KR20040017867A - Alkylsulfanyl)methyl]-1,2-benzenediol, Preparation Method thereof and Antioxidant Agent Containing the same - Google Patents

Abstract

PURPOSE: A novel 4-£(alkylsulfanyl)methyl|-1,2-benzenediol derivative, its preparation method and a composition containing the derivative for an antioxidant, a whitening agent or a cholesterol controller are provided, to obtain the compound which suppresses the generation of melanin and controls the concentration of cholesterol by suppressing the oxidation of low density lipoprotein. CONSTITUTION: The 4-£(alkylsulfanyl)methyl|-1,2-benzenediol derivative is represented by the formula 1, wherein R is an alkyl group of C1-C10. The method comprises the step of reacting the 1,3-benzodioxol-5-yl-methanol of the formula 2 with the alkylthiol represented by R-SH in the presence of an acid, wherein R is an alkyl group of C1-C10. Preferably the acid is selected from the group consisting of AlCl3, FeBr3, FeCl3 and BBr3; and the reaction is performed in at least one solvent selected from the group consisting of dichloromethane, chloroform, acetonitrile, tetrahydrofuran and dimethyl formaldehyde.

Description

4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivatives, preparation method thereof and antioxidant including same {4-[{Alkylsulfanyl) methyl] -1,2-benzenediol, Preparation Method about and Antioxidant Agent Containing the same}

The present invention relates to a novel 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative, more particularly 4-[(alkylsulfanyl) methyl] -1, represented by the following formula (1): The present invention relates to a 2-benzenediol derivative, a pharmaceutically acceptable salt thereof, a method for preparing the same, and a composition for antioxidant, whitening agent and cholesterol control agent containing the compound as an active ingredient.

[Formula 1]

In the above formula, R is a C ₁ to C ₁₀ alkyl group.

All aerobic organisms, including humans, survive through energy metabolism using oxygen (O ₂ ). When oxygen in vivo is subjected to various physical, chemical, and biological stresses, superoxide anion radicals (· O ^2- ; superoxide anion) radicals, hydrogen peroxide (H ₂ O ₂ ), and hydroxy radicals (· OH; hydroxy radicals) such as active oxygen species (active oxygen species) are converted into. Superoxide anions, which are produced during phagocytic cell respiratory bursts, play an important role in destroying bacteria and viruses that invade from the outside, but these reactive oxygen species are more than necessary due to various factors. When produced, it causes fatal physiological disorders in the human body, and in severe cases, causes disease and loses life.

Although living organisms are thought to have evolved by developing antioxidative mechanisms as a self-defense mechanism that removes reactive oxygen species, the development of reactive oxygen species that transcends the defenses of tissues is more sensitive to immunity such as proteins, DNA, enzymes and T cells. Damage to the system factors cause various diseases. In addition, reactive oxygen species attack perunsaturated fatty acids, which are components of cell membranes, to cause peroxidation, and the lipid peroxides accumulated in vivo may cause aging, cancer, multiple atherosclerosis, arthritis, and Parkinson's disease. It is known to cause a variety of diseases, such as disease.

Recently, as the theory that aging and geriatric diseases are caused by reactive oxygen species has been recognized, research on the development of antioxidants known as substances capable of regulating reactive oxygen species has been actively conducted, leading to superoxide dismutase and peroxide. Antioxidants such as oxidase, catalase, and glutathione peroxidase, as well as antioxidants such as tocopherol, ascorbate, carotenoid, and glutathione. Much research has been done on 2,6'-dibutyl-4-hydroxytoluene (BHT), 2,6'-dibutyl-4-hydroxy an Many synthetic antioxidants such as sol (2,6-dibutyl-4-hydroxyanisole, BHA) have been developed and used in medicine and food.

On the other hand, human skin produces melanin to eliminate naturally occurring free radicals, free radicals, and the like, or to prevent ultraviolet rays from penetrating. The starting material for making melanin is a type of amino acid normally present in the human body. Tyrosine. Tyrosine is oxidized by tyrosinase in melanocytes to turn into dopa (3,4-dihydroxyphenylalanine, abbreviated as "DOPA"), and DOPA is further oxidized to DOPA quinone. . DOPA quinones then undergo an auto-oxidation reaction through 5,6-dihydroxyindole to form indole-5,6-quinone, which finally produces a dark brown melanin [Goldsmith, LA] , Physiology, Biochemistry, and Molecular Biology of the Skin, Oxford University Press (1991).

