KR101089637B1 - Method for producing gardenia fruit extract with improved anti-inflammatory activity by enzyme treatment - Google Patents

Abstract

The present invention provides a method for producing a gardenia extract with increased anti-inflammatory activity comprising the step of treating the gardenia extract with β-glucosidase, an anti-inflammatory pharmaceutical composition containing the gardenia extract prepared by the method as an active ingredient and the It relates to a health functional food for the prevention and improvement of inflammation containing the gardenia extract prepared by the method as an active ingredient.

Gardenia extract, β-glucosidase, anti-inflammatory activity, pharmaceutical composition, dietary supplement

Description

Method for producing gardenia fruit extract with improved anti-inflammatory activity by enzyme treatment

The present invention relates to a method for producing a gardenia extract with increased anti-inflammatory activity by enzyme treatment, and more particularly, to a gardenia extract with increased anti-inflammatory activity, comprising the step of treating β-glucosidase to the gardenia extract It relates to a manufacturing method, an anti-inflammatory pharmaceutical composition containing the gardenia extract prepared by the method as an active ingredient, and a health functional food for preventing and improving inflammation containing the gardenia extract prepared by the method as an active ingredient.

In recent years, various lifestyle-related diseases (cancer, arteriosclerosis, diabetes, dementia, arthritis, allergies, etc.) and aging are caused by oxidative substances produced in the body. It is thought that antioxidants can act in the body to prevent or suppress lifestyle diseases and aging.

However, in recent years, the point of view has changed, and it is clear that the anti-inflammatory response caused by dietary ingredients is more involved. Foods contain anti-inflammatory substances that work together to reduce the risk of developing inflammatory diseases. In addition, activation of the intracellular signaling system, which is closely related to the inflammatory response, is widely involved in the development of aging and various lifestyle diseases [Kumar et al. Takada Y, Boriek AM, Aggarwal BB. 2004. Nuclear factor-kappa B: its role in health and disease. J Mol Med. 82: 434-48; Sarkar D, Fisher PB. 2006. Molecular mechanisms of aging-associated inflammation. Cancer Lett. 236; 13-23; Youn HS. 2007. Anti-inflammatory effects of resveratrol, (-)-epigallocatechin-3-gallate and curcumin by the modulation of Toll-like receptor signaling pathway. Kor J Food Sci Technol. 39: 481-487.

A good example of the relationship between lifestyle disease and inflammation is atherosclerosis, which is a life-threatening cause of cardiovascular disease (CVD) and coronary heart disease (CHD). . Among the factors involved in the development of atherosclerosis, inflammation is one of the most important factors [de Winther MPJ, Kanters E, Kraal G, Hofker MH. 2005. Nuclear factro kB signaling in atherogenesis. Arterioscler Thromb Vasc Biol. 25: 904-914.

Natural products that exhibit anti-inflammatory activity include green tea, red pepper, propolis, pomegranate, red pepper, rosemary, cinnamon, aloe, ginger, broccoli, garlic, turmeric, wine, rhubarb, and gardenia. In particular, it has been reported that gardenia exhibits a very good activity for improving dementia and thus can be used to produce antidementia materials.

On the other hand, using the transformation technology of the natural material can effectively improve the physiological activity. In particular, when the sugar chains of glycosides commonly found in natural products are cleaved with enzymes to produce a large amount of aglycones, their usefulness is not only increased in the body but also improved in the small intestine when ingested. This improvement has been reported.

As representative examples thereof, "Oats-derived beta-glucosidase and a method for producing Genistein or Didezein using the same", "Method of low molecular weight of ginseng saponin and functional food production method using ginseng saponin decomposed product", "β-glucosida" Bifidobacterium lactis producing the agent and a method for converting isoflavones into isoflavone nonglycosides using the same, and "method of preparing isoflavones in the form of non-glycosides from bovine".

This prior art has been limited to the improvement of physiological activity through the modification of isoflavones mainly derived from soybean or soybean or saponins derived from ginseng using β-glucosidase or its activity. However, there was no case where the gardenia extract was treated with this enzyme to dramatically improve anti-inflammatory activity.

