KR100462843B1 - Free-radical scavengers from Aspergillus sp. F80161 and its novel use - Google Patents

Abstract

The present invention relates to an active oxygen scavenger derived from the fungus and its novel use, and to the fungus Aspergillus sp. Newly isolated F80161 and the culture of the fungus were subjected to column chromatography, Sephadex LH-20 chromatography, LTC and HPLC using an organic solvent such as ethyl acetate or water to perform butyrolactone with active oxygen scavenging activity. After obtaining (butyrolacton) -based compounds and investigating their chemical structure and physicochemical properties, the lipid peroxidation inhibitory activity was measured by TBA method using rat liver microsomes. There is an excellent effect of providing a new use of the chemical that has a protective effect.

Description

Free radical scavengers derived from molds and their novel uses {Free-radical scavengers from Aspergillus sp. F80161 and its novel use}

The present invention relates to active oxygen scavengers derived from molds and their novel uses. More specifically, the present invention relates to a fungus Aspergillus sp., Which produces free radicals, reactive oxygen species. Activated oxygen scavenging activity and lipid peroxidation inhibitory activity obtained by separating F8016 from soil, extracting active oxygen scavenging compounds from the fungal culture extract, investigating its structural and chemical properties, and examining the lipid peroxidation inhibitory activity of the substance The present invention relates to a butyrolactone compound having a novel use.

All aerobic organisms, including humans, get energy through breathing, which is oxygen as the final electron acceptor. As such, the basic triplet oxygen, which is an oxygen but stable molecular state, which is absolutely necessary for life support, is classified into a superoxide radical (O ₂ ⁻ ) and a hydroxyl radical (H ₂ O) by various physical, chemical and environmental factors. ), _Double- sided oxygen-induced lethal oxygen toxicity when converted to highly reactive free radicals or reactive oxygen species such as hydrogen peroxide (H ₂ O ₂ ) and singlet oxygen ( ¹ O ₂ ) I have it. In other words, these free radicals are non-selective and irreversible destructive action on cell components such as lipids, proteins, sugars and DNA, such as cancer, stroke, Parkinson's disease, Alzheimer's disease and brain diseases, aging, heart disease, ischemia, It is known to cause various diseases such as arteriosclerosis, skin diseases, digestive diseases, inflammation, rheumatism, autoimmune diseases and promote aging.

Lipid peroxides are substances produced by the peroxidation of lipids through various oxidation reactions. The active oxygen and free radicals oxidize the phospholipid of the cell membrane containing a large amount of unsaturated fatty acids, thereby producing lipid peroxides. Accumulation of lipid peroxides on cell membranes results in localized disorders in tissues, such as cell membrane fluidity and functionality, resulting in decreased cellular function and altered cell structure. Therefore, excessive accumulation of lipid peroxides in the human body causes various human diseases including hepatic diseases such as stroke due to cerebrovascular disorders, myocardial infarction, diabetic vascular disorders, hyperlipidemia, acute inflammation, rheumatoid disease, and alcoholic hepatitis. To date, synthetic antioxidants such as butylated hydroxytoluene (BHT) or butylated hydroxyanisole (BHA) have been used to inhibit lipid peroxidation. However, the synthetic antioxidants are excellent in lipid peroxidation inhibitory activity, but are highly likely to cause cancer or malformation and thus cannot be used continuously.

On the other hand, even in normal cells, some free radicals and other free radicals and peroxides are generated during metabolism, but superoxide dismutase (SOD) and catalase as in vivo defenses against them. Along with antioxidant enzymes such as peroxidase and low molecular weight antioxidants such as vitamin E, vitamin C, glutathione, ubiquinone and uric acid, it protects itself from oxidative damage. However, when the biological defense mechanism is abnormal or the production of free radicals exceeds the capacity of the biological defense system by various physical and chemical factors, cell destruction due to oxygen toxicity is caused. Thus, antioxidants such as free radical scavengers or free radical scavengers that can inhibit the production of such free radicals or inhibit the production of peroxides can be used to treat various diseases caused by these oxides and The possibility of developing anti-aging agents is well known.

In particular, as the recent research on natural antioxidants as antioxidant antioxidants and the mechanism of antioxidant defense against their oxidative damage, the development of natural reactive oxygen scavengers for use in the treatment of various diseases caused by oxidative damage Much research is being done to do this. In addition, many studies are focused on the relationship between oxidative damage, antioxidant defense mechanisms, and aging and life extension.

Therefore, research to develop new free radical scavengers that can treat various diseases caused by oxidative stress by eliminating free radicals that cause oxidative damage in search of new methods for human health and life extension Will greatly contribute to that.

