KR100485154B1 - Composition for enhancing immune reaction containing indigestible dextrin - Google Patents

Abstract

The present invention relates to immunoactive compositions comprising indigestible dextrins. In particular, the present invention relates to a composition comprising an indigestible dextrin capable of enhancing nonspecific, specific immune function.

Description

Immunoactive composition comprising indigestible dextrin {COMPOSITION FOR ENHANCING IMMUNE REACTION CONTAINING INDIGESTIBLE DEXTRIN}

[TECHNICAL FIELD OF THE INVENTION]

The present invention relates to an immunoactive composition comprising an indigestible dextrin, and more particularly to a composition comprising an indigestible dextrin that inhibits obesity and atherosclerosis and activates the immune system.

[Private Technology]

Dietary fiber is an indigestible component in plants that is indigestible by the digestive enzymes present in the human body. In addition to gum and mucilage, dietary fiber includes cell wall materials such as cellulose, hemicellulose, lignin, and pectin. .

Dietary fiber is divided into water-soluble and insoluble. Water-soluble dietary fiber, which is widely used as a food raw material, includes pectin, gum, β-glucan, etc. present in apples, carrots, beans, barley, oats, and the like, and cellulose and hemicellulose present in grains, crustaceans, etc. , Lignin and chitin.

Soluble dietary fiber has physiological activities such as lowering blood cholesterol level and suppressing post-prandial blood sugar level. Has action.

Recently, water soluble fiber has been reported to have a positive effect on the improvement of immune function. When soluble dietary fiber, pectin and insoluble dietary fiber, were ingested in rats for 2 weeks, the concentrations of IgA, IgG, and IgM in the intestinal lymph nodes of rats with pectin were significantly increased compared to those with cellulose. The CD4 + T-cell, CD4 + / CD8 + ratio of the intestinal membrane was also observed to increase. Gamma-interferon was also found to be significantly increased, suggesting that soluble dietary fiber enhances the immune function of the gastrointestinal tract. However, it appears that there is a difference in the immune activity effect according to the type of water-soluble dietary fiber, and glucomannan was found to have a much weaker immune function improvement effect than pectin. This activity is believed to be formed by the mechanism by which water-soluble dietary fiber forms short-chain fatty acids in the intestinal tract, and the formed short-chain fatty acids increase the number of T-cells in the gastrointestinal tract.

On the other hand, indigestible dextrin derived from starch is known to inhibit insulin branching (Japanese Patent No. Hei 7-28693) and to have a formal function (Japanese Patent Hei 7-28694). However, the above-mentioned prior arts confirm the blood lipid metabolism improvement, blood sugar control action, and intestinal action of the dietary fiber prepared from the shell of cereals, and do not deviate greatly from the application range of the conventionally known dietary fiber.

In order to solve the problems of the prior art, an object of the present invention is to provide a composition comprising an indigestible dextrin that can reduce weight gain.

It is another object of the present invention to provide a composition comprising an indigestible dextrin capable of increasing the content of high density lipoproteins in the blood and reducing the content of low density lipoproteins.

It is another object of the present invention to provide a composition comprising an indigestible dextrin capable of activating an animal's immune system to promote health.

In order to achieve the above object, the present invention provides an immunoactive composition comprising an indigestible dextrin.

Hereinafter, the present invention will be described in detail.

The present inventors completed the present invention by confirming that the indigestible dextrin derived from starch not only suppresses obesity and atherosclerosis but also improves the immune response in the body.

Indigestible dextrins of the present invention are all kinds of indigestible dextrins, and can be prepared according to the method described in Korean Patent Publication No. 243525, for example. The refractory dextrin manufacturing method includes the steps of predrying the starch with hot air, adding hydrochloric acid to salt treatment, mixing neutralizing agent, dissolving dextrin in hot water, reacting heat-resistant alpha-amylase, and reacting activated carbon with reactant. Adding and heating, filtering to remove activated charcoal, adding anion exchange resin to produce decolorized dextrin, deionizing dextrin by adding ion exchange resin mixed with anion and cation to the sugar solution and concentrating desalted dextrin And spray drying.

