WO2011115314A1 - Antioxidant effect of functional nanodiamond and a use therefor - Google Patents

Antioxidant effect of functional nanodiamond and a use therefor Download PDF

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WO2011115314A1
WO2011115314A1 PCT/KR2010/001688 KR2010001688W WO2011115314A1 WO 2011115314 A1 WO2011115314 A1 WO 2011115314A1 KR 2010001688 W KR2010001688 W KR 2010001688W WO 2011115314 A1 WO2011115314 A1 WO 2011115314A1
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antioxidant
nanodiamond
group
formula
compound
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PCT/KR2010/001688
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French (fr)
Korean (ko)
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이민영
지아영
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나노다이아몬드 주식회사
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Publication of WO2011115314A1 publication Critical patent/WO2011115314A1/en

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    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K15/00—Anti-oxidant compositions; Compositions inhibiting chemical change
    • C09K15/04—Anti-oxidant compositions; Compositions inhibiting chemical change containing organic compounds
    • C09K15/20—Anti-oxidant compositions; Compositions inhibiting chemical change containing organic compounds containing nitrogen and oxygen
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/02—Cosmetics or similar toiletry preparations characterised by special physical form
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/19—Cosmetics or similar toiletry preparations characterised by the composition containing inorganic ingredients
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • C—CHEMISTRY; METALLURGY
    • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
    • C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
    • C11B5/00—Preserving by using additives, e.g. anti-oxidants
    • C11B5/0007—Organic substances
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
    • A61K2800/41—Particular ingredients further characterized by their size
    • A61K2800/413—Nanosized, i.e. having sizes below 100 nm
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
    • A61K2800/52—Stabilizers
    • A61K2800/522—Antioxidants; Radical scavengers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00—Preparations for care of the skin
    • A61Q19/10—Washing or bathing preparations
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q5/00—Preparations for care of the hair
    • A61Q5/02—Preparations for cleaning the hair

Definitions

  • the present invention relates to functional nanodiamonds, and more particularly, to an antioxidant composed of surface functionalized nanodiamond compounds prepared by chemically modifying the surface of nanodiamonds.
  • the antioxidant effect on nanodiamonds is not known yet.
  • the present invention relates to functional nanodiamonds, a new concept of antioxidant, which is not known until now.
  • Oxidation of substances not only causes various diseases in vivo, but also causes aging and is known to shorten the life of many industrial materials.
  • Antioxidants or antioxidants are compounds that inhibit or delay the automatic oxidation of a substance. Antioxidants are classified into preventive antioxidants, primary antioxidants and secondary antioxidants according to their mechanism of action. Prophylactic antioxidants are compounds that prevent the production of free radicals, which are literally the source of antioxidants. Primary antioxidants are chain-breaking antioxidants that quickly eliminate free radicals or peroxy radicals already produced. Secondary antioxidants do not have anti-oxidative effects on their own, but they are also known as antioxidant synergists as compounds that increase antioxidant activity by coexisting with free radical inhibitors.
  • Antioxidants can also be divided into natural and synthetic antioxidants.
  • Vitamin C, tocopherol, natural ascorbic acid, polyphenols, and various plant extracts are used as natural antioxidants, and BHA (butylhydroxyanisole) and butylhydroxytoluene (BHT), which are phenolic compounds, are representatively used as synthetic antioxidants.
  • antioxidants that are chemically synthesized are rarely used. Recently, due to the rapid development of nanotechnology, researches to use nanoparticles such as gold, silver and platinum as antioxidants have been activated, but they are still in the research and development stage due to various problems.
  • the first object of the present invention is to prepare a surface functionalized nanodiamond compound maximized antioxidant effect and to measure their antioxidant effect by the ORAC method.
  • a second object of the present invention is to use the nanodiamond compound as an antioxidant additive such as water, beverages, alcoholic beverages, food, cosmetics, pharmaceutical raw materials, oils and fats, and to prevent oxidation of paints, polymers, ceramics, and metals will be.
  • an antioxidant consisting of surface functionalized nanodiamond compounds represented by the following formula (I) or salts thereof.
  • Formula I ND- (R) n , where n is an integer of 2 or greater.
  • ND represents a nanodiamond single particle, and a plurality of R each represent a moiety chemically bonded to the surface of the nanodiamond, wherein R represents an amine group (NH 2 ), a hydroxyl group (OH), It has a chemical structure containing at least one or more selected from the group consisting of a carboxyl group (COOH), and combinations thereof.
  • R represents an amine group (NH 2 ), a hydroxyl group (OH), It has a chemical structure containing at least one or more selected from the group consisting of a carboxyl group (COOH), and combinations thereof.
  • R may be represented by Formula II.
  • X is a linker for covalently linking an amine group to a nanodiamond surface, and includes a group consisting of one or more methylene groups, carboxyl groups, ether groups, amide groups, ester groups, and combinations thereof. It includes one selected from.
  • R may further include a food nutritional or pharmaceutically acceptable salt of Formula II.
  • the antioxidant may be used in one selected from the group consisting of water, beverages, alcoholic beverages, and coffee to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
  • the antioxidant may be used as a food additive to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
  • the antioxidant may be used as a cosmetic raw material to exhibit an antioxidant, antioxidant, or antioxidant enhancement effect.
  • the antioxidant may be used as a pharmaceutical composition to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
  • the antioxidant may be used in tobacco to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
  • the antioxidant may be used in one selected from the group consisting of soap, shampoo, detergent, bathing articles and can exhibit an antioxidant, antioxidant, or antioxidant enhancement effect.
  • the antioxidant may be used to prevent rancidity of edible fats and oils or industrial oils.
  • the antioxidant can be used as an additive in animal feed.
  • the antioxidant can be used as an additive to preserve the grain.
  • the antioxidant may be used to prevent oxidation of one selected from the group consisting of flowers, fruits, vegetables, meat, poultry, and fish.
  • the antioxidant may be used to prevent one oxidation selected from the group consisting of plastics, rubber, adhesives, fibers, polymer film, and polymer coating agent.
  • the antioxidant may be used to prevent oxidation of one bulk or surface selected from the group consisting of ceramic materials, metal materials, and composite materials in combination thereof.
  • the nanodiamond compounds of the present invention show excellent dispersibility in the liquid phase, so that they can be used as additives in drinking water, beverages, alcohol, food, cosmetics, medicine, coating materials, structural materials, etc. Can be used to show antioxidant effects.
  • Figure 7 Fluorescence attenuation graph according to the concentration of the sample.
  • Nanodiamonds are attracting attention as new nanomaterials because they are not only physically and chemically stable but also have no biotoxicity.
  • particles having a size of 10 nm or less are specifically defined as ultrananocrystalline diamond (UNCD), and their application range is rapidly spreading.
  • UNCD is an ultrafine diamond crystal with a relatively uniform particle size distribution of about 5 nm in diameter, and is mainly synthesized by explosive explosion.
  • due to the problem of dispersibility it has been used in a limited range, and various methods of modifying the surface have been proposed to overcome this problem.
  • nanodiamonds of several nanometers in size can attach a large amount of various functional groups to the surface of nanodiamonds through various chemical synthesis methods in gas phase or liquid phase.
  • the antioxidant effect of such functional nanodiamonds has not been published yet.
  • FRAP ferric reducing / antioxidant power
  • TEAC trolox equivalent antioxidant capacity
  • ORAC oxygen radical absorbance capacity
  • FRAP and TEAC use single electron transfer and ORAC uses hydrogen atom transfer. Since FRAP and TEAC were not developed based on the mechanism of the chain reaction, it is difficult to measure antioxidant activity. Based on the mechanisms that control antioxidant chain reactions, ORAC was first developed in 1993 by researchers at the National Institute of Aging under the NIH, and has since been recognized as a gold standard method of measuring antioxidant effects.
  • Antioxidant effect measurement method using ORAC protocol is as follows.
  • Disodium fluorescein (FL) one of the fluorescent dyes, is fluoresced by the bleaching effect in the presence of an oxidizing agent.
  • Mixing samples that are expected to have an antioxidant effect on the solution prevents the oxidation of FL, resulting in longer fluorescence. That is, the intensity of the fluorescence is reduced when the fluorescent material is oxidized by the oxidizing agent.
  • the presence of antioxidants in the solution decreases the rate of destruction of the fluorescent material and increases the fluorescence intensity.
  • the lower area is calculated from the attenuation curve graph obtained by measuring the intensity of the fluorescence as a function of time.
  • AAPH 2,2'-azobis (2-amidinopropane) dihydrochloride
  • the value obtained by comparing the degree of oxidative destruction of fluorescence with the standard antioxidant Trolox is the ORAC value.
  • a standard curve is created using different Trolox concentrations and then compared to the sample. The final result is expressed as Trolox Equivalent (TE).
  • TE Trolox Equivalent
  • ORAC experiments were performed for the purpose of using functionally-modified nanodiamonds chemically modified on the surface of nanodiamonds as antioxidants or antioxidants.
