Branched dextran, intestinal regulator, cholesterol lowering agent, blood glucose level rise inhibitor, starch retrogradation inhibitor, bread texture improver, raw material, and method for producing branched dextran

Abstract

To provide a dextran having a water-soluble dietary fiber content of a predetermined level or higher.SOLUTION: The present invention provides a branched dextran derived from a lactic acid bacterium belonging to Leuconostoc citreum in which the glycosidic bonds of the branched dextran comprise 50% to less than 99% of α-1, 6 bonds, 1% to less than 40% of α-1,2 bonds, and 0.1 to less than 20% of α-1,3 bonds, the value obtained by dividing the weight average molecular weight of the branched dextran by the number average molecular weight of the branched dextran is 100 to less than 1500, and the branched dextran contains 80% by mass or more of water-soluble dietary fiber.SELECTED DRAWING: None

Description

Technical Field

[0001] The present invention relates to branched dextran, intestinal regulator, cholesterol-lowering agent, blood glucose level increase inhibitor, starch retrogradation inhibitor, bread texture improver, raw material, and method for producing branched dextran.

Background Art

[0002] The polysaccharide produced extracellularly by lactic acid bacteria is called exopolysaccharide (EPS).

[0003] Examples of exopolysaccharide include dextran. Dextran is produced by lactic acid bacteria using sucrose as a raw material, has glucose as the sole constituent, and has a structure mainly composed of α-1,6 glycosidic bonds.

[0004] Examples of lactic acid bacteria that produce dextran include Leuconostoc mesenteroides. Patent Document 1 discloses dextran production by culturing strains belonging to Leuconostoc mesenteroides.

[0005] Known uses of dextran include substitute blood plasma, cosmetics, etc. (Patent Documents 2 to 4).

Prior Art Documents

Patent Documents

[0006]

Patent Document 1

Patent Document 2

[0007] However, there is a need for dextran, which is produced by lactic acid bacteria other than Leuconostoc mesenteroides and has superior functionality.

[0008] Other lactic acid bacteria besides Leuconostoc mesenteroides include those belonging to the Leuconostoc citreum group. Furthermore, a key function required of dextran is a high content of water-soluble dietary fiber.

[0009] This invention has been made in view of the above circumstances, and aims to provide dextran having a water-soluble dietary fiber content of a predetermined level or higher. [Means for solving the problem]

[0010] As a result of our investigations, we have newly discovered that the above problem can be solved by adjusting the ratio of weight-average molecular weight (Mw) and number-average molecular weight (Mn) of branched dextran produced by Leuconostoc citreum, and have completed the present invention. More specifically, the present invention provides the following.

[0011] (1) A branched dextran derived from lactic acid bacteria belonging to Leuconostoc citreum, The glycosidic bonds of the branched dextran consist of α-1,6 bonds accounting for 50% to less than 99% of the total glycosidic bonds, α-1,2 bonds accounting for 1% to less than 40%, and α-1,3 bonds accounting for 0.1% to less than 20%. The value obtained by dividing the weight-average molecular weight of the branched dextran by the number-average molecular weight of the branched dextran is 100 or more and less than 1500. The branched dextran contains 80% by mass or more of water-soluble dietary fiber. Branched dextran.

[0012] (2) The branched dextran according to (1), wherein the water retention rate of the branched dextran is 10% by mass or more.

[0013] (3) An intestinal regulator comprising the branched dextran according to (1) or (2).

[0014] (4) A cholesterol-lowering agent comprising the branched dextran according to (1) or (2).

[0015] (1) A blood sugar level rise inhibitor comprising the branched dextran according to (1) or (2).

[0016] (6) A starch retrogradation inhibitor comprising the branched dextran according to (1) or (2). <于

[0017] (7) A texture improver for bread comprising the branched dextran according to (1) or (2).

[0018] (8) A raw material for food and drink, cosmetics, pharmaceuticals, quasi-drugs or industrial use, comprising the branched dextran according to (1) or (2).

[0019] (9) A raw material comprising Leuconostoc citreum KD3 strain (Accession No. NITE P-03378) for producing the branched dextran according to (1) or (2).

[0020] [End]](10) A culturing step of culturing lactic acid bacteria belonging to Leuconostoc citreum, and a filtering step of recovering a filtrate containing branched dextran by filtering the culture after the culturing step, and in the filtering step, using a porous filtration membrane containing one or more selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide, and having a pore diameter of 0.01 μm or more and 1.00 μm or less, A method for producing branched dextran.

[0021] (11) The method for producing branched dextran according to (10), wherein the lactic acid bacteria belonging to Leuconostoc citreum is Leuconostoc citreum KD3 strain (Accession No. NITE P-03378). [Advantages of the Invention]

[0022] According to the present invention, a dextran having a water-soluble dietary fiber content of a predetermined level or higher is provided. [Modes for carrying out the invention]

[0023] The following describes embodiments of the present invention, but the present invention is not limited thereto.

[0024] <Branching dextran> The branched dextran of the present invention satisfies all of the following requirements. (Requirement 1) It is a branched dextran derived from lactic acid bacteria belonging to Leuconostoc citreum. (Requirement 2) The branched dextran's glycosidic bonds include 50% to less than 99% α-1,6 bonds, 1% to less than 40% α-1,2 bonds, and 0.1% to less than 20% α-1,3 bonds. (Requirement 3) The value obtained by dividing the weight-average molecular weight of branched dextran by the number-average molecular weight of branched dextran is 100 or more and less than 1500, and the branched dextran contains 80% by mass or more of water-soluble dietary fiber.

[0025] (1) Regarding (Requirement 1) The branched dextran of the present invention is derived from lactic acid bacteria belonging to Leuconostoc citreum.

[0026] In this invention, "branched dextran derived from lactic acid bacteria belonging to Leuconostoc citreum" includes branched dextran produced by lactic acid bacteria belonging to Leuconostoc citreum and its degradation products, etc.

[0027] A preferred example of a lactic acid bacterium belonging to the Leuconostoc citreum is the "Leuconostoc citreum KD3" strain, but it is not limited to this strain. This lactic acid bacteria strain was discovered by the inventors and deposited on February 10, 2020, with the Patent Microorganism Depositary Center of the National Institute of Technology and Evaluation (2-5-8 Kazusa-Kamatari, Kisarazu City, Chiba Prefecture 292-0818, Japan, Room 122), and was accepted under the depositary number "NITE P-03378".

[0028] (2) Regarding (Requirement 2) The branched dextran of the present invention mainly has a structure in which glucose is linked to α-1,6 glycosidic bonds. Furthermore, the branched dextran of the present invention has a branched structure by α-1,2 glycosidic bonds and α-1,3 glycosidic bonds. This distinctive sugar chain structure is presumed to be formed through the involvement of at least three enzymes (dextranscrase 1, dextranscrase 2, and alternance) in its synthesis.

[0029] The inventors have discovered a surprising finding: among branched dextrans derived from lactic acid bacteria belonging to Leuconostoc citreum, branched dextrans that satisfy both requirement 2 and requirement 3 (described later) tend to have a high amount of water-soluble dietary fiber.

[0030] Specifically, the branched dextran of the present invention has glycosidic bonds comprising 50% to less than 99% α-1,6 bonds, 1% to less than 40% α-1,2 bonds, and 0.1% to less than 20% α-1,3 bonds relative to the total glycosidic bonds.

[0031] The lower limit of the α-1,6 bond in the branched dextran of the present invention is preferably 60% or more, more preferably 70% or more, relative to the total glycosidic bond. The upper limit of α-1,6 bonds in the branched dextran of the present invention is preferably 90% or less, more preferably 85% or less, relative to the total number of glycosidic bonds.

