Gastric acid-stable probiotic preparation and method for preparing same

Abstract

The present invention relates to a gastric acid-stable probiotic preparation containing edible oils and fats, an emulsifier, and nuts, and a method for preparing same. The probiotic preparation according to the present invention is an excellent probiotic preparation in that, as a result of performing: an experiment on emulsion preparation at various heating temperatures (Experimental Example 1); effects on the proliferation of beneficial gut bacteria (Experimental Example 2); an experiment on lactic acid bacteria viability in artificial gastric juice (pH 2) (Experimental Example 3); an experiment on the efficacy of a composition on improving constipation (Brief Clinical Example 1); an experiment on evaluating preference according to the type of nuts contained in the composition (Brief Clinical Example 2); and the like, it can be confirmed that a lactic acid bacteria preparation of the present invention strongly proliferates beneficial gut bacteria, exhibits excellent viability of lactic acid bacteria in stomach acid, and exhibits more excellent and advanced effects in terms of constipation improvement and preference than existing known lactic acid bacteria preparations. Therefore, the probiotic preparation can be effectively used in various pharmaceutical compositions, food functional foods and health supplements.

Description

Gastric acid-stable probiotic preparation and method for producing the same

[0001] The present invention relates to a gastric acid-stable probiotic preparation containing edible oil, emulsifier and nuts, and a method for producing the same.

[0002] In general, lactic acid bacteria settle in the intestines and exert various physiological effects such as activating intestinal movement, suppressing harmful bacteria, and promoting vitamins and immune-boosting substances. However, due to the structure of the human body, after a person ingests lactic acid bacteria, they pass through the stomach before reaching the intestines. Therefore, the lactic acid bacteria are often killed by the gastric acid secreted in the stomach, preventing them from exerting their original physiological activity functions.

[0003] For this reason, attempts have been made to prevent the death of lactic acid bacteria in the body by coating lactic acid bacteria powder with various proteins or carbohydrates, and accordingly, lactic acid bacteria powder coated with various substances and methods for manufacturing the same have been developed.

[0004] There are approximately 400 species of bacteria present in the human colon and feces, and more than 90% of these are anaerobic strains. These bacteria maintain a certain balance to form intestinal flora, and are affected by not only the human health condition, stress, and physiological changes associated with aging, but also by external food and drugs. For physical health and longevity, the intestinal flora should be maintained with more beneficial bacteria and fewer harmful bacteria. In 2001, the Joint Expert Committee of the World Health Organization (FAO) and the Food and Agriculture Organization of the United Nations (FAO) defined probiotics as "live microorganisms that, when consumed in adequate amounts, are beneficial to health" (Jeong Hye-mi et al., 2011, J Korean Soc. Food. Sci. Nutr. 40(4), pp. 500 508) This refers to a probiotic preparation that can help improve intestinal microflora. Meanwhile, prebiotics refer to substances that can be used by beneficial intestinal microflora and can help the growth and function of beneficial intestinal bacteria. Prebiotics known to date are non-starch polysaccharides and oligosaccharides, and are substances that are not easily broken down by human enzymes but can be utilized by microorganisms (Manning TS. Gibson GR. 2004. Prebiotics. Best Practice & Research Clinical Gastroenterology. 18(2):pp.287-298). Recently, the concept of prebiotics has been broadened as the effect of polyphenols that can have a beneficial effect on intestinal microflora has been recognized (Cardona et al., 2013, Journal of Nutritional biochemistry, 24, pp.1415-1422).

[0005] Meanwhile, lactic acid bacteria are representative bacteria widely used as probiotics. They exist widely in nature and are easily found in the intestines of humans and animals and in fermented foods. They are microorganisms recognized as safe by the U.S. Food and Drug Administration. Lactic acid bacteria attach to intestinal epithelial cells and become parasites, improving the properties of intestinal flora. This helps the host animal in many ways, such as stabilizing the intestinal flora, suppressing the establishment of harmful bacteria, reducing the production of putrefactive products, preventing diseases, activating the immune system, having anticancer effects, and lowering cholesterol. The reason why lactic acid bacteria have growth inhibitory effects on various putrefactive and pathogenic microorganisms is because of several metabolic characteristics. Lactic acid bacteria produce antibacterial factors such as organic acid, hydrogen peroxide, reuterin, diacetyl, acetaldehyde, and bacteriocin as metabolites (Fuller, K., 1989, Probiotics in man and animals, J. Appl. Bacteriol., 66, 365-378; Korean Patent Publication No. 10-2015-0075447).

[0006] Probiotics are currently mainly used as health functional foods, but some are also used for therapeutic purposes. Specifically, they increase the production of immunoglobulin A antibodies and reduce rotavirus shedding, thereby shortening the duration of diarrhea. Therefore, they are actively being applied as preventative and therapeutic agents for antibiotic-associated diarrhea, as well as for treating inflammatory bowel diseases such as ulcerative colitis or Crohn's disease, as agents for treating abdominal distension or constipation, as agents for treating irritable colitis, as agents for treating radiation-induced enteritis, and as adjuvants for treating Helicobacter pylori infection. In particular, they are reported to be effective as agents for treating chronic inflammatory bowel diseases (Korean Patent Publication No. 10-2008-0075971).

[0007] Meanwhile, prior patent documents are as follows.

[0008] Korean Patent Publication No. 10-2022-0068518 (Prior Patent 3, Applicant: Velvet Care Co., Ltd.) discloses a composition for inducing voluntary intake of a solid formulation in a companion animal, characterized in that the composition comprises an oil containing unsaturated fatty acids, an adhesive agent, a sweetener, a mixture of lactic acid bacteria, a flavoring agent, a preservative, and an emulsifier.

[0009] Korean Patent Publication No. 10-1997-0025405 (prior patent 4) discloses a lactic acid bacteria preparation that is not killed by gastric acid and is active in the small and large intestines, wherein a fat and an emulsifier are mixed by heating to about 60°C, and a protective agent and lactic acid bacteria are mixed separately at about 50°C, and the two mixed solutions are emulsified by heating to about 60°C and then spraying the emulsified solution at high pressure into a cooled dispersion at 4°C. The microcapsule preparation of lactic acid bacteria is disclosed to be effective in promoting intestinal health by preventing the death of lactic acid bacteria by gastric acid, thereby increasing the colonization rate of lactic acid bacteria in the intestines.

[0010] Korean Patent Publication No. 10-2023-0001175 (prior patent 5) discloses a complex functionalized enteric soft capsule that can guarantee the number of lactic acid bacteria during the shelf life while containing both high content of lactic acid bacteria and edible oil, and comprises 5.0 to 25.0 wt% of lactic acid bacteria powder, 5.0 to 20.0 wt% of suspending agent, 0.5 to 2.0 wt% of emulsifier, and the remainder of edible oil.

[0011] Korean Patent Publication No. 10-2021-0083423 (prior patent 6) discloses an oral paste containing about 80 to 90% oil, about 0.5 to 20% emulsifier, and about 1 to 2% probiotic lactic acid bacteria effective against dental plaque and tartar.

[0012] However, nowhere in the above literature is there disclosed or described a substitute for a gastric acid stable probiotic preparation containing edible oil, emulsifier and nuts and a method for preparing the same.

[0013] Therefore, the present inventors conducted research to develop a lactic acid bacteria preparation superior to the lactic acid bacteria complex comprising lactic acid bacteria, edible oil, and emulsifier disclosed in the existing prior art patent 5 (Korean Patent Publication No. 10-2023-0001175).

[0014] As a result of conducting experiments on the production of emulsions at different heating temperatures using the lactic acid bacteria preparation of the present invention (Experimental Example 1); experiments on the effects on the proliferation of beneficial bacteria in the intestines (Experimental Example 2); experiments on the viability of beneficial bacteria in artificial gastric juice (pH 2) (Experimental Example 3); experiments on the efficacy of the composition in improving constipation (Simple Clinical Example 1); and experiments on the evaluation of preference according to the types of nuts contained in the composition (Simple Clinical Example 2), it was confirmed that the lactic acid bacteria preparation of the present invention strongly proliferates beneficial bacteria in the intestines, exhibits excellent viability of lactic acid bacteria in gastric acid, and exhibits superior and advanced effects in terms of improving constipation and preference compared to existing known lactic acid bacteria preparations, thereby confirming that an excellent probiotic preparation can be provided, and thus the present invention has been completed.

