HK1176010B - Blood uric acid value lowering agent - Google Patents

Description

Agent for reducing blood uric acid level

Technical Field

The present invention relates to a blood uric acid level lowering agent for preventing and treating hyperuricemia.

Background

Hyperuricemia is a state in which the uric acid concentration in blood (uric acid level in blood) is high. The result of the disruption of the equilibrium between uric acid production and excretion is an abnormal increase in the blood uric acid concentration, and inflammation is caused if uric acid that has not been dissolved in blood is locally crystallized in the body. For example, uric acid crystals cause gout attacks (gouty arthritis) accompanied by intense pain when they accumulate in joints, cause gout nodules when they accumulate subcutaneously, cause renal stones and urinary calculi accompanied by intense pain when they exist in urine, and cause renal damage when they exist in renal tubules and renal interstitium.

In recent years, there have been increasing reports of hyperuricemia itself as an independent risk factor for cardiovascular damage (see non-patent document 1), and it is known that many patients with arteriosclerotic diseases among gout patients (see, for example, non-patent document 2). Furthermore, it is also judged that there is a very good correlation between the blood uric acid level and the risk factor of metabolic syndrome (see non-patent document 3).

As described above, prevention and treatment of hyperuricemia and diseases caused by the hyperuricemia are extremely important.

Currently, drugs for reducing uric acid level include drugs for promoting uric acid excretion such as benzbromarone and drugs for inhibiting uric acid synthesis such as allopurinol, and a prescription can be made according to the type of hyperuricemia of a patient. These drugs, which are prescribed at present, are extremely excellent as uric acid level controlling agents, and the serum uric acid level can be restored to normal by taking the drugs. However, hyperuricemia itself is a disease that is very difficult to cure, and therefore, if the administration of the drug is stopped, the state returns to hyperuricemia again.

The uric acid excretion promoter can inhibit physiological reabsorption of uric acid in renal tubules to improve excretion of uric acid from kidney, thereby lowering serum uric acid level. However, urinary calculi need to be observed frequently during use, and as side effects, gastrointestinal damage, headache, dizziness (ふらつき) and the like are involved. Furthermore, when administered to a patient with a idiosyncratic condition, serious liver damage may occur (non-patent document 4).

Allopurinol is introduced into gout therapy as a uric acid production inhibitory drug and is widely used. The allopurinol blocks xanthine oxidase which acts in the final stage of purine metabolic pathway, lowers serum uric acid level, and also reduces uric acid excretion amount in urine. However, when an excessive amount of allopurinol is administered to a patient with renal insufficiency, a large amount of allopurinol accumulates in the blood, and a fatal poisoning syndrome may be caused (non-patent document 4).

As seen from this, as a therapeutic agent for hyperuricemia, a therapeutic agent that can be taken with confidence for a long period of time and in a reasonable manner, for example, a therapeutic agent derived from natural substances that have been taken daily is desired.

However, few therapeutic agents have been disclosed as natural-derived agents for treating hyperuricemia or agents for reducing the uric acid level in blood. For example, chitosan (patent document 1), water-soluble dietary fiber (patent document 2), activated carbon (patent document 3), and polylysine (patent document 4) are disclosed as substances having an action of regulating the absorption of purines that cause hyperuricemia.

On the other hand, angiotensin converting enzyme inhibitory action (patent documents 5, 6, and 7), anti-inflammatory action (patent document 8), and therapeutic effect on peptic ulcer (patent document 9) have been disclosed for the cheese enzymatic decomposition product and the peptide found from the cheese enzymatic decomposition product. However, there has been no report that a cheese enzymatic hydrolysate and a peptide found from a cheese enzymatic hydrolysate exert an effect of reducing the uric acid level.

Documents of the prior art

Patent document

Patent document 1: japanese patent laid-open publication No. 2001-163788

Patent document 2: japanese patent laid-open publication No. 2005-047828

Patent document 3: japanese patent laid-open No. 2005-187405

Patent document 4: japanese patent laid-open No. 2006 and 001898

Patent document 5: WO2004/032642

Patent document 6: WO2004/047543

Patent document 7: WO2005/061529

Patent document 8: WO2007/080962

Patent document 9: japanese laid-open patent publication No. 2009-120519

Non-patent document

Non-patent document 1: progress in Medicine Vol.14No.12, 1994p.50-53

Non-patent document 2: tokyo physician's Association journal Vol.57No.5, 2004p.35-42

Non-patent document 3: journal of medicine Vol.42No.6, 2006p.125-132

Non-patent document 4: "Abstract edition of treatment guidelines for hyperuricemia and gout" published by the Japan gout and nucleic acid metabolism society (9/1/2002)

Disclosure of Invention

Disclosed is an agent for reducing blood uric acid level, which is derived from a natural substance and can be continuously ingested for a long period of time.

Accordingly, the present inventors have conducted various studies to find a component having a blood uric acid level lowering effect from a natural substance-derived component, particularly a raw material which can be continuously ingested on a daily basis, and as a result, have found that a cheese enzymatic decomposition product has an excellent blood uric acid level lowering effect and active oxygen absorbing ability and can be used as a blood uric acid level lowering agent and a hyperuricemia preventive therapeutic agent, and have completed the present invention.

Namely, the present invention is as follows.