To prevent excessive deposition of melanin in the skin, there are methods to block each step of the melanin biosynthesis pathway, and (i) prevent skin exposure to ultraviolet rays, (ii) use tyrosinase inhibitors, ) Can be classified into methods such as administration of a substance that is specifically toxic to melanocytes and (i) promoting the release of the melanin produced to the outside of the skin.

The substances to prevent hyper pigmentation on the skin are classified according to the mechanism of action, and there are ultraviolet light scattering agents such as ultraviolet absorbers and inorganic pigments to block the ultraviolet rays irradiated to the skin, and vitamin C or kojic acid. Substances that are toxic to melanocytes such as tyrosinase inhibitors such as hydroquinone), free radicals that promote melanogenesis or tocopherols that free radicals, and AHAs that remove melanin by promoting exfoliation. -hydroxyl acid). Among them, arbutin, which binds sugar to hydroquinone, which is known to be toxic to melanocytes, is well known as a tyrosinase inhibitor along with kojic acid and is a form of functional cosmetics for the purpose of inhibiting skin whitening and aging. It is used a lot in. In addition, retinol and ascorbic acid, which are known to inhibit skin aging, are also attracting much attention both at home and abroad, and many products are commercially available. The mechanism of action is not yet known, but studies have shown that the inhibitory action of tyrosinase enzymes is associated with whitening and even anti-aging effects [Curto, EV et al ., Biochemical Pharmacology , 57 , 663]. -672 (1999); Cabanes, J. et al ., J. Pharm. Pharmacol ., 46 , 982-985 (1994)].

As such, tyrosinase inhibitors have been developed and used as various whitening agents, but various problems have been raised at the same time. Indeed, 4-hydroxyanisol and hydroquinone, which are used locally for the treatment of hyperpigmentation such as blemishes, freckles, spots, and hyperpigmentation in pregnancy, have potent melanogenesis inhibitory activities, but at the same time induce degeneration or lethality of pigment cells. Side effects such as impaired original function. In particular, hydroquinone-based compounds have been developed and used as a whitening cream that inhibits melanin biosynthesis, but are known to cause skin irritation or skin diseases due to cytotoxicity, and are currently used only in some countries.

On the other hand, the occurrence of atherosclerosis in humans has been reported to be highly correlated with blood cholesterol levels due to the ideal differentiation of arterial wall cells [Casteli, WP et al ., JAMA., 256, 2835-2845 (1986). ]. The amount of cholesterol delivered to the artery wall by the low density lipoprotein (abbreviated as "LDL"), the main substance that carries blood cholesterol, is dependent on blood concentration. Oxidation of LDL into the artery wall is considered an inflammatory response, which causes monocytes to migrate to this site and macrophages by monocyte-colony stimulating factor (M-CSF), which is secreted by vascular endothelial cells. (macrophage) to differentiate. The macrophages further promote LDL oxidation, and the oxidized LDL accumulates in the macrophages by scavenger receptors, thereby losing the mechanism of macrophages to accumulate cholesterol and accumulating high levels of cholesterol. . At this time, the smooth muscle cells in the middle layer of the artery wall are transferred to the vascular wall endothelial layer, and together with the macrophages, the cells become fat cells, which form fat streak, which is an initial state of arteriosclerosis. The resulting foam cells secrete extracellular matrix to form fibrous caps, and when they form and rupture, plaque forms in the arterial wall, causing myocardial infarction, heart attack or angina. These cholesterol-induced endothelial dysfunctions, ie oxidative modification of LDL, are known to play an important role in the mechanism of arterial wall hyperplasia [Aviram, M. et al ., J. Cardiovascular Pharmacology, 31, 39-45 (1998). Therefore, there is a high possibility of inducing atherosclerosis in the vascular system in which LDL oxidation is actively progressed [Miesenock, G. et al ., Leaf, A. and Webber, PC (ed), Raven Press, New York, vol, 21, p 119-123 (1990). Oxidation of LDL increases lipid peroxide and oxygen free radicals, or oxygen free radicals, and these molecules are toxic to endothelial cells, and oxidized LDL easily attaches to blood vessel walls It damages cells to modify blood vessel tissues and at the same time promotes the division of deformed cells, making it easier to attach surrounding oxidized LDL, platelets and macrophages to the vessel wall [Noguchi, N. et al . Archiv. Biochem. Biophys., 3347, 141-147 (1997).

