KR101057427B1 - Novel L-derived Isigoside Compounds - Google Patents

Abstract

The present invention relates to novel compounds derived from brown algae shells with antioxidant activity, and to pharmaceutical compositions containing them, wherein the novel compounds of formula (1) according to the present invention effectively eliminate DPPH, hydroxyl, superoxide and alkyl radicals. The antioxidant activity is very excellent.

L, antioxidant activity, isigoside

Description

A new ishigoside compound from ishige okamurae

The present invention relates to novel compounds derived from brown algae shells with antioxidant activity, and pharmaceutical compositions containing them.

Free radicals having one or more hole electrons in the outer electron orbit mainly include superoxide anions (O ₂ ^·- ), hydroxyl (HO ·), peroxyl (ROO ·), alkoxyl (RO ·), and nitrogen oxides. And they are oxygen-centered free radicals. More and more studies have shown that free radicals produced by cellular metabolism and induced by exposure to environmental oxidants may lead to chronic and degenerative diseases such as mutations, carcinogenesis, coronary arteries through free radical damage of lipids, nucleic acids and proteins. It has been shown to be involved in heart disease, diabetes, Parkinson's disease and Alzheimer's disease. The fact that such risks of chronic diseases are inversely associated with the intake of dietary antioxidants is facilitating the search for potential antioxidants from vegetables, fruits, herbs, seaweeds and the like.

Shell (Ishige okamurae) is a kind of brown seaweed with leaves, thick skin layer and pointed vertices, belonging to Ishigeaceae and inhabiting rocks in the upper and middle low tide areas of rough open coast. Conventional phytochemical studies on the title algae have revealed the presence of phthalates, sterols, glycerol derivatives, pheophytins, acids and phlorotannins.

In addition, the antibacterial effect of the plaque extract on the dental caries cause has been reported, has been isolated and purified of phospholipase A2 activity inhibitor from the shell extract.

The present inventors earnestly studied to search for bioactive components having antioxidant activity from natural substances, and thus, separated the new glycerol glucolipid compound from the brown algae and led to the present invention.

It is therefore an object of the present invention to provide novel compounds derived from brown algae that have antioxidant activity.

In another aspect, the present invention provides an antioxidant active pharmaceutical composition comprising the compound as an active ingredient.

An object of the present invention as described above is to prepare an extract from brown algae shell, to isolate the novel compound therefrom, and to reveal the structure of the compound through a physicochemical method, their DPPH, hydroxyl, alkyl and superoxide Achieved by identifying radical scavenging activity.

The present invention provides an isoside compound of Formula 1 below.

In the present invention, the compound is isolated from the brown algae, and the compound is based on a comprehensive spectroscopic method including 1D and 2D NMR, MS techniques, and chemical methods. The compound is 1,2-di-O-palmitoyl-3-O- It was found to be a novel glyceroglycolipide, (6-deoxy-6-amino) -aD-glucopyranosyl-glycerol, which was named isigoside.

If the compounds of the present invention have one or more chiral centers, it will be apparent to those skilled in the art that the compounds of the present invention may exist as enantiomers or geometric isomers, or racemic mixtures. Enantiomers, geometric isomers, racemates or mixtures thereof of any possible compounds of Formula 1 of the present invention. For example, optically active forms of the compounds of the present invention can be prepared by chiral chromatographic separation of racemates, synthesis from optically active starting materials, or by asymmetric synthesis based on the processes described below.

It will be apparent to those skilled in the art that certain compounds of the present invention may exist as geometric isomers, such as the E and Z isomers of alkenes. The present invention includes any geometric isomer of the compound of formula (I). It will be apparent or apparent that the present invention includes tautomers of the compound of formula (I).

It will also be apparent to those skilled in the art that certain compounds of the present invention may exist in solvated forms as well as in unsolvated forms. It will also be apparent that the present invention includes such solvated forms of the compound of formula (I).

Salts of compounds of formula 1 are also within the scope of the present invention. In general, pharmaceutically acceptable salts of the compounds of the present invention are standard processes well known in the art, for example, reacting a sufficient amount of a basic compound (eg alkyl) with an appropriate acid (eg HCl or acetic acid). By producing a physiologically acceptable anion. In addition, compounds of the present invention with suitably acidic protons, such as carboxylic acids or phenols, in one aqueous alkali or alkaline earth metal hydroxide or alkoxide (e.g., ethoxide or methoxide) in an aqueous medium, or suitably After reacting with a basic organic amine (e.g., chlorine or meglumine), the corresponding alkali metal (e.g. sodium, potassium or lithium) or alkaline earth metal (e.g. It is also possible to prepare calcium) salts.