(1) A blood uric acid level lowering agent contains enzyme decomposition product of cheese as effective component;

(2) the blood uric acid level lowering agent according to (1), wherein the enzymatic decomposition product of cheese is an enzymatic decomposition product of cheese having an active oxygen absorbing ability;

(3) the agent for reducing blood uric acid level according to item (2), wherein the content of the enzymatic decomposition product of cheese is an amount that shows an active oxygen absorption capacity of 50 μ M water-soluble vitamin E (Trolox) equivalent to or higher than that of 1 day;

(4) the agent for reducing uric acid level in blood according to any one of (1) to (3), wherein the enzymatic decomposition product of cheese is a decomposition product obtained by treating cheese with lipase and protease;

(5) the agent for reducing uric acid level in blood according to (4), wherein the protease is at least 2 selected from the group consisting of endoprotease, exoprotease, exopeptidase/endoprotease complex enzyme, and protease/peptidase complex enzyme;

(6) the agent for reducing blood uric acid level according to (4), wherein the protease is at least 2 selected from the group consisting of endoprotease and protease/peptidase complex enzyme;

(7) the agent for reducing blood uric acid level according to item (6), wherein the endoprotease is an acidic protease derived from Aspergillus saitoi, and the protease/peptidase complex enzyme is a protease/peptidase complex enzyme derived from Aspergillus oryzae (Aspergillus oryzae);

(8) the agent for reducing blood uric acid level according to (4), wherein the lipase is bovine-derived premenstrual esterase;

(9) the agent for reducing blood uric acid level according to any one of (1) to (3), wherein the enzymatic decomposition product of cheese is a decomposition product obtained by treating cheese with lipase, protease and lactic acid bacteria;

(10) the blood uric acid level lowering agent according to (9), wherein the lactic acid bacteria are Lactococcus lactis subsp.

(11) The agent for reducing blood uric acid level according to any one of (1) to (10), which comprises a cheese enzymatic decomposition product alone as an active ingredient;

(12) the agent for reducing blood uric acid level according to any one of (1) to (11), which comprises a cheese enzymatic decomposition product and a formulation coadjuvant only;

(13) an active oxygen absorbent contains enzyme decomposition product of cheese as effective component;

(14) the active oxygen absorber according to (13), which contains only a decomposition product of a cheese enzyme as an active ingredient;

(15) a composition comprises enzymatic decomposition product of cheese for reducing blood uric acid level;

(16) the composition of (15), wherein the enzymatic decomposition product of cheese is an enzymatic decomposition product of cheese having an active oxygen absorbing ability;

(17) the composition according to (16), wherein the content of the enzymatic decomposition product of cheese is an amount that shows an active oxygen absorbing ability of 50 μ M or more in an amount equivalent to that of water-soluble vitamin E administered as an amount per 1 day;

(18) the composition according to any one of (15) to (17), wherein the enzymatic hydrolysate of cheese is a hydrolysate obtained by treating cheese with lipase and protease;

(19) the composition as described in (18), wherein the protease is at least 2 selected from the group consisting of endoprotease, exoprotease, exopeptidase/endoprotease complex enzyme, and protease/peptidase complex enzyme;

(20) the composition according to (18), wherein the protease is at least 2 selected from endoprotease and protease/peptidase complex enzyme;

(21) the composition of (20), wherein the endoprotease is an acidic protease derived from Aspergillus saitoi, and the protease/peptidase complex enzyme is a protease/peptidase complex enzyme derived from Aspergillus oryzae;

(22) the composition of (18), wherein the lipase is bovine-derived forestomach esterase;

(23) the composition according to any one of (15) to (17), wherein the enzymatic hydrolysate of cheese is a hydrolysate obtained by treating cheese with lipase, protease and lactic acid bacteria;

(24) the composition of (23), wherein the lactic acid bacteria are lactococcus lactis subspecies lactis, lactococcus lactis subspecies cremoris, and lactococcus lactis subspecies diacetyl biological mutant strain;

(25) the composition according to any one of (15) to (24), which contains only a decomposition product of a cheese enzyme as an active ingredient;

(26) the composition according to any one of (15) to (25), which contains only a decomposition product of a cheese enzyme and a formulation coadjuvant;

(27) a composition comprises enzymatic hydrolysate of cheese for absorbing active enzyme;

(28) the composition according to (27), which contains only a decomposition product of a cheese enzyme as an active ingredient;

(29) a method for lowering blood uric acid level, which comprises administering an enzymatic decomposition product of cheese;

(30) the method of (29), wherein the enzymatic decomposition product of cheese is an enzymatic decomposition product of cheese having an active oxygen absorbing ability;

(31) the method according to (30), wherein the amount of the enzymatic decomposition product of cheese administered is an amount that shows an active oxygen absorption capacity of 50 μ M or more of water-soluble vitamin E as an amount administered per day;

(32) the method according to (29), wherein the enzymatic hydrolysate of cheese is a hydrolysate obtained by treating cheese with lipase and protease;

(33) the method according to (32), wherein the protease is at least 2 selected from the group consisting of endoprotease, exoprotease, exopeptidase/endoprotease complex enzyme, and protease/peptidase complex enzyme;

(34) the method according to (32), wherein the protease is at least 2 selected from the group consisting of endoprotease and protease/peptidase complex enzyme;

(35) the method according to (34), wherein the endoprotease is an acidic protease derived from Aspergillus saitoi, and the protease/peptidase complex enzyme is a protease/peptidase complex enzyme derived from Aspergillus oryzae;

(36) the method of (32), wherein the lipase is bovine-derived forestomach esterase;

(37) the method according to (29), wherein the enzymatic hydrolysate of cheese is a hydrolysate obtained by treating cheese with lipase, protease and lactic acid bacteria;

(38) the method of (37), wherein the lactic acid bacteria are lactococcus lactis subspecies lactis, lactococcus lactis subspecies cremoris, and lactococcus lactis subspecies diacetyl biological mutant strain;

(39) the method according to (29), wherein substantially only the enzymatic decomposition product of cheese is administered as the active ingredient;

(40) the method according to (39), wherein the enzyme decomposition product of cheese and the preparation coadjuvant are administered alone;

(41) a method for absorbing active oxygen, characterized by administering an enzymatic decomposition product of cheese;

(42) the method according to (41), wherein substantially only the enzymatic decomposition product of cheese is administered as the active ingredient.

The present invention also provides the use of the enzyme decomposition product of cheese for producing a uric acid level lowering agent or an active enzyme absorbent in blood.