[0032] The lower limit of α-1,2 linkages in the branched dextran of the present invention is preferably 5% or more, more preferably 10% or more, relative to the total number of glycosidic links. The upper limit of α-1,2 bonds in the branched dextran of the present invention is preferably 30% or less, more preferably 25% or less, relative to the total number of glycosidic bonds.

[0033] The lower limit of α-1,3 linkages in the branched dextran of the present invention is preferably 0.5% or more, more preferably 1% or more, relative to the total glycosidic linkage. The upper limit of α-1,3 bonds in the branched dextran of the present invention is preferably 10% or less, more preferably 5% or less, relative to the total number of glycosidic bonds.

[0034] The glycosidic bond composition of branched dextran was determined by the sugar chain structure analysis (NMR) shown in the examples. 1 Identified based on the 1H-NMR profile.

[0035] (3) Regarding (Requirement 3) The weight-average molecular weight (Mw) of the branched dextran of the present invention divided by the number-average molecular weight (Mn) of the branched dextran (hereinafter also referred to as "Mw / Mn") is 100 or more and less than 1500. Furthermore, the branched dextran contains 80% by mass or more of water-soluble dietary fiber.

[0036] "Mw / Mn" refers to the molecular weight distribution. Normally, the "Mw" and "Mn" values ​​are the same for a single molecule. Therefore, the closer the "Mw / Mn" ratio of a group of molecules is to 1, the higher the proportion of single molecules. As a result of the inventors' research, it was found that adjusting the molecular weight distribution of branched dextran within a predetermined range, that is, setting "Mw / Mn" to 100 or more and less than 1500, particularly increases its solubility in water. As a result, we found that if the "Mw / Mn" ratio is between 100 and 1500, the amount of water-soluble dietary fiber in branched dextran is a high value of 80% by mass or more.

[0037] The lower limit of "Mw" in the branched dextran of the present invention is preferably 200,000 or more, more preferably 300,000 or more. The upper limit of "Mw" in the branched dextran of the present invention is preferably 1,700,000 or less, more preferably 1,500,000 or less.

[0038] The lower limit of "Mn" in the branched dextran of the present invention is preferably 500 or more, more preferably 700 or more. The upper limit of "Mn" in the branched dextran of the present invention is preferably 5,000 or less, more preferably 4,000 or less.

[0039] The lower limit of "Mw / Mn" in the branched dextran of the present invention is preferably 100 or more, more preferably 110 or more. The upper limit of "Mw / Mn" in the branched dextran of the present invention is preferably 1,500 or less, more preferably 1,300 or less.

[0040] The lower limit of the amount of water-soluble dietary fiber in the branched dextran of the present invention is preferably 75% by mass or more, more preferably 80% by mass or more. The amount of water-soluble dietary fiber in the branched dextran of the present invention is preferably as high as possible, but the upper limit is, for example, 100% by mass or less, or 95% by mass or less.

[0041] The "Mw", "Mn", and "Mw / Mn" of the branched dextran are determined based on gel permeation chromatography (GPC) as shown in the examples.

[0042] The amount of water-soluble dietary fiber in branched dextran is determined based on the methods described in "Analytical Methods for Nutritional Components, etc." (methods listed in column 3 of Appendix 1 of the Nutritional Labeling Standards), "8. Dietary Fiber," and "(2) High-Performance Liquid Chromatography (Enzyme-HPLC Method)" in the "Nutritional Labeling Standards" (Ministry of Health and Welfare Notification No. 146 of May 1996), as shown in the examples.

[0043] (4) Other requirements From the viewpoint of further enhancing functionality, the branched dextran of the present invention may or may not satisfy any or all of the following requirements.

[0044] The lower limit of the water retention rate of the branched dextran of the present invention is preferably 5% by mass or more, and more preferably 10% by mass or more. While a higher water retention rate is preferable for the branched dextran of the present invention, its upper limit may be, for example, 100% by mass or less, or 50% by mass or less.

[0045] The water retention rate of the branched dextran of the present invention is determined based on the sample weight before and after heating under the conditions shown in the examples.

[0046] The lower limit of the intrinsic viscosity (η) of the branched dextran of the present invention is preferably 10 or higher, more preferably 15 or higher. The upper limit of the intrinsic viscosity (η) of the branched dextran of the present invention is preferably 140 or less, more preferably 110 or less.

[0047] The intrinsic viscosity of the branched dextran of the present invention is determined based on the specific viscosity (ηsp) and reduced viscosity (ηred) specified under the conditions shown in the examples.

[0048] The lower limit of the viscosity-average molecular weight (Mv) of the branched dextran of the present invention is preferably 20,000 or more, and more preferably 30,000 or more. The upper limit of the viscosity-average molecular weight (Mv) of the branched dextran of the present invention is preferably 650,000 or less, and more preferably 500,000 or less.

[0049] The viscosity-average molecular weight (Mv) of the branched dextran of the present invention is determined based on the intrinsic viscosity (η) specified under the conditions shown in the examples.

[0050] <Applications of branched dextran> The branched dextran of the present invention has a high water-soluble dietary fiber content and excellent functionality. For example, the branched dextran of the present invention can be used for the following purposes.

[0051] When using the branched dextran of the present invention for various applications, the form of the branched dextran may be isolated branched dextran, or it may be lactic acid bacteria cells or a filtration product of a culture, etc.

[0052] When using the branched dextran of the present invention for various applications, it should be appropriately incorporated into the target substance (pharmaceutical, food, etc.). The method and conditions of incorporation can be those conventionally known, depending on the type of target substance. The branched dextran of the present invention may be incorporated at any stage of the manufacturing process of the target product. For example, if the branched dextran is contained in the cells of lactic acid bacteria, the fermentation process may be carried out after the branched dextran has been incorporated.

[0053] When the branched dextran of the present invention is in the form of lactic acid bacteria cells, it can be used as a starter for lactic acid fermentation. Specifically, a lactic acid fermented product can be prepared by adding the microbial cells to a raw material and allowing fermentation. Through this operation, the branched dextran of the present invention can be incorporated into the desired raw material. The fermentation conditions are not particularly limited, as long as they allow lactic acid bacteria to grow and metabolites (such as branched dextran) to be produced. For example, the fermentation temperature and time can be appropriately set depending on the type of raw material (milk, vegetables, etc.).

[0054] (Intestinal regulator) The branched dextran of the present invention can be prepared as an intestinal regulator. According to the intestinal regulator containing branched dextran of the present invention, for example, short-chain fatty acids (such as propionic acid) can be specifically produced in the large intestine, thereby suppressing the growth of harmful bacteria. As a result, this intestinal regulator can improve the condition of the stomach and intestines and promote intestinal peristalsis, thereby promoting bowel movements and improving constipation.

[0055] The amount of branched dextran in the intestinal regulator of the present invention can be appropriately adjusted depending on the desired effect and the condition of the target patient. The lower limit of the content of branched dextran in the intestinal regulator of the present invention is preferably 1% by mass or more, and more preferably 10% by mass or more, relative to the intestinal regulator. The upper limit of the content of branched dextran of the present invention in the intestinal regulator of the present invention is preferably 100% by mass or less, more preferably 70% by mass or less, relative to the intestinal regulator.

[0056] (Cholesterol-lowering drugs) The branched dextran of the present invention can be prepared as a cholesterol-lowering agent. The cholesterol-lowering agent containing branched dextran of the present invention can, for example, specifically reduce bad cholesterol (non-HDL cholesterol) and lower the total cholesterol concentration in the blood, thereby improving lipid metabolism disorders such as dyslipidemia and arteriosclerosis.