Korean Patent Registration No. 10-0661481 discloses a glycoprotein extracted from gardenia and liver protection, hypolipidemia and anti-inflammatory pharmaceutical composition containing the same.

The present invention has been made in accordance with the requirements as described above, Gardenia fruit extract has an excellent effect on the prevention of dementia or arteriosclerosis has been reported high quality anti-inflammatory activity, but the inactivation of the extract is not high, natural products derived from gardenia It was necessary to develop the material deformation technology that can maximize the economic utility. Therefore, the present invention aimed to confirm the anti-inflammatory activity of gardenia extract by inducing the degradation of glycosides by treating beta-glucosidase derived from the fungus for the purpose of increasing the anti-inflammatory activity of the gardenia extract through material modification.

In order to solve the above problems, the present invention provides a method for producing a gardenia extract with increased anti-inflammatory activity, comprising the step of treating β-glucosidase to the gardenia extract.

The present invention also provides an anti-inflammatory pharmaceutical composition containing the gardenia extract prepared by the method as an active ingredient.

In addition, the present invention provides a health functional food for the prevention and improvement of inflammation containing the gardenia extract prepared by the method as an active ingredient.

According to the present invention, anti-inflammatory activity was remarkably improved by inducing material deformation by enzyme reaction using β-glucosidase in gardenia extract. When used orally ingested as a material of health functional foods, the effect of health promotion is expected by suppressing the inflammatory reactions in the body which are factors of lifestyle-related diseases and aging.

In order to achieve the object of the present invention, the present invention provides a method for producing a gardenia extract with increased anti-inflammatory activity comprising the step of treating β-glucosidase to the gardenia extract.

In the method according to an embodiment of the present invention, the gardenia extract can be extracted by adding a solvent to the gardenia fruit, and processing the microwave. The solvent may be water, ethanol or a mixed solvent thereof, preferably a water-ethanol mixed solvent. For example, the microwave can be processed at 90W for 5 minutes.

In the method according to an embodiment of the present invention, the β-glucosidase may be an enzyme derived from a fungus of the genus Aspergillus . The fungus of the genus Aspergillus is Aspergillus niger , Aspergillus fumigatus , Aspergillus nidulans , Aspergillus terreus , Aspergillus Sparbus ( Aspergillus flavus ) and the like, but is not limited thereto. The treatment time of β-glucosidase may be 12 hours or more, preferably 12 hours to 24 hours. If the treatment time is less than 12 hours, anti-inflammatory activity is not sufficient, and if it exceeds 24 hours, the increase in anti-inflammatory activity compared to the treatment time is not significant.

In the method according to one embodiment of the present invention, the treatment conditions of the β-glucosidase is more preferably 20 unit / ml, pH 4.6, 45 ℃, 24 hours, but is not necessarily limited thereto.

The present invention also provides an anti-inflammatory pharmaceutical composition containing the gardenia extract prepared by the method for producing gardenia extract with increased anti-inflammatory activity as an active ingredient.

Gardenia fruit has been used as a herbal medicine for a long time, the extracts of the present invention extracted therefrom also have no problems such as toxicity and side effects.

The anti-inflammatory pharmaceutical composition of the present invention may include from 0.02 to 50% by weight of the extract based on the total weight of the composition.

Pharmaceutical compositions comprising the extract of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

The pharmaceutical dosage forms of the extracts of the present invention may be used in the form of their pharmaceutically acceptable salts, and may be used alone or in combination with other pharmaceutically active compounds as well as in a suitable collection.

Pharmaceutical compositions comprising extracts according to the invention, in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, oral preparations, suppositories and sterile injectable solutions, respectively, according to conventional methods. Can be formulated and used. Carriers, excipients and diluents that may be included in the composition comprising the extract include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate And various compounds or mixtures including cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil and the like. When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules and the like, and such solid preparations may include at least one excipient such as starch, calcium carbonate and sucrose in the extract. ) Or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Oral liquid preparations include suspensions, solvents, emulsions, and syrups, and may include various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. . Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

Preferred dosages of the extracts of the present invention vary depending on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration and the duration, and may be appropriately selected by those skilled in the art. However, for the desired effect, the extract of the present invention is preferably administered at 0.0001 to 100 mg / kg, preferably 0.001 to 100 mg / kg per day. Administration may be administered once a day or may be divided several times. The dosage does not limit the scope of the invention in any aspect.