Atherosclerosis is the leading cause of death in Western society, and the incidence of cardiovascular disease in Korea is increasing, and attention is being paid to the development of substances that prevent atherosclerosis. Such substances include simvastatin, which lowers plasma cholesterol levels [Aki Nakai et al. : Biol. Pharm. Bull. 19 (9). 1231-1233 (1996)], lovastatin and the like have been used as a therapeutic agent for hyperlipidemia, and probucol for inhibiting LDL oxidation has also been developed and used as a therapeutic agent. In addition, there are antioxidant nutrients such as vitamin C, vitamin E and selenium that can inhibit LDL oxidation in the body.

The present inventors have developed an antioxidant substance capable of effectively regulating noxious oxygen active species, and found that the substance can suppress the induction of melanin production and atherosclerosis caused by the noxious oxygen active species in part. Completed.

An object of the present invention is to provide 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivatives having excellent antioxidant effects, pharmaceutically acceptable salts thereof, and methods for preparing the same.

It is also an object of the present invention to provide a composition for antioxidant, whitening agent and cholesterol control agent containing the compound as an active ingredient.

1 is a graph showing the cytotoxicity test results of the compounds of the present invention.

Figure 2 is a graph showing the results of the melanin secretion test of the compounds of the present invention.

In order to achieve the above object, the present invention provides 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative represented by the following Chemical Formula 1, and a pharmaceutically acceptable salt thereof.

[Formula 1]

Wherein R is a substituted or unsubstituted C ₁ -C ₁₀ straight or branched alkyl group.

The compound of Chemical Formula 1 may be used in the form of a pharmaceutically acceptable salt, and as the salt, an acid addition salt formed by a pharmaceutically acceptable free acid is useful. That is, the compound of formula 1 may form pharmaceutically acceptable acid addition salts according to methods conventional in the art. Organic acids and inorganic acids may be used as the free acid, and hydrochloric acid, bromic acid, sulfuric acid and phosphoric acid may be used as the inorganic acid, and citric acid, acetic acid, lactic acid, tartaric acid, maleic acid, fumaric acid, formic acid, propionic acid, oxalic acid, Trifluoroacetic acid, benzoic acid, gluconic acid, methanesulfonic acid, glycolic acid, succinic acid, 4-toluenesulfonic acid, caluturonic acid, embonic acid, glutamic acid or aspartic acid, and the like can be used.

Preferred examples of the compound of Formula 1 include the following compounds:

4-[(2-propylsulfanyl) methyl] -1,2-benzenediol of the formula (Ia);

[Formula 1a]

4-[(1-butylsulfanyl) methyl] -1,2-benzenediol of the formula (Ib);

[Formula 1b]

4-[(2-butylsulfanyl) methyl] -1,2-benzenediol of the formula (1c);

[Formula 1c]

4-[(1-pentylsulfanyl) methyl] -1,2-benzenediol of the formula (Id);

[Formula 1d]

4-[(isopentylsulfanyl) methyl] -1,2-benzenediol of the formula (1e);

[Formula 1e]

4-[(1-propylsulfanyl) methyl] -1,2-benzenediol of Formula 1f

[Formula 1f]

In the present invention,

To react 1,3-benzodioxol-5-yl-methanol of formula (2) and alkylthiol of formula (3) in the presence of an acid to 4-[(alkylsulfanyl) methyl] -1,2-benzenediol of formula (1) Provided are methods for preparing the derivatives.

[Formula 2]

[Formula 3]

R-SH

In the above formula, R is a C ₁ to C ₁₀ alkyl group.

In this case, the acid may be AlCl ₃ , FeBr ₃ , FeCl ₃ or BBr ₃ and the like .

In addition, the reaction is carried out in an organic solvent such as dichloromethane, chloroform, acetonitrile, tetrahydrofuran or dimethylformamide or a mixed solvent thereof.

In addition, 1,3-benzodioxol-5-yl-methanol of Chemical Formula 2, which is a starting material of the reaction, is oxidized by piperonal of Chemical Formula 4 in the presence of diisobutylaluminum hydride (DIBAL). It is desirable to obtain.

[Formula 4]

In addition, the present invention provides a composition for antioxidant containing 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative of the formula (1) and a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, the present invention provides a composition for a whitening agent containing 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative of Formula 1 and a pharmaceutically acceptable salt thereof as an active ingredient.

The present invention also provides a composition for cholesterol modulator containing 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative of Formula 1 and a pharmaceutically acceptable salt thereof as an active ingredient.

4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivatives of the present invention have excellent antioxidant activity and thus can be used as a whitening agent by inhibiting melanin production, and by inhibiting LDL oxidation to control blood cholesterol concentration. It can also be usefully used as a treatment for atherosclerosis.