[0015] The present invention was conceived to overcome the aforementioned problems. The present invention provides a gastric acid-stable probiotic formulation containing edible oil, an emulsifier, and nuts. The formulation effectively promotes the proliferation of beneficial intestinal bacteria, exhibits excellent survival in gastric acid, and exhibits superior and more advanced efficacy than previously known lactic acid bacteria formulations in terms of improving constipation and palatability. The present invention also provides a method for producing the formulation.

[0016]

[0017] The characteristic configuration of the present invention to achieve the purpose of the present invention as described above and to realize the characteristic effects of the present invention described below is as follows.

[0018] As a preferred embodiment of the present invention, a gastric acid-stable probiotic preparation is provided, which is obtained by performing a process including: a first step of heating 1 to 90 parts by weight of edible oil and fat to 70 to 95°C and then mixing and stirring 0.1 to 5 parts by weight of the emulsifier to produce a mixture of edible oil and emulsifier; a second step of cooling and stirring the mixture of the first step until the temperature reaches 2 to 20°C to produce a gel-like coagulated mixture; and a third step of slowly adding 1 part by weight of lactic acid bacteria to the gel-like coagulated mixture and performing low-speed stirring to produce a lactic acid bacteria mixture.

[0019] As a preferred embodiment of the present invention, a gastric acid-safe probiotic preparation is provided by performing a process further comprising: a fourth step of adding ground nuts to the mixture obtained through the third step and stirring to prepare a mixture of nut lactic acid bacteria; and a fifth step of optionally adding one or more additional ingredients selected from prebiotics and adhesive agents to the mixture obtained through the fourth step and stirring.

[0020] As a preferred embodiment of the present invention, the nut powder has a particle size of 0.01 to 0.5 mm, preferably 0.01 to 0.1 mm, and is characterized in that the nut lactic acid bacteria mixture is prepared by stirring at a stirring speed of 30 to 50 rpm for 30 minutes to 3 hours until all ingredients are evenly mixed.

[0021] As a preferred embodiment of the present invention, the gastric acid stable probiotic preparation is characterized by containing (a) lactic acid bacteria, (b) edible oil, (c) emulsifier, and (d) nuts as essential ingredients.

[0022] As a preferred embodiment of the present invention, the individual composition weight ratios of the gastric acid-stable probiotic preparation are characterized by being composed of (a) 1.0 parts by weight of lactic acid bacteria, (b) 1 to 90 parts by weight of edible oil, (c) 0.1 to 5.0 parts by weight of emulsifier, and (d) 5.0 to 90.0 parts by weight of nuts.

[0023] As a preferred embodiment of the present invention, the (a) lactic acid bacteria is characterized in that it is one or more strains selected from microorganisms of the genus Lactobacillus, the genus Bifidobacterium, the genus Bacillus, the genus Streptococcus, and the genus Enterococcus.

[0024] In a preferred embodiment of the present invention, the (b) edible oil is characterized in that it is at least one component selected from the group consisting of avocado oil, canola oil, evening primrose oil, coconut oil, Brazil nut oil, corn oil, cottonseed oil, flaxseed oil, grapeseed oil, hemp oil, olive oil, palm oil, safflower oil, soybean oil, peanut oil, sunflower seed oil, krill oil, anchovy oil, salmon oil, rice bran oil, brown rice oil, coconut oil, rosehip oil, and tuna oil.

[0025] In a preferred embodiment of the present invention, the (c) emulsifier is characterized by being at least one component selected from the group consisting of propylene glycol fatty acid ester, glycerin fatty acid ester, sucrose fatty acid ester, polyglycerin fatty acid ester, sorbitan fatty acid ester, organic acid monoglyceride, polyoxyethylene fatty acid ester, propylene glycol fatty acid ester, polyoxyethylene fatty acid ester, polysorbate, egg yolk lecithin, soybean lecithin, carboxymethylcellulose, glycerin, soybean phospholipid, and stearyl calcium lactate.

[0026] As a preferred embodiment of the present invention, the (d) nuts are characterized in that they are at least one ingredient selected from the group consisting of peanuts, almonds, walnuts, pine nuts, pistachios, pecans, macadamias, hazelnuts, cashews, Brazil nuts, chestnuts, soybeans, cacao nibs, coconuts, sunflower seeds, and pumpkin seeds.

[0027] In a preferred embodiment of the present invention, the acid-stable probiotic preparation contains, in addition to the essential ingredients consisting of (a) lactic acid bacteria, (b) edible oil, (c) emulsifier, and (d) nuts, at least one additional ingredient selected from (e) prebiotics and (f) adhesive agents.

[0028] As a preferred embodiment of the present invention, the individual components of the gastric acid-stable probiotic preparation are mixed in a weight ratio of (a) 1.0 parts by weight of lactic acid bacteria, (b) 1 to 60 parts by weight of edible oil, (c) 0.1 to 5.0 parts by weight of emulsifier, (d) 5.0 to 90.0 parts by weight of nuts, (e) 0.01 to 5.0 parts by weight of prebiotics, and (f) 0.1 to 5.0 parts by weight of adhesive agent.

[0029] In a preferred embodiment of the present invention, the (e) prebiotics are characterized by being at least one component selected from pentose sugars such as xylose and arabinose; glucose, mannose, fructose, galactose; lactulose, lactitol, sucrose, lactose, maltose, trehalose; fructo-oligosaccharides, raffinose, stachyose, maltodextrin; amylose, amylopectin, starch, cellulose, and pectin.

[0030] As a preferred embodiment of the present invention, the (f) adhesive agent is characterized in that it is at least one component selected from the group consisting of sodium silicoaluminate, xanthan gum, carrageenan, guar gum, diutan gum, cellulose gum, gellan gum, pectin, and carboxymethylcellulose.

[0031]

[0032] Hereinafter, in order to enable a person having ordinary skill in the art to easily practice the present invention, preferred embodiments of the present invention will be described in detail with reference to the attached drawings.

[0033]

[0034] The gastric acid-stable probiotic preparation according to the present invention is manufactured by performing a process including: a first step of heating 1 to 90 parts by weight of edible oil and fat to a temperature range of 70 to 95°C, preferably 75 to 85°C, and then mixing and stirring 0.1 to 5 parts by weight of the emulsifier to produce a mixture of edible oil and emulsifier; a second step of cooling and stirring the mixture of the first step until the temperature reaches 2 to 20°C, preferably 4 to 10°C, to produce a gel-like coagulated mixture; and a third step of slowly adding 1 part by weight of lactic acid bacteria to the gel-like coagulated mixture and performing low-speed stirring to produce a lactic acid bacteria mixture.

[0035] Accordingly, the present invention provides a method for producing a gastric acid-stable probiotic preparation according to the present invention, comprising the steps of: a first step of heating 1 to 90 parts by weight of edible oil and fat to a temperature range of 70 to 95°C, preferably 75 to 85°C, and then mixing and stirring 0.1 to 5 parts by weight of the emulsifier to produce a mixture of edible oil and emulsifier; a second step of cooling and stirring the mixture of the first step until the temperature reaches 2 to 20°C, preferably 4 to 10°C, to produce a gel-like coagulated mixture; and a third step of slowly adding 1 part by weight of lactic acid bacteria to the gel-like coagulated mixture and stirring at low speed to produce a lactic acid bacteria mixture.

[0036] Through this process, the emulsifier is dissolved in the heated edible oil and completely mixed, and then a stably coagulated gel-shaped mixture is created through cooling. Then, by mixing lactic acid bacteria into this mixture, the intestinal retention capacity of the lactic acid bacteria can be significantly improved.

[0037] The gastric acid-stable probiotic preparation according to the present invention may further include a fourth step of preparing a mixture of nut lactic acid bacteria by adding ground nuts to the mixture obtained through the third step and stirring the mixture; and a fifth step of optionally adding one or more additional ingredients selected from prebiotics and adhesive agents to the mixture obtained through the fourth step and stirring the mixture.

[0038] As a preferred embodiment of the present invention, the probiotic preparation manufactured in step 5 may be sealed under nitrogen gas and stored at 25°C or lower. The temperature of the probiotic preparation may be controlled to 30°C or lower, and low-speed stirring may be used to minimize the decrease in lactic acid bacteria content due to temperature and physical friction.

[0039]

[0040] “(a) Lactic acid bacteria” as defined in the present invention means a strain that helps digestion and absorption and inhibits the growth of harmful bacteria in the intestines, thereby maintaining a desirable intestinal environment. The form of the lactic acid bacteria is not limited, but various forms such as dried powder and dried coarse powder are preferred, and the strain size may be preferably in the range of 0.2 to 4.0 ㎛ rather than 0.1 to 8.0 ㎛.