A more preferred method for producing an indigestible dextrin is (a) pre-drying the raw starch with hot air, adding gaseous hydrochloric acid and heat treatment, and (b) mixing the heat-treated dextrin prepared by mixing the neutralizing agent in a dry state to hot water at 70 to 90 ° C. (C) heat-resistant alpha-amylase was added at 0.05 to 0.5% by weight based on heat treated dextrin, and reacted at 80 to 100 ° C. for 100 to 150 minutes, and (d) heat treated activated carbon. About 0.5 to 1.0% by weight of dextrin is added, followed by heating to inactivate the enzyme, (e) filtering the inert liquid to remove activated carbon, and (f) adding an anion exchange resin twice as much volume as the enzyme treatment liquid solid. Stirring at 35 to 45 ° C. for 20 to 30 hours yields a decolorized sugar solution, and (g) a volume of the ion exchange resin mixed with cations and anions in the sugar solution in the enzyme treatment Twice added and stirred to 35 to prepare a sugar solution in demineralized 45 ℃ for 20 to 30 hours and, (h) comprises a sugar solution was desalted and concentrated to a 35 to 45% of that, then dried with a spray drier.

The indigestible dextrin of the present invention inhibits weight gain, prevents obesity, increases blood high density lipoprotein-cholesterol levels and decreases low density lipoprotein-cholesterol levels to inhibit atherosclerosis. Indigestible dextrins also promote health by enhancing specific immune responses as well as nonspecific immune responses.

Accordingly, the present invention provides an immunoactive composition comprising an indigestible dextrin. The composition can also be used for obesity inhibition, arteriosclerosis inhibition or health promotion.

The composition according to the present invention may be a pharmaceutical composition or a food composition, in the present invention is intended to include food, food additives, beverages and beverage additives and the like.

The indigestible dextrin of the present invention can be added to 1 to 100 parts by weight based on the total weight of food, can be prepared as a health functional food such as confectionery, ramen, beverages, mayonnaise, dressings, ice cream, vitamin complexes.

Indigestible dextrins can also be prepared as a medicament by adding 1 to 100 parts by weight to one or more pharmaceutically acceptable carriers relative to the total parts by weight. The carrier may include, but is not limited to, saline, buffered saline, water, glycerol and ethanol, and any suitable agent known in the art (Remington's Pharmaceutical Science (Recent Edition), Mack Publishing Company, Easton PA) may be used. .

The composition may be prepared in the form of a warning agent, granules, powders, syrups, solutions, liquid extracts, emulsions, suspensions, acupuncture, tablets, injections, capsules, creams, troches, pasta preparations, oral or It can be used parenterally. Preferably it is oral. The dosage of the composition is a conventional dosage of dietary fiber, for example, 10 to 50 g of indigestible dextrin may be used per day. The dosage is not limited thereto, and it is preferable to be applied differently according to the age, sex, condition, absorption of the active ingredient, inactivation rate and excretion rate, disease type, and the drug used in the patient.

Hereinafter, examples of the present invention will be described. The following examples are only for illustrating the present invention and the present invention is not limited to the following examples.

Example 1 Preparation of Indigestible Dextrin

Indigestible dextrins were purchased from commercially available heat-treated dextrins (C * Dry Set 08703, Cerestar, USA), or prepared by known methods (Korean Patent Publication No. 243525).

Corn starch (Samyang Genex, South Korea) of 10% moisture was placed in a container of a fluidized bed dryer (Glatt GmbH, Germany) and pre-dried while fluidizing starch at 100 ° C. for 30 minutes with hot air. When starch has a water content of 5%, with the hot air introduced, 36% of hydrochloric acid (Song Yawon Pharmaceutical, Grade 1, Japan) diluted 50% with distilled water is uniformly sprayed with starch dry matter. Spray uniformly for about 10 minutes by 0.5%. The mixture of hydrochloric acid and starch was heat treated at 140 ° C. for 150 minutes based on the temperature of the mixture using a fruit oil boiler (Glatt GmbH, Germany). Subsequently, upon confirmation of the process, the reaction was cooled, and a pH of 6 was maintained by dissolving 0.5 wt% ammonium bicarbonate in distilled water as a neutralizing agent in distilled water, spraying in the same manner as hydrochloric acid, and mixing in a dry state. After cooling for another 30 minutes, heat-treated dextrin was obtained.