The composition of the present invention may be in the form of a solution, suspension or emulsion in oil or an aqueous medium, or in the form of a dry powder which is dissolved in sterile, pyrogen-free water before use, and formulated into an oral dosage form. It may also be formulated into parenteral formulations such as subcutaneous injection, intravenous injection or intramuscular injection.

In the case of oral formulations, for example, tablets, troches, sugar-containing tablets, aqueous or oily suspensions, dispersible powders, etc., by known methods using pharmaceutically acceptable carriers or forming agents. Or in the form of oral dosage forms in the form of particles, emulsions, soft or hard capsules, syrups, elixirs, which may be suitably prepared according to unit dosages, forms and the like.

Parenteral formulations may be injected as a sterile injectable solution or as a suspension in which the active ingredient is suspended in a solvent such as a non-toxic usable diluent or 1,3-butanediol. Excipients or solvents that may be used include water, Ringer's solution, and isotonic saline solution. Cosolvents such as ethanol, polyethylene glycol, and polypropylene glycol may be used. In addition, sterile, nonvolatile oils can be customarily used as solvents or suspending solvents. The suppository form is formulated by mixing the drug with a suitable non-irritating excipient, such as cocoa butter or polyethylene glycol, which is solid at room temperature but liquid at the rectal temperature and will melt in the rectum to release the drug and then enter the rectum. Administration.

When treating a disease using the composition of the present invention, the dose of the compound of formula 1 as an active ingredient is determined by the age, weight, general state of health, sex, meal, time of administration, rate of excretion, combination of drugs, during treatment. Depending on the severity of the disease, depending on the disease can be used daily up to 0.01 ~ 140mg / kg body weight, 0.5mg ~ 7g can be used per day per patient.

On the other hand, the amount of the compound of the present invention mixed with the carrier material to determine one formulation depends on the mode of administration and the patient being treated. For example, formulations intended for oral administration to humans will contain from 5 to 95% by weight of carrier materials and from 0.5 mg to 5 g of active ingredient, and formulations for parenteral administration to humans. Silver contains carrier materials which comprise 5 to 99% of the total composition and 0.1 mg to 2.5 g of active ingredient.

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

Example 1. Synthesis of 4-[(2-propylsulfanyl) methyl] -1,2-benzenediol

(Step 1) Preparation of 1,3-benzodioxol-5-yl-methanol

After dissolving 5 g of piperonal in 340 mL of dichloromethane, 200 mL of a 1M DIBAL solution was added at -78 ° C, stirred for 90 minutes at -78 ° C, 10 mL of methyl alcohol was added, and the temperature of the reaction mixture was raised to room temperature. The reaction mixture was diluted with 250 mL of diethyl ether and washed with 50 mL of brine. The organic layer was dried over anhydrous magnesium sulfate, the solvent was removed, and then the column chromatography was purified using ethyl acetate: n-hexane (1: 3) as eluent. To give 10.92 g of the title compound of the formula (2) (R _f : 0.25, yield: 89%).

Dissolved in CDCl ₃ and TMS and measured by 1 H NMR were as follows:

¹ H NMR δ: 4.578 (s, 2H), 5.95 (s, 2H), 6.79 (m, 3H)

(Step 2) Preparation of 4-[(2-propylsulfanyl) methyl] -1,2-benzenediol

After dissolving 21.9 g of aluminum chloride in 70 mL of anhydrous dichloromethane and adding 150 mL of 2-propanethiol at 0 ° C. and stirring for 20 minutes, 5 g of the compound obtained in (Step 1) was slowly added and stirred at 0 ° C. for 30 minutes, followed by room temperature. Reaction was carried out for 5 hours. After completion of the reaction, dilute with 150 mL of dichloromethane, add ammonium chloride solution to pH 2, extract with dichloromethane and water, dry with anhydrous magnesium sulfate, remove the solvent, and then remove methyl alcohol: dichloromethane (2:98) with eluent. Column chromatography was carried out to obtain 4.78 g of the title compound of Formula 1a (R _f : 0.38, yield: 73%).

Dissolved in CDCl ₃ and TMS and measured by 1H NMR were as follows.

