TW201605463A - The anti-fatigue of black garlic health product - Google Patents

Abstract

The present invention discloses can be used to anti-fatigue of black garlic food. By orally ingesting the anti-fatigue of black garlic the present invention, and therefore the fatigue of muscles be exercise can be controlled, and as a results for improving the ability of exercise can be expected.

Description

Black garlic health care product for resisting fatigue

A black garlic food that can be used to resist fatigue, thereby relieving muscle fatigue caused by exercise, oxidative damage, and black garlic food and its use for improving the activity of antioxidant enzymes in living organisms.

The work and life of today's society bring fatigue and stress to human beings. The proportion of people who are over-tired is gradually expanding. Excessive fatigue not only affects the effectiveness of work, but even harms the health of mind and body. The so-called "overwork death" incident often occurs in the society, and the cause is chronic fatigue, which is excessive labor. Therefore, if you can clearly understand the length of the fatigue caused by excessive fatigue, induce the cause, adjust your life pace, and appropriate rest or timely relief pressure and nutritional supplements, the occurrence of excessive fatigue will be improved.

In recent years, many studies have confirmed that excessive exercise can cause changes in free radical metabolism, which in turn affects the balance of oxidation and anti-oxidation in the human body. When too much reactive oxygen species (ROS) is produced in the body, oxidative stress is formed, which leads to lipid peroxidation (LPO), which destroys the cell membrane and causes cell aging or death. . Therefore, the increase of free radical metabolism is one of the mechanisms of exercise fatigue.

Black garlic is made by fermenting garlic through a special process. Black garlic transforms the stimulating allicin into a garlic-free, low-irritant S-allyl cysteine component. Eating black garlic does not produce an unpleasant tone and stimulates the gastrointestinal sensation. At the same time, S-ene The propylcysteine component also has the effect of preventing myocardial infarction, cerebral infarction and arteriosclerosis. Black garlic is more than 10 times more polyphenolic than raw garlic, and its antioxidant capacity is greatly enhanced; SOD activity is 10 times that of common garlic; free amino acid content is 1.5 times that of raw garlic, which is easier for the body to absorb; Sweet and fruity, it will not produce an unpleasant tone after eating. It is a pure green food without any additives, rich in nutrients and delicious. Professor Sasaki of the Faculty of Medicine, Hirosaki University, Japan - The white rat test proved that the active ingredients of black garlic can improve immunity. In the comparison of more than 300 foods with antioxidant function, black garlic has the highest antioxidant capacity, and its effective anti-cancer ingredients or other effective sulfur-containing compounds will increase three to five times and the antioxidant power will increase tenfold. the above. The Republic of China Patent Application No. 101139793 discloses that "a new use of black garlic food which can increase the activity of an enzyme having alcoholic liver damage" is a technical product of the anti-oxidation ability of black garlic. CN 100506078 C discloses a method for bioprocessing of garlic, which is confirmed by animal experiments to have an anti-fatigue effect, but the invention does not disclose how much food is needed to achieve this effect. CN 101731606 A also discloses a black garlic health product that reveals that anti-aging effect can be achieved by ingesting black garlic.

The invention is modified with reference to the anti-fatigue function evaluation method announced by the Department of Health. The rat swimming endurance test is used as a model, and the black garlic is fed by oral administration, and the changes of the biochemical reaction values in the blood before and after the exercise are tested. The invention discloses that the rats in the black garlic sports group can effectively alleviate the muscle fatigue caused by the exercise process, and reduce the oxidative damage to the living body during the exercise process and improve the anti-oxidase activity in the living body to achieve the anti-free radical effect.

First, the preparation process of black garlic extract

The black garlic was crushed and then extracted with 70% alcohol (1:10, w/v) at 70 ° C for 6 hours. The filtrate was concentrated under reduced pressure and lyophilized to a powder. The powder was called black garlic extract. It can obtain 10.25±0.85g of lyophilized powder per 100g of black garlic, and the extraction rate is about 10%, that is, the lyophilized powder of about 1g of extract powder is used to convert about 10g of black garlic.