The extract of the present invention can be administered to mammals such as rats, mice, livestock, humans and the like in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections.

The present invention also provides a health functional food for preventing and improving inflammation containing the gardenia extract prepared by the method for producing gardenia extract with increased anti-inflammatory activity as an active ingredient.

Examples of the food to which the gardenia extract can be added include various foods, beverages, gums, teas, vitamin complexes, and health functional foods.

It may also be added to foods or beverages for the purpose of preventing and improving anti-inflammatory effects. At this time, the amount of the extract in the food or beverage may be added in 0.01 to 15% by weight of the total food weight, the health beverage composition may be added in a ratio of 0.02 to 5 g, preferably 0.3 to 1g based on 100 ml. have.

The health functional beverage composition of the present invention has no particular limitation on the other ingredients other than the above-mentioned extract as an essential ingredient in the indicated ratio, and may contain various flavors or natural carbohydrates as an additional ingredient such as ordinary beverages. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose and the like; Disaccharides such as maltose, sucrose and the like; And conventional sugars such as polysaccharides such as dextrin, cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents (tauumatin, stevia extract, for example, rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. . The proportion of such natural carbohydrates is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the composition of the present invention.

In addition to the above, the extract of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic flavors and natural flavors, coloring and neutralizing agents (such as cheese and chocolate), pectic acid and salts thereof, alginic acid and its Salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated beverages, and the like. In addition, the extracts of the present invention may contain flesh for the production of natural fruit juices and fruit juice beverages and vegetable beverages. These components may be used independently or in combination, and the proportion of such additives is not so critical but is generally selected in the range of 0 to about 20 parts by weight per 100 parts by weight of the extract of the present invention.

Hereinafter, the present invention will be described in detail by way of examples. However, the following examples are illustrative of the present invention, and the present invention is not limited to the following examples.

The inventors of Aspergillus aimed at effectively inducing sugar chain degradation of glycosides in gardenia berry extract. Β-glucosidase from the genus was treated. Subsequently, in order to evaluate the anti-inflammatory activity of the enzyme-treated gardenia extract, several anti-inflammatory tests using RAW264.7, a mouse macrophage cell line, were performed as follows and the indicators were evaluated.

Experimental Example 1

To prepare the gardenia extract, solvent (3 kinds: water, ethanol, ethanol-water 1: 1 mixture) was added to the gardenia crushed powder. At this time, an atmospheric pressure microwave extraction device was used for effective extraction of useful components. As extraction conditions, 200 mL of three solvents were added to 10 g of the sample, and microwaves were treated for 90 W for 5 minutes. This was filtered through a filter paper, concentrated with a rotary vacuum concentrator, and then adjusted to 50 mL by adding distilled water.

Experimental Example 2

Aspergillus as an enzyme to prepare enzyme reaction of Gardenia extract Beta-glucosidase from the genus was used, mainly from Aspergillus niger (Fluka 49291, U / mL). Almond-derived β-glucosidase (Sigma G4511, U / mL) was used as a control. 5 mL of sodium acetate buffer (pH4.6) containing enzyme (final 0, 0.5, 1, 2, 5, 10, 20 U / mL) was added to 5 mL of the gardenia extract concentrate for the enzymatic reaction of the extract. , 1, 3, 6, 12, 18, 24h) and the temperature (37 ℃, 45 ℃) was treated differently.