After the heat treatment dextrin prepared in 80 ℃ hot water with a constant speed stirrer (Tokyo Chemical Co., Chemy Stirrer B-100, Japan), slowly added to a concentration of 40% by weight to completely dissolve, 1 N NaOH Adjust the pH to 5.8, and add 0.2% by weight of heat-resistant alpha-amylase (Novo Nordisk Bioindustry Ltd., Teramyl 120 LS, Denmark) to the heat-treated dextrin in an electric sugar solution, The solution was reacted for 120 minutes while maintaining the temperature at 95 ° C. 1 N hydrochloric acid solution was added to the obtained sugar solution to reduce the pH to 3. The enzyme was inactivated by adding 0.7% by weight of activated carbon (Samyang Genex, South Korea) of dextrin and heating at 90 DEG C for 30 minutes. The enzyme-inactivated sugar solution was pre-filtered by suction with a vacuum suction pump using a filter paper (Advantec Co., Toyo 5A, Japan) in a Buchner funnel. Thereafter, the filter was filtered through glass filter paper (Whatman International Ltd., GF / B, GF / F, UK) to remove activated carbon, and finally filtered using a membrane filter paper (Gelman Co., Supor-200, USA). . Subsequently, the concentration of the enzyme reaction solution was adjusted to 40% using a vacuum evaporator (Tokyo Chemical Co., NE-1V, Japan), and then activated with 4% NaOH solution (Samyang, WA 30, Korea). ) Was added twice by volume to the enzyme treatment solution solids, and dextrin was decolorized by stirring with a constant speed stirrer for 24 hours in a constant temperature water bath adjusted to 40 ° C. The resin used was removed by filtration using glass filter paper. The filtrate was concentrated to 40% again, and the activated mixed resin obtained by mixing cation (Samyang, SK-1B, Korea) and anion exchange resin (Samyang, WA 30, Korea) in a volume ratio of 1: 2 After adding twice the volume ratio to the solid content it was desalted under the same conditions as decolorization. The desalted sugar solution was concentrated again to a concentration level of 40% using a vacuum concentrator. Then, an indigestible dextrin was finally prepared by drying with a spray dryer (Niro Atomizer Co., Denmark).

Example 2: Experimental Diet

Experimental Animals and Experimental Diet

The animals used in the experiment were Sprague-Dawley rats, and after a week of preliminary breeding, the cellulose group (CD, n = 8) and starch-derived indigestible dextrin group (ID), depending on the type of diet. , n = 8) and breeding for 4 weeks.

The breeding environment of the breeding room was maintained at a temperature of 23 ± 1 ℃, humidity around 50%.

During the breeding period, each animal was free to eat and drink, and the weight was measured once a week and the dietary intake was measured in the morning every day.

The experimental diet was based on AIN-93G, and the content of cellulose or dextrin was added as 5% of the total weight of the diet in terms of the weight of pure fiber (Table 1).

ingredient Cellulose group (g / kg diet) Indigestible dextrin group (g / kg diet) Starch (Samyang Genex) 496 296 Party 124 124 Casein (Samik Oil Processing Co., Ltd.) 180 180 Mineral mixture (AIN-76, Oriental yeast industry) 40 40 Vitamin mixture (AIN-76 pattern) 10 10 Corn oil (Unilever, Haejo) 100 100 Cellulose 50 - Indigestible dextrin - 250 *

All experimental data were treated with mean ± standard deviation, and the significance according to the diet was analyzed by ANOVA method and the mean difference of each group was verified at post-hoc test at P <0.05 level. .

Example 3 Measurement of Weight Gain According to Diet

The weight of the experimental animals of Example 2 was measured to determine the degree of weight gain. After one week of dieting, the body weight of the indigestible dextrin group was lower than that of the cellulose group, and this phenomenon was more pronounced at the second week of diet.

In addition, the average daily dietary intake during the 4-week diet was lower in the indigestible dextrin group than in the cellulose group. As a result of calculating the dietary efficiency in terms of the following formula, the indigestible dextrin group was lower than the cellulose group, but did not show a statistically significant difference (Table 2). In the case of serum triglycerides, the average was 28.0 mg / dl for the cellulose group and 30.8 mg / dl for the dextrin group.