¹ H NMR δ: 1.30 (d, 6H, J = 6.6 Hz), 2.77-2.83 (m, 1H), 3.64 (s, 2H), 6.7-6.79 (m, 2H), 6.85 (s, 1H)

Example 2. Preparation of 4-[(1-butylsulfanyl) methyl] -1,2-benzenediol

To the mixture obtained by dissolving 21.9 g of aluminum chloride in 70 mL of anhydrous dichloromethane, adding 176 mL of 1-butanethiol at 0 ° C. and stirring for 20 minutes, slowly adding 5 g of the compound obtained in (Step 1) of Example 1 for 30 minutes at 0 ° C. After stirring, the mixture was reacted at room temperature for 5 hours. After completion of the reaction, dilute with 150 mL of dichloromethane, add ammonium chloride solution to pH 2, extract with dichloromethane and water, dry with anhydrous magnesium sulfate, remove the solvent, and then remove methyl alcohol: dichloromethane (2:98) with eluent. Column chromatography was carried out to obtain 4.85 g of the title compound of Chemical Formula 1b (R _f : 0.32, yield: 70%).

Dissolved in CDCl ₃ and TMS and measured by 1H NMR were as follows.

¹ H NMR δ: 0.96 (t, 3H, J = 5.8 Hz), 1.29-1.60 (m, 4H), 2.47 (t, 2H, J = 5.8 Hz), 3.63 (s, 2H), 6.7-6.79 (m , 2H), 6.85 (s, 1H)

Example 3. Preparation of 4-[(2-butylsulfanyl) methyl] -1,2-benzenediol

To the mixture obtained by dissolving 21.9 g of aluminum chloride in 70 mL of anhydrous dichloromethane, adding 170 mL of 2-butanethiol at 0 ° C. and stirring for 20 minutes, slowly adding 5 g of the compound obtained in (Step 1) of Example 1 for 30 minutes at 0 ° C. After stirring, the mixture was reacted at room temperature for 5 hours. After completion of the reaction, dilute with 150 mL of dichloromethane, add ammonium chloride solution to pH 2, extract with dichloromethane and water, dry with anhydrous magnesium sulfate, remove the solvent, and then remove methyl alcohol: dichloromethane (2:98) with eluent. Column chromatography was carried out to obtain 5.6 g of the title compound of Formula 1c (R _f : 0.34, yield: 80%).

Dissolved in CDCl ₃ and TMS and measured by 1H NMR were as follows.

¹ H NMR δ: 0.96 (t, 3H, J = 5.8 Hz), 1.22 (d, 3H, J = 6.6 Hz), 1.47-1.61 (m, 2H), 2.63 (q, 1H, J = 6.6 Hz), 3.62 (s, 2H), 6.7-6.79 (m, 2H), 6.85 (s, 1H)

Example 4. Preparation of 4-[(1-pentylsulfanyl) methyl] -1,2-benzenediol

To the mixture obtained by dissolving 5.5 g of aluminum chloride in 30 mL of anhydrous dichloromethane, adding 50 mL of 1-pentanethiol at 0 ° C. and stirring for 20 minutes, 1.28 g of the compound obtained in (Step 1) of Example 1 was slowly added, followed by 30 minutes at 0 ° C. After stirring for 5 hours at room temperature. After completion of the reaction, dilute with 50 mL of dichloromethane, add ammonium chloride solution to pH 2, extract with dichloromethane and water, dry with anhydrous magnesium sulfate, remove the solvent, and then remove methyl alcohol: dichloromethane (2:98) with eluent. Column chromatography was carried out to obtain 1.58 g of the title compound of Chemical Formula 1d (R _f : 0.36, yield: 85%).

Dissolved in CDCl ₃ and TMS and measured by 1H NMR were as follows.

¹ H NMR δ: 0.96 (t, 3H, J = 5.8 Hz), 1.23-1.35 (m, 4H), 1.58 (m, 2H), 2.33 (t, 2H, J = 5.8 Hz), 3.60 (s, 2H ), 6.7-6.79 (m, 2H), 6.85 (s, 1H)

Example 5. Preparation of 4-[(isopentylsulfanyl) methyl] -1,2-benzenediol

To the mixture obtained by dissolving 21.9 g of aluminum chloride in 70 mL of anhydrous dichloromethane, adding 50 mL of isopentane thiol at 0 ° C. and stirring for 20 minutes, 1.123 g of the compound obtained in (Step 1) of Example 1 was slowly added thereto for 30 minutes at 0 ° C. After stirring, the mixture was reacted at room temperature for 5 hours. After completion of the reaction, dilute with 150 mL of dichloromethane, add ammonium chloride solution to pH 2, extract with dichloromethane and water, dry with anhydrous magnesium sulfate, remove the solvent, and then remove methyl alcohol: dichloromethane (2:98) with eluent. Column chromatography was carried out to obtain 2.19 g of the title compound of Formula 1e (R _f : 0.35, yield: 90%).