The extract may be prepared into a pharmaceutical composition by the addition of an excipient which is generally used in the prior art, wherein the excipient comprises a lyophilized powder utilizing the extract and a conventional oral dosage form excipient comprising: bonding Agents, disintegrating agents and lubricants. Adhesives suitable for pharmaceutical compositions and dosage forms: corn starch, potato starch or other starches, sodium alginate, natural and synthetic gums, powdered tragacanth, guar gum, cellulose and its derivatives, ethyl cellulose , cellulose acetate, calcium carboxymethyl cellulose, sodium carboxymethyl cellulose, microcrystalline cellulose, and a mixture of their excipients, the components used in the coating layer include: methyl cellulose (Methyl cellulose), Hydroxymethyl propyl cellulose, Hydroxymethyl propyl cellulose, Polyvinyl alcohol, polyvinyl pyrrolidone, Cellulose acetate phthalate or other material suitable for use in the present invention, methyl cellulose to prepare a pharmaceutical composition.

The black garlic pharmaceutical composition of the present invention may contain a pharmaceutically acceptable additive.

Second, the analysis of black garlic extract

The black garlic extract was analyzed by total phenolics using Folin-Ciocalteau reagent and found to be 232±0.3 mg/g; flavonoids analysis was performed by (2-aminoetheyl) diphenylborate analysis, and the result was 35.5±0.2. Mg/g.

Second, animal experiments

The experimental animals were normal ten-week male Sprague-Dawley (SD) rats. The environmental conditions of the animal breeding room are to control the light and dark of the light according to the sputum and night circulation in nature; the temperature is controlled at 24±2°C, and the humidity is controlled at 50±10%.

The experimental mice were divided into 5 groups on average: control group (not tested for swimming ability and not eating black garlic), exhaustive exercise group (testing for swimming ability and not eating black garlic); in addition, the three groups of black garlic swimming group were Ingest low-, medium-, and high-dose black garlic (testing swimming ability). All experimental animals were calculated according to the group's feed intake (with and without black garlic), and were fed with fixed tubes. In order to avoid the lack of heat caused by excessive labor in the experimental animals, the additional supply of black garlic-free feed was in the animal cage. In order to ensure the source of activity energy of experimental animals.

The experimental animal input dose is designed to be about 5g/kg/day (low dose), 10g/kg/day (medium) Dosage) and 15g/kg/day (high dose). The black garlic dosage conversion algorithm is based on the adult weight of 60 kg divided by the daily recommended intake of the test substance, the recommended intake per kilogram of body weight, and multiplied by the metabolic coefficient of the experimental animal relative to the human body (the mouse relative to the The metabolic coefficient of the human body is 6.25), and the daily intake dose of the experimental animal can be obtained. It is expressed by the formula: the intake dose per kilogram of body weight of the rats = the recommended intake of the human body, the weight of 60 kg × 6.25. The equivalent dose for converting adult weight (60 kg) is approximately 50 g/day (low dose), 100 g/day (medium dose), and 150 g/day (high dose).

After the mice were prepared for feeding, blood samples were collected for analysis, and the rats were sacrificed at the end, and the liver and blood were taken for analysis.

After the mice were prepared for feeding, blood samples were collected for analysis, and the rats were sacrificed at the end, and the liver and blood were taken for analysis.

Third, exercise endurance measurement - swimming test

The evaluation method used in the present invention is mainly modified by referring to the anti-fatigue function evaluation method announced by the Department of Health, and the exercise ability test experiment mode is a swimming test. Different doses of black garlic were used in the study, and mice were fed daily with a feeder. Before the experiment, it was necessary to fast for 12 hours. The experiment was carried out by swimming the rats into water with a water temperature of 25±1°C, a diameter of 60cm and a water depth of 40cm, and began to count until the head of each single mouse was completely submerged. It still can't float on the water in 10 seconds, that is, stop timing.

Fourth, blood collection and blood treatment

After the swimming test, the mice were picked up and rested for 60 minutes. Together with the control group, 2-3 mL of blood was collected from the eye sockets of the rats, and 1 to 1.5 c.c. was loaded into the test tube containing EDTA. Shake to prevent coagulation, and the remaining blood was placed in a test tube containing no EDTA. After standing, it was centrifuged at a rate of 3000 × g per minute for 10 minutes to take serum. The serum collected by each test group was completely reacted with the kit reagent and then measured by a biochemical automatic analyzer.

V. Determination of total polyphenol content and antioxidant capacity in serum Serum pretreatment

The protein was precipitated by adding 20% TCA (0.12 mL/1 mL serum) to the serum, and centrifuged at 12000 × g for 45 minutes, and the supernatant was obtained for subsequent analysis.