[0057] The cholesterol-lowering agent of the present invention can be used in any form for the purpose of reducing cholesterol, and can function as, for example, a lipid absorption inhibitor, a cholesterol excretion promoter, a body fat reducer, a fat accumulation inhibitor, and the like.

[0058] The amount of branched dextran in the cholesterol-lowering agent of the present invention can be appropriately adjusted depending on the desired effect and the condition of the target patient. The lower limit of the content of branched dextran in the cholesterol-lowering agent of the present invention is preferably 1% by mass or more, more preferably 10% by mass or more, relative to the cholesterol-lowering agent. The upper limit of the content of branched dextran of the present invention in the cholesterol-lowering agent of the present invention is preferably 100% by mass or less, more preferably 70% by mass or less, relative to the cholesterol-lowering agent.

[0059] (Blood sugar level suppressant) The branched dextran of the present invention can be prepared as a blood glucose level-increasing agent. According to the present invention, a blood glucose elevation inhibitor containing branched dextran can suppress the degree to which blood glucose levels rise by, for example, taking it before, between, or after meals.

[0060] The amount of branched dextran in the blood glucose-lowering agent of the present invention can be appropriately adjusted depending on the desired effect and the condition of the target patient. The lower limit of the content of branched dextran of the present invention in the blood glucose elevation inhibitor of the present invention is preferably 1% by mass or more, more preferably 10% by mass or more, relative to the blood glucose elevation inhibitor. The upper limit of the content of branched dextran of the present invention in the blood glucose elevation inhibitor of the present invention is preferably 100% by mass or less, more preferably 70% by mass or less, relative to the blood glucose elevation inhibitor.

[0061] (Starch retrogradation inhibitor) The branched dextran of the present invention can be prepared as a starch retrogradation inhibitor. The starch retrogradation inhibitor containing branched dextran of the present invention can suppress the retrogradation of foods mainly composed of starch (such as bread and rice), making it easier to obtain foods with good flavor and texture. Therefore, the branched dextran of the present invention also functions as a texture improver (such as a texture improver for bread).

[0062] The branched dextran in the starch retrogradation inhibitor and texture improver of the present invention may be isolated branched dextran, or it may be lactic acid bacteria cells or a filtration product of a culture.

[0063] The starch retrogradation inhibitor and texture improver of the present invention may be added to the compound, or the branched dextran of the present invention may be produced by culturing or other means and then incorporated into the compound. In the latter embodiment, the starch retrogradation inhibitor and texture improver of the present invention can be incorporated into the formulation by using lactic acid bacteria cells or the filtration product of the culture as a starter. For example, when manufacturing bread, an equal amount of water is mixed with strong flour, and live lactic acid bacteria are added to the resulting mixture as a starter. By fermenting this mixture, a starter (such as a sourdough starter) containing the starch retrogradation inhibitor and the texture improver of the present invention can be obtained.

[0064] The amount of branched dextran of the present invention in the starch retrogradation inhibitor of the present invention can be appropriately adjusted depending on the effect to be obtained and the type of material to be incorporated (bread, rice, etc.). The lower limit of the content of branched dextran of the present invention in the starch retrogradation inhibitor of the present invention is preferably 0.1% by mass or more, more preferably 1% by mass or more, relative to the starch retrogradation inhibitor. The upper limit of the content of branched dextran of the present invention in the starch retrogradation inhibitor of the present invention is preferably 100% by mass or less, more preferably 50% by mass or less, relative to the starch retrogradation inhibitor.

[0065] (Other uses) The branched dextran of the present invention can be used for any other application known to be an application of dextran. For example, the branched dextran of the present invention can be incorporated into food and beverages (including health-oriented food and beverages, such as nutritional supplements, health supplements, nutritional adjustment foods, health functional foods, foods for specified health uses, nutrient function foods, and foods with functional claims), cosmetic compositions, pharmaceutical compositions (pharmaceuticals, quasi-drugs, etc.).

[0066] Examples of food and beverages include human food, infant food (infant formula, weaning food, baby food), feed for non-human animals or aquatic life, pet food, pet treats, etc. Specifically, these include: Fermented foods (lactic acid bacteria beverages, fermented milk, etc.), and their starters, Beverages (Japanese tea, oolong tea, black tea, coffee, juice, processed milk, sports drinks, etc.) Bakery items (bread, pizza, pies, etc.), Western-style confectionery (cookies, crackers, biscuits, cakes, sponge cake, etc.), Noodles (udon, soba, ramen, etc.) Pasta dishes (spaghetti, macaroni, etc.), Snack foods (rice crackers, potato chips, snacks, etc.) Confectionery (hard candy, soft candy, caramel, gum, chocolate, etc.), Frozen desserts (ice cream, sherbet, etc.) Dairy products (cream, cheese, mousse, powdered milk, condensed milk, milk beverages, etc.), Western-style fresh confectionery (jelly, pudding, mousse, yogurt, buttercream, custard cream, etc.), Japanese sweets (gyuhi, uiro, mochi, ohagi, dorayaki, etc.) Processed fruit and vegetable products (jams, marmalades, syrups, candies, etc.), Pastes (flour paste, fruit paste, peanut paste, etc.), Pickles (pickled shallots, pickled vegetables, kimchi, etc.) Compound seasonings (soy sauce, sauce, dip, noodle soup base, dashi stock, soup base, etc.), Condiments (stew mix, soup mix, curry mix, mayonnaise, ketchup, etc.), Retort foods or canned foods (curry, stew, soup, etc.), Frozen or refrigerated foods (ham, sausage, bacon, hamburgers, meatballs, croquettes, dumplings, pilaf, rice balls, etc.), Processed seafood products (chikuwa, kamaboko, etc.), Rice dishes (risotto, sushi, etc.).

[0067] Examples of cosmetic compositions include cleansing agents (soap, body shampoo, cleansing cream, facial cleanser, etc.), cosmetics (lotion, emulsion, serum, etc.), creams (facial foam, vanishing cream, cold cream, emollient cream, massage cream, etc.), masks, bath additives, and the like.

[0068] Examples of pharmaceutical compositions include oral preparations, intravascular preparations, enteral preparations, transdermal preparations, and intraperitoneal preparations. Dosage forms of pharmaceutical compositions include powders, granules, tablets, capsules, pills, suppositories, liquids, and injections. Furthermore, the pharmaceutical composition may contain additives (excipients, bases, emulsifiers, solvents, stabilizers, etc.) depending on the effect to be achieved.

[0069] (Use of branched dextran as a raw material) As described above, the branched dextran of the present invention can be incorporated into various pharmaceutical preparations and food and beverage products. Therefore, the present invention also includes raw materials used for food and beverages, cosmetics, pharmaceuticals, quasi-drugs, or industrial purposes, which include the branched dextran of the present invention.

[0070] <Ingredients for branched dextran> The branched dextran of the present invention is readily obtainable from cultures of Leuconostoc citreum KD3 strain (accession number NITE P-03378). Therefore, the present invention also includes a raw material consisting of Leuconostoc citreum KD3 strain (accession number NITE P-03378) for producing the branched dextran of the present invention.

[0071] <Method for producing branched dextran> The branched dextran of the present invention is obtained from lactic acid bacteria belonging to Leuconostoc citreum.

[0072] The method for preparing branched dextran from lactic acid bacteria is not particularly limited, but typically, the branched dextran of the present invention is obtained by culturing lactic acid bacteria and recovering the culture. The resulting culture may be purified as needed, or it may not be purified.