Dissolved in CDCl ₃ and TMS and measured by 1H NMR were as follows.

¹ H NMR δ: 0.8 (d, 6H, J = 6.6 Hz), 1.45 (t, 2H, J = 5.8 Hz), 1.46-1.62 (m, 1H), 2.45 (t, 2H, J = 5.8 Hz), 3.60 (s, 2H), 6.7-6.79 (m, 2H), 6.85 (s, 1H)

Example 6. Preparation of 4-[(1-propylsulfanyl) methyl] -1,2-benzenediol

To the mixture obtained by dissolving 21.9 g of aluminum chloride in 70 mL of anhydrous dichloromethane and adding 148 mL of 1-propanethiol at 0 ° C. and stirring for 20 minutes, 5 g of the compound obtained in (Step 1) of Example 1 was slowly added thereto for 30 minutes at 0 ° C. After stirring, the mixture was reacted at room temperature for 5 hours. After completion of the reaction, dilute with 150 mL of dichloromethane, add ammonium chloride solution to pH 2, extract with dichloromethane and water, dry with anhydrous magnesium sulfate, remove the solvent, and then remove methyl alcohol: dichloromethane (2:98) with eluent. Column chromatography was carried out to obtain 5.34 g of the title compound of Formula 1f (R _f : 0.37, yield: 82%).

Dissolved in CDCl ₃ and TMS and measured by 1H NMR were as follows.

¹ H NMR δ: 0.98 (t, 3H, J = 5.8 Hz), 1.48-1.66 (m, 2H), 2.43 (t, 2H, J = 5.8 Hz), 3.60 (s, 2H), 6.7-6.79 (m , 2H), 6.85 (s, 1H)

Experimental Example 1 DPPH Scavenging Activity

1,1-Diphenyl-2-picrylhydrazyl (abbreviated as "DPPH") is a purple dye that can measure the antioxidant activity of phenols and aromatic amines. It is a widely used substance [Blois, MS Antioxidant determinations by the use of a stable free radical. Nature , 181, 1990-1200 (1958). DPPH exhibits a strong absorption band due to its odd number of electrons at 520 nm, but when reacted with hydrogen or an electron donor like phenol, it receives electrons or hydrogen radicals from a donor to generate phenoxy radicals. At this time, the absorption disappears and is stabilized to change from the original progressive color to yellow to reduce the absorbance. That is, the radical scavenging activity can be known by measuring the decrease in absorbance of the reaction solution [Yokozawa, T. et al ., Biochemical Pharmacology , 56, 213-222 (1998); Hatano, T. et al ., Chem. Pharm. Bull. , 37 (8), 2016-2021 (1989)].

Determination of the scavenging effect on DPPH radicals of each sample is as follows [Blois, MS Nature , 181, 1990-1200 (1958)].

Each concentration of sample (1.25 ~ 120 ㎍ / mL) was dissolved in methanol and 4 mL each was taken and mixed well with 1 mL of DPPH solution dissolved in methanol at 1.5 × 10 ^-4 M concentration. After leaving the reaction mixture at room temperature for 30 minutes, the absorbance was measured at 520 nm. The free radical scavenging activity was expressed as a percentage and the 50% scavenging concentration (IC ₅₀ ) was calculated as compared to the control without the sample. The measured values are expressed as averages of the results obtained from three replicates [Yoshida et al ., Chem. Pharm. Bull ., 37 (7), 1919-1921 (1989).

As a result of measuring the DPPH radical scavenging effect of the synthetic compounds (see Table 1), the activity of Example 6 was the highest, and the DPPH radical scavenging effect of the remaining compounds was the IC ₅₀ (50% scavenging concentration) value of vitamin C, known as an antioxidant. Lower than In particular, Example 4 was about 55 times stronger than vitamin C.

TABLE 1

Test subject IC ₅₀ (μg / mL) Example 1 0.23 Example 2 0.24 Example 3 0.31 Example 4 0.034 Example 5 0.29 Example 6 0.25 Vitamin C (Control) 1.74