(1) Determination of total polyphenols in serum

Take 20 μL of serum and add 200 μL of sodium carbonate (2%, w/v) for two minutes. Add 10 μL of Folin-Ciocalteau reagent (50%, v/v) and mix well for two hours to use ELISA reader (Molecular Devices, VMax). The absorbance at 650 nm was measured. Use Gallic acid as the calibration curve and convert its equivalent concentration.

(2) The total antioxidant capacity of the hormone (Trolox equivalent antioxodant capacity)

1.5 mL of deionized water, 0.25 mL of ABTS (1000 μM), 0.25 mL of H ₂ O ₂ (500 μM), and 0.25 mL of Peroxidase (40 units/mL) were uniformly mixed. After one hour of reaction, the samples were subjected to blue-green ABTS ⁺ . After the cationic free radicals, 0.25 mL of serum was added, and after lapse of ten minutes, 650 absorbance was measured by E ELISA reader (Molecular Devices, VMax). Trolox was used as a calibration curve to convert its equivalent concentration.

Sixth, blood urea nitrogen test

After 30 minutes of last feeding, do not bear weight swimming in water at a temperature of 25 ± 1 ° C. Minutes, after 60 minutes of rest, blood was collected and serum was taken for use. Using the principle of colorimetric assay, urase reaction was added to the quantitative serum, then ammonia was added as a coloring agent, and a red compound was produced after the action, and the absorbance was measured at a wavelength of 660 nm, and then the blood urea nitrogen concentration was converted.

Seven, blood sugar test

After the last feeding for 30 minutes, the rats were allowed to swim for 90 minutes in the water at a temperature of 25 ± 1 ° C. After 60 minutes of rest, blood was collected and serum was taken for use. Using the principle of enzyme action and colorimetric determination, after adding glucose oxidase to the quantitative serum, 4-aminoantipyrine and 1,7-dihydroxynaphthalene were added. After peroxidase reaction, a red compound was produced, and the absorbance was measured at 555 nm. Converted to its concentration.

Eight, blood lactate test

After 30 minutes of the last feeding, the weight was stopped after swimming for 10 minutes in water at a temperature of 25 ± 1 °C. Blood was collected after swimming and after 20 minutes of rest. After the blood is treated, the lactate oxidase reaction is added to the quantitative serum by the action of enzyme action and colorimetric determination, and then 4-aminoantipyrine and 1,7-dihydroxy-naphthalene are added, and a red compound is produced by peroxidase at 540 nm. The absorbance is measured at a wavelength, and the concentration of lactic acid is converted.

Nine, liver glycan analysis method

After 60 minutes of the last feeding, the SD rats were stunned with CO2, and the liver was immediately dissected, rinsed repeatedly with physiological saline, and then blotted dry with filter paper. Accurately weigh 0.45 g of liver, add 2 mL of 30% KOH, heat at 100 ° C for 20 minutes, remove 200 μl of homogenate into each tube after heating, add 1 mL of absolute ethanol to each tube, and centrifuge at 4000 × g for 10 minutes. , the upper layer is poured off, the precipitate is added to 0.5mL DDW, and finally added After fully mixing with 1 mL of antherone reagent, the absorbance was measured at 620 nm using the principle of enzyme action and colorimetric measurement, and the concentration was further converted.

Nine, the antioxidant state of red blood cells

Antioxidant enzymes include: superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GRD), reduced glutathione/oxidized glutathione (GSH/ GSSG). Red blood cells obtained from whole blood centrifugation were washed three times with physiological saline at 4 ° C. Then, 100 μL of red blood cells were taken into a 1.5 mL microtube, and 300 μL of deionized water was added to shake at 4 ° C. After centrifugation at 4 ° C and 12000 rpm for 10 minutes. The supernatant was taken for the following analysis (1) to (5), and total hemoglobin was used for quantification.