[0073] (Method for culturing lactic acid bacteria) The method for culturing lactic acid bacteria belonging to Leuconostoc citreum is not particularly limited, but any conditions used for culturing lactic acid bacteria can be adopted.

[0074] The lactic acid bacteria used for culture may be live bacteria, resting bacteria, or both.

[0075] As the lactic acid bacteria used for cultivation, Leuconostoc citreum KD3 strain (accession number NITE P-03378) is preferred from the viewpoint of easily obtaining the branched dextran of the present invention.

[0076] As a carbon source in the culture medium, for example, sugars (galactose, glucose, fructose, mannose, sorbose, mannitol maltose, sucrose, trehalose, starch hydrolysates, molasses, etc.) can be used depending on their assimilation properties.

[0077] As nitrogen sources in the culture medium, for example, ammonia, ammonium salts (ammonium sulfate, ammonium chloride, ammonium nitrate, etc.), and nitrates can be used.

[0078] Examples of inorganic salts that can be used in the culture medium include sodium chloride, potassium chloride, potassium phosphate, magnesium sulfate, calcium chloride, calcium nitrate, manganese chloride, ferrous sulfate, and organic components (peptone, sake lees, whey, soybean flour, defatted soybean meal, meat extract, yeast extract, etc.).

[0079] As the culture medium, pre-prepared media (such as MRS medium and its modified media) can be used.

[0080] The culture temperature can be any temperature within the range in which lactic acid bacteria can grow, and is usually suitable in the range of 20 to 40°C.

[0081] The pH during cultivation is preferably 5.0 to 8.0.

[0082] The incubation time is preferably about 10 to 30 hours.

[0083] (Purification of cultures) When purifying a culture, it is preferable to remove all or part of the bacterial cells, lactic acid bacteria fermented products, and culture medium components from the culture by membrane filtration. The resulting filtrate is collected as a fraction containing branched dextran.

[0084] The purification method by membrane filtration is not particularly limited, but examples include ultrafiltration, microfiltration, coarse filtration, and reverse osmosis. When performing membrane filtration, it is preferable to circulate the material to be filtered at a temperature of 20 to 50°C.

[0085] The types of membrane filtration are not particularly limited, but include porous filtration membranes and hollow fiber membranes. Of these, it is preferable to use at least a porous filtration membrane, from the viewpoint of easily obtaining the branched dextran of the present invention.

[0086] Purification by membrane filtration may be performed using a single method or a combination of two or more methods. Furthermore, the same method may be performed multiple times. For example, ultrafiltration using a hollow fiber membrane may be performed before or after ultrafiltration using a porous filtration membrane.

[0087] The components of the porous filtration membrane are not particularly limited as long as they can be used for ultrafiltration, etc., but from the viewpoint of easily obtaining the branched dextran of the present invention, it is preferable that it contains one or more selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide. Such a porous filtration membrane may also be called a "ceramic membrane". When a ceramic membrane is used as a porous filtration membrane, pressure is easily applied to the material being filtered, and shear force is applied during filtration. As a result of this shear force being applied to the branched dextran, the molecular weight distribution of the branched dextran becomes broader, making it easier to obtain branched dextran that satisfies the requirements of the present invention. In the following, "one or more selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide" will also be referred to as "ceramic components."

[0088] The porous filtration membrane may contain, preferably, 90% by mass or more, and more preferably 95% by mass or more, of ceramic components relative to the entire porous filtration membrane. If the porous filtration membrane contains two or more ceramic components, the total amount of these components must be within the above range.

[0089] The structure of the porous filtration membrane is not particularly limited and can be any membrane that functions as a filtration membrane. For example, the porous filtration membrane may be single-layered or multi-layered. If the porous filtration membrane is multi-layered, it may include, for example, a filtration membrane layer and a support layer.

[0090] From the viewpoint of easily obtaining the branched dextran of the present invention, it is preferable that the pore size of each pore in the porous filtration membrane is in the range of 0.01 μm to 1.00 μm. The lower limit of the pore size of the porous filtration membrane is more preferably 0.02 μm or larger, and even more preferably 0.05 μm or larger. The upper limit of the pore size of the porous filtration membrane is more preferably 0.9 μm or less, and even more preferably 0.8 μm or less.

[0091] The components of the hollow fiber membrane are not particularly limited as long as they can be used for ultrafiltration, etc., but from the viewpoint of easily obtaining the branched dextran of the present invention, those containing polyacrylonitrile and / or polysulfone are preferred.

[0092] From the viewpoint of easily obtaining the branched dextran of the present invention, the molecular weight of the fractionated hollow fiber membrane is preferably in the range of 1,000 to 100,000. The lower limit of the fractional molecular weight of the hollow fiber membrane is more preferably 2,000 or more, and even more preferably 3,000 or more. The upper limit of the molecular weight cutoff of the hollow fiber membrane is more preferably 90,000 or less, and even more preferably 80,000 or less.

[0093] (Other processing) The cultured product and its purified form may be subjected to various treatments as needed. Such treatments include concentration (ultrafiltration, etc.), separation (precipitation separation with organic solvents, gel filtration chromatography, centrifugation, etc.), hydrothermal treatment, ultrasonic disruption, enzyme (dextran-degrading enzyme, etc.) addition, removal of impurities (using activated carbon, ion exchange resin, synthetic adsorbents, etc.), and drying (freeze-drying, spray drying, drum drying, etc.). The methods and conditions for each of the above processes can be set as appropriate depending on the desired effect.

[0094] (Preferred method for producing branched dextran) A preferred method for producing branched dextran according to the present invention is: A culture process for culturing lactic acid bacteria belonging to Leuconostoc citreum, The process includes a filtration step in which the culture is filtered after the culture step to recover a filtrate containing branched dextran, In the filtration process, a porous filtration membrane is used that contains a ceramic component (one or more selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide) and has a pore size of 0.01 μm or more and 1.00 μm or less. The conditions for the culture and filtration processes can be appropriately adopted from the conditions described above.

[0095] In the above manufacturing method, the lactic acid bacteria belonging to Leuconostoc citreum is preferably Leuconostoc citreum KD3 strain (accession number NITE P-03378). [Examples]

[0096] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples.

[0097] <Example Test 1: Isolation and Evaluation of Leuconostoc citreum> Leuconostoc citreum was isolated and its properties evaluated using the following method. This experiment was conducted in accordance with "Classification and Identification of Microorganisms" (edited by Takeji Hasegawa, Academic Publishing Center, 1985).

[0098] (1) Isolation of bacterial strains Various vegetables and pickles were suspended in 0.85% sterile saline solution. The resulting suspension was spread onto de Man, Rogosa, and Sharpe (MRS) agar medium ("Difco® Lactobacillus MRS Broth," manufactured by Becton Dickinson) and cultured anaerobically at 30°C for 2-3 days using "Anelopack Kenki" (manufactured by Mitsubishi Gas Chemical Company). After culturing, the grown microbial colonies were randomly picked. The collected microbial colonies were streaked onto agar plates ("BCP-Added Plate Count-Agar," manufactured by Nissui Pharmaceutical Co., Ltd.), and the presence or absence of acid production was confirmed based on the yellowing around the formed colonies. Strains that showed acid production were used as candidate lactic acid bacteria strains in the following tests.