In one embodiment of the invention, the compound of formula 1 is a pharmaceutically acceptable salt or solvate thereof, in particular acid addition salts such as hydrochloride, hydrobromide, phosphate, acetate, fumarate, maleate, tart It may also be converted to laterate, citrate, methanesulfonate or p-toluenesulfonate. Specific examples of the present invention include the following compounds, pharmaceutically acceptable salts, hydrates, solvates, optical isomers, and combinations thereof.

In the present invention, the scavenging activity of DPPH, hydroxyl, superoxide, and alkyl radicals of the compound of Formula 1 was investigated by ESR, and the antioxidant activity of the compound was found.

Therefore, the present invention provides a pharmaceutical composition having an antioxidant activity containing the compound of Formula 1 as an active ingredient.

In the present invention, the term "active ingredient" refers to a substance or a group of substances (a pharmacologically active ingredient or the like, which is expected to express the efficacy or effect of the drug directly or indirectly by intrinsic pharmacological action). It means containing a main component).

The novel compound of formula (I) isolated from the brown algae shell according to the present invention can effectively eliminate DPPH, hydroxyl, superoxide and alkyl radicals, which has very good antioxidant activity.

Hereinafter, preferred embodiments of the present invention will be described in more detail with reference to Examples. However, the scope of the present invention is not limited to these examples.

Example 1 Preparation of L Extract and Isolation of Bioactive Components

¹ H-NMR, ¹³ C-NMR, and 2D-NMR were recorded on a JEOL JNM-ECP 400 spectrometer, and chemical shifts (δ) were expressed in ppm with respect to TMS. IR spectra were measured with a Bucker FT-IR model IFS-88 spectrometer. MS spectra were measured with a JEOL JMX-700 mass spectrometer. Optical rotation was measured with a Perkin-Elmer-341 polarimeter. GC-MS experiments were performed on a Shimadzu GC-MS-QP5050A equipped with a flame ionization detector.

Free radical scavenging activity was measured using a JESFA ESR spectrometer (JEOL, Tokyo, Japan). 2,2-diphenyl-1-picrylhydrazyl (DPPH), 5,5-dimethyl-1-pyrroline-N-oxide (DMPO), FeSO ₄ , H ₂ O ₂ , 2,2-azobis- (2-Amidinopropane) -hydrochloride hydrochloride (AAPH), a- (4-pyridyl-1-oxide) -Nt-butylnitron (4-POBN) are available from Sigma Chemical Co .; St. Louis, MO, USA). Silica gel 60 (0.036-0.2 mm, Merck), Sephadex LH-20 (Sephadex LH-20; Pharmacia Biotech AB, Uppsala, Sweden), and ODS-A gel (DAISO CO., LTD., Japan) were column chromatographed. Used for photography. Thin layer chromatography was performed on plates precoated with silica gel 60 F ₂₅₄ (Kieselgel 60 F ₂₅₄ 5715, Merck). Fractions were monitored by TLC and the points visualized by spraying 5% vanillin as well as 10% alcohol, H ₂ SO ₄ , and then heated at 105 ° C. for 4 minutes.

The plaques were collected in October 2007 on the coast of Busan, South Korea. A voucher specimen (No. 20071003) was deposited in the laboratory. The sample was washed three times with tap water to remove salts, plant parasites, and sand from the surface of the sample. Finally, the samples were air dried and ground in a coffee grinder and the algae powder obtained was stored at -20 ° C in a freezer until use.

Statistical analysis of all experimental results was expressed as mean ± standard deviation, and statistical comparisons were made by Student's t test. p <0.05 was considered significant.