[0041] In the present invention, the content of each component is expressed in weight parts based on 1 weight part of lactic acid bacteria.

[0042] Preferably, as a representative strain, any lactic acid bacteria widely used as probiotics, which are readily available in the art, for example, strains available from the American Type Culture Collection (ATCC), the Korean Collection for Type Cultures (KCTC), the Korean Culture Center of Microorganisms (KCCM), the Korean Agricultural Culture Collection (KACC), the Rural Development Administration, etc.

[0043] Specifically, a microorganism of the genus Lactobacillus, the genus Bifidobacterium, the genus Bacillus, the genus Streptococcus, or the genus Enterococcus; preferably a strain of the genus Bifidobacterium, the genus Lactobacillus, or the genus Streptococcus, and more preferably a strain of the genus Bifidobacterium or the genus Streptococcus; More preferably, a strain of the genus Bifidobacterium, such as Bifidobacterium longum, Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium animalis ssp. lactis, Bifidobacterium adolescentis, Bifidobacterium pseudocatenulatum, Bifidobacterium catenulatum or Bifidobacterium infantis, Bifidobacterium thermophilum; Lactobacillus plantarum, Lactobacillus pentosus, Lactobacillus casei, Lactobacillus casei ssp. paracasei, Lactobacillus rhamnosus, Lactobacillus acidophilus, Lactobacillus delbrueckii, Lactobacillus delbrueckii ssp. bulgaricus.bulgaricus), Lactobacillus delbrueckii ssp. delbrueckii, Lactobacillus fermentum, Lactobacillus gasseri, Lactobacillus reuteri, Lb. brevis, Lb. cellobiosus, Lb. crispatus, Lb. GG, Lb. johnsonii, Lb. lactis, Lb. reuteri, Lb. salivarius Lactobacillus strains such as Bacillus cereus toyoi or Bacillus cereus strains; Leuconostoc species such as Leuconostoc citreum and Leuconostoc mesenteroides; Pediococcus species such as Pediococcus acidilactici and Pediococcus pentosaceus; Propionibacterium species such as Propionibacterium acidifaciens and Propionibacterium acidipropionici; Streptococcus thermophilus, S. cremoris, S. infantarius, S. intermediusStreptococcus species such as S. intermedius, S. lactis, S. salivarius subsp. thermophilus; Enterococcus species such as Enterococcus faecalis and E. faecium; Saccharomyces species such as Saccharomyces cerevisiae, Saccharomyces boulardii, etc., preferably Lactobacillus strains or Bifidobacterium strains, most preferably one or more single or mixed strains such as Lactobacillus acidophilus, Lactobacillus rhamnosus GG, Lactobacillus plantarum, Bifidobacterium longum, Bifidobacterium animalis ssp. Lactis, more preferably Lactobacillus acidophilus, Lactobacillus rhamnosus GG), Lactobacillus plantarum, Bifidobacterium longum and Bifidobacterium animalis ssp. Lactis.

[0044]

[0045] “(b) Edible fat” as defined in the present invention includes one or more ingredients selected from the group consisting of avocado oil, canola oil, evening primrose oil, coconut oil, Brazil nut oil, corn oil, cottonseed oil, flaxseed oil, grapeseed oil, hemp oil, olive oil, palm oil, safflower oil, soybean oil, peanut oil, sunflower seed oil, krill oil, anchovy oil, salmon oil, rice bran oil, brown rice oil, coconut oil, rosehip oil, and tuna oil.

[0046] The above edible oil acts to protect the lactic acid bacteria complex from gastric acid by enveloping it. The edible oil is contained in an amount of 1 to 90 parts by weight, and preferably 15 to 30 parts by weight is mixed. First, the edible oil is heated to 75 to 95°C, preferably 75 to 85°C, and then the emulsifier is mixed and stirred to produce a mixture of edible oil and emulsifier, which is characterized by being stirred in a circulating cooling water tank maintained at 0°C to 10°C, preferably 0 to 5°C, until the temperature of the mixture reaches 2 to 20°C, preferably 4 to 10°C, thereby forming a stable gel-like aggregate.

[0047]

[0048] The “(c) emulsifier” defined in the present invention includes at least one component selected from the group consisting of propylene glycol fatty acid ester, glycerin fatty acid ester, sucrose fatty acid ester, polyglycerin fatty acid ester, sorbitan fatty acid ester, organic acid monoglyceride, polyoxyethylene fatty acid ester, propylene glycol fatty acid ester, polyoxyethylene fatty acid ester, polysorbate, egg yolk lecithin, soybean lecithin, carboxymethylcellulose, glycerin, soybean phospholipid, and stearyl calcium lactate.

[0049] The above emulsifier plays a role in increasing the miscibility between the compositions in the lactic acid bacteria complex. When the edible oil is heated to 75 to 95°C, preferably 75 to 85°C, and the emulsifier is mixed, the emulsifier melts and the miscibility with the edible oil is greatly increased.

[0050] Although a process of heating edible oil to 60℃ and mixing an emulsifier has been known, there was a problem in that the emulsifier did not melt and mix even when the edible oil was heated to 60℃.

[0051] The results of this experiment confirmed that emulsifiers dissolve in edible oils only when heated to at least 70°C. Heat exceeding 100°C can lead to oxidation. Therefore, it is more preferable to heat edible oils to a temperature of 75 to 85°C. (See Figure 1)

[0052] The above emulsifier content may be comprised in an amount of 0.1 to 5.0 parts by weight, preferably 0.20 to 1.60 parts by weight, and more preferably 0.30 to 1 part by weight, of the total weight of the lactic acid bacteria complex. At this time, if the emulsifier content is less than 0.1 part by weight, there may be a problem of poor miscibility between the lactic acid bacteria mixture powder and edible oil, and even if it is used in excess of 5.0 parts by weight, there is no further increase in miscibility between the compositions, so it is appropriate to use it within the above range.

[0053]

[0054] According to a preferred embodiment of the present invention, a gastric acid-stable probiotic preparation containing (a) lactic acid bacteria, (b) edible oil, (c) emulsifier, and (d) nuts as essential ingredients is provided.

[0055] The above probiotic preparation is characterized in that the mixing weight ratios of the individual components are (a) 1.0 parts by weight of lactic acid bacteria, (b) 1 to 90 parts by weight of edible oil, (c) 0.1 to 5.0 parts by weight of emulsifier, and (d) 5.0 to 90.0 parts by weight of nuts.

[0056] Among these, descriptions of (a) lactic acid bacteria, (b) edible oil, and (c) emulsifier are omitted as they have been described above.

[0057]

[0058] “(d) Nuts” as defined in the present invention include one or more ingredients selected from the group consisting of peanuts, almonds, walnuts, pine nuts, pistachios, pecans, macadamias, hazelnuts, cashews, Brazil nuts, chestnuts, soybeans, cacao nibs, coconuts, sunflower seeds, and pumpkin seeds, as additives for forming a suitable viscosity of the composition, increasing flavor / texture, and further enhancing the activity of lactic acid bacteria.

[0059] The above nuts are essential ingredients that contain a large amount of unsaturated fatty acids and are beneficial to various health functions including blood circulation, and are preferably in the form of a pulverized product, and are characterized by being pulverized to a degree that the particle size of the pulverized product is 0.01 to 0.5 mm, preferably 0.01 to 0.1 mm, and exhibits a paste shape.

[0060] The content of the above nuts is mixed in an amount of 5.0 to 90.0 parts by weight, preferably 10.0 to 30.0 parts by weight.

[0061] The above nuts not only improve the palatability and sensory properties of probiotic preparations, but also have been experimentally confirmed to be excellent in maintaining the viability of lactic acid bacteria in gastric acid.

[0062]

[0063] The present invention provides a gastric acid stable probiotic preparation containing, in addition to the above essential ingredients, one or more additional ingredients selected from (e) prebiotics and (f) adhesive agents.

[0064] At this time, the mixing weight ratio of the individual components of the probiotic preparation is composed of (a) 1.0 parts by weight of lactic acid bacteria, (b) 1 to 90 parts by weight of edible oil, (c) 0.1 to 5.0 parts by weight of emulsifier, (d) 5.0 to 90.0 parts by weight of nuts, (e) 0.01 to 5.0 parts by weight of prebiotics, and (f) 0.1 to 5.0 parts by weight of adhesive agent.