(1) Red blood cell SOD activity

Antioxidant enzyme SOD activity was measured using a commercially available reagent set (SD 125; Randox Laboratories, Antrim, UK). After appropriate dilution of various tissue supernatants treated with antioxidant enzyme extraction, 50 μL of sample dilution or different active SOD standard solution, 1.7 mL of reaction substrate solution (50M xanthine, 25M INT), 250μL of enzyme solution (80U/L xanthine oxidase), after mixing, take 1mL into the quartz colorimetric tube, record the change of absorbance within 3 minutes at 37 ° C, wavelength 505nm, the lower the change in absorbance value means The higher the inhibition rate of the reaction, the higher the activity of SOD. The measured value is substituted into the standard curve, multiplied by the dilution factor of the sample, and finally quantified by the hemoglobin contained, and the specific activity (U/g Hb) of the SOD is calculated.

(2) red blood cell CAT activity

The change in antioxidant enzyme CAT activity was measured according to the method of Beers and Sizer (1952) (Beers and Sizer, 1952). After appropriately diluting the supernatant treated with the antioxidant enzyme extraction, take 30 μL of the sample diluent, 570 μL of the secondary water, 300 μL of the reaction solution (59 mM H 2 O 2 in 50 mM potassium phosphate, pH 7.0), mix and take 1 mL was added to a quartz colorimetric tube, and the change in absorbance value was recorded at a wavelength of 240 nm for 3 minutes. The CAT activity unit (1 U) is defined as a 1 mole reduction in H2O2 per minute. Substituting the measured value into the formula, [(△A240nm/min)×1000]/43.6, multiplying by the dilution factor of the sample, and finally quantifying with the hemoglobin contained, the specific activity of CAT can be calculated (U/g Hb)

(3) Red blood cell GRD activity

Analysis of antioxidant enzyme GRD activity was measured using a commercially available reagent set (GF2368; Randox Laboratories, Antrim, UK). After appropriately diluting the supernatant treated with the antioxidant enzyme extraction, take 40 μL of the sample diluent, 1 mL of the mixed reagent (2.2 mM GSSG), 200 μL of 0.17 mM NADPH, and sequentially add the quartz colorimetric tube. After mixing, the change in absorbance was recorded at 37 ° C and a wavelength of 340 nm for 5 minutes. The GRD activity unit (1 U) is defined as a 1 mole reduction in NADPH per minute. The specific value of the GRD (U/g Hb) can be calculated by substituting the measured value into the formula 4983 × ΔA 340 nm / min, multiplying by the dilution factor of the sample, and finally quantifying with the contained heme.

(4) Red blood cell GSH/GSSG ratio

First, the oxidized form of GSH (ie, GSSG) in the sample is reduced to GSH, and finally the concentration of all GSH in the sample is determined. Total GSH content analysis is based on Tietze Method (1969) (Tietze, 1969). Take 10 μL of sample solution or different concentrations of standard solution (0~100M GSH) into 96 well assay plate, add 95 μL of No. 1 reagent (2U/nL glutathione reductase, 200M NADPH, 2mM EDTA in 50mM phosphate buffer, pH 7.2), after mixing evenly, add No. 2 reagent (10 mM DTNB in 50 mM phosphate buffer, pH 7.2), and read the change of absorbance within 5 minutes at a wavelength of 405 nm with a microplate reader (Labsystem Multiskan RC, Finland). The concentration of total GSH in the sample solution was calculated according to the standard curve.

The analysis of the GSSG content was performed according to the method of Griffith (1980) and then determined (Griffith, 1980). First, take 70μL of sample solution or different concentration of standard solution (0~100M GSSG) to 96 well analysis plate, add 4μL of M2VP solution, and let stand for 1 hour at room temperature. This step is to transform GSH in the reduced state. Into other derivatives. Then, take 10 μL of the sample solution after the end of the reaction to another new 96 well assay plate, and add 95 μL of No. 1 reagent (2 U/mL glutathione reductase, 200 M NADPH, 2 mM EDTA in 50 mM phosphate buffer, pH 7. 2) After mixing well, add No. 2 reagent (10 mM DTNB in 50 mM phosphate buffer, pH 7.2), and read the change of absorbance within 5 minutes at a wavelength of 405 nm with a microplate reader (Labsystem Multiskan RC, Finland). The concentration of GSSG in the sample solution was calculated according to the standard curve.

Calculation of GSH/GSSG ratio: GSH/GSSG ratio=(total GSH-2GSSG)/GSSG

X. Statistical analysis

The experimental data were analyzed by SPSS/PC statistical analysis software for analysis of variance (ANOVA). The differences between the experimental groups were tested. If there were differences, Duncan’s multiple range test was used for further analysis. In addition, Student T-test was also used to compare the differences between the experimental groups.

