyl acetate layer was removed after centrifugation, and the remaining filtrate layer was extracted three times with the same method. The ethyl acetate layers were collected, washed with distilled water and concentrated at a maximum temperature of 40 ° C. using a vacuum evaporator. The concentrated material (4.8 g) was finally redissolved in ethyl acetate and filtered through a 0.45-μm PTFE filter. Using the bran extract obtained by filtration according to the above process as a sample, as described in Example 2 Salmonella typhimurium , Escherichia coli , Listeria monocytogenes , Staphylococcus aureus ) Or 150 μl of the sample was put into each agar well plate in which Bacillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 5).

Looking at the results shown in Table 4, wheat germ extract (1.5 g fresh weight) of this Example 3-3 (150W fresh weight) of the antimicrobial activity of the food and cosmetic harmful bacteria analysis results, It can be seen that it shows even antimicrobial activity against Gram-positive and Gram-negative bacteria, in particular Staphylococc us aureus (approximately 22.75mm growth inhibition zone) and Bacillus cereus (approximately 16.47mm It can be seen that it has a strong growth inhibitory effect on the growth inhibition zone).

Table 4

In addition, looking at the results shown in Table 5, the wheat bran extract (1.5g fresh weight (FW; / 150μl) of this Example 3-3) is the antimicrobial activity analysis results for five harmful bacteria and food products , It can be seen that it shows even antimicrobial activity against Gram-positive and Gram-negative bacteria, especially Staphylococcus aureus (approximately 19.42mm growth inhibition zone) and Bacillus cereus (approximately 13.89mm It can be seen that it has a strong growth inhibitory effect on the growth inhibition zone).

Table 5

These results strongly suggest that wheat germ extract or wheat bran extract of the present invention can be used as a useful antimicrobial material for food poisoning prevention. That is, the wheat germ extract or wheat bran extract of the present invention can be confirmed that the material is applicable as an antiseptic for the safe production of food or drink and can be used as an antibacterial and antibiotic.

Example 3-4 Preparation of Wheat Germ or Wheat Bran Extract Using Carbitol and Antimicrobial Efficacy Test

First, the prepared wheat germ was pulverized and 1 L of diethylene glycol monoethyl ether (carbitol) solution was added to 100 g of the ground wheat germ, stirred for 30 minutes, and the solvent layer was recovered by centrifugation. At this time, the diethylene glycol monoethyl ether (carbitol) layer was removed after centrifugation, and the remaining precipitate was subjected to diethylene glycol monoethyl ether (carbitol) extraction three times in the same manner. The diethylene glycol monoethylether (carbitol) layers were collected and concentrated at a maximum temperature of 40 ° C. The concentrated material (4.9 g) was finally filtered through a 0.45-μm PTFE filter. Salmonella (Salmonella typhimurium), as described in Example 2 by using the wheat germ extract obtained is filtered, treated according to the process as a sample, E. coli (Escherichia coli), Listeria monocytogenes Zenith (Listeria monocytogenes), Staphylococcus aureus (Staphylococcus aureus ) or 150 μl of samples were put into each agar well plate in which Bacillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 6).

Further, the prepared wheat bran was pulverized, and 1 L of diethylene glycol monoethyl ether (carbitol) solution was added to 100 g of the crushed wheat bran, stirred for 30 minutes, and the solvent layer was recovered by centrifugation. At this time, the diethylene glycol monoethyl ether (carbitol) layer was removed after centrifugation, and the remaining precipitate was subjected to diethylene glycol monoethyl ether (carbitol) extraction three times in the same manner. Diethylene glycol monoethylether (carbitol) layers were collected and concentrated at a maximum temperature of 40 ° C. The concentrated material (7.2 g) was finally filtered through a 0.45-μm PTFE filter. Using the bran extract obtained by filtration according to the above process as a sample, as described in Example 2 Salmonella typhimurium , Escherichia coli , Listeria monocytogenes , Staphylococcus aureus ), Or 150 μl of each sample was put into an agar well plate in which Bacillus cereus was cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 7).