The agent for lowering blood uric acid level of the present invention has an excellent effect of lowering blood uric acid level, is excellent in taste and high in safety, and therefore can be orally ingested continuously for a long period of time, can control blood uric acid level for a long period of time, and can be used for the prevention and treatment of hyperuricemia and the prevention and treatment of gout, as compared with conventional medicines.

Drawings

FIG. 1 is a graph showing the effect of lowering blood uric acid level of enzymatic decomposition products of cheese.

Detailed Description

The effective component of the blood uric acid level lowering agent and the active oxygen absorbent of the present invention is a cheese enzymatic decomposition product. The enzymatic hydrolysate of cheese is obtained by decomposing fat and protein in cheese by allowing enzyme selected from lipase and protease to act on cheese. More specifically, it is obtained by adding various enzymes derived from microorganisms such as lipase, acidic and neutral proteases, and animal sources to raw cheese, natural cheese after or during ripening, and processed cheese to promote ripening.

The cheese to be used as a raw material for the enzymatic decomposition of cheese may be any of various cheeses including raw cheese, natural cheese and processed cheese. The cheese material is not particularly limited, and milk may be milk of various mammals such as sheep and goats, or human milk may be used as the material. The method for producing the cheese is not limited, and it is needless to say that cheese obtained from raw milk or powdered milk as a raw material and unripe cheese may be fermented or ripened by various microorganisms such as lactic acid bacteria and mold. For example, dada cheese, cheddar cheese, skim cheese, etc. can be used, but are not limited thereto.

As the enzyme for treating cheese, enzymes such as lipase and protease which decompose fat and protein in cheese can be used, but the kind is not limited. As the lipase, not only triacylglycerol lipase existing in gastric juice but also various lipases isolated from human or mammalian body fluids or tissues thereof, insects, plants, bacteria, and the like can be used. As the protease, various proteases isolated from human or mammalian body fluids or tissues thereof, insects, plants, bacteria, and the like, such as pepsin, trypsin, and chymotrypsin secreted from the digestive tract, can be used. In addition, treatment with a protease alone is preferable to treatment with a lipase alone, and treatment with both a lipase and a protease is more preferable.

Proteases include endo-type and exo-type depending on the cleavage site of the amino acid sequence. The endo form cleaves proteins from the middle, and the exo form cleaves amino acids from the C-terminus or N-terminus one by one. The optimum pH and temperature, the proteolytic ability, and the amino acid cleavage site vary depending on the type of protease. From this fact, in the case of the treatment with the protease, it is preferable to combine 2 or more, more preferably 3 or more, proteases having different types such as endo-type, exo-type, proteolytic ability, optimum pH, and optimum temperature.

Among them, 2 or more, particularly 3 or more selected from endoproteases, exoproteases, exopeptidase/endoprotease complex enzymes, and protease/peptidase complex enzymes are preferably used in combination. Further, it is preferable to use 2 or more, particularly 3 or more, selected from endoproteases and protease/peptidase complex enzymes in combination.

Examples of the endoprotease include trypsin (CAS No.9002-07-7, EC3.4.21.4, from bovine pancreas, product No. T8802, SIGMA), pepsin (CASNO. 9001-75-6, EC3.4.4.1, from porcine gastric mucosa, SIGMA), chymotrypsin (Novo, Boehringer), Proleather FG-F (from Bacillus subtilis; Philadelphia), Bioprase (from Bacillus subtilis, Nagase industries), Papain-40 (Philagine), rennin (EC3.4.23.4, Maxiren, from modified yeast Kluyveromyces lactis, GIST-BROCADES N.V.), Alcalasrr (from Bacillus licheniformis, Novo), Molsin F (from Aspergillus saitoi, Kikkoman), Esperase (from Bacillus lentus, Novo), Newlase R (from Bacillus subtilis, Novo), Protamex PTN6.0S, and trypsin (from porcine pancreas), novo), protease S "Amano" G (from Bacillus stearothermophilus, Amano, Inc.), and the like.

Examples of exoproteases include pancreatic carboxypeptidase and aminopeptidase at the brush border of the small intestine.

Examples of the endo/exo-protease complex enzyme include Newlase A (from Aspergillus niger, Setaria), and examples of the exo-peptidase/endo-protease complex enzyme include flavourzyme (from Aspergillus oryzae, Novo), and the like.

Examples of protease/peptidase complexes that can be used include protease A "Amano" SD (from Aspergillus oryzae, Seikagase Corp.), and Umamizyme G (peptidase and protease from Aspergillus oryzae, Seikagase Corp.).

In the present invention, it is preferable to use 2 or more selected from endoproteases and protease/peptidase complexes, it is more preferable to use 2 or more endoproteases and protease/peptidase complexes in combination, and it is particularly preferable to use 3 or more. More specifically, it is particularly preferable to use 2 or more, particularly 3 or more, selected from the group consisting of proteases A "Amano" SD, Molsin, Newlase and Umamizyme G in combination.

Examples of the lipase include Sumizyme NLS (from Aspergillus niger, New Nissan chemical industry), lipase A "Amano" 6 (from Aspergillus niger, Setaria Auricularia.), lipase R "Amano" (from Penicillium rocarpuloides, Setaria Auricularia), and lipase "Pegaster enzyme" (from bovine RenconewZealand), but Pegaster enzyme derived from bovine is particularly preferable.

The conditions for the enzyme treatment are not particularly limited as long as the enzyme is not inactivated. The optimum conditions for the enzyme to be used are preferred from the viewpoint of promoting decomposition, but the pH, reaction temperature, concentration of the enzyme or substrate (cheese), and reaction time in the reaction may be appropriately adjusted depending on the effect of the enzyme to be used in the treatment of cheese and the target enzymatic decomposition product of cheese. When the treatment is carried out using 2 or more enzymes, the treatment may be carried out with all the enzymes at once, or the treatment may be carried out with each enzyme in sequence. The reaction temperature of the enzyme treatment in the present invention is usually suitably 20 to 60 ℃, and preferably 30 to 50 ℃. The reaction time of the enzyme treatment in the present invention is generally about 1 to 8 days, preferably about 2 to 4 days. The amount of the enzyme to be added varies depending on the titer of the enzyme to be used, but is usually preferably 0.01% or more, preferably 0.1 to 10% per unit substrate (cheese).