[0099] (2) Identification of the bacterial strain For each candidate strain of lactic acid bacteria, a partial base sequence of the 16S ribosomal RNA gene on the genome (approximately 600 bp including the V1-V3 hypervariable region) was deciphered. The determination of whether each partial base sequence was a lactic acid bacterium was made by searching for it using the "Basic Local Alignment Search Tool (BLAST)" (URL: https: / / blast.ncbi.nlm.nih.gov / Blast.cgi). As a result, 92 strains were identified as lactic acid bacteria. It was inferred that the majority of these lactic acid bacteria belonged to the genus Leuconostoc. Of the obtained strains, Leuconostoc citreum KD3 strain (accession number NITE P-03378) was selected and subjected to the following tests.

[0100] (3) Mycological properties of lactic acid bacteria The following mycological characteristics were confirmed to be present in the Leuconostoc citreum KD3 strain. The colonies on de Man, Rogosa, and Sharpe (MRS) agar medium ("Difco® Lactobacillus MRS Broth," manufactured by Becton Dickinson) were smooth, circular, white colonies. The colonies on MRS agar medium containing 5% sucrose exhibited a viscous shape. The cell shape was typical of lactic acid bacteria in the genus Leuconostoc (a slightly oval-shaped coccus). Other characteristics observed included absolute heterolactic fermentation (with significant gas production), Gram staining positivity, catalase test negation, and non-spore formation.

[0101] (4) The sugar utilization ability of lactic acid bacteria We confirmed the ability of Leuconostoc citreum KD3 strain to utilize sugars (49 types). Leuconostoc citreum KD3 strain was cultured statically in 10 ml of MRS liquid medium until the stationary phase of growth occurred. The culture was centrifuged, and the resulting cells were washed with an appropriate amount of sterile water. The cells were then collected by centrifugation again. The collected bacterial cells were suspended in 10 ml of sterile water, and the resulting suspension (1 ml) was added to "API 50CHL medium" (manufactured by bioMérieux). Each suspension was dispensed in an appropriate amount into each well of the "API 50CH Kit" (manufactured by bioMérieux). The bottom of each well was coated with 49 different sugars. Next, after culturing at 33°C for 72 hours, the presence or absence of assimilation was determined by observing the change in color in each well. The results are shown in Table 1. In the table, the "Positive Rate" column indicates the probability that bacteria belonging to Leuconostoc citreum possess the ability to assimilate the target sugar after being cultured at 36°C ± 2°C for 48 ± 6 hours. In the "Assimilation Ability" section, "+" indicates that the Leuconostoc citreum KD3 strain can assimilate the target sugar, and "-" indicates that it cannot assimilate the target sugar.

[0102] [Table 1]

[0103] <Test Example 2: Preparation of Branched Dextran> Using the following method, five branched dextrans were prepared using Leuconostoc citreum KD3 strain.

[0104] (1) Preparation of "Branched Dextran-1" A frozen stock of Leuconostoc citreum strain KD3 was added at 1% to the seed culture medium and incubated at 33°C for at least 9 hours. Then, the culture medium was added at 1% to the pre-culture medium (same composition as the seed culture medium) and cultured further under the same conditions as the seed culture. The obtained culture medium was inoculated into branched dextran production medium (2 L) and then cultured with stirring at 30°C and a speed of 150 rpm for 24 hours. The pH during culture was controlled to be 6.5 or higher using 24% caustic soda. The culture medium was heated at 90°C for 1 hour and then cooled to below 25°C. After cooling, the culture medium was ultrafiltered using an ultrafiltration apparatus equipped with a ceramic membrane (0.05 μm UF). The filtrate was washed with water until the Brix of the filtration-side discharged liquid was 0.1% or less, and then concentrated using a ceramic membrane (0.05 μm UF). The concentrate was heat-treated at 90°C for 1 hour, frozen in a freezer, and then freeze-dried in a freeze-dryer. The dried material was pulverized using a 60-mesh pass to obtain "branched dextran-1" as a powder. The yield was approximately 12 g per liter of culture medium.

[0105] The ceramic membrane used in the limit filtration in this example corresponds to a porous filtration membrane and contains one or more elements selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide, with a pore size of approximately 0.05 μm.

[0106] (2) Preparation of "Branched Dextran-2" Except for performing microfiltration of the culture medium before ultrafiltration, the procedure was the same as in "(1) Preparation of Branched Dextran-1" to obtain Branched Dextran-2 as a powder. The yield was approximately 12 g per liter of culture medium. Microfiltration was performed by adding water (at least three times the volume of the culture medium) to the culture medium and using a ceramic membrane (0.8 μm MF). The resulting filtrate was subjected to ultrafiltration.

[0107] The ceramic membrane used in the microfiltration in this example corresponds to a porous filtration membrane and contains one or more elements selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide, with a pore size of approximately 0.8 μm.

[0108] (3) Preparation of "Branched Dextran-3" Except for adding a dextran-degrading enzyme (dextranase) derived from the genus Chaetomium erraticum when inoculating the culture medium into a branched dextran production medium (2 L), and performing ultrafiltration and subsequent concentration using a hollow fiber membrane, the same procedure as in "(2) Preparation of Branched Dextran-2" was performed to obtain "Branched Dextran-3" as a powder. The yield was approximately 7 g per 1 L of culture medium. As the hollow fiber membrane, "Microza UF" (Asahi Kasei Corporation) with a fractional molecular weight cutoff of 3,000 was used. After ultrafiltration, the effluent was washed with water until the Brix was 0.1% or less, and the branched dextran was concentrated using "Microza UF".

[0109] (4) Preparation of "Branched Dextran-4" Except for performing sonication and microfiltration on the culture medium before ultrafiltration, the procedure was the same as in "(1) Preparation of Branched Dextran-1" to obtain Branched Dextran-4 as a powder. The yield was approximately 10 g per liter of culture medium. Ultrasonic treatment was performed using an ultrasonic shredder, and after treatment, the bacterial cells were removed by centrifugation, and the supernatant was subjected to microfiltration. Microfiltration was performed by adding water (at least three times the volume of the culture medium) to the culture medium and using a ceramic membrane (0.8 μm MF). The resulting filtrate was subjected to ultrafiltration.

[0110] The ceramic membrane used in the microfiltration in this example corresponds to a porous filtration membrane and contains one or more elements selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide, with a pore size of approximately 0.8 μm.

[0111] (5) Preparation of "Branched Dextran-5" Except for performing centrifugation instead of ultrafiltration and freeze-drying the resulting supernatant to obtain the powder, the procedure was the same as in "(1) Preparation of Branched Dextran-1" to obtain Branched Dextran-5 as a powder. The yield was approximately 150g per liter of culture medium.

[0112] Table 2 shows the filtration methods used in the preparation of each branched dextran.

[0113] [Table 2]

[0114] <Example 3: Analysis of branched dextran> The branched dextrans obtained in Experiment 2 were analyzed using the following method.

[0115] (1) Measurement of weight-average molecular weight (Mw) and number-average molecular weight (Mn) Each branched dextran was dissolved in deionized water, then filtered through a 0.2 μm filter, and the filtrate was prepared as the sample solution. Next, gel permeation chromatography (GPC) was performed under the following conditions. Using commercially available software, a calibration curve was created from molecular weight standard samples, and the weight-average molecular weight (Mw) and number-average molecular weight (Mn) of branched dextran in each sample solution were measured. In addition, the weight-average molecular weight divided by the number-average molecular weight of the branched dextran (Mw / Mn) was also calculated. The results are shown in Table 3.