  • ORAC experiments were performed on pristine nanodiamonds, which showed a certain level of antioxidant effect despite the high dispersibility.
  • the degree of antioxidant effect of functional nanodiamonds depends on the type of functional groups attached to the surface, especially nanodiamonds having a moiety attached to amine groups (hereinafter referred to as aminated nanodiamonds) exhibit the best antioxidant effects. Appeared.
  • These antioxidant nanodiamonds may be applied to various fields such as food, cosmetics, medicine, agriculture, petrochemicals, polymers, ceramics and metals.
  • FIG. 1 is a schematic diagram of one of the nanodiamond (ND) compounds having an amine group.
  • the central portion of the schematic diagram represents nanodiamonds, and amine groups are attached to the surface in large amounts by covalent bonds.
  • the aminated nanodiamond compound is synthesized from the pristine nanodiamond in a liquid phase through a multi-step manufacturing process as shown in various examples below.
  • These functionalized nanodiamonds show excellent dispersibility compared to pristine nanodiamonds, and have a very high antioxidant effect, i.e., an ORAC index.
  • ND- (COOH) n compound 100 mg was added to 30 mL of BH 3 / THF, followed by sonication for 1 hour, followed by the same filtration and drying process as in Example 1, where a plurality of moieties including hydroxyl groups were present on the surface of the nanodiamond. Obtain ND- (CH 2 OH) n attached.
  • ND- (COOH) n powder 100 mg was dissolved in 50 mL of ethylenediamine, and then 50 mg HATU reagent was added and sonicated for 4 hours to prepare nanodiamonds having an amide group.
  • the reaction was diluted with 200 mL of methanol, filtered and dried to obtain ND- (CONHCH 2 CH 2 NH 2 ) n compound powder having a plurality of moieties including amide groups and amine groups on the surface of nanodiamonds. .
  • ND- (CH 2 OH) n powder 100 mg was sonicated in 30 ml THF for 30 minutes to introduce an amine group (NH 2 ) to the nanodiamond surface.
  • 10 mg of diethylazodicarboxylate coupling agent and 50 mg of phthalimide were added thereto, sonicated for 2 hours, 300 mL of methanol was poured out, diluted, filtered and dried.
  • This powder was placed in 50 mL of trifluoroacetic acid (TFA), sonicated for 3 hours, filtered and dried to provide ND- (CH 2 NH 2 ) n powder having a plurality of moieties containing amine groups on the surface of nanodiamonds.
  • TFA trifluoroacetic acid
  • the surface modification of the surface functionalized ND compounds was analyzed by making samples in the form of KBR pellets using FTIR (Varian).
  • the strong peaks appearing at 1670 cm ⁇ 1 in the spectrum of the ND- (COOH) n compound of FIG. 2 are due to C ⁇ O stretching.
  • the C ⁇ O stretching peak disappears and peaks appear at 2923 cm ⁇ 1 and 2845 cm ⁇ 1 , showing the CH stretching oscillation mode in the methylene group.
  • the IR spectrum of the ND- (CONHCH 2 CH 2 NH 2 ) n compound shows a CN stretching oscillation mode at 1030 cm ⁇ 1 .
  • the vibration mode corresponding to carbonyl stretching appears at 1679 cm -1 .
  • the ND- (CH 2 NH 2 ) n compound has an in-plane bending mode of amine group at 1624 cm ⁇ 1 and an out-of-plane bending mode at 737 cm ⁇ 1 .
  • Figure 3 shows the particle size distribution of pristine nanodiamonds and aminated nanodiamonds in aqueous solution.
  • the measurement for the particle size analysis was performed using a dynamic light scattering device (Scatteroscope, Qudix).
  • Pristine nanodiamonds show a bimodal pattern with average particle diameters of 16 nm and 20 ⁇ m, indicating that a large number of nanodiamonds are aggregated in the liquid phase.
  • nanodiamonds have an average particle diameter of 8 nm, showing that they are distributed as single particles without forming aggregates in aqueous solution.
  • the chemical modification of the surface of the nanodiamond has the effect of providing the nanodiamond with excellent dispersibility as well as chemical functionality.
  • Table 1 shows the pH of the aqueous solution measured in the concentration range of ND- (CH 2 NH 2 ) n 0.001 to 10 ⁇ M.
  • ND- (CH 2 NH 2 ) n 0.001 to 10 ⁇ M As a molecular weight value of the nanodiamond required for the concentration calculation, 100,000, which is a value calculated in consideration of the particle size, was used.
  • the pH of the aqueous solution is not linearly proportional to the log 10 value of the ND- (CH 2 NH 2 ) n concentration, which is a typical characteristic of the weakly basic solute.
  • the results shown in Table 1 indicate that the aminated nanodiamonds can be added to water, liquor, or food to increase the pH, and can be used to convert acidic or neutral water into weakly alkaline if necessary. For example, adding 10 mg of aminated nanodiamonds to 1 L of neutral water can produce an aqueous solution of acidity close to 7.4, the physiological pH value.
  • ORAC assay was carried out by the following method. Fluorescence measurement was performed using a fluorescence spectrophotometer (F-4500, Hitachi). The excitation and fluorescence wavelengths were 480 nm and 520 nm, respectively, and all experiments were performed at room temperature.
  • fluorescein (FL) was dissolved in 1 ml of phosphate buffer at pH 7.4 to make 70 ⁇ M of stock solution. This stock solution was stored in the dark and diluted with 70 nM of solution prior to measurement. 0.1356 g of 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) was dissolved in phosphate buffer to form a 50 mM aqueous solution. 5 mM Trolox stock solution was prepared by dissolving 0.0125 g of Trolox in 10 mL of phosphate buffer. It was diluted to 1 ⁇ 5 ⁇ M and used to calculate trolox calibration curve.
  • AAPH 2,2'-azobis (2-amidinopropane) dihydrochloride
  • the calibration curve of trolox was measured after incubation of FL solution (240 ⁇ L), AAPH (240 ⁇ L), and trolox (120 ⁇ L) for 10 minutes at 37 ° C. That is, after measuring the fluorescence intensity over time of the buffer solution of FL and AAPH, a series of fluorescence attenuation graphs of the solution in which the trolox solution is mixed to 5 ⁇ M at 1 ⁇ M intervals are obtained in series.
  • the final total solution used for fluorescence measurement was 600 ⁇ L and the fluorescence intensity was measured up to 3,600 seconds (60 minutes) at 25 second intervals.
  • FL solution (240 ⁇ L), AAPH (240 ⁇ L), and antioxidant solution (120 ⁇ L) were mixed and incubated at 37 ° C. for 10 minutes to measure fluorescence for 60 minutes.
  • Blank was measured after incubation of a mixture of buffer (120 ⁇ L), fluorescein solution (240 ⁇ L), and AAPH (240 ⁇ L) at 37 ° C. for 10 minutes.
  • AUC was calculated using the following equation after peak normalization of all fluorescence attenuation graphs to calculate the area under curve (AUC) of the fluorescence attenuation graph.
  • f 0 is the fluorescence intensity at time 0 and n is the number of data for obtaining the fluorescence attenuation graph.
  • the relative ORAC value (Trolox equivalent) from the AUC is calculated by the following equation.
  • ORAC [(AUC sample -AUC blank ) / (AUC sample -AUC blank )]
  • FIG. 4 shows a FL fluorescence attenuation graph according to trolox concentration
  • FIG. 5 is a trolox calibration curve calculated therefrom.
  • x is the trolox concentration ( ⁇ M)
  • y is the AUC.
  • the correlation coefficient (R 2 ) is above 0.99, indicating a very satisfactory level.
  • FIG. 6 is a graph showing fluorescence attenuation of pristine nanodiamonds.
  • the molecular weight value of the pristine diamond required for the concentration calculation 100,000 was calculated based on the size of the nanoparticles.
  • the AUCs were calculated for each of the concentration ranges from 1 to 5 ⁇ M from FIG. 6 and the average value of the relative ORAC values obtained therefrom was 5.2.
  • pristine nanodiamonds exhibit antioxidant effects, they appear to be mainly due to the radical scavenging function of the surface sp 2 structure, but the detailed mechanism is not yet known.
  • Figure 7 shows the fluorescence attenuation graph of vitamin C, clove, surface functionalized nanodiamonds. Since the antioxidant effect of vitamin C is well known, it was selected as a control. Clove was selected for relative comparison because it is known to have the highest ORAC level among natural products known to date. For the relative comparison of their antioxidant levels, a graph of fluorescence attenuation of each sample, especially at 5 ⁇ M, is shown in FIG. 8. At this concentration, the aminated nanodiamond shows little change in fluorescence intensity for 60 minutes and remains constant. 9 shows the AUC of each sample by measurement concentration.
  • Table 2 shows the ORAC values calculated by the above formula for each concentration, and their average values are also shown. Although the molecular weight of the nanodiamond may be slightly changed through surface functionalization, 100,000, which is the same value as the pristine nanodiamond, was used to maintain consistency.