[0065] More preferably, it is characterized by (a) 1.0 parts by weight of lactic acid bacteria, (b) 10 to 30 parts by weight of edible oil, (c) 0.2 to 1.6 parts by weight of emulsifier, (d) 15.0 to 30.0 parts by weight of nuts, (e) 0.5 to 2.0 parts by weight of prebiotics, and (f) 2.0 to 5.0 parts by weight of adhesive agent.

[0066]

[0067] The "(e) prebiotics" defined herein are ingredients that are used as food for lactic acid bacteria and are not digested by digestive enzymes in animals, but rather cause the growth of lactic acid bacteria such as bifidobacteria, which are intestinal bacteria, in the lower small intestine, thereby exhibiting various physiological effects, including intestinal regulation. Monosaccharides, disaccharides, oligosaccharides, and polysaccharides that are well known in the art to be usable as prebiotics are possible, and preferably, monosaccharides containing 5-carbon sugars such as xylose and arabinose, or 6-carbon sugars such as glucose, mannose, fructose, and galactose; Disaccharides such as lactulose, lactitol, sucrose, lactose, maltose, and trehalose; oligosaccharides such as fructo-oligosaccharide, raffinose, stachyose, and maltodextrin; polysaccharides such as amylose, amylopectin, starch, cellulose, and pectin; More preferably, it comprises a sugar compound such as glucose, fructose, galactose, sucrose, lactose, fructo-oligosaccharide, and more preferably, glucose, fructose, lactulose, lactitol, or fructo-oligosaccharide.

[0068] The content of the above (e) prebiotics component can be increased or decreased in a dosage well known in the art depending on the type and content of the above (a) lactic acid bacteria, and it is generally preferable to add 0.01 to 5 parts by weight, preferably 0.5 to 2.0 parts by weight, per 1 weight of lactic acid bacteria.

[0069] The above "(f) adhesive agent" defined herein is contained in an amount of 0.1 to 5 parts by weight based on the total weight of the composition and serves to impart adhesiveness to the composition. It is preferably composed of at least one selected from the group consisting of sodium silicoaluminate, xanthan gum, carrageenan, guar gum, diutan gum, cellulose gum, gellan gum, pectin, and carboxymethylcellulose.

[0070] If the content of the above-mentioned adhesive agent is less than 0.1 parts by weight, the above-mentioned effect is minimal, and if the content of the above-mentioned adhesive agent exceeds 5 parts by weight, the viscosity may increase excessively, resulting in reduced workability.

[0071] In addition, the present invention can additionally add one or more additives selected from an appropriate amount of pigment, fragrance, etc. to the above composition within a range of about 0 to 10 parts by weight relative to 1 weight of the total lactic acid bacteria.

[0072]

[0073] Meanwhile, the present invention provides a pharmaceutical composition, health functional food, and health supplement for promoting the growth of intestinal beneficial bacteria, which contain a gastric acid-stable probiotic preparation manufactured by the above-mentioned manufacturing method as an active ingredient.

[0074] The probiotic preparation of the present invention was tested for the production of an emulsion at different heating temperatures (Experimental Example 1); for the effect on the proliferation of beneficial intestinal bacteria (Experimental Example 2); for the viability of lactic acid bacteria in artificial gastric juice (pH 2) (Experimental Example 3); for the effect of the composition on improving constipation (Simple Clinical Example 1); and for the evaluation of preference according to the type of nuts contained in the composition (Simple Clinical Example 2). As a result, it was confirmed that the probiotic preparation of the present invention strongly proliferates beneficial intestinal bacteria, exhibits excellent viability of lactic acid bacteria in gastric acid, and exhibits superior and more advanced effects in terms of improving constipation and preference than existing known lactic acid bacteria preparations, and is therefore useful as an excellent probiotic preparation, a pharmaceutical composition, a health functional food, and a health supplement.

[0075] To achieve the above purpose, the present invention provides a pharmaceutical composition containing the probiotic preparation as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing and treating diseases caused by the growth of harmful intestinal bacteria.

[0076] “Diseases caused by the proliferation of harmful intestinal bacteria” as defined herein include, but are not limited to, antibiotic-associated diarrhea, inflammatory bowel diseases such as ulcerative colitis and Crohn’s disease, abdominal bloating, constipation, acute or chronic irritable bowel syndrome, radiation-induced enteritis, Helicobacter pylori infection, etc.

[0077] The composition of the present invention comprises the probiotic preparation in an amount of 0.01 to 99% by weight based on the total weight of the composition.

[0078] However, the composition as described above is not necessarily limited to this and may vary depending on the patient's condition and the type and progression of the disease.

[0079] A composition comprising the probiotic preparation of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

[0080] The composition including the probiotic preparation according to the present invention can be formulated and used in the form of oral formulations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., external preparations, suppositories, and sterile injection solutions, respectively, according to conventional methods, and carriers, excipients, and diluents that can be included therein include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, magnesium stearate, and mineral oil. When formulated, it is usually prepared using diluents or excipients such as fillers, bulking agents, binders, wetting agents, disintegrating agents, and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, and capsules, and these solid preparations are prepared by mixing the probiotic preparation with at least one excipient, such as cotton, starch, calcium carbonate, sucrose or lactose, gelatin, etc. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral administration include suspensions, oral solutions, emulsions, and syrups, and in addition to commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, and suppositories. Non-aqueous solutions and suspensions can include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate.Suppository bases that can be used include witepsol, macrogol, tween 61, cocoa butter, laurin butter, and glycerogelatin.

[0081] The preferred dosage of the probiotic preparation of the present invention varies depending on the patient's condition and weight, the severity of the disease, the drug form, the route of administration, and the duration of administration, but can be appropriately selected by those skilled in the art. However, for desirable effects, the probiotic preparation is preferably administered at a daily dose of 0.01 mg / kg to 10 g / kg, preferably 1 mg / kg to 1 g / kg. Administration may be administered once a day or divided into several doses. Therefore, the above dosage does not limit the scope of the present invention in any way.

[0082] The composition of the present invention can be administered to mammals, such as mice, rats, livestock, and humans, via various routes. Any method of administration is conceivable, including oral, rectal, or intravenous administration.

[0083] In addition, the present invention provides a treatment method for treating a patient with a disease caused by the promotion of the growth of beneficial intestinal bacteria or the growth of harmful intestinal bacteria, comprising administering the probiotic preparation to the patient with a disease caused by the promotion of the growth of beneficial intestinal bacteria or the growth of harmful intestinal bacteria.

[0084] In addition, the present invention provides a use of the probiotic preparation for producing a drug for promoting the growth of beneficial intestinal bacteria or treating a disease caused by the growth of harmful intestinal bacteria.

[0085] In addition, the present invention provides a health functional food containing a probiotic preparation as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing and improving diseases caused by the growth of harmful intestinal bacteria.

[0086] The composition of the present invention comprises the probiotic preparation in an amount of 0.01 to 99% by weight based on the total weight of the composition.

[0087]

[0088] However, the composition as described above is not necessarily limited to this and may vary depending on the patient's condition and the type and progression of the disease.

[0089] A composition comprising the probiotic preparation of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

[0090]

[0091] The composition including the probiotic preparation according to the present invention can be formulated and used in the form of oral formulations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., external preparations, suppositories, and sterile injection solutions, respectively, according to conventional methods, and carriers, excipients, and diluents that can be included therein include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, magnesium stearate, and mineral oil. When formulated, it is usually prepared using diluents or excipients such as fillers, bulking agents, binders, wetting agents, disintegrating agents, and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, and capsules, and these solid preparations are prepared by mixing the probiotic preparation with at least one excipient, such as cotton, starch, calcium carbonate, sucrose or lactose, gelatin, etc. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral administration include suspensions, oral solutions, emulsions, and syrups, and in addition to commonly used simple diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, and suppositories. Non-aqueous solutions and suspensions can include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate.Suppository bases that can be used include witepsol, macrogol, tween 61, cocoa butter, laurin butter, and glycerogelatin.

[0092] The preferred dosage of the probiotic preparation of the present invention varies depending on the patient's condition and weight, the severity of the disease, the drug form, the route of administration, and the duration of administration, but can be appropriately selected by those skilled in the art. However, for desirable effects, it is recommended to administer the probiotic preparation at a daily dose of 0.01 mg / kg to 10 g / kg, preferably 1 mg / kg to 1 g / kg. Administration may be administered once a day or divided into several doses. Therefore, the above dosage does not limit the scope of the present invention in any way.

