KR20060007104A - Organic constituents of wood vinegar from konara having the effects of cure and prevention of cancer, antibacterial and antioxidant - Google Patents

Abstract

The present invention relates to woody vinegar organics having an excellent effect on cancer treatment and prevention, antibacterial and antioxidant.

The present invention (a) carbonizing oak chips to capture the smoke at a predetermined flue temperature to extract the wood vinegar, excluding the solids and tar tar from the extracted wood vinegar, and aged for a predetermined period of time while leaving the excluded wood vinegar Wood liquor carbonization process; (b) a wood liquor distillation step of removing harmful low-boiling components from the carbonized wood liquor by distillation under reduced pressure, adsorbing and filtering the distilled distilled liquor to separate egg separation impurities, and obtaining wood liquor from the filtrate; (c) adding an organic solvent selected from methylene chloride or ethyl acetate to the wood vinegar, which has been subjected to the distillation, and shaking the mixture to first separate the organic solvent layer and the wood vinegar layer, and to the wood vinegar layer, an organic solvent selected from methylene chloride or ethyl acetate. Adding and then shaking to separate the organic solvent layer and the wood vinegar layer to obtain the first and the second organic solvent layer wood vinegar organic material extraction step; And (d) an organic solvent removing step of removing the organic solvent from the mixed organic solvent layer.

The present invention can prevent cancer by treating cancer or at the same time inhibiting mutations, and has an effect that can be used as an additive in health food by utilizing antibacterial and antioxidant effects.

Wood vinegar, organic matter, anticancer effect, antimutagenic effect, antibacterial effect, antioxidant effect

Description

Organic constituents of wood vinegar from Konara having the effects of cure and prevention of cancer, antibacterial and antioxidant}

1 is a graph showing the antimutagenic activity of the mutant MNNG of wood vinegar organic matter according to the present invention,

Figure 2 is a graph showing the antimutagenic activity for mutant NQO of the wood vinegar organic matter according to the present invention,

3 is a graph showing the lipid peroxidation inhibitory effect of wood vinegar organics according to the present invention,

4 is a graph showing the DPPH radical scavenging effect of the wood vinegar organic matter according to the present invention.

The present invention relates to a wood vinegar organic matter having an excellent effect on cancer treatment and prevention, antibacterial and antioxidant, and more particularly, purified wood vinegar in high purity, and then extracted organic matter from the purified wood vinegar solution is excellent anti-cancer effect, The present invention relates to a pharmaceutical composition or a dietary supplement comprising a wood-based organic matter having an excellent effect of inhibiting mutation, which is a major inducing factor, and having an antimicrobial activity against various kinds of strains, an inhibitory effect on lipid peroxidation, and a radical scavenging effect. .

Wood vinegar is a crude wood vinegar, which collects and cools the smoke generated by the pyrolysis polymerization of wood in the process of directly carbonizing oak wood. The carbonization temperature is important for obtaining good wood vinegar. It is known that the smoke (gas) produced at the temperature of less than 150 ℃ produced.

And, when aged and aged for 6 months or more in this state, the crude oil in the upper layer, middle vinegar in the middle layer, tarryul in the lower layer is divided into three layers of water, soluble in water in the middle layer is called wood vinegar solution for agricultural, industrial as well as pharmaceutical It is widely used for health drinks.

Usually, 80 ~ 90% of wood vinegar is water, and 10 ~ 20% is organic acid and acidic liquid whose main component is acetic acid. It has a smell of phosphorus.

In addition, wood vinegar is widely used as a folk remedy in Korea, Japan, China, etc. It is mainly known that it has a liver protection effect, anti-diabetic effect, alcohol detoxification effect, anti-cancer effect, etc. Hundreds are registered, but mainly developed for agriculture (pesticide substitute, fertilizer mixture, etc.) or industrial (deodorant, fungicide, pollutant decomposition, food additives, etc.). Etc. are known.

In addition, studies on the anticancer effect of wood vinegar as a study on the immunomodulatory effect and anticancer effect of oak vinegar [Kim Dong-Hee et al. Effect of Wood Vinegar and Anticancer Herbal Compounds on Anticancer and Antimetastatic Activity. Korean Journal of Oriental Physiology and Pathology 2001, 15 (1), 136-142] and a study on the anticancer effects of extracts from several kinds of anticancer herbal medicines with wood vinegar [Kim, Dong-Hee et al. A Study on the Immunoregulatory and Anticancer Effects of Oak Tree Vinegar The Korean Journal of Oriental Physiology and Pathology 2001, 15 (6), 881-886.