Example  1. Enzyme and Solvent Treatment Tooth extract NO  Inhibitory Activity Comparison

(1) Experimental method

Dispense 200 uL into 96 wells at a cell density of 1 × 10 ⁶ cells / ml in DMEM containing 10% (v / v) FBS, 100 units / ml penicillin, 100 ug / ml streptomycin using a RAW264.7 cell line. The culture time was 37 ℃, 5% CO ₂ incubator. Then, the gardenia extract reacted with the above treatment was added at different amounts (0.313, 0.625, 1.25, 2.5, 5.0 uL), incubated for 1 hour, and treated with LPS (1㎍ / ml) for 24 hours. It was. Resveratrol was used as a positive control and the extract was used as an experimental group. For NO analysis, the amount of nitrate produced during cell culture was measured. That is, 50ul of the cell culture supernatant was taken and reacted with Griess reagent (50µl) for 5 minutes, and the absorbance was measured at 540nm using an ELISA reader. At this time, by comparing the relative amount of the finally produced NO based on the standard NaNO ₂ , the NO inhibitory activity by treatment was obtained and used as a material of anti-inflammatory activity.

(2) Experiment result

A) Comparison of anti-inflammatory activity according to beta-glucosidase enzyme type

When comparing the applicability of A. niger- derived and almond-derived enzymes as a treatment enzyme for gardenia extract (pH 4.6, 37 ° C, 48h shaking, 25 units / ml of enzyme), the latter was compared to the former. NO inhibitory activity was not high (Fig. 1). In addition, A. niger also treated with beta-glucosidase enzyme from A. fumigatus. By showing a very similar tendency to the derived enzyme (data omitted), it was found that the enzyme derived from the genus Aspergillus was very effective in increasing the anti-inflammatory activity of the gardenia extract.

B) Change of anti-inflammatory activity according to enzyme concentration

Gardenia extract enzyme reaction obtained by reacting A. niger- derived beta-glucosidase (0, 0.5, 1, 2, 5, 10, 20 unit / ml) in gardenia extract at pH 4.6, 45 ° C. for 24 hours Was applied to the NO inhibitory activity test. As a result, IC ₅₀ (expressed as the amount of reactant added in units of μl) of each of the enzyme reactions was ND, 4, 1.5, 0.9, 0.8, 0.75, and 0.4, respectively, which strongly inhibited NO production in proportion to the enzyme treatment concentration. were (Fig. 2), this translates to IC ₅₀ values as key quantities respectively corresponding to ND, 600, 225, 135, 120, 113, about 60 ug / ml. However, in the treatment of enzymes of 2U / ml or more, when a large amount of enzyme reaction was administered to the cultured cells (5 μl), a phenomenon of decreasing NO inhibitory activity was observed. However, as a whole, it was found that the NO inhibitory activity of the gardenia extract was increased by several orders of magnitude.

C) Changes in anti-inflammatory activity with time of enzyme treatment

A. niger-derived beta-glucosidase was added to the gardenia extract and the change of anti-inflammatory activity was investigated over time with pH 4.6, 45 ° C., and reaction for 24 hours. As a result, the NO inhibitory activity increased in proportion to the time up to 12 hours (increasing the relative activity compared to the IC ₅₀ standard), but afterwards the activity tended to decrease slightly. Therefore, in this condition, the enzyme reaction time was found to be more than 12 hours (Fig. 3).

D) Review of optimum conditions for enzyme treatment

What kind of extraction solvent (water, 50% ethanol, 100% ethanol) was used in the gardenia extract for anti-inflammatory activity, and if the enzyme treatment was performed on these three extracts, the effect (inactivity) differed to some extent. The optimal condition was examined by comparing the presence of the present condition. At this time, the reaction conditions were enzyme addition 20 unit / ml, pH 4.6, 45 ℃, 24 hours. As a result, the extraction solvent was 50% ethanol>water> 100% ethanol in order, and the inactivation was increased more than 10 times in all cases treated with enzyme (Fig. 4). Therefore, it was judged that the anti-inflammatory activity was the best by treating enzymes derived from Aspergillus spp. On gardenia extract prepared with 50% ethanol as the extraction solvent. At this time, IC ₅₀ of the gardenia extract enzyme reaction was 0.5 μl, which was about 70 ug / ml in terms of dry weight.