The protease treatment may be accompanied by fermentation with lactic acid bacteria. As the lactic acid bacterium, lactic acid bacteria belonging to the genus lactococcus can be preferably used. Specific examples of the species of bacteria belonging to the genus Lactococcus include Lactococcus garvieae (Lactococcus garvieae), Lactococcus lactis (Lactococcus lactis), Lactococcus lactis subsp (Lactococcus lactis subsp.lactis), Lactococcus lactis subsp.cremoris (Lactococcus lactis subsp.cremoris), Lactococcus lactis subsp.hominis (Lactococcus lactis subsp.hordinae), Lactococcus lactis diacetyl mutant strain (Lactococcus lactis diacetyl microorganism), Lactococcus pisci (Lactococcus pisciosus), Lactococcus plantarum (Lactococcus plantula), Lactococcus raffinosus (Lactococcus raffinosus), and the like, but are not limited to these examples. In particular, 3 species of lactococcus lactis subspecies lactis, lactococcus lactis subspecies cremoris, lactococcus lactis subspecies lactis diacetyl biological mutant strain are more preferably used. The pH in the enzyme treatment step can be maintained on the acidic side by lactic acid fermentation. In place of lactic acid fermentation, organic acids and inorganic acids commonly used in pharmaceuticals and foods, such as citric acid, gluconic acid, hydrochloric acid, acetic acid, lactic acid, and phosphoric acid, may be used to adjust the pH. When the pH is maintained at the acidic side, the pH is set to 3.5 to 6.0, preferably 4.0 to 5.6.

The enzymatic hydrolysate of cheese used in the present invention is particularly preferably a hydrolysate treated with lipase, protease and lactic acid bacteria. The reaction temperature for these treatments is usually suitably 20 to 60 ℃ and preferably 30 to 50 ℃. The reaction time of these treatments is generally about 1 to 8 days, preferably about 2 to 4 days. The amount of the enzyme to be added varies depending on the titer of the enzyme to be used, but is usually preferably 0.01% or more, preferably 0.1 to 10% per unit substrate (cheese).

The obtained enzymatic decomposition product of cheese has an excellent effect of lowering the blood uric acid level, and is useful as a prophylactic and therapeutic agent for hyperuricemia or a prophylactic and therapeutic agent for gout, as shown in examples described below. In addition, the enzymatic decomposition product of cheese has excellent active oxygen absorbing ability and can be used as an active oxygen absorbent. The excellent blood uric acid level lowering effect of the enzymatic decomposition product of cheese is considered to be related to the active oxygen absorbing ability, i.e., the antioxidant ability.

In the case of the enzymatic decomposition product of cheese, although the enzymatic decomposition product of cheese may be used alone, it is provided in the form of various compositions together with a substantially nontoxic carrier in most cases. Here, the term "substantially nontoxic" means that the composition does not exert a bad effect even when ingested by a human or an animal, and the effect of a decomposition product of cheese enzyme is rarely reduced, and various pharmaceutically or hygienically acceptable carriers can be used. Examples of such carriers (hereinafter, also referred to as formulation adjuvants) include various starches such as corn starch, excipients such as lactose, crystalline cellulose, glucose, mannitol, sucrose, sorbitol, gelatin, gum arabic, calcium hydrogen phosphate, calcium dihydrogen phosphate, sodium phosphate, and sodium carbonate; lubricants such as stearic acid, zinc stearate, calcium stearate, or magnesium stearate; binders such as sucrose, polyethylene glycol, polyvinylpyrrolidone, and the like; pH adjusting agents such as suitable organic acids, inorganic acids or alkali metal (e.g., sodium or potassium) salts thereof, e.g., benzoic acid, citric acid, tartaric acid, succinic acid, phosphoric acid, and hydrochloric acid; a flavoring agent; a flavoring agent; a colorant; a disintegrant; a cosolvent; a suspension; coating agents, and the like. Thus, the enzyme-decomposed cheese product can be provided together with these carriers in the form of an orally administered pharmaceutical composition such as powder, granule, fine granule, dry syrup, tablet, capsule, liquid, or beverage according to a conventional method. Further, other components such as vitamins, minerals, saccharides, amino acids, peptides, etc. may be added in appropriate amounts. The composition of the present invention is preferably a composition containing substantially only a decomposed product of cheese enzyme as an active ingredient, and more specifically, may be a composition containing only a decomposed product of cheese enzyme and a formulation auxiliary adjuvant.

The agent for lowering blood uric acid level of the present invention may be administered simultaneously with other active ingredients, for example, a composition having a blood uric acid level lowering effect different from the action mechanism of the enzymatic decomposition product of cheese, or may be administered as a mixture. Examples of such a composition having an action of lowering a uric acid level in blood with a different mechanism of action include chitosan which adsorbs nucleic acids ingested as food and inhibits the absorption of nucleic acids from the intestinal tract, water-soluble dietary fibers, activated carbon, -polylysine, Urtica dioica which increases urine volume and promotes the excretion of uric acid, an extract of celery seed, lactic acid bacteria which degrades purines ingested from food and reduces the absorption in the body to reduce the uric acid level in serum, and adenine which increases the amount of purine reutilizing enzyme to inhibit the excessive formation of uric acid. In addition, propolis having xanthine oxidase inhibitory activity, Filipendula ulmaria, Cinnamomum cassia, citronella, rhodiola rosea, Alpinia galanga, Myristica fragrans, St.