[0093] The composition of the present invention can be administered to mammals such as rats, mice, livestock, and humans via various routes. All modes of administration are conceivable, and for example, administration can be performed orally, rectally, or intravenously.

[0094]

[0095] In addition, the present invention provides a health functional food containing the above probiotic preparation as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing and improving diseases caused by the growth of harmful intestinal bacteria.

[0096] “Health functional food” as defined herein means a food manufactured and processed using raw materials or ingredients with functionality useful to the human body as defined in Act No. 6727 on Health Functional Foods, and “functionality” means consumption for the purpose of obtaining a beneficial effect for health purposes such as regulating nutrients for the structure and function of the human body or physiological effects.

[0097] The health functional food of the present invention for promoting the growth of beneficial intestinal bacteria or preventing and improving diseases caused by the growth of harmful intestinal bacteria contains the above-mentioned component in a weight percentage of 0.01 to 95%, preferably 1 to 80%, based on the total weight of the composition.

[0098] Moreover, the composition of the present invention may be a health food for the purpose of promoting the proliferation of beneficial intestinal bacteria or treating or improving diseases caused by the proliferation of harmful intestinal bacteria.

[0099] In addition, it is possible to manufacture and process it into a pharmaceutical dosage form such as powder, granule, tablet, capsule, pill, suspension, emulsion, syrup, or a health functional food in the form of tea bag, infused tea, or health drink for the purpose of promoting the growth of beneficial intestinal bacteria or preventing and improving diseases caused by the growth of harmful intestinal bacteria.

[0100] In addition, the present invention provides a health supplement containing the above probiotic preparation as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing and improving diseases caused by the growth of harmful intestinal bacteria.

[0101] Among the health supplements defined herein, the main ingredient means that the content of the active ingredient relative to the total composition weight is in the range of about 30% to 99%, preferably in the range of 50% to 99%, and more preferably in the range of 70% to 99%.

[0102] The health functional beverage composition of the present invention contains the probiotic preparation as an essential ingredient in the indicated proportion, and has no particular limitations on other ingredients, and may contain various flavoring agents or natural carbohydrates as additional ingredients, like conventional beverages. Examples of the aforementioned natural carbohydrates include monosaccharides such as glucose, fructose, etc.; disaccharides such as maltose, sucrose, etc.; and polysaccharides such as dextrin, cyclodextrin, etc., and conventional sugars, and sugar alcohols such as xylitol, sorbitol, erythritol, etc. In addition to the aforementioned flavoring agents, natural flavoring agents (thaumatin, stevia extracts (e.g., rebaudioside A, glycyrrhizin, etc.)) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The proportion of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g, per 100 ml of the composition of the present invention.

[0103] When the probiotic preparation according to the present invention is used as a food additive, the probiotic preparation may be added as is or used together with other foods or food ingredients, and may be used appropriately according to a conventional method. Examples of foods to which the above substances may be added include dairy products including meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes, and include all health foods in the conventional sense.

[0104] In addition to the above, the composition of the present invention may contain various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and thickening agents (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. In addition, the composition of the present invention may contain fruit pulp for the production of natural fruit juice and fruit juice drinks and vegetable drinks. These components may be used independently or in combination. The proportion of these additives is not so critical, but is typically selected in the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

[0105] In addition, the probiotic preparation of the present invention may be added to food or beverages for the purpose of promoting the growth of beneficial intestinal bacteria or preventing diseases caused by the growth of harmful intestinal bacteria. In this case, the amount of the probiotic preparation in the food or beverage may be added in an amount of 0.01 to 15% by weight of the total food weight, and in the case of a health beverage composition, the amount may be added in an amount of 0.02 to 5 g, preferably 0.3 to 1 g, based on 100 ml.

[0106] In addition, the present invention provides a food or food additive containing the above probiotic preparation as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing and improving diseases caused by the growth of harmful intestinal bacteria.

[0107] Examples of items listed in the above "Food Additives Codex" include chemically synthesized products such as ketones, glycine, potassium citrate, nicotinic acid, and cinnamic acid; natural additives such as persimmon pigment, licorice extract, crystalline cellulose, and guar gum; and mixed preparations such as sodium L-glutamate preparations, alkaline agents for noodles, preservative preparations, and tar color preparations.

[0108] Functional foods containing the probiotic preparation of the present invention include confectionery such as bread, rice cakes, dried fruits, candies, chocolates, chewing gums, and jams; ice cream products such as ice cream, frozen desserts, and ice cream powders; dairy products such as milk, low-fat milk, lactose-hydrolyzed milk, processed milk, goat milk, fermented milk, buttermilk, condensed milk, milk cream, buttermilk, natural cheese, processed cheese, powdered milk, and whey; meat products such as processed meat products, processed egg products, and hamburgers; fish products such as processed fish products such as fish cakes, ham, sausage, and bacon; noodles such as ramen, dried noodles, fresh noodles, fried noodles, luxurious dried noodles, improved boiled noodles, frozen noodles, and pasta; lactic acid bacteria drinks such as fruit drinks, vegetable drinks, carbonated drinks, soy milk, and yogurt; and mixed drinks such as soy sauce, soybean paste, red pepper paste, and soybean paste. Seasonings such as cheonggukjang, mixed sauces, vinegar, sauces, tomato ketchup, curry, dressings, margarine, shortening, and pizza are included, but are not limited to these.

[0109] In the process of manufacturing the above health functional food, the content of the probiotic preparation according to the present invention added to food, including beverages, can be appropriately increased or decreased as needed.

[0110]

[0111]

[0112] As a result of conducting experiments on the preparation of an emulsion according to heating temperature (Experimental Example 1); an experiment on the effect on the proliferation of beneficial bacteria in the intestines (Experimental Example 2); an experiment on the viability of lactic acid bacteria in artificial gastric juice (pH 2) (Experimental Example 3); an experiment on the efficacy of the composition in improving constipation (Simple Clinical Example 1); and an experiment on the preference according to the type of nuts contained in the composition (Simple Clinical Example 2), it was confirmed that the lactic acid bacteria preparation of the present invention strongly proliferates beneficial bacteria in the intestines, exhibits excellent viability of lactic acid bacteria in gastric acid, and exhibits an excellent and advanced effect in terms of improving constipation and preference compared to existing known lactic acid bacteria preparations.

[0113] Therefore, the present invention can provide an excellent gastric acid stable probiotic preparation and a method for producing the same.

[0114]

[0115] Figure 1 shows the results of an experiment observing the properties of the emulsifier of the present invention at various temperatures;

[0116] Figure 2 is an experimental result measuring the effect of the lactic acid bacteria preparation of the present invention on strain growth in a selective medium (MRS agar plate);

[0117] Figure 3 shows the results of a test on the viability of lactic acid bacteria in artificial gastric fluid (pH 2) of the present invention.

[0118] It will be apparent to those skilled in the art that various modifications and variations can be made to the compositions, uses, and manufactures of the present invention without departing from the spirit or scope of the invention.

[0119] However, the following examples and experimental examples are merely illustrative and do not limit the scope of the present invention, and the content of the present invention is not limited by the following examples and experimental examples.

[0120] Example 1. Preparation of lactic acid bacteria preparation with added emulsifier (Composition: Table 1)

[0121] 20g of sunflower oil (vegetable oil, Sajodaerim Co., Ltd.) was placed in a stirring heater (Hotplate Stirrer. MSH-20D) and heated to 75 to 85℃, then 0.4g of glycerin fatty acid ester (emulsifier, ES food ingredient) was added and mixed while stirring.

[0122] The vegetable oil and emulsifier mixture of the above step was stirred and emulsified in a circulating cooling water bath (low-temperature circulating constant-temperature water bath, SH-WB-7CDR) maintained at 0 to 4°C until the temperature of the mixture reached 4 to 10°C. (Test-1 40°C, 60°C, 85°C, 100°C, respectively. The experimental process and experimental results for manufacturing emulsions at each temperature are described in Experimental Example 1 below.)