Example 2. Evaluation of Other Anti-Inflammatory Indicators

(1) Experimental method

In the anti-inflammatory activity test of the RAW264.7 cell line, we investigated the inhibitory activity of NO production induced by LPS treatment in this case. For example, changes in inflammatory cytokines, changes in enzyme expression involved in intracellular signaling, changes in NF-kB activity, and the like.

A) Analysis of Inflammatory Cytokines

In order to analyze the degree of cytokine production by the treatment of gardenia extract, RAW 264.7 cells were dispensed in 12 well plates at a concentration of 1 × 10 ⁶ cells / ml (or 1 × 10 ⁵ cells / ml; for TNF-a) and incubated for 2 hours. After each sample was treated by concentration and incubated for 1 hour, and then treated with LPS (1㎍ / ml) was incubated in 37 ℃, 5% CO ₂ incubator for 24 hours. Each cytokine (IL-6, IL-1b, TNF-a) generated in the supernatant obtained at this time was analyzed according to the instructions using OptEIA ^™ Mouse ELISA set (BD Sci. Co.), respectively.

B) Western blot analysis

After extracting the protein of the cell was centrifuged to take the supernatant. The supernatant was quantified for protein concentration using Bradford reagent to take 40 μg of protein. The extracted protein was subjected to 10% SDS-PAGE and the separated protein was transferred to the nitrocellulose membrane. After treatment with 5% skim milk for one night, iNOS and COX-2 antibodies at a ratio of 1: 500 were left at room temperature for 4 hours, and then washed twice with TTBS at 15 minute intervals. Secondary antibodies diluted at a ratio of 1: 1000 were left at room temperature for 1 hour. After washing three times with TTBS every 15 minutes, it was developed with ECL.

C) Analysis of NF-kB inhibitory activity using luciferase assay

To evaluate the inhibitory activity of NF-kB activity, the firefly luciferase plasmid and renilla luciferase plasmid (control, Promega, E2231) obtained by mass propagation in E. coli and purified by Endofree plasmid Maxi kit, respectively, 0.5 ㎍ / well, RAW264.7 cells were co-transfected to add 0.005 μg / well. At this time, the cells were cultured by dispensing into 96 wells at a density of 1x10 ⁵ cells / well the previous day. After 6 hours of transfection, medium replacement was performed once with complete media. Thereafter, 18 hours of incubation, the samples were treated by concentration, and 1 hour later, LPS (1 µg / ml) was treated and cultured for 23 hours. Then, according to the Dual-Glo Luciferase Analysis System (Promega, E2940), the luminescence level obtained by treating the reagent was measured by a luminometer, and the result was expressed as RRR (Relative Response ratio).

(2) Experiment result

A) Changes in cytokine production by treatment of gardenia extract with enzyme reactant

In the experimental system, the effect of the enzyme reaction treatment of the gardenia extract on the production of inflammatory cytokines (IL-1b, IL-6, and TNFa) was investigated (FIG. 5). As a result, IL-1b did not differ significantly from the activity of unenzymatic activity (inhibition of cytokine secretion), but the production of inflammatory cytokines was inhibited from the enzyme treatment of IL-6 and TNFa. Therefore, it was also confirmed here that the gardenia extract has high anti-inflammatory activity by enzyme treatment.

B) Inhibition of Inflammation-related Enzyme Production by Extract Treatment

Next, changes in the inhibitory activity of COX-2 and iNOS production were investigated. 50% ethanol extracts of gardenia and rhubarb showed strong concentration-dependent COX-2 inhibitory activity. The intensity of activity was much higher in the enzyme-treated gardenia extract (FIG. 6). In addition, iNOS production inhibition experiments by the extract treatment showed a similar tendency to that of COX-2 (data omitted). Therefore, the production inhibitory activity of these inflammation-related enzymes also showed good agreement with the NO inhibitory activity investigated previously.

C) Inhibition of Transcription Factor NF-kB Activity by Extract Treatment

It is well known that external signals through TLRs, such as LPS, are transmitted into cells to activate NF-kB, which then migrates into the nucleus and is involved in transcription, leading to the production of inflammatory cytokines and the like and ultimately triggering inflammatory responses. Therefore, we tried to observe the inhibition of NF-kB activation induced by the treatment of gardenia extract.