Looking at the results shown in Table 6, the wheat germ extract (1.5g fresh weight (FW; 150μl)) of this Example 3-4 was analyzed by the antimicrobial activity of five harmful food and cosmetic bacteria, It can be seen that it exhibits even antimicrobial activity against Gram-positive and Gram-negative bacteria, especially Staphylococcus aureus (approximately 25.59 mm growth inhibition zone) and Bacillus cereus (approximately 13.49 mm growth). It can be seen that it has a strong growth inhibitory effect on the zone of inhibition).

Table 6

In addition, looking at the results shown in Table 7, the wheat bran extract (1.5g fresh weight (FW; 150μl) of this Example 3-4) of the antimicrobial activity analysis results for five food and cosmetic harmful bacteria , It can be seen that it shows even antimicrobial activity against Gram-positive and Gram-negative bacteria, in particular Staphylococcus aureus (approximately 18.18mm growth inhibition zone) and Bacillus cereus (approximately 15.19mm It can be seen that it has a strong growth inhibitory effect on the growth inhibition zone).

TABLE 7

These results strongly suggest that wheat germ extract or wheat bran extract of the present invention can be used as a useful antimicrobial material for food poisoning prevention. That is, the wheat germ extract or wheat bran extract of the present invention can be confirmed that the material is applicable as an antiseptic for the safe production of food or drink and can be used as an antibacterial and antibiotic.

Example 3-5: Preparation of Wheat Germ or Wheat Bran Extract and Antimicrobial Efficacy Test Using Supercritical Fluid Extraction Method

The supercritical fluid extraction method used in this example is Extraction method using a supercritical fluid that is a fluid that exceeds the critical temperature and boundary pressure. The supercritical fluid extraction method is more efficient than the liquid extraction method. Since the solvent power is strongly dependent on the density, the extraction conditions suitable for the target material can be obtained by changing the operating temperature or the pressure. Generally, inert, tasteless, odorless, and harmless carbon dioxide is used at room temperature, so it can be safely used in foods and medicines. At normal pressure, only the extract component is easily obtained because the fluid vaporizes.

In this example, the antimicrobial activity of the wheat germ extract or wheat bran extract extracted using the supercritical fluid extraction method was evaluated by the disk diffusion method (see Table 8 and Table 9).

First, after drying the wheat germ powder or wheat bran powder to control the extractor (extractor) temperature to 50 ~ 80 ℃ during supercritical fluid extraction, maintain the separator temperature (30 ~ 60 ℃), the temperature of the cooler The temperature was maintained at -1 to -5 ° C. Inject 100 g of wheat germ powder or wheat bran powder into a supercritical fluid extractor, adjust the pressure of the extractor to 60 to 300 bar, and adjust the flow rate of CO ₂ to 10 to 80%. Ethyl alcohol (alcohol) was injected at a flow rate of 0.5 to 2.0 ml / min to extract the raw materials at 30-minute intervals for 20 to 300 minutes when the pressure reached 200 to 300 bar. The extracted sample was again mixed with alcohol and filtered through a 0.45-㎛ PTFE filter.

As described in Example 2, the wheat germ extract or wheat bran extract obtained by filtration according to the above procedure was used as a sample, as described in Example 2, Salmonella typhimurium , Escherichia coli , Listeria monocytogenes , 150 µl of each sample was added to agar well plates in which Staphylococcus aureus or Bacillus cereus were cultured, and the antimicrobial activity was measured using the well diffusion method (see Table 8 and Table 9).

Looking at the results shown in Table 8, the wheat germ extract (1.5g fresh weight (FW; 150μl)) of the present Example 3-5 was analyzed by the antimicrobial activity of five harmful bacteria and food, cosmetics, It can be seen that it exhibits even antimicrobial activity against Gram-positive and Gram-negative bacteria, especially Staphylococcus aureus (approximately 26.77 mm growth inhibition zone) and Bacillus cereus (approximately 13.01 mm growth). It can be seen that it has a strong growth inhibitory effect on the zone of inhibition).