Plaque powder (500 g) was extracted three times at room temperature using 2 L methanol. After evaporating off the solvent, the extract (96 g) was suspended in H ₂ O and then successively partitioned with hexane, dichloromethane, EtOAc and n-BuOH. Hexane solvent (4 L) on EtOAC-soluble fraction (22 g) silica gel, hexane and ethyl acetate mixture (20: 1, 10: 1, 5: 1, 1: 1, v / v, 5L each), dichloromethane (3L ) And column chromatography by eluting with a mixture of dichloromethane and methanol (10: 1, 5: 1, 1: 1, v / v, 5 L each) to give 29 fractions based on TLC analysis (fraction 1). To 29). Fraction 5 was further chromatographed on silica gel and Sephadex LH-20 to give compound 3 (7 mg). Fraction 7 was separated by eluting with chloroform-methanol (10: 1 v / v) on silica gel to give six subfractions (fractions 7A-7F). Fraction 7D was further purified on Sephadex LH-20 to give Compound 2 (16 mg). Fraction 22 was separated by eluting with chloroform-methanol-H ₂ O (5: 1: 0.1 v / v / v) on silica gel to give four subfractions (fractions 22A-22D). After further purification on the fraction 22C Sephadex LH-20 column, the ODS column (eluted with methanol-H ₂ O / 40: 60) was purified to give Compound 1 (22 mg) of Formula 1 It was named (ishigoside).

Ichigoside (Compound 1) obtained as a white amorphous powder had the formula C ₄₁ H ₇₉ NO ₉ as determined by mass spectrum and NMR spectral data and further confirmed by HREIMS at m / z 729.5775. IR bands at 3430 and 1737 cm ⁻¹ showed the presence of hydroxyl and amino groups in the molecule, respectively. Detailed analysis of the ¹ H, ¹³ C NMR, ¹ H- ¹ H COZY, HMQC, and HMBC spectra revealed that the structure of Compound 1 was 1,2-di-O-palmitoyl-3-O- (6-deoxy-6 -Amino) -aD-glucopyranosyl-glycerol.

Three spin systems appeared in the ¹ H NMR spectrum of Compound 1 (see Table 1). The first spin system is due to saturated fatty acids, which are two terminal protons at d _H 0.85 (6H, t, J = 6.6 Hz), broad methylene protons at d _H 1.23 (48H, br s), d _H 1.49 ( 4H, br s) it is shown by a couple of methylene protons associated with carbonyl groups in the two methylene protons and d _H 2.28 (4H, m) in the. These data were consistent with proton mythology of two palmitic acid residues. The second spin system consists of glycerol residues [d _H 3.39 (1H, dd, J = 10.4, 5.8 Hz), 3.88 (1H, dd, J = 10.8, 5.8 Hz), 4.14 (1H, dd, J = 12.0, 7.5 Hz ), 4.33 (1H, doublet of doublets, J = 12.0, 2.9 Hz), 5.13 (1H, m)]. The third spin system indicates the presence of glycosyl residues, where the proton signals of the four oxymethines are seen at d _H 2.94-3.77 (m) and ano at d _H 4.57 (1H, d, J = 3.3 Hz). Unshielded methylene was seen in the mer proton, d _H 2.56 (1H, dd, J = 13.7, 6.7 Hz), 2.88 (1H, dd, J = 13.7, 4.6 Hz). ^{The 13} C NMR spectrum (see Table 1) shows the carbonyl carbon signal of two fatty acid ester groups at d _C 172.5 and 172.4, two methyl carbons at d _C 13.9, 28 methylene carbons at zone d _C 22.1-33.5, d _C Three carbon signal properties of glycerol residues at 62.6, 64.6, and 69.7, and carbon signals of six glycerol residues at d _C 54.7, 68.5, 71.6, 72.9, 74.3, and 98.3.

Abnormal upfield shift H-6 '(d _H 2.56 and 2.88) ¹ H NMR resonance and C-6' (d _C 54.7) ¹³ C-NMR resonance suggest that the glycosyl residue is an amino sugar. The amino group will be attached to C-6 '. The atomic arrangement at the C-1 'of the sugar was determined as α by the relatively small coupling constant (J = 3.3 Hz) of the anomer protons. Compared to the reported values, the glycosyl residues of compound 1 had the same ¹³ C-NMR data of 6-deoxy-6-amino-aD-glucopyranosyl residues. Also, when Compound 1 was hydrolyzed in NaOMe / MeOH, only methyl palmitate was detected by GC-MS from nonpolar organic extract after fractionation, confirming that both acylated fatty acids are palmitic acid. gave. HMBC cross-peaks of H-1 'and C-3, H-2 and C-1'', and H-1 and C-1''' include sugars in C-2 of glycerol and palmitic acid in C- The conclusion is that they are located at 1 and C-2.