(formula)

2 weeks of weight gain / total dietary intake = dietary efficiency

Cellulose group (mean ± standard deviation, n = 8) Indigestible dextrin group (mean ± standard deviation, n = 8) Weight gain after diet for 2 weeks (g) 327.8 ± 3.7 ^a 310.0 ± 3.5 ^a Daily Dietary Intake (g) 19.6 ± 0.12 ^a 17.6 ± 0.60 ^a Dietary efficiency 0.19 ± 0.008 ^b 0.18 ± 0.005 ^b

( ^a : significant difference (p <0.05), ^b : no significant difference)

Example 4 Measurement of Cholesterol Level Change According to Diet

After four weeks of diet, each animal was fasted for at least 18 hours, anesthetized with diethyl ether, and blood collected from the abdominal vena cava. Blood was stored at 4 ° C. for 30 minutes and centrifuged at 2000 × g for 10 minutes to separate serum and stored at −80 ° C. until analysis.

Serum total cholesterol, high-density lipoprotein (HDL) -cholesterol and low-density lipoprotein (LDL) -cholesterol concentrations were measured using a fully automated blood analyzer (Bayer Co., Chiron Diagnostics) and the results are shown in Table 3 below. It was.

Cellulose group (mean ± standard deviation, n = 8) Indigestible dextrin group (mean ± standard deviation, n = 8) Total Cholesterol (mg / dL) 103.9 ± 3.76 ^a 102.5 ± 7.60 ^a HDL-cholesterol (mg / dL) 22.8 ± 1.16 ^a 28.8 ± 1.02 ^a LDL-cholesterol (mg / dL) 10.5 ± 1.09 ^b 8.8 ± 0.91 ^b

( ^a : significant difference (p <0.05), ^b : no significant difference)

In Table 3, no significant differences were observed between the groups when comparing total cholesterol, but HDL-cholesterol increased by about 26% in the indigestible dextrin group compared to the cellulose group. LDL-cholesterol was observed to be no significant difference, the indigestible dextrin group showed a lower tendency than the cellulose group.

Thus, indigestible dextrins can inhibit atherosclerosis by increasing HDL-cholesterol concentrations that reduce vascular cholesterol accumulation and by decreasing LDL-cholesterol concentrations involved in vascular cholesterol accumulation.

Example 5: Ig Concentration Measurement

Changes in immune response due to the indigestible dextrin diet were observed by measuring IgE and IgA.

IgA is an antibody that is involved in mucosal immunity such as the gastrointestinal tract and respiratory tract. When IgA enters the mucous membrane, a virus or bacterium interferes with the attachment of these foreign substances to host cells. In order to evaluate the immune activity function of the indigestible dextrin dietary component, the concentration change of IgA was measured to evaluate the change of immune function in the gastrointestinal tract after ingestion.

IgE is also an antibody that mediates immediate type hypersensitivity, the first of four types of allergic reactions (or allergic reactions). That is, when IgE detects target cells by binding to mast cells or basophils, specific antigens (allergens) cross-link with IgE and induce degranulation of target cell granules to secrete allergic reactions. appear. Therefore, the decrease in the concentration of antibody (IgE) involved in hay fever, asthma, urticaria, food allergy, eczema, etc. means that the sensitivity to allergic reactions is reduced. Measured.

Serum Ig concentration was measured by ELISA. IgE is based on purified IgE from rats (Chemicon, USA) and reacted with purified mouse anti-rat IgE monoclonal antibodies (Beckton Dickinson, USA) followed by biotin conjugated mouse anti-rat IgE monoclonal The antibody was reacted with a detection antibody and the IgE concentration was measured at 405 nm. IgA was also measured in the same manner as IgE concentration measurement method, and the antibody was used as IgA monoclonal antibody. The concentrations of IgE and IgA measured in serum collected from each group of 4 weeks of experimental diet are shown in Table 4 below.

Cellulose group (mean ± standard deviation, n = 8) Indigestible dextrin group (mean ± standard deviation, n = 8) Ig E (g / mL) 48.51 ± 15.14 40.02 ± 5.53 Ig A ((g / mL) 48.8 ± 1.70 50.2 ± 2.52

The concentration of IgE was lower in the indigestible dextrin group than in the cellulose group, indicating that the immune response induced by helper T cells was improved.The concentration of IgA, which inhibits antigen invasion and prevents allergic reaction, is indigestible dextrin group. Appeared high.

Example 6: Distribution of NK Cells by Indigestible Dextrin

Mesenteric lymph nodes (MLNs) were collected from the dietary animals and placed in RPMI1640 culture medium (FCS 10%), homogenized, and then adjusted to 4 x 10 ⁶ / mL cells.