However, until now, there are no known cytotoxicity studies using various cancer cells, antimutagenic activity, antimicrobial activity and antioxidative activity against organic matter of wood vinegar.

In addition, cancer is one of diseases increasing with the development of civilization, and methods for treating the same are known as surgical surgery, radiation therapy, chemotherapy by administration of anticancer substances, and the like. However, most of the treatment methods are limited to the treatment of early cancer patients or specific cancers, and death due to cancer is steadily increasing. In particular, radiation therapy or chemotherapy causes many side effects.

Therefore, there is a demand for the development of anticancer drugs that can increase anticancer rates while reducing side effects, and as an alternative, attempts have been actively made to develop anticancer drugs from conventional natural products, particularly herbal medicines.

For example, in the case of Salvia miltiorrhiza, 18 terpene compounds isolated from the methylene chloride fraction show anticancer effects, and have ED ₅₀ values of about 20 to 500 ㎍ / ml depending on the cancer cells. (Planta Medica 63, 1997, Georg Thieme Verlag, Germany, SY Ryu, CO Lee, SU Choi, In vitro cytotoxicity of tanshinones from Salvia miltiorrhiza, pp. 339-342), and Magnolia obovata Compounds with six different structures isolated from ethyl acetate fractions also have anti-cancer effects and have been reported to have ED ₅₀ values ranging from 12.5 to 20.2 µg / ml depending on the cancer cells (Planta Medica 65, 1999, Georg Thieme). Verlag, Germany, YK Kim, SY Ryu, Cytotoxic components from stem bark of Magnolia obovata, pp. 291-292).

In addition, eight compounds having various structures isolated from methylene chloride fractions of Artemisia princeps showed anti-cancer effects and reported that they had ED ₅₀ values of 28.5 to 45.8 µg / ml depending on cancer cells. (Planta Medica 63, 1997, Georg Thieme Verlag, Germany, SY Ryu, JO Kim, SU Choi, Cytotoxic components of Artemisia princeps, pp. 384-385).

However, the conventional herbal extracts such as the anticancer drugs were able to reduce the side effects because they are herbal extracts, but the anticancer effect was weak and not effective in the treatment of cancer. Moreover, in the case of such conventional herbal extract anticancer drugs, no research has been conducted on whether the cancer-causing mutations can be suppressed, and thus, it was not possible to confirm whether they could prevent cancer.

On the other hand, in recent years, there is a growing interest in antioxidants that convert or remove reactive oxygen species (ROS), which are harmful substances generated after oxygen participates in metabolic processes, into safe substances.

This is because active oxygen species are known to be involved as a direct cause of pathophysiology such as cerebral neurological diseases such as cancer, various adult diseases including cancer, aging, Parkinson's disease, Alzheimer's disease and ischemic nerve injury.

For example, it is known from research that loss of regulation of ROS production and degradation mechanism is caused by excessive ROS inducing lipid peroxidation of cell membrane lipids, resulting in damage to biological tissues and various diseases.

Accordingly, the present inventors have tried to find a natural composition that can simultaneously treat or prevent the adult diseases, neurological diseases and pathophysiological phenomena, and has little side effects, and the organic matter of wood vinegar in the natural composition includes various types of active ingredients including polyphenol components. In view of the fact that it contains, it was determined that it can be used as an antioxidant as well as prevention and treatment of various diseases including cancer.

Thus, in order to solve the problems described above, the present inventors have developed a method different from the conventional method for producing the wood vinegar, in order to produce a wood vinegar containing more content and types of active ingredients (polyphenol components, etc.) than conventional wood vinegar, The organic component of the wood vinegar prepared by the method developed above was to confirm that the excellent excellent in the treatment and prevention of cancer, antibacterial and antioxidant effect, the wood vinegar according to the production method of the present invention prevents and treats cancer, microbial infection, The present invention has been completed by proving that it has an excellent effect of improving various symptoms and diseases caused by oxidation of biological components.                         