  • Figure 10 shows the ORAC index of vitamin C, clove and functional nanodiamonds. Aminized nanodiamonds exhibited 25-fold antioxidant activity at the same concentrations as compared to vitamin C and also showed higher ORAC values than cloves.

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Abstract

Provided is an antioxidant or oxidation inhibitor made using a surface-functionalized nanodiamond compound. The compound comprises functional nanoparticles resulting from the chemical modification of the nanodiamond surfaces. The antioxidant effect has been measured using the oxygen radical absorbance capacity (ORAC) method. The compound has been confirmed to have an antioxidant effect overall, albeit that some variation exists depending on the chemical structure of the moiety which is chemically bound to the surface of the nanodiamond. More particularly, a nanodiamond compound having an amine group has exhibited an ORAC value which is 25 times the level of vitamin C at the same water concentration. The nanodiamond compound enhances the antioxidant properties of water, soft drinks, alcoholic drinks, foods, cosmetics, medicinal products, edible fats/oils or grains, or can be used in order to prevent oxidation of industrial oils, polymers, ceramics or metals (Figure 7).

Description

기능성 나노다이아몬드의 항산화 효과 및 그 응용Antioxidant Effects of Functional Nanodiamonds and Their Applications
본 발명은 기능성 나노다이아몬드에 관한 것이며, 특히 나노다이아몬드의 표면을 화학적으로 개질하여 제조된 표면기능화 나노다이아몬드 화합물로 이루어진 항산화제에 관한 것이다.The present invention relates to functional nanodiamonds, and more particularly, to an antioxidant composed of surface functionalized nanodiamond compounds prepared by chemically modifying the surface of nanodiamonds.
기존의 항산화제가 갖는 열적안정성 및 생체독성문제를 해결하기 위한 새로운 항산화제 개발이 요구되고 있는바, 나노기술에 기반을 둔 항산화제가 이를 해결할 수 있는 유력한 대안으로 제시되고 있다. 금, 은, 백금 나노입자를 항산화제로 이용하려는 시도가 있어 왔지만 분산성, 안전성, 가격 등이 해결해야 할 문제로 남아 있다. It is required to develop new antioxidants to solve the thermal stability and biotoxicity problems of existing antioxidants, and nanotechnology-based antioxidants have been suggested as a viable alternative. Attempts have been made to use gold, silver, and platinum nanoparticles as antioxidants, but dispersibility, safety, and price remain problems to be solved.
나노다이아몬드에 대한 항산화 효과는 아직 알려진 바가 없는데 본 발명은 이제까지 알려 지지 않은 새로운 개념의 항산화제인 기능성 나노다이아몬드에 관한 것이다. The antioxidant effect on nanodiamonds is not known yet. The present invention relates to functional nanodiamonds, a new concept of antioxidant, which is not known until now.
산화의 기본 메커니즘은 자유라디칼(free radical)의 형성으로 시작되는데, 자유라디칼은 짝을 이루지 못한 전자들을 가지고 있는 분자로서 매우 불안정이기 때문에 다른 분자를 공격하여 화학 구조를 변화시킬 수 있다. 자유라디칼의 대부분은 활성산소종(reactive oxygen species)으로 하이드록시 라디칼(hydroxyl radical), 수퍼옥사이드 라디칼(superoxide radical), 일중항 산소(singlet oxygen), 알코옥시 라디칼(alkoxy radical) 등이 이에 해당한다. 물질의 산화는 생체 내에서 다양한 질병을 일으킬 뿐만 아니라 노화의 원인이기도 하며 많은 산업 소재의 수명을 단축시키는 요인으로 알려져 있다. The basic mechanism of oxidation begins with the formation of free radicals, which are unstable molecules with unpaired electrons that can attack other molecules and change their chemical structure. Most of the free radicals are reactive oxygen species, such as hydroxy radicals, superoxide radicals, singlet oxygen, and alkoxy radicals. . Oxidation of substances not only causes various diseases in vivo, but also causes aging and is known to shorten the life of many industrial materials.
항산화제(antioxidant) 또는 산화방지제란 물질의 자동 산화를 억제하거나 지연시키는 화합물이다. 항산화제는 작용기전에 따라 예방적 항산화제, 일차 항산화제, 이차 항산화제로 구분된다. 예방적 항산화제는 글자 그대로 항산화의 요인이 되는 자유 라디칼의 생성을 미연에 방지하는 화합물이다. 일차 항산화제는 이미 생성된 자유 라디칼(free radical)이나 퍼록시 라디칼 (peroxy radical)을 빠르게 소거하는 연쇄절단형 항산화제 (chain-breaking antioxidant)를 말한다. 이차 항산화제는 스스로 산화방지 효과는 없지만 자유라디칼 저해제와 공존하여 항산화작용을 증가시키는 화합물로서 항산화 상승제라고도 한다.Antioxidants or antioxidants are compounds that inhibit or delay the automatic oxidation of a substance. Antioxidants are classified into preventive antioxidants, primary antioxidants and secondary antioxidants according to their mechanism of action. Prophylactic antioxidants are compounds that prevent the production of free radicals, which are literally the source of antioxidants. Primary antioxidants are chain-breaking antioxidants that quickly eliminate free radicals or peroxy radicals already produced. Secondary antioxidants do not have anti-oxidative effects on their own, but they are also known as antioxidant synergists as compounds that increase antioxidant activity by coexisting with free radical inhibitors.
항산화제는 또한 천연 항산화제와 합성 항산화제와 나눌 수 있다. 천연 항산화제로는 비타민 C, 토코페롤, 천연 아스코르빈산, 폴리페놀, 각종 식물 추출물 등이 사용되고 있으며 합성 항산화제로는 페놀계 화합물인 BHA (butylhydroxyanisole), BHT (butylhydroxytoluene) 등이 대표적으로 사용되고 있다. Antioxidants can also be divided into natural and synthetic antioxidants. Vitamin C, tocopherol, natural ascorbic acid, polyphenols, and various plant extracts are used as natural antioxidants, and BHA (butylhydroxyanisole) and butylhydroxytoluene (BHT), which are phenolic compounds, are representatively used as synthetic antioxidants.
항산화제의 중요성에도 불구하고 화학적으로 합성되어 사용되는 합성 항산화제는 종류가 많지 않은 편이다. 최근 들어 나노기술의 비약적인 발전에 의해 금 은 백금등의 나노입자를 항산화제로 이용하려는 연구가 활성화되고 있지만 여러 가지 문제점 때문에 아직 연구 개발단계에 머물러 있다.Despite the importance of antioxidants, synthetic antioxidants that are chemically synthesized are rarely used. Recently, due to the rapid development of nanotechnology, researches to use nanoparticles such as gold, silver and platinum as antioxidants have been activated, but they are still in the research and development stage due to various problems.
본 발명의 첫 번째 목적은 항산화 효과가 극대화된 표면기능화 나노다이아몬드 화합물을 제조하여 이들의 항산화 효과를 ORAC 방법으로 측정하는 것이다.The first object of the present invention is to prepare a surface functionalized nanodiamond compound maximized antioxidant effect and to measure their antioxidant effect by the ORAC method.
본 발명의 두 번째 목적은 상기 나노다이아몬드 화합물을 물, 음료, 주류, 식품, 화장품, 의약품 원료, 유지류 등의 항산화 첨가제로서, 그리고 도료, 폴리머, 세라믹, 금속 등의 산화방지 용도로 사용할 수 있도록 하는 것이다.A second object of the present invention is to use the nanodiamond compound as an antioxidant additive such as water, beverages, alcoholic beverages, food, cosmetics, pharmaceutical raw materials, oils and fats, and to prevent oxidation of paints, polymers, ceramics, and metals will be.
본 발명에 따르면, 하기 화학식 I로 표시되는 표면 기능화 나노다이아몬드 화합물(surface functionalized nanodiamond compounds) 또는 그의 염으로 이루어지는 항산화제를 제공한다.According to the present invention, there is provided an antioxidant consisting of surface functionalized nanodiamond compounds represented by the following formula (I) or salts thereof.
화학식 I: ND-(R)n, 여기서 n은 2 이상의 정수이다.Formula I: ND- (R) n , where n is an integer of 2 or greater.
상기 화학식 I에서, ND는 나노다이아몬드 단일입자를 나타내며, 복수개의 R은 각각 상기 나노다이아몬드 표면에 화학결합된 부분(moiety)를 나타내되, 상기 R은 아민기(NH2), 수산기(OH), 카르복실기(COOH), 및 이들의 조합으로 이루어진 일군에서 선택된 하나를 적어도 1개 이상 포함하는 화학구조를 가진다.In Formula I, ND represents a nanodiamond single particle, and a plurality of R each represent a moiety chemically bonded to the surface of the nanodiamond, wherein R represents an amine group (NH 2 ), a hydroxyl group (OH), It has a chemical structure containing at least one or more selected from the group consisting of a carboxyl group (COOH), and combinations thereof.
다른 방법으로, 상기 R은 하기 화학식 II일 수 있다.Alternatively, R may be represented by Formula II.