Table 8

In addition, looking at the results shown in Table 9, the wheat bran extract (1.5g fresh weight (FW; / 150μl) of this Example 3-5) is the antimicrobial activity analysis results for five food and cosmetic harmful bacteria , It can be seen that it shows even antimicrobial activity against Gram-positive and Gram-negative bacteria, especially Staphylococcus aureus (approximately 29.84mm growth inhibition zone) and Bacillus cereus (approximately 13.43mm It can be seen that it has a strong growth inhibitory effect on the growth inhibition zone).

Table 9

These results strongly suggest that wheat germ extract or wheat bran extract of the present invention can be used as a useful antimicrobial material for food poisoning prevention. That is, the wheat germ extract or wheat bran extract of the present invention can be confirmed that the material is applicable as an antiseptic for the safe production of food or drink and can be used as an antibacterial and antibiotic.

Example 4: Food preservation confirmation test using milk and soy milk

As shown in Table 10 and Table 11, after artificially inoculating Staphylococcus aureus at a concentration of 10 ⁵ CFU per ml of soy milk or milk, the wheat germ extract or wheat bran extract of the present invention as described above Treatment was performed to determine the growth inhibition against Staphylococcus aureus in milk or soymilk.

Table 10

Table 11

Looking at the results shown in Figures 1 and 2, it was confirmed that the wheat germ extract of the present invention has a strong growth inhibitory effect against Staphylococcus aureus concentration-dependent, and also the results shown in Figures 4 and 5 Looking at them, it was confirmed that wheat bran extract of the present invention also has a strong growth inhibitory effect against Staphylococcus aureus concentration-dependent. From this, it could be confirmed once again that the wheat germ extract or wheat bran extract of the present invention is applicable as an antiseptic for the safe production of foods or beverages and can be used as an antibacterial and antibiotic.

Example 5: Staphylococcus aureus killing rate measurement result

Staphylococcus aureus ( Staphylococcus aureus ) was added to the liquid medium medium of the concentration of about 10 ⁴ CFU / ml by the concentration of the wheat bran extract of the present invention was incubated for 24 hours to measure the killing of bacteria. As a result, as shown in Figure 3, it was confirmed that the killing of Staphylococcus aureus occurred within one hour after adding the bran extract of the present invention to the Staphylococcus aureus liquid medium.

Example 6 Antiseptic Test Using Cosmetic Base

Staphylococcus aureus , Escherichia coli , Pseudonomas aeruginosa , Candida albicans and Aspergillus niger were incubated at 37 ° C and 25 ° C for 18 hours and then 10 ^-3 After diluting and adding 1 ml of each culture to 9 g of the cosmetic base (Table 12) sample, the mixture was stirred, and then the wheat germ extract or wheat bran extract of the present invention was added to each sample at a concentration of 0.5-2.0% to 25 ° C. The number of bacteria in each sample was measured on days 0, 7, 14 for the antiseptic test of the stored samples.

Table 12: Cosmetic Base / Formula: MM 6420

As a result of the experiment, in the case of the treatment group of wheat germ extract of the present invention as confirmed in Table 13, it was found that the number of bacteria significantly decreased from the 7th day.

Table 13

In addition, in the case of the treatment group of wheat bran extract of the present invention as confirmed in Table 14, it was found that the number of bacteria significantly decreased from the seventh day.

Table 14

Therefore, it was confirmed that the wheat germ extract or wheat bran extract of the present invention can be used as food and cosmetics as a safe and inexpensive natural antimicrobial agent against food-borne pathogens. In particular, wheat germ and wheat bran are inexpensive and easily available in large quantities as a by-product of the flour production process, so that wheat germ extract or wheat bran extract can be widely used in food, beverages, cosmetics, etc. There is an advantage that can be used as an antimicrobial agent that can replace antibiotics.