Accordingly, an object of the present invention is (a) wood grass liquor carbonization process; (b) wood liquor distillation process; And (c) provides a method for producing a new wood vinegar consisting of the wood vinegar organic matter extraction process, to provide a wood vinegar organic matter obtained therefrom.

In order to achieve the above object, the present invention (a) carbonized oak chips to collect smoke at the flue gas temperature of 75 ~ 85 ℃ extract the wood vinegar, remove the solids and wood tar from the extracted wood vinegar, Wood liquor carbonization step of ripening for 6 to 12 months while leaving the excluded liquor liquor; (b) a wood liquor distillation step of removing harmful low-boiling components from the carbonized wood liquor by distillation under reduced pressure, adsorbing and filtering the distilled distilled liquor to separate egg separation impurities, and obtaining wood liquor from the filtrate; (c) adding methylene chloride, an organic solvent, to the wood vinegar after distillation, followed by shaking to separate the organic solvent layer and the wood vinegar layer, and adding ethyl acetate, the organic solvent, to the wood vinegar layer, followed by shaking. A wood-choice organic matter extraction step of secondly separating the solvent layer and the wood-choice layer to obtain the first and second organic solvent layers; And (d) an organic solvent removal step of removing the organic solvent from the organic solvent layer obtained.

In addition, in order to achieve another object, the present invention provides a pharmaceutical composition for preventing and treating cancer, containing the wood vinegar organic matter as an active ingredient.

In addition, in order to achieve another object, the present invention provides a pharmaceutical composition for antimicrobial containing the wood vinegar organic matter as an active ingredient.

Furthermore, the present invention provides a dietary supplement containing the wood-based organic matter as an active ingredient.

Hereinafter, the present invention will be described in detail.

At this time, if there is no other definition in the technical terms and scientific terms used, it has a meaning commonly understood by those of ordinary skill in the art.

 In addition, repeated description of the same technical configuration and operation as in the prior art will be omitted.

Compared with the organic components contained in other wood vinegar, the wood vinegar organic matter of the present invention exhibits an excellent effect on antibacterial and antioxidant activity and the ability to treat and prevent cancer.

This is because there is a difference in the active ingredient according to the extraction process of wood vinegar, and also in the content of ingredients such as tar or cresol harmful to the body depending on the purification process.

To this end, the production method for obtaining the wood vinegar organic matter of the present invention comprises the following steps.

(a) wood liquor carbonization process

The oak chips are carbonized to capture smoke at a predetermined flue temperature, to extract wood vinegar, to remove solids and tar from the extracted wood vinegar, and aged for a predetermined period of time while leaving the removed wood vinegar.

At this time, it is preferable to extract the wood vinegar at the flue gas temperature of 75 ~ 85 ℃, which is less than 75 ℃ when the toxic content of tar, methanol, phenol, etc. in the wood vinegar is increased, the effective ingredient is reduced when it exceeds 85 ℃ Because it can be.

In addition, it is desirable to ripen for 6 to 12 months while standing in order to precipitate the remaining harmful impurities in the extracted wood vinegar, which can reduce the cost of removing impurities due to mass production, and when aged less than 6 months to precipitate impurities It is difficult to remove it sufficiently, and if it exceeds 12 months, there is no difference in aging efficiency compared to the excess period.

(b) wood liquor distillation process

After removing the harmful low boiling point component from the wood vinegar, which has been subjected to the step (a) by distillation under reduced pressure, the vacuum distilled wood vinegar is adsorbed and filtered to separate the egg separation impurities, and then the wood vinegar of the filtrate is obtained.

At this time, the distillation under reduced pressure is preferably distilled at a temperature of 40 ~ 60 ℃ under a pressure of -752mmHg, which is less than 40 ℃ to remove enough low-boiling components of tar, methanol, phenol contained in a small amount in wood vinegar, 60 It is because there is almost no difference in removal efficiency of the low boiling point components when the temperature is exceeded.

In addition, in order to more highly remove the low boiling point component, the vacuum distillation, adsorption, and filtration may be performed one or more times.

(c) Wood Vinegar Organic Extraction Process

After adding the methylene chloride as an organic solvent to the wood vinegar after the step (b), the organic solvent layer and the wood vinegar layer are separated by primary shaking, and the ethyl acetate, which is an organic solvent, is added to the wood vinegar layer, followed by shaking. Separation of the second layer and the wood vinegar layer to obtain the organic solvent layer of the primary and secondary.