Experimental result First, swimming test

The improvement of exercise endurance is the most direct and objective indicator of strengthening anti-fatigue ability. The length of swimming time can reflect the degree of fatigue of animals. In this study, after the feeding of black garlic, the swimming exhaustion test was carried out. The results are shown in Table 1. The swimming group of the black garlic swimming group has significantly improved the swimming tolerance time compared with the exhaustive exercise group. This phenomenon indicates that the feeding black The garlic head can effectively improve the aerobic exercise and delay the fatigue of the mouse. The reason is that the black garlic has the antioxidant ability to effectively reduce the fatigue of the muscle during exercise.

Second, the biochemical reaction value test results

The results in Table 2 show that the total polyphenol content in the serum of the ingested black garlic swimming group increased compared with the exhaustive exercise group with the prolonged feeding time.

The results in Table 3 show that the total antioxidant capacity in the serum of the ingested black garlic swimming group increased compared with the exhaustive exercise group with prolonged feeding time.

◎Ingestion of black garlic to reduce muscle fatigue test during exercise

Blood urea nitrogen is a metabolite of protein and amino acid. Under normal physiological conditions, nitrogenous substances such as proteins and amino acids are catabolized, first removing amino groups, ammonia is converted into urea in the liver, and blood urea nitrogen is passed through the blood. The circulation is excreted from the kidneys. Blood urea nitrogen is related to biochemical indicators such as bodily functions and fatigue, and can be used as one of the criteria for assessing the intensity of exercise load. The present invention discloses that the blood urea nitrogen concentration in the blood of the black garlic swimming group is changed before and after swimming as shown in Table 5. In Table 5, there is a significant difference in blood urea nitrogen values between the black garlic swimming group and the exhaustive exercise group (p<0.05). From the rising rate, it can be clearly seen that the black garlic has reduced motion. The ability of blood urea nitrogen, and the rate of increase of urea nitrogen in high-dose black garlic rats was comparable to that of the control group.

Blood sugar itself is available as a nutrient for cells and is a major source of cellular energy. The blood sugar concentration of the human body needs to be maintained within a certain range. When the blood sugar is too high or too low, it will have a certain degree of influence on the body. Therefore, during a long period of intense exercise, it may lead to the exhaustion of liver sugar in the muscle, causing fatigue and reducing exercise performance, thereby forming a phenomenon of hypoglycemia and affecting the central nervous system. From the results in Table 5, it was found that the ingested black garlic swimming group compared with the exhaustive exercise group, the former slowed down the blood sugar caused by the exhaustion exercise, and the result showed that feeding black garlic helps to increase the tolerance of the rats.

Lactic acid is an indicator of the intensity of exercise training and is often used in the laboratory as a measure of the degree of anaerobic and aerobic metabolism. During exercise, the production of lactic acid, which spreads from the active muscle cells into the blood, is a product of energy metabolism during muscle activity. Because the accumulation of lactic acid interferes with the conduction of nerve impulses and the contraction of muscles, and these disturbances lead to fatigue, blood lactic acid is closely related to the load intensity, so blood lactic acid is one of the important indicators for judging exercise intensity or fatigue, and also represents after exercise. Fatigue and recovery (Gladden LB, 2001). The concentration of lactic acid during exercise depends on the intensity of exercise, the amount of muscle applied, and the length of exercise time. During intense exercise, the skeletal muscles contract and the muscle glycogen is rapidly decomposed, causing the lactic acid and blood lactic acid to increase rapidly, and the accumulation of lactic acid in the body is excessive, thereby dissociating the hydrogen ions (H ⁺ ) and lowering the blood pH. Increased hydrogen ion concentration inside the muscle, affecting the release of Ca ²⁺ in the sarcoplasmic reticulum and reducing the contractility of muscle fibers, reducing the phosphofructoside (PFK) activity of glycolysis, and reducing the ability of muscle cells to produce ATP, which is motility One of the important reasons for fatigue, the elimination of lactic acid is beneficial to the relief of fatigue (Yu Qian, 1999).