화학식 II: X-NH2 Formula II: X-NH 2
상기 화학식 II에서, X는 나노다이아몬드 표면에 아민기를 공유 결합으로 연결시키기 위한 링커(linker)로서, 하나 또는 다수의 메틸렌기, 카르복실기, 에테르기, 아마이드기, 에스테르기, 및 이들의 조합으로 이루어진 일군에서 선택된 하나를 포함한다.In Formula II, X is a linker for covalently linking an amine group to a nanodiamond surface, and includes a group consisting of one or more methylene groups, carboxyl groups, ether groups, amide groups, ester groups, and combinations thereof. It includes one selected from.
또 다른 방법으로, 상기 R은 상기 화학식 II의 식품영양학적 또는 약학적으로 허용 가능한 염을 더 포함할 수 있다.In another method, R may further include a food nutritional or pharmaceutically acceptable salt of Formula II.
또 다른 방법으로, 상기 항산화제는 물, 음료, 주류, 커피로 이루어진 일군에서 선택된 하나에 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타낼 수 있다.As another method, the antioxidant may be used in one selected from the group consisting of water, beverages, alcoholic beverages, and coffee to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
또 다른 방법으로, 상기 항산화제는 식품 첨가제로 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타낼 수 있다.Alternatively, the antioxidant may be used as a food additive to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
또 다른 방법으로, 상기 항산화제는 화장품 원료로 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타낼 수 있다.Alternatively, the antioxidant may be used as a cosmetic raw material to exhibit an antioxidant, antioxidant, or antioxidant enhancement effect.
또 다른 방법으로, 상기 항산화제는 약제 조성물로 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타낼 수 있다.Alternatively, the antioxidant may be used as a pharmaceutical composition to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
또 다른 방법으로, 상기 항산화제는 담배에 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타낼 수 있다.Alternatively, the antioxidant may be used in tobacco to exhibit antioxidant, antioxidant, or antioxidant enhancement effects.
또 다른 방법으로, 상기 항산화제는 비누, 샴푸, 세정제, 입욕용품으로 이루어진 일군에서 선택된 하나에 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타낼 수 있다.In another method, the antioxidant may be used in one selected from the group consisting of soap, shampoo, detergent, bathing articles and can exhibit an antioxidant, antioxidant, or antioxidant enhancement effect.
또 다른 방법으로, 상기 항산화제는 식용유지류 또는 산업용오일의 산패 방지를 위해 사용될 수 있다.Alternatively, the antioxidant may be used to prevent rancidity of edible fats and oils or industrial oils.
또 다른 방법으로, 상기 항산화제는 동물사료에 첨가제로 사용될 수 있다.Alternatively, the antioxidant can be used as an additive in animal feed.
또 다른 방법으로, 상기 항산화제는 곡물의 보존을 위해 첨가제로 사용될 수 있다.Alternatively, the antioxidant can be used as an additive to preserve the grain.
또 다른 방법으로, 상기 항산화제는 꽃, 과일, 채소, 육류, 가금류, 및 어류로 이루어진 일군에서 선택된 하나의 산화 방지를 위해 사용될 수 있다.Alternatively, the antioxidant may be used to prevent oxidation of one selected from the group consisting of flowers, fruits, vegetables, meat, poultry, and fish.
또 다른 방법으로, 상기 항산화제는 플라스틱, 고무, 접착제, 섬유, 폴리머막, 및 폴리머코팅제로 이루어진 일군에서 선택된 하나의 산화 방지를 위해 사용될 수 있다.Alternatively, the antioxidant may be used to prevent one oxidation selected from the group consisting of plastics, rubber, adhesives, fibers, polymer film, and polymer coating agent.
또 다른 방법으로, 상기 항산화제는 세라믹재료, 금속재료, 및 이들이 조합된 복합재료로 이루어진 일군에서 선택된 하나의 벌크 또는 표면의 산화방지를 위해 사용될 수 있다.Alternatively, the antioxidant may be used to prevent oxidation of one bulk or surface selected from the group consisting of ceramic materials, metal materials, and composite materials in combination thereof.
본 발명에 따른 표면기능화 나노다이아몬드 화합물이 분산된 수용액에 대하여 ORAC 방법을 이용하여 항산화효과를 측정한 결과 나노다이아몬드의 표면에 화학적으로 결합된 부분(moiety)의 화학구조에 따른 다소의 차이는 있으나 전반적으로 항산화효과를 가지고 있음을 확인하였다. 특히 말단기가 아민으로 부착된 화합물이 가장 우수한 항산화 효과를 나타내었다. 표면기능화 시키지 않은 프리스틴(pristine) 나노다이아몬드와 비교하여, 본 발명의 나노다이아몬드 화합물은 액상에서 우수한 분산성을 보여 주므로 먹는 물, 음료, 주류, 식품, 화장품, 의약, 코팅재료, 구조재료 등에 첨가제로 사용되어 항산화 효과를 나타낼 수 있다.As a result of measuring the antioxidant effect of the aqueous solution of the surface functionalized nanodiamond compound according to the present invention using ORAC method, there are some differences depending on the chemical structure of the moiety chemically bonded to the surface of the nanodiamond. It was confirmed that it has an antioxidant effect. In particular, compounds having terminal groups attached with amines showed the best antioxidant effects. Compared to pristine nanodiamonds without surface functionalization, the nanodiamond compounds of the present invention show excellent dispersibility in the liquid phase, so that they can be used as additives in drinking water, beverages, alcohol, food, cosmetics, medicine, coating materials, structural materials, etc. Can be used to show antioxidant effects.
도 1. 아민기를 갖는 표면기능화 나노다이아몬드 (ND) 화합물 중 하나의 모식도.Schematic diagram of one of the surface functionalized nanodiamond (ND) compounds having an amine group.
도 2. 표면기능화 나노다이아몬드 화합물의 FTIR 스펙트럼. 2. FTIR spectra of surface functionalized nanodiamond compounds.
도 3. 프리스틴 다이아몬드와 표면기능화 나노다이아몬드 화합물의 입도 분포.3. Particle size distribution of pristine diamond and surface functionalized nanodiamond compound.
도 4. Trolox 농도에 따른 fluorescein 형광 감쇄 그래프.Figure 4. Graph of fluorescein fluorescence attenuation according to Trolox concentration.
도 5. ORAC assay에 있어서 trolox의 표준 그래프.Figure 5. Standard graph of trolox in ORAC assay.
도 6. 프리스틴 나노다이아몬드의 형광 감쇄 그래프. Figure 6. Fluorescence attenuation graph of pristine nanodiamonds.
도 7. 시료의 농도에 따른 형광 감쇄 그래프. Figure 7. Fluorescence attenuation graph according to the concentration of the sample.
도 8. 5 uM 표면기능화 나노다이아몬드 화합물에 의한 형광 감쇄 그래프. 8. Graph of fluorescence attenuation by 5 uM surface functionalized nanodiamond compound.
도 9. 농도 및 시료에 따른 형광 감쇄 그래프의 면적 비교. 9. Area comparison of fluorescence attenuation graph with concentration and sample.
도 10. 비타민 C, 정향, 및 기능성 나노다이아몬드 화합물의 ORAC 지수.10. ORAC index of vitamin C, cloves, and functional nanodiamond compounds.
나노다이아몬드는 물리적, 화학적으로 안정할 뿐만 아니라 생체 독성이 없기 때문에 새로운 나노소재로 각광받고 있다. 특히 나노다이아몬드 중에서도 10 nm 이하의 크기를 갖는 입자들을 특별히 Ultrananocrystalline diamond(UNCD) 정의하여 이의 응용 범위가 급속도로 확산되고 있다. UNCD는 직경이 5 nm 내외의 비교적 균일한 입도분포를 가지는 초미세 크기의 다이아몬드 결정으로서 주로 폭약 폭발법에 의해 합성된다. 그러나 분산성의 문제로 인하여 제한적인 범위에서 사용되어 왔는데 이를 극복하기 위하여 표면을 개질하는 다양한 방법이 제시되어 왔다. 수 나노미터 크기의 나노다이아몬드는 큰 표면적으로 인하여 기체상 또는 액상에서 여러 화학적 합성 방법을 통해 다양한 기능성기를 나노다이아몬드의 표면에 다량으로 부착시킬 수 있다. 이러한 기능성 나노다이아몬드의 항산화 효과에 대하여는 아직 발표된 바가 없다. Nanodiamonds are attracting attention as new nanomaterials because they are not only physically and chemically stable but also have no biotoxicity. Particularly, among nanodiamonds, particles having a size of 10 nm or less are specifically defined as ultrananocrystalline diamond (UNCD), and their application range is rapidly spreading. UNCD is an ultrafine diamond crystal with a relatively uniform particle size distribution of about 5 nm in diameter, and is mainly synthesized by explosive explosion. However, due to the problem of dispersibility, it has been used in a limited range, and various methods of modifying the surface have been proposed to overcome this problem. Due to the large surface area, nanodiamonds of several nanometers in size can attach a large amount of various functional groups to the surface of nanodiamonds through various chemical synthesis methods in gas phase or liquid phase. The antioxidant effect of such functional nanodiamonds has not been published yet.