This process is to maximize the type and amount of the active ingredient by dissolving the organic matter in the wood vinegar in an organic solvent layer and then extracted, the mixing ratio of wood vinegar and organic solvent is 2: 1 to 3: 1 It is preferable to isolate and concentrate. This means that when the amount of organic solvent used is less than one third of the amount of wood vinegar, the organic matter of wood vinegar is not sufficiently dissolved in the organic solvent layer. When the amount of the organic solvent is more than half, the amount of the organic solvent is increased and the extraction cost is increased. This is because the extraction effect is insignificant.

(d) organic solvent removal process

The organic solvent is removed from the organic solvent layer subjected to the step (c).

At this time, it is preferable to use a reduced pressure concentration method to remove the organic solvent.

In this way, the wood vinegar organics obtained through the steps (a), (b), (c) and (d) are used as the wood vinegar organics of the present invention.

On the other hand, the wood vinegar organic compounds of the present invention as a result of in vitro experiments completely inhibited the growth of all cancer cells at a concentration of 100 ㎍ / ㎖, in particular, melanoma completely inhibits the cancer cells even at a concentration of 30 ㎍ / ㎖.

In addition, the wood vinegar organic matter of the present invention showed 86.9% of the antimutagenic activity at the concentration of 1 mg / plate of the wood vinegar organic component to NPD (4-nitro- o- phenylenediamine) which is a direct mutagenesis, NaN ₃ (sodium) azide) showed 78.8% anti-mutant activity, while SOS chromotest divides β-galactoxylase activity by alkaline phosphatase activity and maintains a low enzyme activity ratio to prevent cancer.

In addition, the wood-based organic matter of the present invention has antimicrobial activity to both Gram-positive bacteria and Gram-negative bacteria, and as a result of the experiment, Gram-positive bacteria have a high antimicrobial effect, especially to strains of the genus Bacillus, and do not have an antimicrobial effect to lactic acid bacteria (lactic acid bacteria) The microorganisms effective in the organism have no antimicrobial activity.

In addition, gram Corynebacterium salmonella which can cause food poisoning or enteritis among (Corynebacterium) and gram-negative bacteria (Salmonella) causing the infection in the skin or mucous membrane of a mammal of the positive bacteria, Enterococcus (Enterococcus), Enterobacter (Enterobacter) , Antibacterial to E. coli .

In addition, the wood vinegar organic matter of the present invention greatly suppresses the formation of lipid peroxide in the living tissue, the concentration of 0.1mg / ㎖, 0.5mg / ㎖, 1.0mg / ㎖ DPPH (1,1-diphenyl-2-picrylhydrazyl) radicals Have 35.6%, 68.8%, and 82.4% radical scavenging effect, respectively, and have antioxidant properties.

Therefore, the present invention can be usefully used for the prevention and treatment of cancer and the antimicrobial pharmaceutical composition and health supplement prepared and developed according to the present invention.

More specifically, medicines or health supplements containing the wood-based organic matter of the present invention as an active ingredient may be prepared in any dosage form. Specifically, the form may be in the form of tablets, capsules, powders, ointments, solutions, gels, pastes, granules. It can be prepared by.

In addition, the effective amount of the wood vinegar organic matter according to the present invention is usually preferably taken 0.1 to 0.3 mg [0.1 to 0.3 mg / kg / day] per 1 kg body weight per day, more preferably 0.2 mg / kg / Day is to be taken. However, it is to be understood that the actual dosage should be determined in light of several related factors such as oral dosage form, age, sex, weight of the patient and severity of the patient, and therefore the dosage should in any way limit the scope of the invention. It is not.

In addition, to administer according to the formulation within the above range, it can be prepared by further comprising a variety of excipients, carriers, diluents and the like that are well known to those skilled in the art and can be easily selected by those skilled in the art.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in detail, and the scope of the present invention is not limited to these examples.

EXAMPLES Preparation of Wood Vinegar Organics According to the Present Invention

The oak chips are put in a kiln and carbonized at a kiln temperature of 350-400 ° C. while collecting smoke within the range of flue gas 75-85 ° C. to extract wood vinegar, and the solids and tar visible to the naked eye are removed from the extracted wood vinegar. Aged for about 6 months while standing.