Meanwhile, the present invention uses chloroform, methylene chloride, ethyl acetate or diethylene glycol monoethyl ether as an example for extracting antimicrobial substances from wheat germ or wheat bran, but the present invention is not limited to these solvents, but various food fields. And of course, it does not exclude known organic solvents that can be used for the extraction of active ingredients in the medical field.

Although the present invention has been described with reference to the above embodiments, the present invention is not limited thereto. Those skilled in the art can make modifications and changes without departing from the spirit and scope of the present invention, and it will be appreciated that such modifications and changes also belong to the present invention.

Claims (14)

1. An antimicrobial composition comprising wheat germ extract or wheat bran extract having an antimicrobial effect as an active ingredient.
2. The method of claim 1,

   The wheat germ extract or wheat bran extract is an antimicrobial composition, characterized in that extracted with at least one solvent selected from the group consisting of chloroform, methylene chloride, ethyl acetate and diethylene glycol monoethyl ether.
3. The method of claim 2,

   The wheat germ extract or wheat bran extract is characterized in that the antimicrobial composition obtained by further filtering after extraction with the solvent.
4. The method of claim 1,

   The wheat germ extract or wheat bran extract is antimicrobial composition, characterized in that extracted by the supercritical fluid extraction method.
5. The method of claim 4, wherein

   The supercritical fluid used in the supercritical fluid extraction method is carbon dioxide and the co-solvent is ethyl alcohol.
6. The method of claim 5,

   The wheat germ extract or wheat bran extract is antimicrobial composition, characterized in that obtained by further filtering after mixing with the co-solvent.
7. The method according to any one of claims 1 to 6,

   The wheat germ extract or wheat bran extract is characterized in that the antimicrobial composition is prepared in the form of a dry powder through a drying process.
8. The method according to any one of claims 1 to 6,

   An antimicrobial composition comprising from 0.01% to 100% by weight of the wheat germ extract or wheat bran extract relative to the total weight of the composition.
9. The method according to any one of claims 1 to 6,

   An antimicrobial composition, characterized in that used as an antimicrobial substance to replace the antiseptics added to food preservatives added to food, beverages or cosmetics or antibiotics added to animal feed.
10. (a) grinding the prepared wheat germ or wheat bran and treating the ground wheat germ or wheat bran with at least one solvent selected from the group consisting of chloroform, methylene chloride, ethyl acetate and diethylene glycol monoethyl ether or wheat germ extract or Obtaining wheat bran extract,

    (B) a method for producing an antimicrobial composition comprising the step of re-dissolving the wheat germ extract or wheat bran extract obtained in the step (a) in the solvent.
11. The method of claim 10,

    The method of claim (a) further comprises the step of evaporating the wheat germ extract or wheat bran extract to a maximum temperature of 40 ℃ in a vacuum evaporator.
12. The method of claim 11,

    The step (b) is a wheat germ extract or wheat bran extract obtained in step (a), or the evaporated concentrate of wheat germ extract or wheat bran extract redissolved in the solvent and then through a PTFE (polytetrafluoroethylene) filter A method for producing an antimicrobial composition, characterized in that the filtration treatment.
13. (a) pulverizing and drying the prepared wheat germ or wheat bran and supercritical fluid extraction method using carbon dioxide as a supercritical fluid and ethyl alcohol as a co-solvent for milled or dried wheat germ or wheat bran to extract wheat germ or Obtaining wheat bran extract,

    (B) a method for producing an antimicrobial composition comprising the step of mixing the wheat germ extract or wheat bran extract obtained in the step (a) with the co-solvent and filtering.
14. The method of claim 13,

    The step (b) is a method for producing an antimicrobial composition, characterized in that the wheat germ extract or wheat bran extract obtained in the step (a) is mixed with the co-solvent and filtered through a PTFE (polytetrafluoroethylene) filter.

Images