항산화 효과를 측정하는 방법으로서 FRAP(ferric reducing/antioxidant power)와 TEAC(Trolox equivalent antioxidant capacity), ORAC(oxygen radical absorbance capacity) 등이 알려져 있다. FRAP과 TEAC은 single electron transfer 를 이용하는 방법이며 ORAC은 hydrogen atom transfer를 이용하는 방법이다. FRAP과 TEAC은 연쇄 반응의 메커니즘에 근거를 두고 발전된 것이 아니기 때문에 반드시 항산화 활성을 측정한다고 보기 어렵다. 항산화 연쇄반응을 제어하는 메커니즘에 근거를 둔 ORAC은 1993년 미국 NIH 산하 국립 노화 연구소의 연구자들에 의해 처음 개발되었으며, 그 이후로 항산화 효과를 측정하는 골드 스탠다드 방법으로 인식되고 있다. As a method of measuring antioxidant effects, ferric reducing / antioxidant power (FRAP), trolox equivalent antioxidant capacity (TEAC), oxygen radical absorbance capacity (ORAC), and the like are known. FRAP and TEAC use single electron transfer and ORAC uses hydrogen atom transfer. Since FRAP and TEAC were not developed based on the mechanism of the chain reaction, it is difficult to measure antioxidant activity. Based on the mechanisms that control antioxidant chain reactions, ORAC was first developed in 1993 by researchers at the National Institute of Aging under the NIH, and has since been recognized as a gold standard method of measuring antioxidant effects.
ORAC 프로토콜을 이용한 항산화 효과 측정 방법은 다음과 같다. 형광염료중의 하나인 disodium fluorescein(FL)은 산화제의 존재하에서 표백 효과에 의하여 형광이 소거된다. 이 용액에 항산화 효과가 있으리라고 기대되는 시료를 같이 혼합하게 되면 FL의 산화를 막아 주게 되어 형광이 더 오래 지속한다. 즉 형광물질이 산화제에 의해 산화 반응이 일어나게 되면 형광의 강도가 감소한다. 그러나 용액 속에 항산화제가 존재하게 되면 형광 물질의 파괴 속도가 감소하여 형광 강도는 증가한다. 형광의 세기를 시간의 함수로 측정하여 얻는 감쇄 곡선 그래프로부터 하부 면적을 계산한다. 실제 실험에서는 산화제로는 2,2’-azobis(2-amidinopropane) dihydrochloride (AAPH)를 사용한다. AAPH의 반감기는 중성 pH 및 37 ˚C에서 175시간 정도이다. 형광 세기를 측정하는 시간은 조건에 따라 다르지만 대략 30-60분 정도이다. Antioxidant effect measurement method using ORAC protocol is as follows. Disodium fluorescein (FL), one of the fluorescent dyes, is fluoresced by the bleaching effect in the presence of an oxidizing agent. Mixing samples that are expected to have an antioxidant effect on the solution prevents the oxidation of FL, resulting in longer fluorescence. That is, the intensity of the fluorescence is reduced when the fluorescent material is oxidized by the oxidizing agent. However, the presence of antioxidants in the solution decreases the rate of destruction of the fluorescent material and increases the fluorescence intensity. The lower area is calculated from the attenuation curve graph obtained by measuring the intensity of the fluorescence as a function of time. In the actual experiment, 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) was used as the oxidant. The half-life of AAPH is about 175 hours at neutral pH and 37 ° C. The time to measure the fluorescence intensity varies depending on the conditions but is approximately 30-60 minutes.
형광의 산화적 파괴 정도를 표준 항산화제인 Trolox(항산화제인 수용성 비타민 E 유사물질)와 비교하여 얻는 값이 ORAC 값이다. 상이한 Trolox 농도를 사용하여 표준 곡선을 만든 다음 이것과 샘플을 비교한다. 최종 결과를 Trolox Equivalent(TE)로 표시한다. ORAC 지수는 현재 전세계적으로 가장 주목받고 있는 항산화 효과 측정 방법이며, ORAC 수치가 높을 수록 활성 항산화력이 높다는 것을 의미한다. The value obtained by comparing the degree of oxidative destruction of fluorescence with the standard antioxidant Trolox (an antioxidant water-soluble vitamin E analog) is the ORAC value. A standard curve is created using different Trolox concentrations and then compared to the sample. The final result is expressed as Trolox Equivalent (TE). The ORAC index is currently the world's most popular method of measuring antioxidant effects, and the higher the ORAC level, the higher the active antioxidant activity.
본 발명에서는 나노다이아몬드의 표면을 화학적으로 개질한 기능성 나노다이아몬드를 항산화제 또는 산화방지제로 사용할 목적으로 ORAC 실험을 수행하였다. 먼저 프리스틴 나노다이아몬드에 대한 ORAC 실험을 수행하였는데, 분산성이 높지 않음에도 불구하고 일정 수준의 항산화 효과가 있음을 보여주고 있다. 기능성 나노다이아몬드의 항산화 효과의 정도는 표면에 부착되어 있는 작용기의 종류에 따라 달라지는데, 특히 아민기가 부착된 부분(moiety)을 갖는 나노다이아몬드 (이하 아민화 나노다이아몬드라 한다)가 가장 우수한 항산화 효과를 보이는 것으로 나타났다. 이러한 항산화 나노다이아몬드는 식품, 화장품, 의약, 농업, 석유화학제품, 폴리머, 세라믹, 금속 등 다방면의 분야에 응용될 수 있을 것으로 보인다.In the present invention, ORAC experiments were performed for the purpose of using functionally-modified nanodiamonds chemically modified on the surface of nanodiamonds as antioxidants or antioxidants. First, ORAC experiments were performed on pristine nanodiamonds, which showed a certain level of antioxidant effect despite the high dispersibility. The degree of antioxidant effect of functional nanodiamonds depends on the type of functional groups attached to the surface, especially nanodiamonds having a moiety attached to amine groups (hereinafter referred to as aminated nanodiamonds) exhibit the best antioxidant effects. Appeared. These antioxidant nanodiamonds may be applied to various fields such as food, cosmetics, medicine, agriculture, petrochemicals, polymers, ceramics and metals.
도 1은 아민기를 갖는 나노다이아몬드 (ND) 화합물 중 하나의 모식도이다. 상기 모식도의 중심부는 나노다이아몬드를 나타내며 표면에는 아민기가 공유결합에 의하여 다량으로 부착되어 있다. 본 발명에서, 아민화 나노다이아몬드 화합물은 액상에서 프리스틴 나노다이아몬드로부터 하기 여러 실시예들에서 보여 주는 바와 같이 여러 단계의 제조 공정을 거쳐 합성한다. 이러한 기능화 나노다이아몬드는 프리스틴 나노다이아몬드와 비교하여 우수한 분산성을 보여주며, 항산화 효과 즉 ORAC 지수가 대단히 높다.1 is a schematic diagram of one of the nanodiamond (ND) compounds having an amine group. The central portion of the schematic diagram represents nanodiamonds, and amine groups are attached to the surface in large amounts by covalent bonds. In the present invention, the aminated nanodiamond compound is synthesized from the pristine nanodiamond in a liquid phase through a multi-step manufacturing process as shown in various examples below. These functionalized nanodiamonds show excellent dispersibility compared to pristine nanodiamonds, and have a very high antioxidant effect, i.e., an ORAC index.
이하의 실시예는 본 발명의 이해를 돕기 위한 구체적인 예를 상세히 설명하는 것으로서 본 발명의 범위가 이들 실시예에 의해 한정되는 것은 아니다.The following examples illustrate specific examples to aid in understanding the present invention in detail, and the scope of the present invention is not limited to these examples.
<실시예 1><Example 1>
HNO3(70%)와 H2SO4(98%)를 1:3 중량비로 배합한 강산 용액에 직경 5 nm 내외의 프리스틴 나노다이아몬드를 첨가하여 40˚C에서 3시간 초음파 처리한다. 이 용액을 증류수로 희석한 후 막 필터를 사용하여 여과시킨 후 80˚C 오븐에서 4시간 건조시켜 나노다이아몬드 표면에 복수개의 카르복실기가 부착된 ND-(COOH)n 화합물 분말을 얻는다. To a strong acid solution containing HNO 3 (70%) and H 2 SO 4 (98%) in a 1: 3 weight ratio, prestained nanodiamonds with a diameter of about 5 nm were added and sonicated at 40 ° C. for 3 hours. The solution was diluted with distilled water, filtered using a membrane filter, and then dried in an oven at 80 ° C. for 4 hours to obtain ND- (COOH) n compound powder having a plurality of carboxyl groups attached to the nanodiamond surface.