Ishigoside (1) : white amorphous powder. [a] 22 D = + 34.2 (c 0.305, MeOH). IR (neat) g _max : 3430, 2914, 1737 cm ^-1 . HREIMS m / z 729.5775 [M] ⁺ , Calc. for C ₄₁ H ₇₉ NO ₉ , 729.5755. ¹ H NMR (DMSO-d ₆ , 400 MHz), ¹³ C NMR (DMSO-d ₆ , 100 MHz), COZY and HMBC (see Table 1).

TABLE 1

NMR spectral data of isigosides in DMSO- d ₆ ^*

*: ¹ H and ¹³ C NMR were recorded at 400 and 100 MHz, respectively.

On the other hand, two known compounds were identified by comparing their spectral data and literature data for palmitic acid and methyl palmitate.

Ichigoside (Compound 1 , 6.0 mg) was dissolved in 2 mL of methanol and treated with 2% NaOMe-MeOH solution (1.5 mL) with stirring at room temperature for 3 hours. The reaction mixture was neutralized with ion exchange resin (Dowex 50 W × 8) and the resin was filtered off. The filtrate was extracted with hexane and the hexane layer was concentrated under reduced pressure to give methyl palmitate (4.0 mg).

The methyl palmitate was identified by GC-MS based on a comparison with a certified sample. The MeOH layer was dried under reduced pressure to give a residue, which was separated by (SiO ₂ , CHCl ₃ / MeOH / H ₂ O, 4: 1: 0.1) to give monoglucosyl glycerol (1.5 mg).

Microalgae can be used to produce a wide range of first and second metabolites, including proteins, lipids, carbohydrates, carotenoids, vitamins and other biologically active compounds. Glyceroglycolipids and their homologues containing 2-deoxy-2-amino are widely distributed in red, brown and green algae as well as other marine organisms. However, glyceroglycolipids with 6-deoxy-6-amino glycerides are rare in nature. To the best of our knowledge, the novel glyceroglycolipids mentioned above have been separated from algae three times, with the exception of two homologues isolated from green algae Avrainvillea nigricans and unidentified marine algae.

Example 2 Measurement of Free Radical Scavenging Activity

The activity of DPPH radicals was measured using the method described by Nanjo et al. Briefly, 30 mL sample solution (or ethanol itself as a control) was added to 30 mL of DPPH (60 mM) in ethanol solution. After vigorously mixing for 10 seconds, the solution was transferred to a 100 mL quartz capillary tube and the scavenging activity for DPPH radicals was measured with a JESFA ESR spectrometer (JEOL, Tokyo, Japan). After exactly 2 minutes the spin adduct was measured on an ESR spectrometer. The experimental conditions were as follows: magnetic field, 336.5 ± 5 mT; Power, 5 mW; Modulation frequency, 9.41 GHz; Amplitude, 1 × 1000; Cleaning time, 30 s. DPPH radical scavenging activity was calculated according to the following formula.

DPPH clearing activity (%) = (1-A / A ₀ ) × 100,

Wherein A is the relative peak height obtained using the sample,

A ₀ is the relative peak height obtained without using a sample.

% Scavenging activity was plotted against the sample concentration to obtain an EC ₅₀ value.

Hydrogen radicals were generated by the Fenton HaberWeiss reaction, and the resulting hydroxyl radicals were reacted rapidly with nitron spin trap DMPO. The resulting DMPO-OH adduct was detected by an ESR spectrometer. The phlorotannin solution (20 mL) was mixed with DMPO (0.3 M, 20 mL), FeSO4 (10 mM, 20 mL) and H ₂ O ₂ (10 mM, 20 mL) in phosphate buffer solution (pH 7.4). Transfer to a 100 L quartz capillary tube. After 2.5 minutes, ESR spectra were recorded using an ESR spectrometer. The experimental conditions were as follows: magnetic field, 336.5 ± 5 mT; Power, 1 mW; Modulation frequency, 9.41 GHz; Amplitude, 1 × 200; Cleaning time, 4 min. Radical scavenging activity was calculated according to the following formula.

% Scavenging activity = (1-A / A ₀ ) × 100,

Wherein A is the relative peak height obtained using the sample,

A ₀ is the relative peak height obtained without using a sample.