Experimental Example 2 Cytotoxicity Test

3,4,5-dimethyl thiazole-3,5-diphenyl tetrazolium bromine (3,4,5-dimethyl thiazole-3,5- to examine the cytotoxicity of the compounds prepared in Examples 1-6 diphenyl tetrazolium bromide, hereafter abbreviated as "MTT") test was conducted by Mosmann [Mosmann, T .: J. Immunol. Methods 65, 55-63, (1983)] was tested for the toxicity to melanocytes. The cytotoxic effects of drugs are examined by MTT assay, which is based on the ability of live cells to break down tetrazolium salts, preventing the accumulation of MTT in the cells. 5 mg / ml of MTT stock solution is dissolved in phosphate buffer saline (abbreviated as "PBS") and then filtered through a 0.22 μm filter to remove formazan crystals and stored in a dark room at room temperature. Stored. For the experiment, B16 melanoma cells were transferred to 96-well plates at a concentration of 3,000 cells / well, and the compounds prepared in Examples 1-6 were treated. After a certain incubation period, 10-fold diluted MTT was added to each well, and after 4 hours, the plate was centrifuged at 275 × g for 5 minutes and 100 μl of the supernatant was collected. To remove formazan crystals, DMSO was added and shaken until the DMSO was completely dissolved in a microplate shaker, followed by a microplate ELISA reader (El 312e, Bio-Tek). Absorbance was measured at a wavelength of 560 nm. Cell viability in each drug treatment group was expressed as% control compared to the control (see FIG. 1).

According to FIG. 1, arbutin and vitamin C, which are known to have a whitening effect, showed almost 100% cell viability up to a concentration of 10 µg / ml, but hydroquinone, as known, was highly cytotoxic. On the contrary, the compounds of Examples 1-6 were generally cytotoxic, but at a concentration of 10 µg / ml, they showed some cytotoxicity. However, at the concentration of 1 μg / ml, the compounds of Examples 1-6 all showed cell survival rates close to 100%.

<Experiment 3> Melanin secretion inhibitory effect test using B16 veloma cell line

The whitening effect of the drugs was investigated using whitened B16 mouse melanoma cells. B16 cells were incubated with Dulbecco's modified Eagle's medium containing 10% heat inactivated fetal calf serum, 100 mg / ml streptomycin and 100 U / ml penicillin. B16 cells cultured for the experiment were treated with trypsin, detached and dispensed into a 24-well tissue culture flask at a concentration of 1 × 10 ⁵ cells / ml. After one day, the cell cultures were replaced with modified dulcocco eagle medium without phenol red containing 2 μM α-melanocyte stimulating hormone and 2 mM theophylline. The drug was added at various concentrations and incubated for 7 days. The whitening effect of the added drug was determined by the amount of melanin pigment secreted from B16 cells. After 7 days of drug treatment, the cell culture solution of each well was collected, the cells were removed by centrifugation, and the absorbance was measured by ELISA reader at 490 nm. The amount of melanin taken was measured. The inhibitory effect of each substance was calculated using the following formula:

As a result, arbutin, vitamin C, and hydroquinone showed a tendency to inhibit melanin secretion from B16 melanoma cell line in a whitening effect. Compounds of Examples 1-6 showed a stronger secretion inhibitory effect than arbutin, in particular the compounds of Examples 1 and 6 showed 100% secretion inhibitory effect at a concentration of 0.1 ㎍ / ml or more. Taken together with the cytotoxicity results by the previous MTT assay, it was found that these compounds 100% inhibition of melanin secretion without cytotoxicity in the concentration range of 0.1 ~ 1.0 ㎍ / ml (see Figure 2).

Experimental Example 4 Measurement of LDL Oxidation Inhibition Rate

LDL (Sigma No. L-5402) was purchased and dialyzed with PBS for 24 hours, and then sterilized by passing through a 0.45 μm sterile filter-mounted syringe. Protein concentration of LDL was quantified by Lowry method using bovine serum albumin (hereinafter abbreviated as "BSA") as a standard.

To the reaction solution containing 0.1 mg protein / ml LDL and the compound prepared in Examples, the final concentration of Cu ²⁺ was added to 5M and oxidized at 40 ° C. for 4 hours. 1% oxidized LDL reaction solution and malondialdehyde (hereinafter abbreviated as "MDA") prepared by concentration in 0.4% thiobarbituric acid (hereinafter abbreviated as "TBA") standard reaction solution, 15 1 ml of a solution of thiobarbituric acid reactive substances (TBARS) containing 2.5% trichloroacetic acid (hereinafter abbreviated as "TCA") and 2.5% HCl was mixed, and the reaction mixture was reacted for 20 minutes in a 95-100 ° C water bath. The hot sample was vortexed to remove bubbles, cooled in cold water, centrifuged at 2000 rpm for 10 minutes, and the supernatant was measured for absorbance at 532 nm. The concentration of TBARS in the sample was obtained from the standard curve of MDA made of MDA and expressed as nmole of MDA.