The present invention discloses the rate of change of lactic acid concentration in the blood of mice after different swimming doses of black garlic, respectively, as shown in Table 5. It can be seen from the table that the results of the low, medium and high feeding doses can significantly inhibit the increase of lactic acid concentration in the blood and achieve statistical difference (p<0.05), and have a dose effect. Therefore, it is known that the black garlic head significantly reduces the blood lactic acid content after exercise, so that the body acidity is lowered, and the fatigue after exercise is reduced, indicating that the black garlic has a good effect in reducing physical fatigue.

Hepatic glycogen is the main source of energy in energy metabolism. The amount of hepatic glycogen in the organism itself can reflect its anti-fatigue ability. When the liver stores more glycogen, its ability to withstand fatigue is stronger. Therefore, it can be seen from Table 5 that low-medium-high doses of exercise-type mice can effectively store hepatic glucose in the liver. This phenomenon indicates that rats fed black garlic help liver glycogen stored in the liver and glucose during exercise. The form is released into the blood to maintain a constant blood sugar level, effectively improve the aerobic exercise of the mouse, reduce the blood lactate value, and delay the appearance of fatigue.

◎Ingestion of black garlic reduces the oxidative damage to the organism during exercise and increases the activity of antioxidant enzymes in the body during exercise

From the experimental results, it was found that exercise caused superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GRD), reduced glutathione/oxidized glutathione in mouse red blood cells. The ratio of (GSH/GSSG) was significantly reduced (see Table 6), indicating that exercise caused an imbalance in the antioxidant state in the body. Past studies have shown that exercise causes a large amount of free radicals in the body, which further increases the oxidative stress (Groussard et al., 2003; Margaritis et al., 2003). On the other hand, during intense exercise, GSH is oxidized to form GSSG (Voces et al., 2004), and exercise also causes an increase in GSSG and a decrease in GSH in red blood cells (Bloomer et al., 2005). These findings are consistent with the results of the reduced GSH/GSSG ratio caused by exercise in this experiment. In addition, the present invention discloses that a mouse fed a black garlic does have an ability to increase the antioxidant enzyme of red blood cells, and its antioxidant capacity is positively correlated with the amount of black garlic consumed by the mouse.

The black garlic of the present invention or an extract thereof can be appropriately administered to an animal such as a human, a dog, a cat, a pig, a cow, a sheep, or other mammal. The route of administration may include an oral or similar route of administration, preferably for oral administration, taking into account convenience.

The black garlic or the extract component thereof of the present invention may be solvent extracted (including, for example, water, alcohol, acetone, etc.), but considering convenience and consumer habits, it is preferably directly orally, and after commercialization, it is made into an aqueous solution, syrup, capsule. There are no restrictions on the type of tablet and the like.

The black garlic pharmaceutical composition of the present invention can be used as a medicine or health food for slowing down muscle fatigue, oxidative damage caused by exercise, and increasing antioxidant enzyme activity in a living body.

The black garlic food of the present invention may be an anti-fatigue health food.

The specific embodiments of the present invention are described in detail below, but the following embodiments are only used to further disclose the technical content of the present invention, and should not limit the scope of the invention.

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Claims (9)

An anti-fatigue black garlic food is used in an amount of at least 50 grams per day for more than 7 days of continuous feeding. A black garlic food for relieving muscle fatigue, oxidative damage caused by exercise, and increasing the activity of antioxidant enzymes in the living body, and the dosage is at least 50 grams per day for more than 7 days. A use of black garlic for the preparation of a pharmaceutical composition for relieving muscle fatigue, oxidative damage and enhancing the activity of antioxidant enzymes in the body during exercise. A black garlic use for preparing a pharmaceutical composition for slowing down muscle fatigue during exercise and enhancing blood urea nitrogen metabolism during exercise. A black garlic use for preparing a pharmaceutical composition that slows down muscle fatigue during exercise and maintains blood glucose constant during exercise. A black garlic use for preparing a pharmaceutical composition for slowing the production of lactic acid produced by muscles during exercise. A black garlic use for preparing a pharmaceutical composition for reducing fatigue of muscles during exercise and for increasing liver glycogen content. For example, the black garlic use of the scope of claims 3-7 is used for at least 50 grams of continuous feeding for more than 7 days per day. A black garlic use for preparing a pharmaceutical combination for enhancing superoxide dismutase activity, enhancing glutathione reductase activity, and reducing reduced glutathione/oxidized glutathione (GSH/GSSG) ratio The amount of the food is at least 50 grams per day for more than 7 days of continuous feeding.