[0116] [GPC measurement conditions] GPC device: HLC-8220 GPC This column: Three TSKgel GMPW tubes in series. Guard column: PWH Eluent: 200 mM sodium nitrate aqueous solution Flow rate: 1.0mL / min Detector: RI detector Column temperature: 40℃ Sample concentration: 0.5%~2.0% Injection volume: 100μL Analysis time: 40 minutes Molecular weight standard samples: glucose, maltose, maltopentaose, maltoheptaose, standard pullulan (P-5, P-10, P-20, P-50, P-100, P-200, P-400, P-800 (Shodex))

[0117] (2) Measurement of water-soluble dietary fiber content Based on the methods described in "Analytical Methods for Nutritional Components, etc." (the method listed in column 3 of Appendix 1 of the Nutritional Labeling Standards), "8. Dietary Fiber," and "(2) High-Performance Liquid Chromatography (Enzyme-HPLC Method)" in the "Nutritional Labeling Standards" (Ministry of Health and Welfare Notification No. 146 of May 1996), the amount of water-soluble dietary fiber in each branched dextran was measured using the following method. The results are shown in Table 3.

[0118] (2-1) Method for preparing analytical samples Each branched dextran (1g) was weighed out, and 40ml of 50mM MES-TRIS buffer (pH 6.3, 24℃) was added and dispersed by ultrasonic fracturing. For ultrasonic fracturing, a "Sonifier 450" (Branson, output memory 4, 10min, 1 / 8” tapered microchip) was used. The obtained solution (4 ml) was dispensed into a test tube, and 0.02 ml of the heat-stable α-amylase reagent included with the Dietary Fiber Assay Kit (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) was added. The test tube was then covered with aluminum foil. The mixture was then reacted in a boiling water bath for 30 minutes, stirring every 5 minutes, and then cooled. To the resulting reaction solution, water (1 ml) was added while rinsing the walls of the test tube. After cooling to 60°C, the protease solution (0.02 ml) and amyloglucosidase solution (0.02 ml) included in the kit were added simultaneously, and the mixture was covered with aluminum foil. The mixture was then allowed to react for 30 minutes while shaking in a 60°C water bath, and then cooled. The resulting reaction solution (approximately 5 ml) was desalted by passing it through an ion exchange resin at an SV of 1.0 or less. As the ion exchange resin, a mixture of "DOWEXTM22 (Cl type)" (manufactured by Dow Chemical) and "DOWEXTM88 (Na type)" (manufactured by Dow Chemical) in a 3:2 ratio was used. Furthermore, the solution was eluted with approximately three times the volume of deionized water, and the total volume of the eluate was adjusted to approximately 20 ml. The obtained eluate was diluted to 25 ml in a volumetric flask, filtered through a 0.22 μm pore size membrane filter, and the resulting filtrate was collected as the analytical sample.

[0119] (2-2) Calculation of water-soluble dietary fiber content In each analytical sample, undigested branched dextran that remained undegraded even after enzymatic treatment was considered water-soluble dietary fiber. Based on these premises, the amount of glucose (mg) in the analytical sample was quantified using the conventional glucose oxidase method, and the amount of water-soluble dietary fiber was calculated based on Equations 1 and 2. [Formula 1] Water-soluble dietary fiber content (mg) = Solid content in the analytical sample (mg) - Glucose content in the analytical sample (mg) [Formula 2] Water-soluble dietary fiber content (mass%) = Total water-soluble dietary fiber content in the analytical sample (mg) / Total solid content in the analytical sample (mg) × 100

[0120] (3) Glycan structure analysis The structure of each branched dextran was analyzed under the following conditions, and the composition of glycosidic bonds was measured. The results are shown in Table 4.

[0121] (3-1) 1 H-NMR measurement conditions 1 H-NMR equipment: ECX-400P, JEOL Tokyo, Japan

[0122] (3-2) Structural analysis method After dissolving each branched dextran in heavy water (D2O), 1 Using an H-NMR spectrometer, 1The 1H-NMR spectrum was measured. Furthermore, the composition and ratio of glycosidic bonds in each branched dextran were determined from the following spectrum, which is characteristic of α-glycosidic bonds appearing at 4.8-5.2 ppm. 4.85 ppm; α-1,6 4.98 ppm; α-1,2 5.07 ppm; α-1,6 / 2,1 5.20 ppm; α-1,3

[0123] [Table 3]

[0124] [Table 4]

[0125] As shown in Tables 3 and 4, when the glycosidic bond composition and "Mw / Mn" met the requirements of the present invention, each branched dextran had a high water-soluble dietary fiber content of 90% or more.

[0126] <Test Example 4: Calculation of Water Retention Rate of Branched Dextran> The water retention rate of each branched dextran obtained above was calculated based on the following method. The results are shown in Table 5.

[0127] For each branched dextran, the following values ​​were measured: "W0" (weight in grams) after heating and drying at 95°C for 5 hours; "W1" (weight in grams) after leaving it at 35°C for 24 hours under 100% relative humidity (RH); and "W2" (weight in grams) after leaving it at 35°C for 24 hours under 43% relative humidity (RH). Next, based on the measurement results of each weight (g), the water retention rate was calculated using Equation 3. [Formula 3] Water retention rate (%)=((W2-W1) / (W1-W0))×100

[0128] <Test Example 5: Measurement of Intrinsic Viscosity and Viscosity-Average Molecular Weight of Branched Dextran> The intrinsic viscosity (η) and viscosity-average molecular weight (Mv) of each branched dextran obtained above were calculated based on the following method. The results are shown in Table 5.

[0129] Each branched dextran was prepared as an aqueous solution at 0, 0.1, 0.2, 0.3, 0.4, and 0.5 (w / v)% concentrations at 30°C. Next, an Ubbelohde viscometer was placed in a water tank filled with 30°C water, and the flow time (s) of each aqueous solution was measured. The specific viscosity was calculated using Equation 4, based on the flow time "t" of each sample and the flow time "t0" of the pure solvent (water). [Formula 4] Specific viscosity (ηsp)=(t-t0) / t0 Furthermore, the specific viscosity (ηsp) was divided by the mass concentration (c) to calculate the reduced viscosity (ηred). The relationship between the solution concentration and the reduced viscosity (ηred) was then plotted, and the y-intercept value (extrapolated value) of the approximate straight line was identified as the intrinsic viscosity (η).

[0130] Furthermore, regarding the intrinsic viscosity (η), equation 5 holds true (where "K" (Flory-Fox constant) and "α" are constants that depend on the combination of polymer and solvent and the temperature). [Formula 5] Intrinsic viscosity (η) = KMα (Mark-Kuhn-Houwink formula) In this study, the values ​​for amylose (K=0.0132, α=0.68) were used to calculate the viscosity-average molecular weight (Mv) equivalent to amylose.

[0131] [Table 5]

[0132] <Test Example 6: Evaluation of the intestinal regulating effect of branched dextran intake> The bowel-regulating effects of each branched dextran obtained above were evaluated based on the following method. The results are shown in Table 6.

[0133] Healthy men and women aged 20 to 40 who tended to be constipated were randomly divided into two groups (10 people each). In this study, a tendency towards constipation was defined as having a bowel movement six days or less per week. After a two-week pre-observation period, subjects in each group were given either branched dextran-2 or branched dextran-5, ingesting 6g per day for two weeks (n=10 in each group). Participants were asked to self-report their condition at each point in time, before intake and two weeks after intake, using a questionnaire (number of days with bowel movements, amount of stool, and post-test comments (free-response)). The amount of stool was recorded as the number of ping-pong balls.

[0134] [Table 6]

[0135] As shown in Table 6, ingestion of branched dextran (branched dextran-2) that satisfies the requirements of the present invention increased the number of days with bowel movements and showed an increasing trend in stool volume. In post-ingestion feedback, many users reported an improvement in their constipation tendencies when taking "branched dextran-2" compared to when taking "branched dextran-5".