In order to examine the inhibitory effect on the LDL oxidation of the compounds prepared in Examples, 1, 5, 25, and 50 µg / mL of the compounds were added to the LDL reaction solution. Compared to the effect. The TBARS concentration of oxidized LDL (40 ° C., 4hrs) was 30.35 ± 5.60 nmole MDA / mg protein LDL. The results were repeated three times with the mean ± standard deviation. Samples were repeated three times for each experiment.

As shown in Table 2, the compounds prepared in Examples 1-6 show potent antioxidant capacity against LDL oxidation. Antioxidant effect of the compounds of the present invention was lowered by about 72% to 80% inhibitory effect of Preparation Example 1-6 compared to about 21% of the vitamin C showed LDL oxidation inhibitory effect when the compound 1㎍ / mL added The antioxidant effect was also significantly higher than that of vitamin C.

TABLE 2

Test subject LDL oxidation inhibition rate (%) 1 μg / mL 5 μg / mL 25 μg / mL 50 μg / mL Vitamin c -20.56 ± 17.13 -38.99 ± 9.07 -49.12 ± 3.34 -50.51 ± 1.10 Example 3 -76.85 ± 9.01 -88.74 ± 7.43 -88.07 ± 2.43 -90.86 ± 2.46 Example 4 -79.39 ± 3.55 -87.09 ± 6.48 -88.98 ± 0.84 -92.83 ± 0.74 Example 5 -79.65 ± 1.47 -76.15 ± 13.67 -73.93 ± 19.35 -79.04 ± 13.23 Example 6 -77.54 ± 1.46 -82.72 ± 3.83 -83.12 ± 6.55 -83.05 ± 6.34 Example 1 -73.27 ± 3.28 -81.96 ± 6.04 -81.96 ± 2.86 -83.95 ± 0.19 Example 2 -72.41 ± 8.34 -80.51 ± 9.75 -87.61 ± 1.34 -82.60 ± 4.60

Although the present invention has been described through the preferred embodiments so far, those skilled in the art can make various modifications without departing from the spirit and scope disclosed in the present invention. Such modifications are included in the scope of the present invention.

As described above, the present invention exhibits excellent antioxidant activity, effectively inhibits melanin production, and therefore can be used as a whitening agent, and can also be useful as a cholesterol modulator because it also inhibits oxidation of LDL.

Claims (9)

4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative represented by the following formula (1) and a pharmaceutically acceptable salt thereof. [Formula 1] In the above formula, R is a C ₁ to C ₁₀ alkyl group. According to claim 1, wherein the compound of Formula 1 4-[(2-propylsulfanyl) methyl] -1,2-benzenediol; 4-[(1-butylsulfanyl) methyl] -1,2-benzenediol; 4-[(2-butylsulfanyl) methyl] -1,2-benzenediol; 4-[(1-pentylsulfanyl) methyl] -1,2-benzenediol; 4-[(isopentylsulfanyl) methyl] -1,2-benzenediol; And 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivatives and pharmaceuticals, characterized in that selected from the group consisting of 4-[(1-propylsulfanyl) methyl] -1,2-benzenediol Acceptable salts thereof. 4-[(alkylsulfanyl) methyl] -1,2-benzenediol, characterized by reacting 1,3-benzodioxol-5-yl-methanol of formula 2 with alkylthiol of formula 3 in the presence of an acid Process for the preparation of derivatives. [Formula 2] [Formula 3] R-SH In the above formula, R is a C ₁ to C ₁₀ alkyl group. The method of claim 3, wherein The acid is selected from the group consisting of AlCl ₃ , FeBr ₃ , FeCl ₃ and BBr ₃ . The method of claim 3, wherein The reaction is carried out in a single solvent or a mixed solvent selected from the group consisting of dichloromethane, chloroform, acetonitrile, tetrahydrofuran and dimethylformamide. The method of claim 3, wherein 1,3-benzodioxol-5-yl-methanol of Chemical Formula 2 is obtained by oxidizing piperonal of the following Chemical Formula 4 in the presence of diisobutylaluminum hydride (DIBAL) . [Formula 4] An antioxidant comprising the 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative of claim 1 and a pharmaceutically acceptable salt thereof as an active ingredient. The composition for whitening agent containing 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative of Claim 1, and its pharmaceutically acceptable salt as an active ingredient. A composition for cholesterol control comprising the 4-[(alkylsulfanyl) methyl] -1,2-benzenediol derivative of claim 1 and a pharmaceutically acceptable salt thereof as an active ingredient.