Alkyl radicals were generated by 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) according to the method of Hiramoto et al. Briefly, 20 mL of 40 mM AAPH was added to 20 mL of phosphate buffered water (PBS), 20 mL of 40 mM (4-pyridyl-1-oxide) -N-tert-butylnitron (4-POBN) and the test sample at the specified concentration. Mix with 20 mL. The mixture was vortexed and incubated at 37 ° C. for 30 minutes. The reaction mixture was then transferred to a sealed capillary tube and the spin adduct was recorded at controlled spectroscopic conditions: modulation frequency, 100 KHz; Microwave power, 10 mW; Microwave frequency, 9441 MHz; Magnetic field, 336.5 ± 5 mT and cleaning time, 30 s. Alkyl radical scavenging activity was calculated according to the following formula.

% Scavenging activity = (1-A / A ₀ ) × 100,

Wherein A is the relative peak height obtained using the sample,

A ₀ is the relative peak height obtained without using a sample.

Superoxide radicals were generated with a UV irradiated riboflavin / EDTA system. The reaction mixture containing 0.3 mM riboflavin, 1.6 mM EDTA, 800 mM DMPO, and the test sample at the indicated concentration was irradiated with a 365 nm UV lamp for 1 minute. The reaction mixture was transferred to a 100 mL quartz capillary tube on an ESR spectrometer. The experimental conditions were as follows: magnetic field, 336.5 ± 5 mT; Power, 10 mW; Modulation frequency, 9.41 GHz; Amplitude, 1 × 1000; Cleaning time, 1 min. Superoxide radical scavenging activity was calculated according to the following formula.

% Scavenging activity = (1-A / A ₀ ) × 100,

Wherein A is the relative peak height obtained using the sample,

A ₀ is the relative peak height obtained without using a sample.

At present, ESR is known to be the most useful spectrometer that can accurately measure the level of radicals remaining in the reactants. ESR is widely used as the most useful method for measuring radical species due to its convenience, high sensitivity and short term consumption. Therefore, as described above, the free radical scavenging activity of the isigoside against DPPH, superoxide, hydroxyl, and alkyl radicals was measured using an ESR spectrometer, and the radical scavenging activity of the isigoside is shown in FIG. 1.

DPPH radicals are stable organic free radicals, which can be reduced to non-radical form DPPH by reaction when accepting electrons or hydrogen in the presence of a hydrogen-donating antioxidant. It is widely used for screening anti-radical activity because it can prepare a large number of samples in a short time and is sensitive enough to detect the active ingredient even at low concentration. The DPPH scavenging activity of Ishigoside is shown in Figure 1a. Igosides exhibited Igoside DPPH clearance in a dose-dependent manner. When the concentration of isigosides increased from 6.25 mM to 100 mM, DPPH scavenging activity increased from 15.4% to 95.4%; DPPH scavenging activity was statistically significant (p <0.05) compared to the control at a concentration of 12.5 mM or more. However, no DPPH scavenging activity was observed at concentrations above 100 mM. EC ₅₀ , defined as the concentration required for the radicals produced by the reaction system to eliminate Ishigoside 50%, was not significantly different than α-tocopherol (p> 0.05), but DPPH radical valent activity was overwhelming than α-tocopherol It was.

The hydroxyl radical (HO.) Is the most reactive ROS, which can react quickly with most biological molecules and may be involved in the pathology of many human diseases. Hydrogen radicals were generated in the Fenton system and HO was identified because of the ability to form nitroxide byproducts from commonly used DMPO spin traps. The spin trap has greater oxygen-centric radical trapping capability than other nitron spin traps. By-product DMPO-OH radicals exhibit a characteristic ESR reaction, which can be detected by an ESR spectrophotometer. Inhibition of isigosides against hydroxyl radicals produced by the Fenton system was measured and shown in FIG. 1B. Concentration-dependent inhibition of hydroxyl radicals was observed in isigosides and EGCG, respectively. Compared with EGCG, hydroxyl radical scavenging activity of Igoside was greater, and EC50 (16.7 mM) of Igoside and EC50 (41.0 mM) of ECGC were significantly different. Even at lower concentrations of 12.5 mM, Igosides significantly inhibited the generated radicals by 43.2% (p <0.05), while EGCG inhibited only 16.7% of the hydroxyl radicals at these concentrations, and moreover The scavenging effect was not statistically significant compared to the control (p> 0.05).