[0136] <Test Example 7: Evaluation of blood glucose fluctuations due to branched dextran intake> The effect of branched dextran (branched dextran-2) obtained above on blood glucose fluctuations was evaluated based on the following method. The results are shown in Table 7.

[0137] As a sample, a 1.25 g / 50 ml aqueous solution of glucose or branched dextran was prepared. Next, each sample was administered as a single dose of 0.5 g per kg of body weight to mice (C57BL / 6JJcl (male, 10 weeks old)) (n=6 in each group). Blood glucose levels were measured directly from the tail vein using "StatStrip Express 900" (Siemens Healthineers) at each time point after administration (0, 30, 60, 90, or 120 minutes post-administration). For statistical analysis of blood glucose levels, a paired two-way ANOVA was performed, followed by a simple main effects test (* p < 0.001). The Mann-Whitney U test was used for AUC (Area Under the Curve) values ​​from 0 to 120 minutes (* p < 0.01).

[0138] [Table 7]

[0139] As shown in Table 7, ingestion of branched dextran (branched dextran-2) that satisfies the requirements of the present invention suppressed the rise in blood glucose levels 30 minutes or more after administration compared to the glucose administration group. In particular, the AUC value 120 minutes after administration was significantly lower in the branched-dextran group, at approximately 40% of the glucose group's value of approximately 100 mg·h / dL.

[0140] <Test Example 8: Functional Evaluation of Branched Dextran Intake> The functionality of each branched dextran obtained above was evaluated based on the following method. The results are shown in Table 8.

[0141] As samples, we prepared a diet (D12451) with 2.5% (w / w) of each branched dextran obtained above added. As a control sample, we prepared a diet with 2.5% (w / w) of corn starch added. Next, each sample was administered to mice (C57BL / 6J (male, 6 weeks old)) for 10 weeks (including a 1-week pre-feeding period) (n=6 in each group, except for the control group which had n=7).

[0142] The following data was collected during the intake period. (feces analysis) Fecal weight: Daily fecal weight (mg) IgA amount: Daily amount of IgA in feces (μg / day) (cecal content analysis) Contents: Weight of cecal contents (mg) Propionic acid content: Amount of propionic acid per mouse (μmol / mouse) Propionic acid concentration: Propionic acid concentration per 1g of cecum (μmol / g) (Blood analysis) TC: Total cholesterol (mg / dL) non-HDL-C: non-HDL cholesterol concentration (mg / mL)

[0143] All data are expressed as mean ± standard error, and the presence or absence of a statistically significant difference was determined using the Tukey-Kramer test with a significance level of 0.05 (* p<0.05).

[0144] [Table 8]

[0145] As shown in the fecal analysis results, the intake of branched dextran that satisfies the requirements of the present invention significantly increased the amount of fecal IgA per day compared to the control group.

[0146] As shown in the cecal contents analysis, ingestion of branched dextran that satisfies the requirements of the present invention significantly increased the amount of propionic acid per animal and the amount of propionic acid per gram of cecum compared to the control group.

[0147] As shown in the blood analysis results, ingestion of branched dextran that satisfies the requirements of the present invention resulted in a decreasing trend in total cholesterol concentration and non-HDL cholesterol concentration compared to the control group. In particular, the intake of branched dextran-2 significantly reduced non-HDL cholesterol levels compared to the control group.

[0148] <Test Example 9: Effect of branched dextran formulation on improving bread quality> The quality improvement effect of incorporating the branched dextran (branched dextran-2) obtained above into bread was evaluated based on the following method. The results are shown in Table 9.

[0149] The ingredients listed in the "Composition" section of Table 9 were mixed, and bread (white bread) was produced using a household bread maker ("Automatic Home Bakery SD-BT-103," manufactured by National) by sequentially performing the following steps: kneading, adding dry yeast, resting, kneading, fermentation, baking, and cooling. In the table, the values ​​shown in the "Added Ingredients" column represent the percentage (weight %) of branched dextran or commercially available dextran relative to the weight (g) of "strong flour". For the "commercially available dextran," we used "Dextran from Leuconostoc mesenteroides" (manufactured by Sigma-Aldrich), which has an average molecular weight of 1,500,000 to 2,800,000. For each loaf of bread, on the day of preparation and after one day of storage at room temperature, it was sliced ​​into 20mm thick slices (5 slices), and the central portion of the three inner slices was cut out to obtain a 40mm x 40mm crumb section. Each of the obtained crumb portions was subjected to the following evaluations.

[0150] (Measuring the hardness of the crumb) Using a creep meter (RE2-3305, manufactured by Yamaden Co., Ltd.) equipped with a cylindrical plunger (30 mm in diameter), uniaxial compression was performed at a compression speed of 1 mm / sec, with the deformation limit set to 60% of the thickness of the cramb. The stress (gf) at a strain rate of 30% was determined as the hardness of the cramb.

[0151] (Measurement of change in crumb hardness) Based on the average value obtained by dividing the hardness of the crumbs on the day of preparation and after one day of storage at room temperature by the standard error, the change in hardness due to storage was calculated using Equation 6. [Formula 6] Change (Δgf) = Hardness of crumb after 1 day of storage at room temperature (gf) - Hardness of crumb on the day of preparation (gf)

[0152] (Evaluation of bread softness) Based on the crumb hardness value on the day the bread was made, the softness of the bread was evaluated according to the following evaluation criteria. [Criteria for evaluating bread softness] 70gf or more: × (0 points) 60gf or more but less than 70gf: △ (1 point) 50gf or more but less than 60gf: ○ (3 points) Less than 50gf: ◎ (5 points)

[0153] (Evaluation of bread rise) The volume of the bread on the day of preparation was measured using the rapeseed substitution method, and the rise of the bread was evaluated according to the following evaluation criteria. [Criteria for evaluating bread rise] Less than 1.50L: × (0 points) 1.50L or more: △ (1 point) 1.60L or more: ○ (3 points) 1.70L or more: ◎ (5 points)

[0154] (Evaluation of the effect of inhibiting starch staling in bread) The effect of inhibiting starch staling in bread was evaluated based on the degree of decrease in crumb hardness compared to the control group, according to the following evaluation criteria. [Evaluation criteria for the effect of inhibiting starch staling in bread] Less than 30% decrease: × (0 points) A decrease of 30% to less than 50%: △ (1 point) A decrease of 50% to less than 70%: ○ (3 points) 70% or more reduction: ◎ (5 points)

[0155] (Overall evaluation of the bread) Based on the sum of the evaluation results for softness, rise, and starch staling inhibition, the bread was comprehensively evaluated according to the following evaluation criteria. [Overall evaluation criteria for bread] 0~3 points: × 4~7 points:△ 8~11 points: ○, 12~15 points:◎

[0156] [Table 9]

[0157] As shown in Table 9, by incorporating branched dextran (branched dextran-2) that satisfies the requirements of the present invention, bread was obtained that showed less change in hardness than the control, and furthermore, had good softness, rise, and starch retrogradation inhibitory effects.

[0158] Furthermore, the branched dextran that met the requirements of the present invention received higher evaluations in all categories compared to commercially available dextrans.

[0159] <Test Example 10: Effect of branched dextran in improving the quality of cooked rice> The quality improvement effect of the branched dextran (branched dextran-2) obtained above, when incorporated into cooked rice, was evaluated based on the following method. The results are shown in Table 10.