The agent for reducing blood uric acid level of the present invention can be used not only in the form of a pharmaceutical product but also in the form of a drink or food. For example, it is expected that hyperuricemia can be prevented or treated by taking the extract as a food for special use such as a specific health food or a nutritional functional food. In addition, in the case of preparing a food or drink, the form of liquid, paste, solid, powder or the like is not limited, and the food or drink may be added to various foods (e.g., milk, processed milk, milk drink, refreshing drink, fermented milk, yogurt, cheese, bread, biscuit, cracker, pizza, ice cream, candy, prepared milk powder, liquid food, food for patients, milk powder for infants, food such as milk powder required by lactating women, nutritional food, nutritional supplement food, additive, frozen food, processed food, other commercially available food, etc.) and ingested.

The drink and food of the present invention can be produced by mixing water, protein, saccharide, lipid, vitamins, minerals, organic acids, inorganic acids, organic bases, inorganic bases, fruit juices, flavors, thickeners, and the like with the enzymatic decomposition product of cheese. Examples of the protein include animal and plant proteins such as whole milk powder, skim milk powder, partially skim milk powder, casein, whey protein powder, whey protein, concentrated whey protein, whey protein isolate, α -casein, β -casein, κ -casein, β -lactoglobulin, α -lactalbumin, lactoferrin, soybean protein, egg protein, meat protein, and decomposed products thereof. Examples of the saccharide include saccharides such as sucrose and maltose, processed starch (soluble starch, british starch, oxidized starch, starch ester, starch ether, and the like, in addition to dextrin), dietary fiber, and the like. Examples of the lipid include animal fats and oils such as lard, fish oil, and the like, and fractionated oils thereof, hydrogenated oils, and transesterified oils thereof; vegetable oils and fats such as palm oil, safflower oil, corn oil, rapeseed oil, coconut oil, fractionated oil thereof, hydrogenated oil, and transesterified oil thereof. Examples of the vitamins include vitamin a, carotenes, vitamin B, vitamin C, vitamin D, vitamin E, vitamin K, vitamin P, vitamin Q, nicotinic acid, pantothenic acid, biotin, inositol, choline, and folic acid. Examples of the minerals include calcium, potassium, magnesium, sodium, copper, iron, manganese, zinc, selenium, and whey minerals. Examples of the organic acid include malic acid, citric acid, lactic acid, and tartaric acid, and examples of the inorganic acid include hydrochloric acid and phosphoric acid. Examples of the thickener include agar, gelatin, carrageenan, guar gum, and xanthan gum. Further, various milk-derived components such as butter, whey minerals, cream, whey, non-protein nitrogen, sialic acid, phospholipids, and lactose; peptides such as casein phosphopeptide; arginine, lysine and the like. In the production of the food and drink of the present invention, these ingredients may be synthetic products, natural products, or a large amount of food containing these as a raw material. More than 2 of these ingredients may be used in combination.

The agent for lowering uric acid level in blood of the present invention is administered to a human or an animal showing hyperuricemia or a human or an animal predicted to have an elevated uric acid level, and comprises an effective amount of a decomposition product of cheese enzyme. The administration amount is about 0.01mg/kg to 2000mg/kg, preferably 1mg/kg to 1000mg/kg, and more preferably 10mg/kg to 500mg/kg per day as a cheese enzymatic hydrolysate. The administration amount is appropriately changed depending on the age, weight, and symptoms of the human or animal to be administered, the site to be treated, the kind and amount of the food to be ingested.

In addition, since the enzymatic decomposition product of cheese has an active Oxygen-absorbing action as described above, the dose can be limited based on the active Oxygen-absorbing Capacity (ORAC: Oxygen Radical adsorption Capacity).

The active oxygen absorption capacity is an index of antioxidant efficacy developed by the national aging institute of the national academy of agricultural affairs, usa. In the presence of fluorescent substancesWherein a predetermined active oxygen species is generated, the fluorescence intensity of the fluorescent substance decomposed thereby is measured, and when the fluorescence intensity decreases with time is plotted, the rate of decrease in the fluorescence intensity of the fluorescent substance is retarded if the antioxidant coexists in the reaction system. Based on this principle, the difference (net AUC) between the Area Under the Curve of fluorescence intensity in the presence of the sample (or standard substance) (AUC: Area Under the Curve) and the AUC in the absence of the sample (or standard substance) (blank) was calculated. For the net AUC of the samples, the standard substance (water-soluble vitamin E, Trolox) was determined relative to the known concentration^TM: 6-hydroxy-2, 5,7, 8-tetramethylchroman-2-carboxylic acid; trolox is a registered trademark) of the net AUC. Based on the relative value, the antioxidant effect of the sample was determined by converting the concentration of the water-soluble vitamin E. (unit. mu. mol TE/g: micromole water-soluble vitamin E equivalent/g)

The amount of the agent for lowering blood uric acid level of the present invention to be administered may be an amount which shows an active oxygen absorbing ability of 50. mu.M or more of the equivalent amount of water-soluble vitamin E, preferably 100. mu.M or more of the equivalent amount of water-soluble vitamin E, more preferably 1000. mu.M or more of the equivalent amount of water-soluble vitamin E as an amount to be administered for 1 day.

Examples

The present invention will be described in detail below based on the following examples. It should be noted that the present invention is not limited to the following examples.

[ example 1 ]

(preparation of culture broth of fermentation agent (スタ - タ))

20g of skim milk powder was dissolved in 200ml of distilled water and sterilized to prepare a 10w/v% skim milk powder medium. About 0.1g of 3 kinds of starter bacteria (lactococcus lactis subspecies lactis, lactococcus lactis subspecies cremoris, lactococcus lactis subspecies lactis diacetyl biological mutant strain) were inoculated thereto, and cultured at 37 ℃ for 16 hours.