The alkyl radical scavenging ability of the isigoside was tested using the ESR technique. Alkyl radicals were formed by decomposing AAPH, a water soluble peroxyl radical inhibitor, which was then trapped with 4-POBN and the spin adduct formed with ESR was measured according to the method described above. All test samples were observed for scavenging activity in a dose-dependent manner (FIG. 1C). When the concentration of isigoside was increased to 10 mM, significant scavenging activity of the alkyl radical was shown, and 82.8% of the alkyl radical was scavenged at the 80 mM concentration. The EC ₅₀ of Igoside and EGCG were 22.7 mM and 21.6 mM, respectively, which indicates that the alkyl radical scavenging activity of Igoside is comparable to that of EGCG because there is no significant difference between EC ₅₀ between Igoside and EGCG (p> 0.05). Suggest.

Finally, in the present invention, the scavenging activity of the superoxide radical of isigoside was studied. Superoxide radicals form early in the oxidation reactions of the chain and, despite being weak oxidants, they can be converted into strong and dangerous hydroxyl radicals in the presence of metals such as iron and copper, leading to cell damage and ROS-related diseases. have. UV irradiation of the riboflavin / EDTA system produced superoxide radicals. In the present invention, the superoxide scavenging ability of the l-derived igoshiside was measured by an ESR spectrometer. 1D shows the superoxide radical scavenging activity of various concentrations of isigoside and EGCG. All of these exhibited strong superoxide radical scavenging activity in a dose-dependent manner, but EGCG used as a positive control showed the largest superoxide radical scavenging activity. Weaker superoxide scavenging activity was observed compared to EGCG, and EC ₅₀ (26.8 mM) of Igoside and EC ₅₀ (0.05 mM) of EGCG were significantly different (p <0.05). Igosides significantly inhibited hydroxyl radicals produced at concentrations of only 20 mM and above, while Igosides scavenged 79.4% of hydroxyl radicals at the highest concentration of 80 mM, while EGCG had 89.7% of hydroxyl radicals at these concentrations. Could be eliminated (p <0.01).

Antioxidants can be classified into four categories: prophylactic antioxidants, radical scavenging antioxidants, repair and neonatal antioxidants, and adaptive antioxidants. Of these, radical scavenging antioxidants are known to be the most beneficial. As mentioned above, the inventors have found that Igoside exhibits a broad spectrum of radical scavenging activity against DPPH, hydroxyl, alkyl and superoxide radicals. The EC ₅₀ of isigosides for the DPPH, hydroxyl, alkyl and superoxide radicals was 31.2 mM, 16.7 mM, 22.7 mM, and 26.8 mM, respectively. Comparing the EC ₅₀ of the isigoside, the hydroxyl radical was most effectively scavenged by the isigoside, while the DPPH radical was most ineffectively quenched, which is believed to be due to the poor hydrogen-donating ability of the isigoside. The results of the present invention also showed that Igoside is an excellent antioxidant with free radical scavenging activity.

In conclusion, in the present invention, Igoside, a novel glycerol lipid with amino sugars, was isolated from the plaques by a series of column chromatography, and the structure of Igoside was determined based on 1D and 2d NMR, MS techniques and chemical methods. Came out. ECR spectroscopy showed that Igosides eliminated DPPH, hydroxyl, alkyl and superoxide radicals in a dose-dependent manner. Of the four free radicals, Igoside was able to effectively eliminate hydroxyl radicals with an EC ₅₀ of 16.7 mM. Further studies on the physiological activity of these siliceous compounds are currently underway.

1 is a graph showing the free radical scavenging activity of the l-derived isigoside measured by ESR, (a) DPPH scavenging activity, (b) hydroxyl radical scavenging activity, (c) alkyl radical scavenging activity, (d) superoxide Radical scavenging activity is shown and each value is expressed as mean ± SD over 3 replicates.

Claims (2)

1,2-di-O-palmitoyl-3-O- (6-deoxy-6-amino) -α-D-glucopyranosyl-glycerol (hereinafter, referred to as "isigoside") compound of Formula 1 :

An antioxidant active pharmaceutical composition comprising the isigoside compound of claim 1 as an active ingredient.

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