<실시예 2><Example 2>
BH3/THF 30mL에 상기 ND-(COOH)n 화합물 100mg을 첨가한 후 1시간 동안 초음파 처리한 후 상기 실시예 1과 동일한 여과 건조 과정을 거쳐 나노다이아몬드 표면에 수산기를 포함한 부분(moiety)이 복수개 부착된 ND-(CH2OH)n를 얻는다. 100 mg of the ND- (COOH) n compound was added to 30 mL of BH 3 / THF, followed by sonication for 1 hour, followed by the same filtration and drying process as in Example 1, where a plurality of moieties including hydroxyl groups were present on the surface of the nanodiamond. Obtain ND- (CH 2 OH) n attached.
<실시예 3><Example 3>
100 mg의 상기 ND-(COOH)n 분말을 50 mL의 ethylenediamine에 녹인 후 50 mg HATU 시약을 첨가하고 4시간 동안 초음파 처리하여 아마이드기를 갖는 나노다이아몬드를 제조한다. 상기 반응물을 200 mL의 methanol을 사용하여 희석시킨 후 여과, 건조하여 나노다이아몬드 표면에 아마이드기 및 아민기를 포함한 부분(moiety)이 복수개 부착된 ND-(CONHCH2CH2NH2)n 화합물 분말을 얻는다. 100 mg of the ND- (COOH) n powder was dissolved in 50 mL of ethylenediamine, and then 50 mg HATU reagent was added and sonicated for 4 hours to prepare nanodiamonds having an amide group. The reaction was diluted with 200 mL of methanol, filtered and dried to obtain ND- (CONHCH 2 CH 2 NH 2 ) n compound powder having a plurality of moieties including amide groups and amine groups on the surface of nanodiamonds. .
<실시예 4><Example 4>
나노다이아몬드 표면에 아민기(NH2)를 도입하기 위하여 100 mg의 상기 ND-(CH2OH)n 분말을 30 ml THF 내에서 30분간 초음파 처리한다. 여기에 10 mg의 diethylazodicarboxylate coupling agent와 50 mg의 phthalimide를 첨가하여 2시간 동안 초음파 처리한 후 300 mL의 methanol를 부어 희석시킨 후 여과, 건조한다. 이 분말을 50 mL의 trifluoroacetic acid (TFA)에 넣은 후 3시간 동안 초음파 처리한 후에 여과, 건조하여 나노다이아몬드 표면에 아민기를 포함한 부분(moiety)이 복수개 부착된 ND-(CH2NH2)n 분말을 얻는다.100 mg of the ND- (CH 2 OH) n powder was sonicated in 30 ml THF for 30 minutes to introduce an amine group (NH 2 ) to the nanodiamond surface. 10 mg of diethylazodicarboxylate coupling agent and 50 mg of phthalimide were added thereto, sonicated for 2 hours, 300 mL of methanol was poured out, diluted, filtered and dried. This powder was placed in 50 mL of trifluoroacetic acid (TFA), sonicated for 3 hours, filtered and dried to provide ND- (CH 2 NH 2 ) n powder having a plurality of moieties containing amine groups on the surface of nanodiamonds. Get
<실시예 5> Example 5
상기 표면기능화 ND 화합물들의 표면 개질 여부는 FTIR (Varian)을 이용하여 KBR pellet 의 형태로 시료를 만들어 분석하였다. The surface modification of the surface functionalized ND compounds was analyzed by making samples in the form of KBR pellets using FTIR (Varian).
도 2는 ND-(R)n 화합물들의 IR 스펙트럼들을 도시한 것이다. 2 shows IR spectra of ND- (R) n compounds.
도 2의 ND-(COOH)n 화합물의 스펙트럼상 1670 cm-1에서 나타나는 강한 peak는 C=O stretching에 의한 것이다. ND-(CH2OH)n 화합물에서는 C=O stretching 피크가 사라지고 methylene group에서의 C-H stretching 진동모드를 보여주는 2923 cm-1와 2845 cm-1에서 피크가 나타난다. ND-(CONHCH2CH2NH2)n 화합물의 IR 스펙트럼을 보면 C-N stretching 진동모드가 1030 cm-1에서 나타난다. 또한 carbonyl stretching에 해당하는 진동 모드가 1679 cm-1에서 나타남을 알 수 있다. 마지막으로, ND-(CH2NH2)n 화합물은 아민기의 in-plane bending mode가 1624 cm-1에서, out-of-plane bending mode가 737 cm-1에서 관찰됨을 알 수 있다. The strong peaks appearing at 1670 cm −1 in the spectrum of the ND- (COOH) n compound of FIG. 2 are due to C═O stretching. In the ND- (CH 2 OH) n compound, the C═O stretching peak disappears and peaks appear at 2923 cm −1 and 2845 cm −1 , showing the CH stretching oscillation mode in the methylene group. The IR spectrum of the ND- (CONHCH 2 CH 2 NH 2 ) n compound shows a CN stretching oscillation mode at 1030 cm −1 . In addition, it can be seen that the vibration mode corresponding to carbonyl stretching appears at 1679 cm -1 . Finally, it can be seen that the ND- (CH 2 NH 2 ) n compound has an in-plane bending mode of amine group at 1624 cm −1 and an out-of-plane bending mode at 737 cm −1 .
<실시예 6> <Example 6>
도 3은 수용액에서 프리스틴 나노다이아몬드와 아민화 나노다이아몬드의 입도 분포를 보여준다. 입도 분석에 대한 측정은 동적 광산란 장치(Scatteroscope, Qudix)를 이용하였다. 프리스틴 나노다이아몬드는 평균 입자의 직경이 16 nm와 20 μm에서 나타나는 bimodal 패턴이 나타나는데 이는 다수의 나노다이아몬드가 액상에서 응집체로 존재하고 있음을 의미한다. 반면에, 나노다이아몬드는 평균 입자의 직경이 8 nm로서 수용액에서 응집체를 형성함이 없이 단일 입자들로 분포함을 보여주고 있다. 결론적으로, 나노다이아몬드 표면의 화학적 개질은 나노다이아몬드에 화학적 기능성은 물론 우수한 분산성도 함께 제공하는 효과가 있다. Figure 3 shows the particle size distribution of pristine nanodiamonds and aminated nanodiamonds in aqueous solution. The measurement for the particle size analysis was performed using a dynamic light scattering device (Scatteroscope, Qudix). Pristine nanodiamonds show a bimodal pattern with average particle diameters of 16 nm and 20 μm, indicating that a large number of nanodiamonds are aggregated in the liquid phase. On the other hand, nanodiamonds have an average particle diameter of 8 nm, showing that they are distributed as single particles without forming aggregates in aqueous solution. In conclusion, the chemical modification of the surface of the nanodiamond has the effect of providing the nanodiamond with excellent dispersibility as well as chemical functionality.
<실시예 7> <Example 7>
아민기가 수용액상에서 염기성을 나타내므로 아민화 나노다이아몬드를 물에 분산시키면 염기성을 나타낸다. 표 1은 ND-(CH2NH2)n 0.001~10 μM 농도 범위에서 측정한 수용액의 pH이다. 농도 계산에 필요한 나노다이아몬드의 분자량 값은 입자의 크기를 고려하여 산정한 값인 100,000을 사용하였다. Since the amine group shows basicity in aqueous solution, dispersing the aminated nanodiamond in water shows basicity. Table 1 shows the pH of the aqueous solution measured in the concentration range of ND- (CH 2 NH 2 ) n 0.001 to 10 μM. As a molecular weight value of the nanodiamond required for the concentration calculation, 100,000, which is a value calculated in consideration of the particle size, was used.
표 1
Conc. (μM) 0.001 0.05 0.1 0.5 1 10
pH 7.06 7.27 7.39 7.51 7.83 7.99
Table 1
Conc. (μM) 0.001 0.05 0.1 0.5 One 10
pH 7.06 7.27 7.39 7.51 7.83 7.99
표 1을 참조하면, 수용액의 pH는 ND-(CH2NH2)n 농도의 log10 값에 선형적으로 비례하지는 않음을 알 수 있는바, 이는 약염기성 용질의 전형적인 특징이다. 표 1에서 보여주는 결과는 아민화 나노다이아몬드를 물이나 주류, 또는 식품 등에 첨가하여 pH를 높일 수 있으며, 필요한 경우 산성 또는 중성의 물을 약알칼리성으로 변환시키는데 사용될 수 있음을 의미한다. 한 예로, 중성의 물 1 L에 아민화 나노다이아몬드 10 mg 을 첨가하면 생리적인 pH 값인 7.4 에 가까운 산도의 수용액을 만들 수 있다. Referring to Table 1, it can be seen that the pH of the aqueous solution is not linearly proportional to the log 10 value of the ND- (CH 2 NH 2 ) n concentration, which is a typical characteristic of the weakly basic solute. The results shown in Table 1 indicate that the aminated nanodiamonds can be added to water, liquor, or food to increase the pH, and can be used to convert acidic or neutral water into weakly alkaline if necessary. For example, adding 10 mg of aminated nanodiamonds to 1 L of neutral water can produce an aqueous solution of acidity close to 7.4, the physiological pH value.