To 100 µl of each cell solution, 500 µl of PBS buffer solution (HEPES 1.191 g, 5 ml of 10% FCS, 0.1 g Na-Az, 0.1 ml, 10 ml of PBS, 500 ml with distilled water) was added and centrifuged at 2000 xg for 1 minute. Separated. Phycoerythrin (PE) anti-rat CD4 (PharMingen, USA) and biotin anti-rat CD8 (PharMingen, USA) diluted 200-fold in the sediment; fluorescein isothiocyanate (FITC) anti-rat CD45RA (PharMingen, USA) and 20 μl of biotin anti-mouse NKRPIA (PharMingen, USA) was added. It was left on ice for 20 minutes, 700 μl of PBS buffer was added, and then centrifuged at 2000 × g for 1 minute. Add 10 μl of streptavidin-FITC (Beckton Dickinson, USA) or staleptavidin-PE diluted 100-fold to the precipitate, and leave on ice again for 20 minutes, then add 700 μl of PBS to 1 minute at 2000 xg. Centrifuged. Precipitates were suspended in 500 μl of PBS and analyzed by distribution of NK cells on flow cytometry. Natural Killer (NK) cells belong to the innate immune system. That is, NK cells have a nonspecific immune function that acts directly on target cells and destroys target cells.

Cellulose group (mean ± standard deviation, n = 8) Indigestible dextrin group (mean ± standard deviation, n = 8) NK cells (%) 1.84 ± 0.09 ^a 2.90 ± 0.272 ^b

( ^a : significant difference (p <0.05), ^b : no significant difference)

In Table 5, the indigestible dextrin group showed a higher distribution ratio of NK cells than the cellulose group. Thus, indigestible dextrins enhance the nonspecific immunomodulatory function of animals.

Example 7: Concentrations of Gamma-Interferon and Interleukin-4

Spleen and intestinal lymph nodes of the experimental animals of Example 2 were extracted and the concentrations of gamma-interferon and interleukin-4 were measured.

Mesenteric lymph nodes (MLNs) were collected from the dietary animals and placed in RPMI1640 culture medium (FCS 10%), homogenized, and then adjusted to 4 x 10 ⁶ / mL cells.

Into each 300 ㎕ each cell solution in the 12-well plate, 100 ㎕ of Con A (25 g / ㎖) and LPS (50 g / ㎖) during insert 72 hours unleavened state CO ₂ 5%, at 37 ℃ conditions Incubated.

Interleukin-4 concentrations were measured using the OptEIA ^™ mouse interleukin-4 set (PharMingen, USA), and gamma-interferon concentrations were measured using the mouse gamma-interferon ELISA (Endogen, USA) commercial kit. The measured results are shown in Table 6.

Con A Cellulose group (mean ± standard deviation, n = 8) Indigestible dextrin group (mean ± standard deviation, n = 8) Gamma-Interferon (pg / mL) - - - + 9.2 ± 3.53 ^b 12.7 ± 3.60 ^b Interleukin-4 (pg / mL) - - + 13.2 ± 2.72 ^a 19.7 ± 4.13 ^a

( ^a : significant difference (p <0.05), ^b : no significant difference)

Without stimulating the intestinal lymph node cells with Con A, neither interleukin-4 and gamma-interferon were expressed in both groups. However, after stimulation with Con A, gamma-interferon was measured in the intestinal lymph nodes, but there was no significant difference between the two groups, but the indigestible dextrin group expressed higher concentrations of gamma-interferon than the cellulose group.

Interleukin-4 was significantly expressed in the indigestible dextrin group compared with the cellulose group upon ConA stimulation.

Thus, indigestible dextrins increase the expression of cytokines that activate the immune response when mesenteric lymph node immune cells undergo external stimulation.

As mentioned above, the immunoactive composition comprising the indigestible dextrin of the present invention can be widely used in health functional food materials and pharmaceutical applications by improving nonspecific and specific immune function.

Claims (5)

An immune response activating composition comprising an indigestible dextrin, The immune response is immune response activation composition is selected from the group consisting of non-specific immune response, specific immune response and hyperimmune reaction. The composition of claim 1, wherein the indigestible dextrin is derived from starch. The composition of claim 1, wherein the immune response activating composition is a food or food additive. The composition of claim 3, wherein the food is confectionary, ramen, beverage, mayonnaise, dressing, ice cream or vitamin complex. delete