Then, the wood vinegar except for the precipitated impurities was subjected to dark distillation at 40-60 ° C. under a pressure of −752 mmHg, and the remaining wooden vinegar was adsorbed and filtered with filter paper (Whatman 2).

Then, the vacuum distillation, adsorption and filtration were performed once more.

Then, 500 ml of methylene chloride was added to 1 L of the purified high-purity wood vinegar, and placed in a separatory funnel, followed by strong shaking 10 times, and left for 1 hour to give two layers, followed by the upper aqueous layer (wood herb layer) and the lower layer. The methylene chloride layer was separated. The methylene chloride layer was concentrated under reduced pressure with a rotary concentrator to remove methylene chloride to obtain methylene chloride soluble organics.

In addition, ethylacetate soluble organics were obtained in the same manner as above except that ethyl acetate was added to the aqueous layer.

Then, the organic matter obtained from the two organic solvents were mixed to complete the wood vinegar organic matter of the present invention.

Experimental Example 1 Anticancer Effect Measurement

The cancer cell line was used as a cell line in culture at the Korea Research Institute of Chemical Technology, and the cell culture was maintained at 37 ° C., 100% humidity, and 10% CO ₂ concentration. Example In the day before administration of the organic material (hereinafter referred to as sample), the D-MEM (full name) containing 10% of serum was added to each well of 1.0 ml in a 24-well plate, and the cells in the culture were trypsinized and harvested. Cell concentration was calculated with a cell counter. Then, the cells were diluted in concentrations by D-MEM and plated in each well by varying the number of cells. After 18 hours of cell plating, the medium was aspirated, and then 1.0 mL of D-MEM containing no serum was added to each well, and the sample was diluted by concentration with D-MEM containing no serum and administered to each well. After 4 hours of administration, all of the medium was removed, and then 2.0 mL of D-MEM containing 10% of serum was added thereto. After 6 days of sample administration, all of the medium was removed, stained with crystal violet for 3 minutes, dried, and observed under an anatomical microscope to compare the number of colonies with the number of plated cells. . The results are shown in Table 1.

From Table 1, it can be seen that the wood vinegar organic compound of the present invention has an excellent anticancer effect on human cancer cells. In other words, the growth of all cancer cells was completely inhibited at a concentration of 100 ㎍ / ㎖, in particular, melanoma was found to completely inhibit the cancer cells even at a concentration of 30 ㎍ / ㎖.

Experimental Example 2 Anti-mutation Effect Measurement

2-1. Ames test

The histidine-required mutants Salmonella typhimurium TA100 and TA98 [passage cultured from Korea Collection for Type Cultures of the Institute of Biotechnology, Korea Advanced Institute of Science and Technology] were tested. As a negative control, DMSO, a solvent in which a sample was dissolved, was used. As a positive control, mutation rates were compared using a mutagen NPD (4-nitro- o- phenylenediamine) and NaN ₃ (sodium azide).

Meanwhile, before the experiment, the strains were regularly used as test strains after confirming genotypes such as histidine requirement, deep rough character, UV sensitivity, and R factor.

The medium for anti-mutation experiments and the preparation of the necessary reagents were performed according to the method of Maron and Ames.

That is, 0.1 ml of the strain cultured overnight (1-2 × 10 ⁹ cells / ml), 0.1 ml of mutagen (containing 10 µg of NPD, 1 µg of NaN ₃ ), 0.5 ml of sodium phosphate buffer and the wood-vinegar organic matter of the example 1 After addition to the test tube at the concentration of mg / plate, it was lightly vortexed and preincubated for 20 minutes at 37 ℃. Then, 2 ml of top agar stored at 45 ° C. were poured into tubes, vortexed for 3 seconds, plated on minimal glucose agar plate, and incubated at 37 ° C. for 48 hours. The number of return mutations was counted.

As a result, as shown in Table 2, the wood vinegar organic matter according to the present invention for the mutagenic NPD in Salmonella typhimurium TA98 showed an antimutagenic activity of 86.9% at a concentration of 1 mg / plate.

In addition, as shown in Table 3, the herbaceous organic matter according to the present invention for the mutant NaN ₃ in Salmonella typhimurium TA100 showed 78.8% anti-mutant activity.