[0123] Into the emulsified oil formed in the form of a gel as described above, mixed lactic acid bacteria, i.e., Lactobacillus acidophilus DDS-1 (Ollab Co., Ltd.), Lactobacillus rhamnosus GG (Ollab Co., Ltd.), Lactobacillus plantarum VI-07 (Bytech Co., Ltd.), Bifidobacterium longum BL5 (Bytech Co., Ltd.), and Bifidobacterium animalis ssp. lactis BLC1 (Ollab Co., Ltd.), were added the same number of bacteria (2x10 9 1 g of mixed lactic acid bacteria evenly mixed in a ratio of 100 cfu / g was used. (For the observation of the viability of Test-2 lactic acid bacteria, the activity experiment process and results in artificial gastric fluid (pH 2) are shown in Experimental Example 2 below.)

[0124] As additives to form a suitable viscosity of the composition, enhance flavor / texture, and further enhance the activity of lactic acid bacteria, natural sweetener fructooligosaccharide (food for lactic acid bacteria; prebiotics) and peanuts (nuts, Daechang Food), which contain a large amount of unsaturated fatty acids and are beneficial to various health factors including blood circulation, were added and stirred with a mixing mixer (OFM-1507B, Opel Co., Ltd.) at a speed of 30 to 50 rpm for more than 1 hour until all ingredients were evenly mixed.

[0125] At this time, the best way to use nuts is to grind them with a grinding device (ultra-high-speed vacuum blender, KCB-001W, Kucham Co., Ltd.) so that the particle size is 0.01 to 0.1 millimeters (mm) (approximately the size of a paste). (Test-3: The reason for selecting peanuts with the highest preference was the evaluation. For the results of the experiment in which peanuts, almonds, and walnuts were mixed into the composition and evaluated for swallowing and preference, refer to Experimental Example 3.)

[0126] Composition of lactic acid bacteria preparationComposition*Composition AComposition BComposition COlive oil 20 g20 g95.5 gEmulsifier 0.3 g-1.5 gFructooligosaccharide 1 g1 g1 gCrushed nuts 76.7 g77 gMixed lactic acid bacteria 2 g2 g2 gTotal 100 g100 g100 g*Composition A:Olive oil+Emulsifier+Fructooligosaccharide+Crushed nuts+Mixed lactic acid bacteria / Composition B:Olive oil+Fructooligosaccharide+Crushed nuts+Mixed lactic acid bacteria / Composition C:Olive oil+Emulsifier+Fructooligosaccharide+Mixed lactic acid bacteria

[0127] Example 2. Preparation of a lactic acid bacteria preparation without added emulsifier (Composition: Table 1)

[0128] Without using the emulsification step using vegetable oil and emulsifier described in the method for preparing composition A of Example 1 above, sunflower oil (vegetable oil), five types of mixed lactic acid bacteria, and the remaining additives (fructooligosaccharides and ground peanuts) were simply mixed under the same conditions to prepare a lactic acid bacteria preparation without added emulsifier (composition B), which was used as a sample in the following experimental example.

[0129]

[0130] Example 3. Preparation of comparative lactic acid bacteria preparation

[0131] A comparative enteric lactic acid bacteria complex containing 10.0 wt% of lactic acid bacteria powder, 11.5 wt% of sulfur (suspending agent), 1.0 wt% of soy lecithin (emulsifier), and the remainder of rTG type purified fish oil (edible oil) disclosed in the existing literature (Korean Patent Publication No. 10-2023-0001175) was prepared using the manufacturing method of Example 1 of the existing literature, and used as a sample in the following experimental example.

[0132]

[0133]

[0134] Experimental Example 1. Experiment on the production of emulsions at different temperatures

[0135] 2.0 g of emulsifier (glycerin fatty acid ester, ES Food Ingredients) was added to 100 g of sunflower oil (vegetable oil, Sajo Daerim Co., Ltd.) and heated to four temperatures of 40 ℃ / 60 ℃ / 85 ℃ / 100 ℃, and the mixture was stirred and emulsified in a circulating cooling water bath (low-temperature circulating constant-temperature water bath, SH-WB-7CDR) maintained at 0 to 4 ℃ until the temperature of the mixture reached 4 to 10 ℃, and the optimal temperature suitable for producing an emulsion was measured.

[0136] As a result of the above experiment, as shown in Fig. 1, each photograph shows the shape of the emulsion formed at each temperature. At 40°C and 60°C, the emulsifier was not sufficiently dissolved, so the emulsion was not properly formed. However, at 85°C and 100°C, the emulsifier was sufficiently dissolved, so the emulsion was evenly formed. (See Fig. 1: Experiment on the properties of the emulsion at various temperatures of the emulsifier)

[0137]

[0138] Experimental Example 2. Effects on the proliferation of beneficial intestinal bacteria

[0139] In order to confirm the effect of the sample of the present invention obtained in the above example on the proliferation of intestinal beneficial bacteria, the following experiment was conducted with reference to the lactic acid bacteria count test method notified in the Food Code Microbiological Test Method Commentary (published by the National Institute of Food and Drug Safety Evaluation, Ministry of Food and Drug Safety).

[0140]

[0141] 2-1. Experimental strains

[0142] The strain was provided by the Korean Culture Center of Microorganisms (KCCM, Republic of Korea), and Lactobacillus acidophilus ATCC 4356 (ATCC: American Type Culture Collection) was used as the intestinal beneficial bacteria. Lactobacilli MRS broth (288130, BD difco, USA) was used as the growth medium for culturing the intestinal beneficial bacteria, and the strain was activated by subculture at least three times before use in the experiment.

[0143]

[0144] 2-2. Experimental process

[0145] The number of lactic acid bacteria produced was measured by counting the number of lactic acid bacteria colonies using MRS plate measurement medium, and multiplying the average number of colonies by the dilution factor to calculate the number of viable bacteria per mL of culture solution. 10 mL of the test sample was homogenized by adding a 9-fold dilution solution (sterile saline solution) (10 -1 solution). Test solution (10 -1 Add the dilution solution to 1 mL of solution to make 10 mL. -2 After making the test solution, dilute it using the same procedure.

[0146] Each diluted test solution was inoculated at 1 (v / v)% onto a Lactobacillus selective medium (MRS agar plaste), and anaerobically cultured at 37°C for 72±3 hours. The plate that produced 15 to 300 colonies per plate was selected to count the number of colonies, and the number of bacteria was calculated by multiplying the measured number of colonies by the dilution factor.

[0147]

[0148] 2-3. Experimental Results

[0149] Based on the viability results of the above-mentioned examples, it was confirmed that the best method for manufacturing the emulsion was at 85°C. (See Table 2 and Figure 2)

[0150] Effect of strain growth on selective medium (MRS agar plate) of lactic acid bacteria preparation Temperature of emulsion Number of bacteria 40℃ 2.7×10 2 cfu / g60℃7.7×10 2 cfu / g85℃1.7×10 3 cfu / g100℃1.5×10 3 cfu / g

[0151] Experimental Example 3. Lactic acid bacteria survival test in artificial gastric fluid (pH 2)

[0152] In order to confirm the effect of the sample of the present invention obtained in the above example on the viability of lactic acid bacteria in artificial gastric fluid (pH 2), the following experiment was conducted by applying the method disclosed in the literature (Jeon et al., 2007, Identification and Characterization of Lactic Acid Bacteria Starters Isolated from the Commercial Drink-Yogurt Products, Korean J. Food Sci. Ani. Resour. 27(4), 509-516).

[0153]

[0154] 3-1. Experimental strains and experimental process

[0155] To confirm the viability of lactic acid bacteria in the composition, composition A was stirred in artificial gastric fluid (pH 2) for a certain period of time (1 hour), and the collected composition was immediately plated on a lactobacillus selective medium (MRS agar plate, MB-M1024-P50, KisanBio) and anaerobically cultured at 37℃ for 72±3 hours. The plate that produced 15 to 300 colonies per plate was selected, the number of colonies was counted, and the number of bacteria was calculated by multiplying the measured number of colonies by the dilution factor. For comparison, a composition control group in which a mixture of lactic acid bacteria was mixed in non-emulsified oil was used.

[0156]

[0157] 3-2. Experimental Results

[0158] The above experimental results demonstrate that the emulsified oil and lactic acid bacteria mixture exhibited superior survival rates compared to the control group. Furthermore, when peanut powder was added as an additive to the emulsified oil and cultured under identical conditions, the lactobacillus survival rate in the lactic acid bacteria mixture containing more peanut powder was significantly higher. (See Table 3 and Figure 3.)