Accordingly, the inventors of the present invention, while searching for a new high value-added active substance from various microbial cultures inhabiting domestic soil, aspergillus sp. From the culture extract of F80161, butyrolactone-based compounds were isolated and their chemical structures, physicochemical properties, and biological activities were identified. These compounds showed active oxygen scavenging activity and a novel use of lipid peroxidation inhibitory activity. The invention was completed.

Therefore, an object of the present invention is a fungus Aspergillus sp. To provide a novel butyrolactone-based active oxygen scavenging substance having a novel lipid peroxidation inhibitory activity obtained from F80161 and its fungi extract with little toxicity or side effects.

Another object of the present invention is to provide a composition containing an active oxygen scavenger having an excellent lipid peroxidation inhibitory activity as an active ingredient.

The object of the present invention is a fungus Aspergillus sp. Which produces free-radical, ie free radical scavengers. F80161 was isolated from the soil, and the culture extract was treated with silica gel column chromatography, Sephadex LH-20 column chromatography, TLC and HPLC using ethyl acetate, an organic solvent such as acetone or water, and the like. After obtaining the scavengers butyrolactone compounds 161A, 161B and 161c and measuring their respective ¹ H NMR, ¹³ C NMR and HMBC spectra, their chemical structures were revealed, and their lipid peroxidation inhibitory activity using the rat liver microsomes of 161A and 161B was obtained. Achieved by measurement using acid (TBA) method.

Hereinafter, the configuration of the present invention will be described in detail.

1 is a fungal strain of the present invention Aspergillus sp. Schematic diagram showing the process of separating the active oxygen scavengers 161A, 161B and 161C of the present invention from the metabolite of F80161.

FIG. 2 shows HPLC profiles of compounds 161A and 161B of the present invention analyzed using HPLC columns under conditions of YMC-ODS column, 4.6 × 250 mm, 50% ACN, 0.8 mL / min, UV 220 nm.

Figure 3a is a result of measuring the ¹ H (300MHz) NMR spectrum of 161A in CD ₃ OD to identify the chemical structure of the compound 161A of the present invention.

Figure 3b is the result of measuring the ¹³ C (75MHz) NMR spectrum of 161A in CD ₃ OD to identify the chemical structure of the compound 161A of the present invention.

Figure 4a is the result of measuring the HMBC spectrum to identify the complete chemical structure of the compound 161A of the present invention.

Figure 4b shows the ¹ H- ¹³ C long-range correlation shown in the HMBC spectrum analysis to identify the complete chemical structure of the compound 161A of the present invention.

Figure 5a is a result of measuring the ¹ H (300MHz) NMR spectrum of 161B in CD ₃ OD to identify the chemical structure of the compound 161B of the present invention.

Figure 5b is the result of measuring the ¹³ C (75MHz) NMR spectrum of 161B in CD ₃ OD to identify the chemical structure of the compound 161B of the present invention.

Figure 6a is the result of measuring the HMBC spectrum to identify the complete chemical structure of the compound 161B of the present invention.

Figure 6b shows the ¹ H- ¹³ C long-range correlation shown in the HMBC spectrum analysis to identify the complete chemical structure of the compound 161B of the present invention.

Figure 7a is a result of measuring the ¹ H (300MHz) NMR spectrum of 161C in CD ₃ OD to identify the chemical structure of the compound 161C of the present invention.

Figure 7b is the result of measuring the ¹³ C (75MHz) NMR spectrum of 161C in CD ₃ OD to identify the chemical structure of the compound 161C of the present invention.

Figure 8 shows the ¹ H- ¹³ C long-range correlation shown in the HMBC spectrum analysis to identify the complete chemical structure of the compound 161C of the present invention.

9 is a result of measuring the lipid peroxidation inhibitory activity using the rat liver microsomes of the compounds 161A and 161B of the present invention using thiobabituric acid (TBA) method.

The present invention is a fungus strain Aspergillus sp. To produce a free radical scavenging active substance extracted by ethyl acetate. Separating F80161 from the soil and culturing in yeast-malt extraction medium (YM); The acetone extracts of the culture and the cells were separated by acetate extraction and separated by silica gel column chromatography, and the fractions showing free radical scavenging activity were separated by Sephadex LH-20 column chromatography, and the three fractions FrI, Obtaining FrII and FrIII; Purifying the compounds FrI, FrII and FrIII fractions in an acetonitrile solvent to purify compounds 161A, 161B and 161C; Investigating the physicochemical properties, molecular weights and molecular formulas of the purified compounds 161A, 161B and 161C of the present invention by performing EI-MS analysis, melting point investigation, IR spectrum investigation and nuclear magnetic resonance spectrum data analysis; Illuminating the chemical structures of the present inventions 161A, 161B and 161C by measuring the ¹ H NMR, ¹³ C NMR and HMBC spectra of the compounds 161A, 161B and 161C of the present invention; And measuring the lipid peroxidation inhibitory activity of the chemicals 161A and 161B of the present invention using thiobabituric acid (TBA) method using rat liver microsomes. The compounds of the present invention can be obtained by combining alone or suitably known methods used for separation and extraction of mold-containing components.