(preparation of enzymatic hydrolysate of cheese 1)

Sterilized water was added to domestic dada cheese, a starter culture solution and sodium chloride were added thereto, and after stirring, protease a "Amano" SD (protease from aspergillus oryzae, manufactured by tianwazyme) and lipase (pre-gastric esterase, manufactured by renco new zealand) were added and the mixture was shaken at 34 ℃ to decompose the components. After 48 hours, the pH was adjusted to 4.1 with citric acid, and Molsin F (acidic protease from Aspergillus saitoi, Kikkoman Co., Ltd.) and Umamizyme G (protease from Aspergillus oryzae, Nakayase Co., Ltd.) were added thereto, and the mixture was shaken at 34 ℃ to decompose the components. After 4 days, the pH was adjusted to 5.0 with sodium hydroxide and the enzyme was inactivated by heating at 110 ℃ for 15 minutes to obtain a cheese enzymatic hydrolysate. The amount of the cheese component to be blended per 100g of the domestic cheese is as follows.

[ fitting chart ]

Domestic Gaoda cheese 100g

50.5g of sterilized water

18g of starter culture solution

2.2g of sodium chloride

Protease A "Amano" SD 0.6g

Lipase 0.2g

Citric acid 3.5g

Molsin F 0.2g

Umamizyme G 0.3g

Sodium hydroxide 1.5g

[ example 2 ]

(preparation of enzymatic hydrolysate of cheese 2)

Sterilized water was added to domestic cheddar cheese, the starter culture solution prepared in example 1 and sodium chloride were added thereto, and after stirring, protease a "Amano" SD (from aspergillus oryzae, gammazyme) and lipase (RencoNewZealand) were added thereto, and the mixture was shaken at 34 ℃. After 48 hours, the pH was adjusted to 4.1 with citric acid, and Newlase R (from Bacillus subtilis, Novo) and UmamizymeG (protease from Aspergillus oryzae, manufactured by Amazozyme) were added thereto, respectively, and the mixture was shaken at 34 ℃ to decompose the protein. After 4 days, the pH was adjusted to 5.0 with sodium hydroxide and the enzyme was inactivated by heating at 110 ℃ for 15 minutes to obtain a cheese enzymatic hydrolysate. The amount of the cheese in each 100g of the domestic cheddar is as follows.

[ fitting chart ]

Domestic Cheddar cheese 100g

50.5g of sterilized water

20g of starter culture solution

Sodium chloride 0.3g

Protease A "Amano" SD 0.6g

Lipase 0.2g

Citric acid 3.3g

Newlase R 0.2g

Umamizyme G 0.3g

Sodium hydroxide 1.5g

[ example 3 ]

(preparation of enzymatic hydrolysate of cheese 3)

Distilled water was added to Danish defatted cheese (aged for 6 months), the starter culture prepared in example 1 and sodium chloride were added thereto, and after stirring, protease S "Amano" G (from Bacillus stearothermophilus, Ganshire) was added thereto, and the mixture was shaken at 34 ℃ to decompose the components. After 48 hours, the pH was adjusted to 4.1 with citric acid, and Newlase A (from Aspergillus niger, Seamase) and Umamizyme G (from Aspergillus oryzae, Seamase) were added, respectively, and the mixture was shaken at 34 ℃ to decompose the protein. After 5 days, the pH was adjusted to 5.0 with sodium hydroxide and the enzyme was inactivated by heating at 110 ℃ for 15 minutes to obtain a cheese enzymatic hydrolysate. The amount of the defatted cheese blended per 100g of Danish is as follows.

[ fitting chart ]

Danish defatted cheese 100g

74g of distilled water

20g of starter culture solution

Sodium chloride 0.2g

Protease S "Amano" G0.6G

Citric acid 3.2g

Newlase A 0.3g

Umamizyme G 0.3g

Sodium hydroxide 1.4g

Example 4 Effect of cheese enzymatic decomposition products on hyperuricemia caused by Oxazinic acid

(animal experiments)

Rats (Wistar, male, 7 weeks old, Japan SLC) were purchased and fed with AIN-93G powder during the acclimation period. After the acclimation period, blood was collected in the morning and the uric acid level was measured (day 0). Based on the values, the test subjects were divided into a negative group, a control group, a 1 st test subject group, a 2 nd test subject group, and a 3 rd test subject group, and 5 groups (1 group and 6 subjects) were counted. The enzymatic cheese degradation product prepared by the method for producing an enzymatic cheese degradation product according to example 1 was used.

Further, AIN-93G was composed of a standard purified feed for use in Nutrition studies for mice and rats published in 1993 by the American institute of Nutrition (American national institute of Nutrition).

Negative group: feeding AIN-93G, water for injection (10 mL/kg)

Control group: feed AIN-93G mixed with 2.5% Potassium Oxazinate, Water for injection (10 mL/kg)

Test group 1: feed AIN-93G mixed with 2.5% Potassium Oxonate, cheese enzymatic hydrolysate (300 mg/kg, 10 mL/kg)

The 2 nd subject group: feed AIN-93G mixed with 2.5% Potassium Oxonate, cheese enzymatic hydrolysate (1G/kg, 10 mL/kg)

Test group 3: feed AIN-93G mixed with 2.5% Potassium Oxonate, cheese enzymatic hydrolysate (3G/kg, 10 mL/kg)

[ formal rearing (test) ]

From the time of the grouping, the negative groups were fed with AIN-93G and the other groups were fed with AIN-93G mixed with 2.5% oteracil potassium (test diet). The cheese enzymatic hydrolysate was administered 1 time per morning from day 1 to day 15. Partial blood collection was performed on day 2 (2 hours after administration), day 5 (2 hours after administration), day 8 (2 hours after administration), day 12 (2 hours after administration), and day 15 (2 hours after administration) in all the examples, and serum uric acid values were measured by the phosphotungstic acid method. The post-observation period was set from the administration of the enzymatic decomposition product of cheese on day 15 to day 22. Blood was collected in the morning on day 22, and serum uric acid level was measured by phosphotungstic acid method.

[ measurement and examination, etc. ]

Observation of general conditions and measurement of body weight

Body weight measurements were performed on days 0, 1, 5, 8, 12, 15 and 22.