<실시예 8> <Example 8>
ORAC assay는 다음 방법으로 진행되었다. 형광 측정은 형광 분광 광도계(F-4500, Hitachi)를 사용하였다. 여기 및 형광측정 파장은 각각 480 nm 와 520 nm를 사용하였고 모든 실험은 상온에서 수행되었다. ORAC assay was carried out by the following method. Fluorescence measurement was performed using a fluorescence spectrophotometer (F-4500, Hitachi). The excitation and fluorescence wavelengths were 480 nm and 520 nm, respectively, and all experiments were performed at room temperature.
0.0263 g의 fluorescein(FL)을 pH 7.4의 phosphate buffer 1 ml에 녹여서 70 μM 의 stock solution을 만들었다. 이 저장 용액(stock solution)은 어두운 곳에서 보관한 후 측정 전에 70 nM의 용액으로 희석하여 사용하였다. 0.1356 g의 2,2’-azobis(2-amidinopropane) dihydrochloride (AAPH)를 phosphate buffer에 녹여서 50 mM의 수용액을 만들었다. 트롤록스(Trolox) 0.0125 g을 10mL의 phosphate buffer에 녹여서 5 mM의 트롤록스 저장 용액을 준비하였다. 이를 1~5μM로 희석하여 트롤록스 calibration curve를 구하는데 사용하였다. 트롤록스의 calibration curve는 FL 용액 (240 μL), AAPH (240 μL), trolox (120 μL) 혼합 용액을 37 ℃ 에서 10분간 incubation한 후 측정하였다. 즉 FL과 AAPH의 buffer 용액의 시간에 따른 형광 세기를 측정한 후, 트롤록스 용액을 1μM 간격으로 5 μM 까지 혼합한 용액의 형광 감쇄 그래프를 일련적으로 얻는다. 0.0263 g of fluorescein (FL) was dissolved in 1 ml of phosphate buffer at pH 7.4 to make 70 μM of stock solution. This stock solution was stored in the dark and diluted with 70 nM of solution prior to measurement. 0.1356 g of 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) was dissolved in phosphate buffer to form a 50 mM aqueous solution. 5 mM Trolox stock solution was prepared by dissolving 0.0125 g of Trolox in 10 mL of phosphate buffer. It was diluted to 1 ~ 5μM and used to calculate trolox calibration curve. The calibration curve of trolox was measured after incubation of FL solution (240 μL), AAPH (240 μL), and trolox (120 μL) for 10 minutes at 37 ° C. That is, after measuring the fluorescence intensity over time of the buffer solution of FL and AAPH, a series of fluorescence attenuation graphs of the solution in which the trolox solution is mixed to 5 μM at 1 μM intervals are obtained in series.
형광 측정에 사용한 최종 전체 용액의 부피는 600 μL 이며 형광 세기는 25초 간격으로 3,600초(60분)까지 측정하였다. 항산화제의 형광 감쇄 그래프를 얻을 때의 부피비는 FL: AAPH: sample = 2:2:1을 사용하였으며 형광 측정 전에 37˚C에서 10분간 incubation 하였다. 형광을 찍기 위하여 FL 용액(240 μL), AAPH (240 μL), antioxidant 용액 (120 μL)을 혼합하여 37 ℃ 에서 10분간 incubation 하여 형광을 60분간 측정하였다. Blank는 buffer (120 μL), fluorescein solution (240 μL), 및 AAPH (240 μL)의 혼합액을 37 ℃ 에서 10분간 incubation 한 후 측정하였다.The final total solution used for fluorescence measurement was 600 μL and the fluorescence intensity was measured up to 3,600 seconds (60 minutes) at 25 second intervals. When the fluorescence attenuation graph of the antioxidant was obtained, the volume ratio of FL: AAPH: sample = 2: 2: 1 was used and incubated at 37 ° C for 10 minutes before fluorescence measurement. For fluorescence, FL solution (240 μL), AAPH (240 μL), and antioxidant solution (120 μL) were mixed and incubated at 37 ° C. for 10 minutes to measure fluorescence for 60 minutes. Blank was measured after incubation of a mixture of buffer (120 μL), fluorescein solution (240 μL), and AAPH (240 μL) at 37 ° C. for 10 minutes.
<실시예 9>Example 9
형광 감쇄 그래프의 하부곡선 면적(area under curve;AUC)을 계산하기 위하여 모든 형광 감쇄 그래프를 최대 규격화(peak normalization)한 후 AUC는 다음 식을 사용하여 계산하였다.  AUC was calculated using the following equation after peak normalization of all fluorescence attenuation graphs to calculate the area under curve (AUC) of the fluorescence attenuation graph.
Figure PCTKR2010001688-appb-I000001
Figure PCTKR2010001688-appb-I000001
여기서, f 0 는 시간 0에서의 형광 세기이며 n은 형광 감쇄 그래프를 얻기 위한 데이터 개수이다. Here, f 0 is the fluorescence intensity at time 0 and n is the number of data for obtaining the fluorescence attenuation graph.
AUC로부터 상대적 ORAC 수치(Trolox equivalent)는 다음 식으로 계산된다. The relative ORAC value (Trolox equivalent) from the AUC is calculated by the following equation.
ORAC = [(AUCsample-AUCblank)/(AUCsample-AUCblank)]ORAC = [(AUC sample -AUC blank ) / (AUC sample -AUC blank )]
x (molarity of trolox / molaraity of sample) x (molarity of trolox / molaraity of sample)
도 4는 트롤록스 농도에 따른 FL 형광 감쇄 그래프를 보여주며, 도 5는 이로부터 계산한 trolox calibration curve이다. 최소 자승법으로 얻은 caclibration curve는 y = 4.45x - 7.56이다. 여기서 x는 트롤록스 농도(μM)를, y는 AUC를 나타낸다. 상관계수 (R2) 값은 0.99 이상으로서 아주 만족할 만한 수준의 값을 보여 주고 있다.4 shows a FL fluorescence attenuation graph according to trolox concentration, and FIG. 5 is a trolox calibration curve calculated therefrom. The caclibration curve obtained by the least squares method is y = 4.45x-7.56. Where x is the trolox concentration (μM) and y is the AUC. The correlation coefficient (R 2 ) is above 0.99, indicating a very satisfactory level.
<실시예 10> <Example 10>
도 6은 프리스틴 나노다이아몬드의 형광 감쇄 그래프이다. 농도 계산에 필요한 프리스틴 다이아몬드의 분자량 값은 나노입자의 사이즈를 고려하여 계산한 100,000을 사용하였다. 도 6으로부터 1~5μM 농도 범위에서 AUC를 각각 계산하였으며 이로부터 얻은 상대적 ORAC 수치의 평균값은 5.2이다. 프리스틴 나노다이아몬드가 항산화 효과를 보여주는 것은 주로 표면의 sp2 구조에 의한 라디칼 제거 기능에 의한 것으로 보이지만 자세한 메카니즘은 아직 알려져 있지 않다.6 is a graph showing fluorescence attenuation of pristine nanodiamonds. As the molecular weight value of the pristine diamond required for the concentration calculation, 100,000 was calculated based on the size of the nanoparticles. The AUCs were calculated for each of the concentration ranges from 1 to 5 μM from FIG. 6 and the average value of the relative ORAC values obtained therefrom was 5.2. Although pristine nanodiamonds exhibit antioxidant effects, they appear to be mainly due to the radical scavenging function of the surface sp 2 structure, but the detailed mechanism is not yet known.
<실시예 11> <Example 11>
도 7은 비타민 C, 정향(clove), 표면기능화 나노다이아몬드들의 형광 감쇄 그래프를 보여준다. 비타민 C의 항산화 효과는 잘 알려져 있으므로 하나의 대조군(control)으로 선택하였고, 정향은 지금까지 알려진 천연물 중에 가장 ORAC 수치가 높다고 알려져 있으므로 상대적 비교를 위해 선택하였다. 이들의 항산화 정도의 상대적인 비교를 위하여 특별히 5 μM에서의 각 샘플의 형광 감쇄 그래프를 도 8에 나타내었다. 이 농도에서 아민화 나노다이아몬드는 60분간 형광 세기가 거의 변하지 않고 일정한 값을 유지하고 있음을 보여준다. 도 9는 각 샘플의 AUC를 측정 농도별로 도시한 것이다. Figure 7 shows the fluorescence attenuation graph of vitamin C, clove, surface functionalized nanodiamonds. Since the antioxidant effect of vitamin C is well known, it was selected as a control. Clove was selected for relative comparison because it is known to have the highest ORAC level among natural products known to date. For the relative comparison of their antioxidant levels, a graph of fluorescence attenuation of each sample, especially at 5 μM, is shown in FIG. 8. At this concentration, the aminated nanodiamond shows little change in fluorescence intensity for 60 minutes and remains constant. 9 shows the AUC of each sample by measurement concentration.