2-2. SOS Chromotest

SOS chromotest using MNNG (N-methyl-N'-nitrosoguadine) and NQO (4-nitroquinoline-1-oxide) as mutagens to more clearly confirm the antimutagenic effect of the wood-choice organic matter of Experimental Example 2-1 Was carried out in the following manner.

First, 50 ml of E. coli PQ37 (passage cultured in the laboratory) was cultured and 50 ml of Liria Bertani medium (10 g of tryptone, 5 g of yeast extract, 10 g of NaCl, 1.0 l of distilled water). pH 7.0) was incubated for 4 hours at 37 ℃ (OD = 0.4 ~ 0.8). In addition, 600 μl of the culture solution, 20 μl of mutagen (MNNG, NQO) (1 μg / assay) and 10 μl (100 μg / assay) of the wood vinegar organic matter of the present invention were put into a test tube to induce an SOS reaction. After standing at 37 ° C. for 2 hours, alkaline phosphatase enzyme activity was measured.

In addition, β-galactoxytase activity was 1.8 ml of B buffer (composition: Na ₂ HPO ₄ 5.5 g, KCl 0.75 g, MgSO ₄ 0.25 g, SDS 1.0 g, β-mercaptoethanol 2.7 ml, distilled water 1,000 ml, pH 7.0) was added, and the mixture was allowed to stand at 37 DEG C for 5 minutes to uniform the temperature. Then, 0.4 ml of o -nitrophenyl-β-galactoxytase (hereinafter referred to as ONGP) solution was added to maintain color development for 10 minutes. After reacting by adding 1.4 ml of 2.0 M Na ₂ CO ₃ solution, the absorbance at 420 nm was measured.

Alkaline phosphatase activity was measured using P buffer (composition: tris-base 121 g, SDS 1.0 g, distilled water 1,000 mL, pH 8.8) instead of B buffer, and measuring p-nitrophenyl-phosphogalactoxidase activity instead of ONGP solution. It was. To stop the enzymatic reaction, 0.7 ml of 2.5M HCl was added and then the reaction was stopped for 5 minutes, and 0.7 ml of 2.0M Tris-HCl buffer was added.

The activity of each enzyme was measured using the formula of 1000 × A ₄₂₀ / t (wherein A ₄₂₀ represents the absorbance at ₄₂₀ nm and t represents the reaction time in minutes). Was calculated by dividing β-galactoxytase activity by alkaline phosphatase activity.

As a result, as shown in Figures 1 and 2, the same as the Ames test, wood herb organics according to the present invention showed a low enzyme activity ratio value, the fact that the wood herb organics of the present invention by effectively inhibiting mutations The preventive effect of cancer was very good.

Experimental Example 3 Antibacterial Effect Measurement

 Indicator bacteria for measuring the antimicrobial effect were incubated overnight in the corresponding liquid medium and then plated on the same solid medium. Several holes with a diameter of about 6 mm were drilled on the solid medium, and 50 μl of the wood vinegar organic matter (1 mg / ml) of the present invention was added thereto and left at 4 ° C. for 2 hours, followed by incubation overnight at the optimum growth temperature of the bacteria. As a result, the degree of antimicrobial activity was determined by measuring the size of the growth zone around the hole. A total of 48 strains were used for the experiment.

As a result, as shown in Table 4, the wood vinegar organic matter of the present invention showed an antimicrobial effect on both Gram-positive bacteria and Gram-negative bacteria. Among the Gram-positive bacteria, the strains of Bacillus genus showed high antibacterial effect, but not to lactic acid bacteria (lactic acid bacteria). Therefore, taking wood vinegar does not affect the body's lactobacillus. Also it exhibited a high antimicrobial activity on Gram Corynebacterium (Corynebacterium) causing the infection in the skin or mucous membrane of a mammal of the positive bacteria, gram-negative bacteria from among the Salmonella that cause food poisoning or enteritis (Salmonella), Enterococcus (Enterococcus ), Enterobacter, and E. coli showed antimicrobial effects.