[0160] 150g of uncooked rice (Akita Komachi rice from Akita Prefecture, polished) was rinsed and transferred to a rice cooker, and water was added to make a total of 368g. Next, branched dextran was added in the proportions shown in the "Added Ingredients" section of Table 10, lightly stirred, and then cooked in a rice cooker. In the table, the values ​​shown in the "Additives" column represent the ratio (weight %) of the weight (g) of branched dextran to the weight (g) of raw rice. After cooking, the rice was lightly loosened and left to steam for 15 minutes on the keep-warm setting. It was then wrapped in plastic wrap in 130g portions to a thickness of about 2cm, allowed to cool at room temperature for 1 hour, and then refrigerated. The following evaluations were conducted on cooked rice before and after storage.

[0161] (Measurement of the hardness of cooked rice) Using a creep meter (RE2-3305, manufactured by Yamaden Co., Ltd.) equipped with a cylindrical plunger (20 mm in diameter), the maximum load (gf) when uniaxial compression was performed at a compression speed of 1 mm / sec, with the deformation limit set to 50% of the thickness of cooked rice, was determined as the hardness of the cooked rice. The measurement was repeated 10 times, and the average of the 8 measurements was calculated after excluding the lowest and highest values.

[0162] (Measurement of the change in hardness of cooked rice) Based on the average value obtained by dividing the standard error by the hardness of the cooked rice on the day of preparation and after refrigeration for one day, the change in hardness of the cooked rice due to storage was calculated using Equation 7. [Formula 7] Change (Δgf) = Hardness of cooked rice after refrigeration for 1 day (gf) - Hardness of cooked rice on the day of preparation (gf)

[0163] (Evaluation of the effect of inhibiting starch retrogradation in cooked rice) The effect of inhibiting starch retrogradation in cooked rice was evaluated based on the degree of decrease in the hardness of cooked rice compared to a control, according to the following evaluation criteria. [Evaluation criteria for the effect of inhibiting starch retrogradation in cooked rice] Decreased by 20% or more: △ 30% or more decrease:○ 40% or more decrease:◎

[0164] [Table 10]

[0165] As shown in Table 10, by incorporating branched dextran (branched dextran-2) that satisfies the requirements of the present invention, cooked rice was obtained with less change in hardness and better starch retrogradation inhibition effect compared to the control. In particular, when 0.3% by weight of branched dextran that satisfies the requirements of the present invention was added, a remarkable starch retrogradation inhibitory effect was observed.

[0166] <Test Example 11: Making Sourdough Bread> Sourdough bread is a type of bread made using dough co-cultured with multiple types of microorganisms. In this example, sourdough bread was made using yeast along with lactic acid bacteria (Leuconostoc citreum KD3 strain) as follows.

[0167] After uniformly mixing 50g of strong flour and 50g of sterile water, add 1ml of Leuconostoc citreum KD3 strain cell suspension and mix well. The initial number of viable cells is 1 × 10⁶. 8 The concentration was adjusted to approximately cfu / ml. The resulting mixture was fermented at 30°C for 24 hours to obtain sourdough starter. Next, referring to the bread composition described in Table 9 of "Test Example 9" above, 100g of the total 420g of strong flour and water was replaced with the above-mentioned sourdough starter, mixed with the other ingredients, and sourdough bread (white bread) was produced by sequentially performing the following steps using a home bread maker ("Automatic Home Bakery SD-BT-103", manufactured by National): kneading, adding dry yeast, resting, kneading, fermentation, baking, and cooling. The resulting sourdough bread was a high-quality bread in which the starch was modified by branched dextran contained in the sourdough starter, suppressing staling, maintaining softness, and improving texture by giving it a chewy feel.

[0168] <Test Example 12: Making Pickles> Commercially available Chinese cabbage was cut into 5cm cubes, soaked in 100ppm sodium hypochlorite (Purelax S (Oyalax Co.)) for 10 minutes, and then rinsed with tap water to sterilize the cabbage. After sterilization, 50g portions of soft and hard leaf tissue were placed in resealable plastic bags in roughly equal proportions. 150ml of salt / sucrose-containing water (prepared to have a 5% sucrose concentration and a 1% salt concentration) and 200μl of Leuconostoc citreum KD3 strain cell suspension were added to each bag, and the mixture was kept warm at 15°C to begin fermentation. We confirmed that the pH reached below 4.0 on the 2nd to 3rd day after the start of fermentation. Pickles inoculated with Leuconostoc citreum KD3 strain as a fermentation starter possessed the characteristic flavor of lactic acid fermentation, and furthermore, the branched dextran of the present invention gave the pickling liquid a thicker consistency, resulting in high-quality pickles with enhanced body.

[0169] <Example 13: Preparation of Amazake> 200g of rice koji, 200g of cooked rice, and 600ml of water were mixed and kept warm at 60°C in a yogurt maker for 10-12 hours to ferment. Next, 1ml of Leuconostoc citreum KD3 strain cell suspension was added to the resulting fermented product and mixed well. This was then fermented at 30°C for 24 hours to produce lactic acid fermented amazake. The resulting amazake, produced using Leuconostoc citreum KD3 strain as a fermentation starter, was a high-quality amazake with a clean taste due to the characteristic flavor of lactic acid fermentation, and enhanced body due to the branched dextran produced.

[0170] <Example 14: Yogurt Preparation> Add 10 ml of Leuconostoc citreum KD3 strain cell suspension to 1000 ml of milk, mix well, and the initial number of viable cells is 1 × 10⁶. 9 The concentration was adjusted to approximately cfu / ml. This was then kept warm in a yogurt maker at 30°C and fermented for 10-15 hours to produce yogurt. The resulting yogurt was a high-quality yogurt with a smooth texture due to the branched dextran produced by using Leuconostoc citreum KD3 strain as a fermentation starter.

Claims

1. A branched dextran derived from lactic acid bacteria belonging to Leuconostoc citreum, The glycosidic bonds of the branched dextran consist of α-1,6 bonds accounting for 50% to less than 99% of the total glycosidic bonds, α-1,2 bonds accounting for 1% to less than 40%, and α-1,3 bonds accounting for 0.1% to less than 20%. The value obtained by dividing the weight-average molecular weight of the branched dextran by the number-average molecular weight of the branched dextran is 100 or more and less than 1500. The branched dextran contains 80% by mass or more of water-soluble dietary fiber. Branched dextran.

2. The branched dextran according to claim 1, wherein the water retention rate of the branched dextran is 10% by mass or more.

3. An intestinal regulator comprising branched dextran according to claim 1 or 2.

4. A cholesterol-lowering agent comprising branched dextran according to claim 1 or 2.

5. A blood glucose level elevation inhibitor comprising branched dextran according to claim 1 or 2.

6. A starch retrogradation inhibitor comprising branched dextran according to claim 1 or 2.

7. A bread texture improver comprising branched dextran as described in claim 1 or 2.

8. Leuconostoc citreum KD3 strain (accession number NITE P-03378) for producing branched dextran according to claim 1 or 2.

9. A culture process for culturing lactic acid bacteria belonging to Leuconostoc citreum, The process includes a filtration step in which the culture is filtered after the culture step to recover a filtrate containing branched dextran, In the filtration step, a porous filtration membrane is used that contains one or more selected from the group consisting of aluminum oxide, zirconium oxide, and titanium oxide, and has a pore size of 0.01 μm or more and 1.00 μm or less. A method for producing branched dextran according to claim 1 or 2.

10. The method for producing branched dextran according to claim 9, wherein the lactic acid bacterium belonging to Leuconostoc citreum is Leuconostoc citreum KD3 strain (accession number NITE P-03378).

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