[0159] Results of the experiment on the survival of lactic acid bacteria in artificial gastric fluid (pH 2) Composition composition*Bacterial countOil (O) + lactic acid bacteria (L) mixture 1.2×10 2 cfu / g oil (O) + emulsifier (E) + lactic acid bacteria (L) mixture 1.7×10 3 cfu / g oil (O) + emulsifier (E) + lactic acid bacteria (L) mixture + peanut powder (P) 2.4 × 10 3 cfu / g*oil (O) / emulsifier (E) / lactic acid bacteria (L) / peanut powder (P) are expressed respectively.

[0160]

[0161] Simple clinical example 1. Experimental study on the constipation-improving efficacy of the composition.

[0162] In order to confirm the constipation improvement efficacy of the sample of the present invention obtained in the above example, the following experiment was conducted.

[0163] Composition A, used for testing to improve constipation, was evaluated for its effect on improving constipation in 50 volunteers in the Seoul area, aged between 20 and 50, who were experiencing inconvenience in their daily lives due to constipation.

[0164] The experimental subjects were ingested Composition A twice a day (morning / evening) at 5g per serving for ten days and answered the following questions. In addition, Composition B was ingested by 50 control subjects in the same manner as the experimental subjects.

[0165] As a result of the above experiment, it was confirmed that the composition A of the present invention has an excellent effect in improving constipation. (See Tables 4 and 5)

[0166] After taking Composition A (lactic acid bacteria composition made with emulsified oil), the effect of improving constipation was very good. Slightly good. No change. Number of bowel movements: 40 people, 7 people, 3 people, 50 people. Bowel movements time: 35 people, 12 people, 3 people, 50 people. Abdominal distension: 31 people, 15 people, 4 people, 50 people. Feeling of incomplete evacuation: 36 people, 10 people, 4 people, 50 people.

[0167] Effect of improving constipation after taking Composition B (lactic acid bacteria composition made with non-emulsified oil) Very good Slightly good No change Number of bowel movements 11 people 26 people 13 people 50 people Bowel movements time 15 people 28 people 7 people 50 people Abdominal distension 7 people 21 people 22 people 50 people Feeling of incomplete evacuation 16 people 23 people 11 people 50 people

[0168] Simple clinical example 2. Preference evaluation experiment according to the type of nuts contained in the composition.

[0169] In order to confirm the preference according to the type of nuts contained in the composition of the sample of the present invention obtained in the above example, an experiment was conducted as follows.

[0170] After preparing a composition using peanuts, almonds, and walnuts as ground nuts, the taste, aroma, texture, and overall preference of each type of nut were evaluated by having 20 adult men and women consume 3g of each.

[0171] The criteria for the preference evaluation were based on a 5-point scale, with the average value indicated (5 points for very good, 4 points for good, 3 points for average, 2 points for bad, 1 point for very bad) (see Table 6).

[0172] Results of the preference evaluation by nut Type Preference evaluation Taste Aroma Texture Overall preference Peanut 4.7 4.6 4.5 4.60 Almond 4.4 4.4 4.6 4.47 Walnut 4.1 3.9 4.6 4.20

[0173] As a result of the above experiment, it was confirmed that the preference for the composition using peanut powder was the best.

[0174]

[0175] As a result of the above experiments, the effect of the lactic acid bacteria preparation of the present invention on the proliferation of beneficial bacteria in the intestines (Experimental Example 2); the experiment on the viability of lactic acid bacteria in artificial gastric fluid (pH 2) (Experimental Example 3); the experiment on the efficacy of the composition in improving constipation (Simple Clinical Example 1); and the experiment on the preference according to the type of nuts contained in the composition (Simple Clinical Example 2) were performed, and it was confirmed that the lactic acid bacteria preparation of the present invention strongly proliferated beneficial bacteria in the intestines, exhibited excellent viability of lactic acid bacteria in gastric acid, and exhibited superior and more advanced effects than existing known lactic acid bacteria preparations in terms of improving constipation and preference.

[0176]

[0177] Below, an example of a composition comprising the probiotic preparation of the present invention is described, but the present invention is not intended to be limited thereto, but is merely intended to be described in detail.

[0178]

[0179] Preparation Example 1. Preparation of a powder

[0180] Composition A ----------------------------------------------- 20 mg

[0181] Lactose --------------------------------------------------- 100 mg

[0182] Talc ---------------------------------------------------- 10 mg

[0183] The above ingredients are mixed and filled into a sealed bag to prepare a powder.

[0184]

[0185] Preparation Example 2. Preparation of tablets

[0186] Composition A ---------------------------------------------- 10 mg

[0187] Cornstarch -------------------------------------------- 100 mg

[0188] Lactose -------------------------------------------------- 100 mg

[0189] Magnesium stearate ------------------------------------- 2 mg

[0190] After mixing the above ingredients, tablets are manufactured by pressing them according to the usual tablet manufacturing method.

[0191]

[0192] Preparation Example 3. Preparation of capsules

[0193] Composition A ---------------------------------------------- 10 mg

[0194] Crystalline cellulose --------------------------------------- 3 mg

[0195] Lactose --------------------------------------------- 14.8 mg

[0196] Magnesium stearate --------------------------------- 0.2 mg

[0197] The above ingredients are mixed according to the conventional capsule manufacturing method and filled into a gelatin capsule to manufacture a capsule.

[0198]

[0199] Preparation Example 4. Preparation of Injectable

[0200] Composition A ----------------------------------------- ---- 10 mg

[0201] Mannitol ------------------------------------------------ 180 mg

[0202] Sterile distilled water for injection ----------------------------------- 2974 mg

[0203] Na2HPO4,12H2O ---------------------------- 26 mg

[0204] According to the manufacturing method of a conventional injection, it is manufactured with the above ingredient content per 1 ampoule (2 ml).

[0205]

[0206] Preparation Example 5. Preparation of liquid preparation

[0207] Composition A ----------------------------------------------- 20 mg

[0208] Isunghwadang ------------------------------------------------- 10 g

[0209] Mannitol ---------------------------------------------------- 5 g

[0210] Purified water --------------------------------------------------- Appropriate amount

[0211] According to the usual method of manufacturing a liquid, each ingredient is added to purified water and dissolved, an appropriate amount of lemon flavor is added, the above ingredients are mixed, purified water is added, and the total volume is adjusted to 100 ml. Then, the liquid is filled into a brown bottle and sterilized to manufacture the liquid.

[0212]

[0213] Example 6. Manufacturing of health food

[0214] Composition A --------------------------------------------- 1000 mg

[0215] Vitamin Mixture -------------------------------------------- Appropriate amount

[0216] Vitamin A Acetate ------------------------------------- 70 μg

[0217] Vitamin E ----------------------------------------------- 1.0 mg

[0218] Vitamin B1 --------------------------------------------- 0.13 mg

[0219] Vitamin B2 --------------------------------------------- 0.15 mg

[0220] Vitamin B6 ---------------------------------------------- 0.5 mg

[0221] Vitamin B12 --------------------------------------------- 0.2 μg

[0222] Vitamin C ------------------------------------------------ 10 mg

[0223] Biotin -------------------------------------------------- 10 μg

[0224] Nicotinamide ----------------------------------------- 1.7 mg

[0225] Folic acid ---------------------------------------------------- 50 μg

[0226] Calcium pantothenate ------------------------------------------ 0.5 mg

[0227] Mineral mixture -------------------------------------------- Appropriate amount

[0228] Ferrous sulfate --------------------------------------------- 1.75 mg

[0229] Zinc oxide ---------------------------------------------- 0.82 mg

[0230] Magnesium carbonate ------------------------------------------ 25.3 mg

[0231] Monopotassium phosphate --------------------------------------------- 15 mg

[0232] Dicalcium phosphate --------------------------------------------- 55 mg

[0233] Potassium citrate ---------------------------------------------- 90 mg

[0234] Calcium carbonate ----------------------------------------------- 100 mg

[0235] Magnesium chloride ------------------------------------------ 24.8 mg

[0236] The composition ratio of the above vitamin and mineral mixture is a preferred example of a mixture of ingredients suitable for relatively healthy foods, but the mixing ratio may be arbitrarily modified, and the above ingredients may be mixed according to a conventional health food manufacturing method, granules may be manufactured, and used to manufacture a health food composition according to a conventional method.