As a result of analyzing the luciferase gene as a reporter gene in RAW264.7 cells (FIG. 7), the inhibitory activity of the enzyme extract of Gardenia jasminoides was observed in a concentration dependent manner, but the inhibitory activity was almost suppressed in the gardenia extract without enzyme treatment. Did not appear. This result was also in good agreement with the results of the NO inhibitory activity investigated above.

Therefore, the overall results are summarized as follows. Gardenia extract showed weak activity in experimental system using RAW264.7 cell line which is widely used for anti-inflammatory test. However, when the gardenia extract treated with beta-glucosidase was treated in the experimental system, it was confirmed that the anti-inflammatory activity significantly increased more than 10 times. Therefore, the health functional material produced from gardenia in this way can be utilized to suppress various lifestyle diseases and aging that can be caused by inflammation in the human body.

1 Induced by LPS in RAW264.7 cells by tooth extract (50% EtOH + microwave extraction) treated with β-glucosidase from A. niger and almond for 48 hours at 25U / ml, pH 4.6, 37 ° C Results in NO production inhibition. RV: resveratrol, A. nig. enz .: from A. niger β-glucosidase , Almond enz .: β-glucosidase from almonds.

Figure 2 shows LPS-induced NO in RAW264.7 cells by tooth extract (water + microwave extraction) treated with β-glucosidase derived from A. niger for 24 hours at 0-20U / ml, pH 4.6, 45 ° C. Results in production inhibition. RV: resveratrol, A. nig. enz .: from A. niger β-glucosidase .

Figure 3 RAW264 with tooth extract (water + microwave extraction) treated with 20 U / ml of β-glucosidase from A. niger for 1, 3, 6, 12, 18, and 24 h at pH 4.6, 45 ° C. The result is LPS-induced NO production inhibitory activity in 7 cells. The results are shown as relative inhibitory activity of NO production.

Figure 4 LPS in RAW264.7 cells by tooth extract (water, EtOH, 50% EtOH + microwave extraction) treated with β-glucosidase from A. niger for 24 hours at 20U / ml, pH 4.6, 45 ° C. Induced NO inhibition. RV: resveratrol, Gardenia Water: Gardenia extract from distilled water, Gardenia EtOH: Gardenia extract from ethanol, Gardenia extract 50% EtOH: Gardenia extract from 50% ethanol, enz 20U: β-glucosidase from A. niger 20U / ml treatment.

FIG. 5 shows the inhibition of LPS-induced inflammatory cytokine production in RAW264.7 cells by a tooth extract treated with β-glucosidase derived from A. niger for 24 hours at 20 U / ml, pH 4.6, 45 ° C. FIG. Gardenia jasminoides: Gardenia extract, Gardenia A.nig, extracted with 50% ethanol. enz .: Gardenia extract treated with β-glucosidase from A. niger .

FIG. 6 shows the results of inhibition of LPS-induced inflammatory enzyme COX-2 production in RAW264.7 cells by Gardenia extract treated or untreated with β-glucosidase. Resv .: resveratrol, Gardenia Ext .: Gardenia extract extracted with 50% ethanol, Gardenia + enzyme: Gardenia extract treated with β-glucosidase from A. niger .

7 shows the results of inhibition of LPS-induced inflammatory transcription factor NF-kB activity in RAW264.7 cells by gardenia extracts treated or untreated with β-glucosidase. Gardenia jasminoides: Gardenia extract, Gardenia A.nig, extracted with 50% ethanol. enz .: Gardenia extract treated with β-glucosidase from A. niger .

Claims (10)

A mixture of water-ethanol was added to the gardenia fruit and the gardenia extract extracted by treating the microwave at 90W for 5 minutes was treated with β-glucosidase derived from the fungus of Aspergillus at 45 ° C for 12 to 24 hours. Method of producing a gardenia extract with increased anti-inflammatory activity comprising the step. delete delete delete delete delete delete The method according to claim 1, wherein the treatment conditions of β-glucosidase are 20 unit / ml, pH 4.6, 45 ° C, 24 hours. delete delete

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