Experimental Example 4 Antioxidant Effect Measurement

4-1. Lipid Peroxidation Inhibitory Effect

To measure the lipid peroxide content, 0.1 ml of brain tissue homogenate and 0.05 ml of sample were mixed with 0.65 ml of 0.1 M potassium phosphate buffer (pH 7.4) according to the method of Okawa et al. To this, 0.2 ml of 8.1% sodium dodecyl sulfate, 1.5 ml of 20% acetate buffer (pH 3.5) and 1.5 ml of 0.8% 2-thiobarbituric acid solution were added and reacted again at 95 ° C for 1 hour. The mixture was cooled to room temperature, and the resulting red colored material was transferred to an n -butanol: pyridine (15: 1, v / v) mixture, centrifuged at 3,000 rpm for 10 minutes, and the supernatant was absorbed at wavelength 532 nm. Changes were measured and quantified. Lipid peroxide content is the amount of malondialdehyde (MDA) per gram of tissue in nanomoles (nmole).

In addition, as a result of observing in vitro how much the wood-vinegar organic matter of the present invention inhibits lipid peroxidation reaction in cells, as shown in FIG. 3. Compared to 22.5 MDA nmoles / g of control (nanomol of MDA per gram of tissue) in the brain tissue, the organics of the present invention were 21.9, 10.4, and 4.5 at concentrations of 0.1 mg / ml, 0.5 mg / ml and 1.0 mg / ml, respectively. nmoles / g.

From this, it can be seen that the wood vinegar organic matter of the present invention greatly inhibits the production of lipid peroxide in brain breakfast and liver tissue in a dose-dependent manner.

4-2. DPPH radical scavenging effect

The content of DPPH (1,1-diphenyl-2-picrylhydrazyl) radical was measured according to the method of Blosi, and 4 ml of 1.5x10 ^-5 M ethanol solution of DPPH radical was added to 0.1 ml of organic component of wood vinegar at room temperature. After reacting for 30 minutes, absorbance at 520 nm is measured and calculated. Radical scavenging activity of DPPH radical was calculated by the following equation.

Scheme 1

Radical scavenging activity (%) = (ODcontrol-ODsample) / ODcontrol × 100

And, as a result of measuring the extent to which the organic component of the wood vinegar inhibits DPPH radical, as shown in Figure 4, the wood vinegar organic compound of the present invention is radical scavenging effect at the concentration of 0.1mg / ㎖, 0.5mg / ㎖, 1.0mg / ㎖ Were 35.6%, 68.8%, and 82.4%, respectively.

From this, it was found that the wood vinegar organic matter of the present invention was very excellent in the radical scavenging effect.

As described above, the organic matter of the wood vinegar according to the present invention can prevent cancer by treating the cancer or at the same time by inhibiting mutations, and has an effect that can be used as an additive in health foods by utilizing antibacterial and antioxidant effects.

Claims (5)

(a) carbonizing oak chips to capture smoke at the flue temperature of 75 ~ 85 ℃ to extract the wood vinegar, excluding solids and tar tar from the extracted wood vinegar, aged for 6 to 12 months while leaving the excluded wood vinegar Wood liquor carbonization process; (b) a wood liquor distillation step of removing harmful low-boiling components from the carbonized wood liquor by distillation under reduced pressure, adsorbing and filtering the distilled distilled liquor to separate egg separation impurities, and obtaining wood liquor from the filtrate; (c) adding an organic solvent selected from methylene chloride or ethyl acetate to the wood vinegar, which has been subjected to the distillation, and shaking the mixture to first separate the organic solvent layer and the wood vinegar layer, and to the wood vinegar layer, an organic solvent selected from methylene chloride or ethyl acetate. Adding and then shaking to separate the organic solvent layer and the wood vinegar layer to obtain the first and the second organic solvent layer wood vinegar organic material extraction step; And (d) The wood vinegar organic material obtained from the process comprising the organic solvent removal step of removing the organic solvent from the organic solvent layer obtained. The method of claim 1, The distillation under reduced pressure in the wood vinegar distillation process is distilled under reduced pressure at a temperature of 40 ~ 60 ℃ under pressure of -752mmHg wood organic matter organic matter. The method of claim 1, The organic matter of the wood vinegar, characterized in that the step of distillation under reduced pressure and adsorption and filtration in the wood vinegar distillation step. A pharmaceutical composition for preventing and treating cancer, comprising the wood vinegar organic matter of any one of claims 1 to 5 as an active ingredient. A pharmaceutical composition for antimicrobial containing the wood vinegar organic matter of any one of claims 1 to 5 as an active ingredient.

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