<실시예 12><Example 12>
표 2는 상기 식으로 계산한 ORAC 수치를 농도별로 표시한 것이며, 이들의 평균값을 함께 나타내었다. 표면 기능화를 통하여 나노다이아몬드의 분자량이 약간 변할 수 있으나 일관성을 유지하기 위하여 프리스틴 나노다이아몬드와 같은 값인 100,000을 사용하였다. Table 2 shows the ORAC values calculated by the above formula for each concentration, and their average values are also shown. Although the molecular weight of the nanodiamond may be slightly changed through surface functionalization, 100,000, which is the same value as the pristine nanodiamond, was used to maintain consistency.
표 2
Conc. (uM) Vitamin C Clove ND-(COOH)n ND-(CH2OH)n ND-(CONHCH2-CH2NH2)n ND-(CH2NH2)n
1 0.79 17.83 4.94 1.38 8.62 18.12
2 0.78 18.89 1.22 1.17 8.18 18.94
3 0.74 16.76 0.70 1.02 7.88 18.17
4 0.61 14.22 0.76 1.12 5.65 16.47
5 0.51 11.05 0.73 1.01 4.84 12.16
Avg. 0.69 15.75 1.67 1.14 7.03 16.77
TABLE 2
Conc. (uM) Vitamin c Clove ND- (COOH) n ND- (CH 2 OH) n ND- (CONHCH 2 -CH2NH 2 ) n ND- (CH 2 NH 2 ) n
One 0.79 17.83 4.94 1.38 8.62 18.12
2 0.78 18.89 1.22 1.17 8.18 18.94
3 0.74 16.76 0.70 1.02 7.88 18.17
4 0.61 14.22 0.76 1.12 5.65 16.47
5 0.51 11.05 0.73 1.01 4.84 12.16
Avg. 0.69 15.75 1.67 1.14 7.03 16.77
표 2를 참조하면, 본 발명에서 측정한 모든 기능성 나노다이아몬드들이 유의미한 수준의 항산화 효과를 보여주지만 그 중에서 아민기를 가진 나노다이아몬드의 ORAC 값이 가장 크다. 주목할 만한 것으로, 긴 사슬을 갖는 아마이드의 말단에 아민기를 가진 나노다이아몬드 화합물에 비하여, 아마이드기가 없이 표면을 -CH2NH2로 기능화한 아민화 나노다이아몬드가 더 높은 항산화 효과를 나타낸다.Referring to Table 2, all of the functional nanodiamonds measured in the present invention show a significant level of antioxidant effect, among which the ORAC value of the nanodiamonds having the amine group is the largest. Notably, compared to the nanodiamond compound having an amine group at the terminal of the amide having a long chain, the aminated nanodiamond functionalized with -CH 2 NH 2 without an amide group shows a higher antioxidant effect.
도 10는 비타민 C, 정향(clove) 및 기능성 나노다이아몬드의 ORAC 지수를 나타낸 것이다. 아민화 나노다이아몬드는 비타민 C와 비교하여 같은 농도에서 25배의 항산화 효과를 나타내며 또한 정향보다 ORAC 값이 높음을 알 수 있다.Figure 10 shows the ORAC index of vitamin C, clove and functional nanodiamonds. Aminized nanodiamonds exhibited 25-fold antioxidant activity at the same concentrations as compared to vitamin C and also showed higher ORAC values than cloves.

Claims (15)

  1. 하기 화학식 I로 표시되는 표면 기능화 나노다이아몬드 화합물(surface functionalized nanodiamond compouns) 또는 그의 염으로 이루어지는 항산화제. An antioxidant comprising a surface functionalized nanodiamond compouns represented by the following formula (I) or a salt thereof.
    화학식 I: ND-(R)n, Formula I: ND- (R) n ,
    상기 화학식 I에서, n은 2 이상의 정수이고, ND는 나노다이아몬드 단일입자를 나타내며, 복수개의 R은 각각 상기 나노다이아몬드 표면에 화학결합된 부분(moiety)를 나타내며, R은 아민기(NH2), 수산기(OH), 카르복실기(COOH), 및 이들의 조합으로 이루어진 일군에서 선택된 하나를 적어도 1개 이상 포함하는 화학구조를 가진다.In Formula I, n is an integer of 2 or more, ND represents a nanodiamond single particle, a plurality of R each represents a moiety chemically bonded to the surface of the nanodiamond, R represents an amine group (NH 2 ), It has a chemical structure containing at least one selected from the group consisting of hydroxyl group (OH), carboxyl group (COOH), and combinations thereof.
  2. 제 1 항에 있어서, 상기 R은 하기 화학식 II인 것을 특징으로 하는 항산화제.According to claim 1, wherein R is an antioxidant, characterized in that formula II.
    화학식 II: X-NH2, Formula II: X-NH 2 ,
    상기 화학식 II에서, X는 나노다이아몬드 표면에 아민기를 공유 결합으로 연결시키기 위한 링커(linker)로서, 하나 또는 다수의 메틸렌기, 카르복실기, 에테르기, 아마이드기, 에스테르기, 및 이들의 조합으로 이루어진 일군에서 선택된 하나를 포함한다.In Formula II, X is a linker for covalently linking an amine group to a nanodiamond surface, and includes a group consisting of one or more methylene groups, carboxyl groups, ether groups, amide groups, ester groups, and combinations thereof. It includes one selected from.
  3. 제 2 항에 있어서, 상기 R은 상기 화학식 II의 식품영양학적 또는 약학적으로 허용 가능한 염을 더 포함하는 것을 특징으로 하는 항산화제.The antioxidant of claim 2, wherein R further comprises a food nutritional or pharmaceutically acceptable salt of Formula II.
  4. 제 1 항 내지 제 3 항의 어느 하나에 있어서, 물, 음료, 주류, 커피로 이루어진 일군에서 선택된 하나에 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타내는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used for one selected from the group consisting of water, beverages, alcoholic beverages, and coffee, and exhibits antioxidant, antioxidant, or antioxidant enhancement effects.
  5. 제 1 항 내지 3 항의 어느 하나에 있어서, 식품 첨가제로 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타내는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used as a food additive to exhibit antioxidant, antioxidant or antioxidant enhancement effects.
  6. 제 1 항 내지 3 항의 어느 하나에 있어서, 화장품 원료로 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타내는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used as a cosmetic raw material and exhibits antioxidant prevention, antioxidant, or antioxidant enhancement effect.
  7. 제 1 항 내지 3 항의 어느 하나에 있어서, 약제 조성물로 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타내는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used as a pharmaceutical composition to exhibit antioxidant, antioxidant or antioxidant enhancement effects.
  8. 제 1 항 내지 3 항의 어느 하나에 있어서, 담배에 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타내는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used in tobacco to exhibit antioxidant, antioxidant or antioxidant enhancement effects.
  9. 제 1 항 내지 3 항의 어느 하나에 있어서, 비누, 샴푸, 세정제, 입욕용품으로 이루어진 일군에서 선택된 하나에 사용되어 산화 예방, 항산화, 또는 항산화 증진 효과를 나타내는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used for one selected from the group consisting of soaps, shampoos, cleaning agents, and bathing articles, and exhibits antioxidant prevention, antioxidant, or antioxidant enhancement effects.
  10. 제 1 항 내지 3 항의 어느 하나에 있어서, 식용유지류 또는 산업용오일의 산패 방지를 위해 사용되는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used for preventing rancidity of edible fats or oils.
  11. 제 1 항 내지 3 항의 어느 하나에 있어서, 동물사료에 첨가제로 사용되는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used as an additive in animal feed.
  12. 제 1 항 내지 3 항의 어느 하나에 있어서, 곡물의 보존을 위해 첨가제로 사용되는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used as an additive for the preservation of grain.
  13. 제 1 항 내지 3 항의 어느 하나에 있어서, 꽃, 과일, 채소, 육류, 가금류, 및 어류로 이루어진 일군에서 선택된 하나의 산화 방지를 위해 사용되는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used for the oxidation prevention of one selected from the group consisting of flowers, fruits, vegetables, meat, poultry, and fish.
  14. 제 1 항 내지 3 항의 어느 하나에 있어서, 플라스틱, 고무, 접착제, 섬유, 폴리머막, 및 폴리머코팅제로 이루어진 일군에서 선택된 하나의 산화 방지를 위해 사용되는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used for the oxidation prevention of one selected from the group consisting of plastic, rubber, adhesive, fiber, polymer film, and polymer coating agent.
  15. 제 1 항 내지 3 항의 어느 하나에 있어서, 세라믹재료, 금속재료, 및 이들이 조합된 복합재료로 이루어진 일군에서 선택된 하나의 벌크 또는 표면의 산화방지를 위해 사용되는 것을 특징으로 하는 항산화제.The antioxidant according to any one of claims 1 to 3, which is used for the oxidation of one bulk or surface selected from the group consisting of ceramic materials, metal materials, and composite materials in combination thereof.
PCT/KR2010/001688 2010-03-18 2010-03-18 Antioxidant effect of functional nanodiamond and a use therefor WO2011115314A1 (en)

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PL424918A1 (en) * 2018-03-16 2019-09-23 Politechnika Koszalińska Application of anti-oxidative layers with applied detonative nanodiamonds for packages of selected food products, cosmetic products, pharmaceuticals and dietary supplements

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