[0237]

[0238] Preparation Example 7. Preparation of a health drink

[0239] Composition A ----------------------------------------------- 1000mg

[0240] Citric acid ------------------------------------------------- 1000 mg

[0241] Oligosaccharide ------------------------------------------------- 100 g

[0242] Plum concentrate ------------------------------------------------- 2 g

[0243] Taurine ----------------------------------------------------- 1 g

[0244] Add purified water ------------------------------------ Total 900 ml

[0245] The above ingredients are mixed according to a conventional health drink manufacturing method, then stirred and heated at 85°C for about 1 hour, the resulting solution is filtered, placed in a sterilized 2L container, sealed and sterilized, and then stored in a refrigerator, and then used to manufacture the health drink composition of the present invention.

[0246] The above composition ratio is a preferred example of a mixture of ingredients relatively suitable for a preferred beverage, but the mixing ratio may be arbitrarily modified according to regional and ethnic preferences such as demand class, demand country, and intended use.

[0247] As described above, the probiotic preparation of the present invention was tested in an experiment on manufacturing an emulsion at different heating temperatures (Experimental Example 1); an experiment on the effect on the proliferation of beneficial intestinal bacteria (Experimental Example 2); an experiment on the viability of lactic acid bacteria in artificial gastric juice (pH 2) (Experimental Example 3); an experiment on the effect of improving constipation of the composition (Simple Clinical Example 1); an experiment on the preference according to the type of nuts contained in the composition (Simple Clinical Example 2), etc. As a result, it was confirmed that the lactic acid bacteria preparation of the present invention strongly proliferates beneficial intestinal bacteria, exhibits excellent viability of lactic acid bacteria in gastric acid, and exhibits an excellent and advanced effect in terms of improving constipation and preference compared to existing known lactic acid bacteria preparations, and thus can be usefully utilized as an excellent probiotic preparation in various pharmaceutical compositions, health functional foods, and health supplements.

Claims

1. A first step of producing a mixture of edible oil and emulsifier by heating 1 to 90 parts by weight of edible oil to 70 to 95°C and then mixing and stirring 0.1 to 5 parts by weight of the emulsifier; A second step of cooling and stirring the mixture of the first step until the temperature reaches 2 to 20°C to produce a gel-like coagulated mixture; Step 3: Slowly adding 1 part by weight of lactic acid bacteria to a gel-shaped coagulated mixture and stirring at low speed to create a lactic acid bacteria mixture; A method for producing a gastric acid stable probiotic preparation, characterized by performing a process including:

2. In paragraph 1, Step 4: Adding ground nuts to the mixture obtained through Step 3 and stirring to prepare a mixture of nut lactic acid bacteria; A fifth step of adding one or more additional ingredients selected from prebiotics and adhesive agents to the mixture of the fourth step and stirring; A method for producing a gastric acid-safe probiotic preparation characterized by further comprising:

3. In paragraph 2, A method for producing a gastric acid-stable probiotic preparation, characterized in that the nut powder of the above 4 steps has a particle size of 0.01 to 0.5 mm and is stirred at a stirring speed of 30 to 50 rpm for 30 minutes to 3 hours until all ingredients are evenly mixed.

4. A gastric acid stable probiotic preparation containing (a) lactic acid bacteria, (b) edible oil, (c) emulsifier and (d) nuts as essential ingredients.

5. In paragraph 4, The above probiotic preparation is a gastric acid stable probiotic preparation characterized in that the mixing weight ratios between individual components are (a) 1.0 parts by weight of lactic acid bacteria, (b) 1.0 to 90.0 parts by weight of edible oil, (c) 0.1 to 5.0 parts by weight of emulsifier, and (d) 5.0 to 90.0 parts by weight of nuts.

6. In paragraph 4, A probiotic preparation characterized in that the above (a) lactic acid bacteria is one or more strains selected from microorganisms of the genus Lactobacillus, the genus Bifidobacterium, the genus Bacillus, the genus Streptococcus, and the genus Enterococcus.

7. In paragraph 4, A gastric acid stable probiotic preparation characterized in that the above (b) edible oil is at least one ingredient selected from the group consisting of avocado oil, canola oil, evening primrose oil, coconut oil, Brazil nut oil, corn oil, cottonseed oil, flaxseed oil, grapeseed oil, hemp oil, olive oil, palm oil, safflower oil, soybean oil, peanut oil, sunflower seed oil, krill oil, anchovy oil, salmon oil, rice bran oil, brown rice oil, coconut oil, rosehip oil, and tuna oil.

8. In paragraph 4, A gastric acid-stable probiotic preparation characterized in that the above (c) emulsifier is at least one ingredient selected from the group consisting of propylene glycol fatty acid ester, glycerin fatty acid ester, sucrose fatty acid ester, polyglycerin fatty acid ester, sorbitan fatty acid ester, organic acid monoglyceride, polyoxyethylene fatty acid ester, propylene glycol fatty acid ester, polyoxyethylene fatty acid ester, polysorbate, egg yolk lecithin, soybean lecithin, carboxymethylcellulose, glycerin, soybean phospholipid, and stearyl calcium lactate.

9. In paragraph 4, A stomach acid stable probiotic preparation characterized in that the above (d) nuts are at least one ingredient selected from the group consisting of peanuts, almonds, walnuts, pine nuts, pistachios, pecans, macadamias, hazelnuts, cashews, Brazil nuts, chestnuts, soybeans, cacao nibs, coconuts, sunflower seeds, and pumpkin seeds.

10. In paragraph 4, A gastric acid stable probiotic preparation containing, in addition to the above essential ingredients, one or more additional ingredients selected from (e) prebiotics and (f) adhesive agents.

11. In paragraph 10, A probiotic preparation characterized in that the mixing weight ratios of the individual components are (a) 1.0 parts by weight of lactic acid bacteria, (b) 1 to 60 parts by weight of edible oil, (c) 0.1 to 5.0 parts by weight of emulsifier, (d) 5.0 to 90.0 parts by weight of nuts, (e) 0.01 to 5.0 parts by weight of prebiotics, and (f) 0.1 to 5.0 parts by weight of adhesive agent.

12. In paragraph 10, A gastric acid-stable probiotic preparation characterized in that the above (e) prebiotics are at least one component selected from pentose sugars such as xylose and arabinose; glucose, mannose, fructose, galactose; lactulose, lactitol, sucrose, lactose, maltose, trehalose; fructo-oligosaccharides, raffinose, stachyose, maltodextrin; amylose, amylopectin, starch, cellulose, and pectin.

13. In paragraph 10, A gastric acid-stable probiotic preparation characterized in that the above (f) adhesive agent is at least one ingredient selected from the group consisting of sodium silicoaluminate, xanthan gum, carrageenan, guar gum, diutan gum, cellulose gum, gellan gum, pectin, and carboxymethylcellulose.

14. A pharmaceutical composition containing the gastric acid-stable probiotic preparation described in Article 4 as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing and treating diseases caused by the growth of harmful intestinal bacteria.

15. In paragraph 14, A pharmaceutical composition characterized by diseases caused by the proliferation of harmful intestinal bacteria, including antibiotic-associated diarrhea, watery diarrhea, vomiting, inflammatory bowel disease such as ulcerative colitis and Crohn's disease accompanied by nausea, abdominal distension, constipation, acute or chronic irritable colitis, radiation-induced enteritis, or Helicobacter pylori infection.

16. A health functional food containing the gastric acid-stable probiotic preparation described in Article 4 as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing or improving diseases caused by the growth of harmful intestinal bacteria.

17. In paragraph 16, The above health functional food is a health functional food in the form of pharmaceutical dosage forms such as powder, granules, tablets, capsules, pills, suspensions, emulsions, syrups, or tea bags, infused teas, or health drinks.

18. A health supplement containing the gastric acid-stable probiotic preparation described in Article 4 as an effective ingredient for promoting the growth of beneficial intestinal bacteria or preventing or improving diseases caused by the growth of harmful intestinal bacteria.

19. A food additive containing the gastric acid-stable probiotic preparation described in Article 4 as an effective ingredient, which promotes the growth of beneficial intestinal bacteria or prevents or improves diseases caused by the growth of harmful intestinal bacteria.

20. A treatment method for treating a patient with a disease caused by the promotion of the growth of beneficial intestinal bacteria or the growth of harmful intestinal bacteria, comprising administering a gastric acid-stable probiotic preparation described in Article 4 to the patient with a disease caused by the promotion of the growth of beneficial intestinal bacteria or the growth of harmful intestinal bacteria.

21. Use of the gastric acid-stable probiotic preparation described in Article 4 for manufacturing a drug for promoting the growth of beneficial intestinal bacteria or treating diseases caused by the growth of harmful intestinal bacteria.

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