Hereinafter, specific examples of the present invention will be described in detail with reference to Examples, but the scope of the present invention is not limited thereto.

Example 1 Fungal Strains of the Invention Aspergillus sp. Isolation of reactive oxygen scavengers from culture medium of F80161 strain

Aspergillus sp. The active oxygen scavengers 161A, 161B, and 161C compounds of the present invention were isolated from the culture medium of F80161.

The primary microorganisms were selected by investigating the free radical scavenging activity of the acetone extract of the culture medium and the cells prepared by culturing the obtained microbial strains 100 mL each using a Erlenmeyer flask, and ethylacetate was selected from the selected strains. Fungus Aspergillus sp. F80161 strain was selected.

Aspergillus sp. Of the present invention fungi strain producing the active oxygen scavenger. F80161 was deposited on December 4, 2000, with the accession number KCTC 0895BP to the Gene Bank in the Biotechnology Research Institute.

Aspergillus sp. The F80161 strain was seeded at 28 ° C. and 140 rpm for 3 days using yeast-malt extract (YM) medium and 500 ml Erlenmeyer flask, and then filled with 3 L of YM medium in a 5 L fermentor. Inoculated with 3% seed culture was incubated for 7 days. Acetone (acetone) extracts of the culture medium and the cells were separated by ethyl acetate, and concentrated under reduced pressure to remove the solvent, followed by silica gel column chromatography. At this time, the eluting solvent conditions were fractionated by eluting the CHCl ₃ -MeOH 1: 1 mixed solvent to increase the polarity by increasing the ratio of MeOH from the mixed solvent of CHCl ₃ -MeOH 25: 1. Only fractions (1063 mg) showing free radical scavenging activity in each fraction were collected and subjected to Sephadex LH-20 column chromatography saturated with a CHCl ₃ -MeOH 1: 3 mixed solvent. Collected. At this time, the fractions having free radical scavenging activity were divided into three, and each fraction was named Fr I, Fr II, Fr III in the order of elution. Reverse phase (ODS) HPLC was performed using 55% acetonitrile as a solvent for Fr I to purely isolate 105 mg of free radical scavenger 161A. Meanwhile, 161B compound was separated from Fr II by the following process. Fr II was injected into an MPLC column (ODS 30 × 300 m / m) saturated with 20% acetonitrile and eluted with increasing acetonitrile ratio up to 90% acetonitrile for free radical scavenging activity. Fractions having were collected. The active fractions were concentrated and reverse phase (ODS) HPLC was performed using 50% acetonitrile as a solvent to purely separate 28 mg of 161B. From Fr III, reverse phase ODS TLC (50% ACN) was performed to isolate 4.2 mg of pure 161C compound. The separation and purification of the free radical scavenging material is shown in FIG. 1. In addition, the HPLC profile of the 161A and 161B compounds analyzed using an analytical HPLC column is shown in FIG. 2.

Example 2 Physical and Chemical Properties of Free Radical Scavengers 161A, 161B and 161C Compounds of the Invention

The physico-chemical properties of the compounds 161A and 161B of the present invention were investigated and the results are shown in Table 1. Both of these compounds were separated into a sticky colorless prism state and showed the same UV-Visible spectrum showing a maximum absorption peak at 223 and 309 nm, indicating that the chromophores were the same compound.

Physicochemical Properties of Compounds of Active Oxygen Scavengers 161A and 161B 161A 161B Exterior colorless prism colorless prism Molecular Weight 424 356 Molecular formula C ₂₄ H ₂₄ O ₇ C ₁₉ H ₁₆ O ₇ EI-MS ( m / z ) 424 (M ⁺ ) 356 (M ⁺ ) UVλmax ^MeOH nm (logε) 223 (4.18), 309 (4.31) 223, 309 IR ν max (KBr) cm ^-1 OH: 3565, 3480, 3280C = O: 1755, 1740 3390, 3290, 1760, 17401730, 1660 [α] _D ²⁰ + 100 ° ( c = 1.0, EtOH) + 85 ° ( c = 0.7, EtOH) Melting Point 95 ℃ (decomposition) 91 ~ 94 ℃ Solubility Solvent Insolvent MeOH, sodium carbonate solution CHCl ₃ , n -hexame, EtOAc, sodium carbonate solution MeOHCHCl ₃ , n -hexane, EtOAc, water