Measurement of food intake and water intake

The food intake was measured on day 0 (set value), day 5 (residual value, set value), day 8 (residual value, set value), day 12 (residual value, set value), day 15 (residual value, set value), day 19 (residual value, set value), and day 22 (residual value). The water intake was measured on day 0 (set value), day 2 (residual value, set value), day 6 (residual value, set value), day 8 (residual value, set value), day 12 (residual value, set value), day 15 (residual value, set value), day 19 (residual value, set value), and day 22 (residual value).

[ statistical treatment ]

The results are expressed as mean ± standard deviation, comparing the control group with each group given the enzymatic decomposition product of cheese. The variance ratio of the numerical test values was tested by F-test, Student's' st test, Dunnett test or Tukey-Kramer test at the same variance, and Mann-Whitney U test at different variances.

[ results ]

The results of the body weight measurement are shown in table 1, the results of the food intake and water intake are shown in table 2, and the transition of the serum uric acid level is shown in fig. 1 and table 3.

TABLE 1

TABLE 2

TABLE 3

During the test, the weight average of each group increased. No significant difference was confirmed between groups (Tukey-Kramer test).

Feeding with the Potassium Oxonate Mixed feed (control group, test group 1, test group 2, and test group 3) showed a decrease in food intake and an increase in water intake as compared with feeding with AIN-93G (negative group). In the test groups 1, 2 and 3 to which the test substances were administered, no significant change in food intake and water intake was observed compared with the control group (Dunnett test).

By feeding with the Potassium Oxonate mixed feed, the uric acid level was significantly increased to become a hyperuricemia state (negative vs control, p <0.001, Student' st-test). The amount of the enzyme decomposition product of cheese administered is dependent on the amount of the enzyme decomposition product to suppress the increase in uric acid level. The uric acid levels were significantly reduced on days 2 and 12 when 1000mg/kg of the enzyme decomposition product of cheese was administered (Dunnett test, p < 0.05), and on days 2,5, 8, 12, and 15 when 3000mg/kg of the enzyme decomposition product of cheese was administered (3 rd test group) (Dunnett test, p < 0.05), as compared with the control group. After 7 days from the start of the administration of the test substance suspension (day 22), no significant difference in uric acid level was observed among the control group, the 1 st test group, the 2 nd test group, and the 3 rd test group.

Example 5 measurement of active oxygen absorbing ability of enzymatic decomposition product of cheese

Active Oxygen absorption Capacity (ORAC) is a new index of antioxidant efficacy developed by researchers in the United states of America of agricultural sciences (USDA) and the national institute of Aging (national institute of Aging) in 1992, and is excellent as a method for analyzing antioxidant efficacy in food, and is now the most abundant analysis method in databases in food raw materials such as vegetables and fruits, and processed foods.

[ measurement ]

The enzymatic decomposition product of cheese to be a sample was prepared to 30mg/mL by adding 8M urea solution, and then the enzymatic decomposition product of cheese was uniformly dispersed by ultrasonic treatment. 2-fold 10-fold dilutions were made with 75mM potassium phosphate buffer (pH 7.4).

In the presence of a labeling substance (fluorescein: 3',6' -dihydroxyspiro [ isobenzofuran-1 [3H ]],9'[9H]-xanthene]-3-one) was added with 75mM potassium phosphate buffer (pH 7.4) to prepare 117nM fluorescein. In the standard substance (Trolox)^TM: 6-hydroxyGroup-2, 5,7, 8-tetramethylchroman-2-carboxylic acid) to 75mM potassium phosphate buffer (pH 7.4) was added to prepare 50. mu.M of water-soluble vitamin E. To a free radical initiator (AAPH: 2,2 '-azobisisobutylamidine dihydrochloride, 2,2' -azo-bis (2-amidinopropane) dihydrate) was added 75mM potassium phosphate buffer (pH 7.4) to prepare 40mM AAPH. To a 96-well fluorescent plate, 20. mu.l of the sample (or 50. mu.M of a water-soluble vitamin E as a standard substance) and 120. mu.l of 117nM fluorescein were added, and preincubated at 37 ℃ for 15 minutes. Then, 40mM AAPH 60. mu.l of a radical initiator was added, and the fluorescence intensity was measured every 1 minute at an excitation wavelength of 485nm and a fluorescence wavelength of 520 nm.

[ analysis method ]

When a predetermined amount of active oxygen species is generated, the fluorescence intensity of the fluorescent substance decomposed therefrom is measured, and the fluorescence intensity that decreases with time is plotted, the rate of decrease in fluorescence intensity of the fluorescent substance is retarded if the antioxidant coexists in the reaction system. Based on this principle, the difference (net AUC) between the Area Under the curve of fluorescence intensity in the presence of the sample (or standard substance) (AUC: Area Under the curve) and the AUC in the absence of the sample (or standard substance) (blank) was calculated. For the net AUC of the sample, the relative value of the net AUC was calculated with respect to the known concentration of the standard substance (water-soluble vitamin E). Based on the relative value, the antioxidant effect of the sample was determined by converting the relative value into a water-soluble vitamin E concentration. (unit. mu. mol TE/g: micromole water-soluble vitamin E equivalent/g)

[ results ]

The active oxygen absorption capacity of the cheese enzymatic hydrolysate is 189.5 +/-11.0 mu M water-soluble vitamin E equivalent/g.

Example 6 production of fermented milk

(production example 1 of fermented milk)

Using enzyme decomposition product of cheese (Taste Concentrate G (Gada type)) and Lactobacillus bulgaricus (L. Bulgaricus) JCM1002^TLike and likeStreptococcus thermophilus (s. thermophilus) ATCC19258, pure yoghurt was prepared. First, Lactobacillus bulgaricus JCM1002 was prepared using a 10% skim milk powder medium^TAnd Streptococcus thermophilus ATCC 19258. Subsequently, the enzyme-decomposed cheese was mixed with a yogurt mix (solid nonfat milk (SNF): 9.5%, FAT component (FAT): 3.0%), and heat-treated at 95 ℃ for 5 minutes. The mixture after the heat treatment was inoculated with 1% each of the ferments of Lactobacillus bulgaricus JCM1002T and Streptococcus thermophilus ATCC19258, and fermented at 43 ℃ for 4 hours to obtain pure yogurt. The pure yogurt was cooled in a refrigerator (5 ℃ C.), and then the taste and physical properties were confirmed. In this case, both the taste and physical properties were good.

Example 7 production of fermented milk

(production example 2 of fermented milk)

Lactobacillus gasseri OLL 2959 (NITE P-224) and Lactobacillus bulgaricus JCM1002 are lactobacillus with blood uric acid level reducing effect by using cheese enzymatic decomposition product (cheese Taste Concentrate G (Gaoda type)), and lactobacillus gasseri^TStreptococcus thermophilus ATCC19258, pure yogurt was prepared. First, a block fermentation agent of Lactobacillus gasseri OLL 2959, Lactobacillus bulgaricus JCM1002T, Streptococcus thermophilus ATCC19258 was prepared using a 10% skim milk powder medium. Subsequently, the enzyme-decomposed cheese was mixed with a yogurt mix (solid nonfat milk (SNF): 9.5%, FAT component (FAT): 3.0%) and heat-treated at 95 ℃ for 5 minutes. The heat-treated mixture was inoculated with Lactobacillus bulgaricus JCM1002^TFermenting with 1% of a starter of Streptococcus thermophilus ATCC19258 and 5% of a starter of Lactobacillus gasseri OLL 2959 at 43 ℃ for 4 hours to obtain pure yogurt. The pure yogurt was cooled in a refrigerator (5 ℃ C.), and then the taste and physical properties were confirmed. In this case, both the taste and physical properties were good.

Example 8 production of processed cheese

(example of production of processed cheese)

The processed cheese was prepared by adding a enzymatic decomposition product of cheese having an active oxygen absorbing capacity of about 150. mu.M equivalent/g of water-soluble vitamin E to a cheese starting material, which was a cheddar cheese produced in New Zealand, and then adjusting the cheese by the following method. The blending ratio is as follows.

[ fitting chart ]

New Zealand cheddar cheese 8.0kg

0.05kg of enzyme decomposition product of cheese

Molten salt (sodium tripolyphosphate) 0.2kg

1.75kg of water (containing steam for warming)

[ production method ]

The cheese is coarsely ground in advance by a meat grinder. The entire amount of the raw material was charged into a 20-liter vessel type kneader (however, the amount of water was set to an amount excluding the amount of steam used for heating), and while stirring at a rotational speed of 120rpm, steam was blown and the temperature was raised to 85 ℃ in about 10 minutes. 200g each of the melted and fluid cheeses were collected in a container, sealed and cooled overnight in a refrigerator at 5 ℃. The product has no peculiar smell of cheese enzymatic decomposition product, and has good taste and composition.

Claims (16)

1. Use of enzymatic decomposition product of cheese, which is obtained by treating cheese with lipase and protease, for producing agent for lowering uric acid level in blood,

the protease is the combination of endoprotease and protease/peptidase complex enzyme,

the lipase is bovine-derived forestomach esterase,

the addition amount of the lipase and the protease is 0.1-10% per unit substrate.

2. The use according to claim 1, wherein the enzymatic cheese hydrolysate is an enzymatic cheese hydrolysate having active oxygen absorbing ability.

3. The use according to claim 2, wherein the content of the enzymatic decomposition product of cheese is an amount that shows an active oxygen absorption capacity of 50 μ M or more in terms of the amount of water-soluble vitamin E administered as 1 day.

4. The use of claim 1, wherein the endoprotease is an acidic protease derived from Aspergillus saitoi, and the protease/peptidase complex enzyme is a protease/peptidase complex enzyme derived from Aspergillus oryzae.

5. The use according to any one of claims 1 to 3, wherein the enzymatic cheese hydrolysate is a hydrolysate obtained by treating cheese with lipase, protease and lactic acid bacteria.

6. The use according to claim 5, wherein the lactic acid bacteria are lactococcus lactis subspecies lactis, lactococcus lactis subspecies cremoris, and lactococcus lactis subspecies diacetyl biological mutant strains.

7. The use according to any one of claims 1 to 3, wherein only the enzymatic decomposition product of cheese is contained as an active ingredient.

8. The use according to any one of claims 1 to 3, wherein the cheese enzymatic hydrolysate and the formulation coadjuvant are contained only.

9. Use of an enzymatic decomposition product of cheese, which is a decomposition product obtained by treating cheese with a lipase and a protease, in the preparation of a composition for lowering uric acid level in blood,

the protease is the combination of endoprotease and protease/peptidase complex enzyme,

the lipase is bovine-derived forestomach esterase,

the addition amount of the lipase and the protease is 0.1-10% per unit substrate.

10. The use according to claim 9, wherein the enzymatic cheese hydrolysate is an enzymatic cheese hydrolysate having active oxygen absorbing ability.

11. The use according to claim 10, wherein the content of the enzymatic decomposition product of cheese is an amount that shows an active oxygen absorption capacity of 50 μ M water-soluble vitamin E equivalent or more as an administration amount for 1 day.

12. The use of claim 9, wherein the endoprotease is an acidic protease derived from Aspergillus saitoi and the protease/peptidase complex enzyme is a protease/peptidase complex enzyme derived from Aspergillus oryzae.

13. The use according to any one of claims 9 to 11, wherein the enzymatic cheese hydrolysate is a hydrolysate obtained by treating cheese with lipase, protease and lactic acid bacteria.

14. The use according to claim 13, wherein the lactic acid bacteria are lactococcus lactis subspecies lactis, lactococcus lactis subspecies cremoris, and lactococcus lactis subspecies diacetyl biological mutant strains.

15. The use according to any one of claims 9 to 11, wherein only the enzymatic decomposition product of cheese is contained as an active ingredient.

16. The use according to any one of claims 9 to 11, wherein the composition comprises only a cheese enzymatic hydrolysate and a formulation coadjuvant.