As a result of EI-MS analysis of 161A compound, M ⁺ peak appeared at m / z 424, indicating that the molecular weight of this compound was 424. In addition, fragment ions such as m / z 380, 348, 320, 291, 175, 131, 91, and 69 were shown on EI-MS. Compound 161A showed a characteristic that no molecular ion peak was observed on the MS at a temperature higher than 130 ° C. The compound is a weak acid, soluble in sodium carbonate but not in bicarbonate solution. The melting point is unclear, but it starts to melt at 74 ℃ and decomposes at 94∼96 ℃. In the IR spectrum, absorption bands of several types of hydroxyl groups (3565, 3480, 3280 cm ^-1 ) and carbonyl groups (1755, 1740 cm ^-1 ) were observed. This compound showed optical activity of [α] _D ²⁰ = + 100 ^o . Based on the above physicochemical properties and ¹ H, ¹³ C NMR spectrum data analysis, the molecular formula of the 161A compound was determined to be C ₂₄ H ₂₄ O ₇ .

Compound 161B showed a molecular ion peak of m / z 356 on the EI-MS spectrum and a molecular ion peak of m / z 356.0878 (calculated: 356.0894) as a result of measuring high resolution EI-MS, resulting in ¹ H, ¹³ C NMR nuclei. NMR spectrum data analysis showed that the molecular weight was 356 and the molecular formula was C ₁₉ H ₁₆ O ₆ . Examination of the IR spectrum showed absorption bands attributable to hydroxyl groups at 3390 and 3290 cm −1 and carbonyl groups at 1760 and 1740 cm −1. The melting point of the compound was 91-94 ° C., and showed an optical activity of [α] D ²⁰ = + 85 ^o .

The 161C compound was isolated as a yellow powder and exhibited a UV maximum absorption peak at 240, 326 and 370 nm. As a result of measuring EI-MS, a molecular ion peak was observed at m / z 432, and determined as molecular weight 432, molecular formula C ²⁷ H ²⁸ O ⁵ . In the IR spectrum, absorption bands of 3300 cm < -1 > derived from hydroxyl groups and 1700 cm < -1 > derived from carbonyl groups were observed.

Example 3 Chemical Structure of Compound 161A

In order to elucidate the chemical structure of compound 161A, ¹ H NMR, ¹³ C NMR and HMBC spectra of 161A were measured.

As a result of analyzing the ¹ H NMR spectrum (FIG. 3A) of 161A, 13 signals were observed. Of these, two methyl proton peaks were observed as singlets at 1.57 and 1.66 ppm, respectively, and triplet methine at 5.07 ppm, and the respective protons of methylene at 3.07 and 3.09 ppm, respectively. protons were observed, indicating that they form a 3,3-dimethylallyl group. In addition, the aromatic protons of 6.42 ( d , 2.0 Hz), 6.50 ( d , 8.0 Hz) and 6.54 ppm ( d , 2.0, 8.8 Hz) were produced from 1,2,4-tri-substituted benzene rings (1, 2,4-trisubstituted benzene ring) is present. Proton signals, also aromatic protons of 6.88 ( d , 9.0 Hz) and 7.60 ppm ( d , 9.0 Hz), have two protons each superimposed from their integrals, which are 1,4-disubstituted benzenes. It is shown to form a ring (1,4-disubstituted benzene ring) structure. In addition, in the proton spectrum of the compound, AB-type methyl proton (3.41, 3.46 ppm) and O-methyl group were observed.

^{In the 13} C NMR spectrum (FIG. 3b), two methyl carbons (17.7, 25.9 ppm), methine carbon (123.5 ppm), which form a 3,3-dimethylallyl (isoprenyl) group, and methylene carbon (28.6 ppm) was identified. Aromatic carbons 125.1 (C-6), 132.3 (C-7), 128.4 (C-8), 154.9 (C-9), 115.0 (C-10) and 129.7 (C-11) ppm carbons are 1, A 2,4-tri-substituted benzene ring was formed, and the chemical shift value of -154.9 ppm of carbon was found to be bonded to -OH group. On the other hand, two other aromatic carbons, 13,6 ppm (C-18 and C-22) and 116.6 ppm (C-19 and C-21), 123.3 ppm (C-17) and 159.1 ppm (C-20) The carbon of) forms a 1,4-di-substituted benzene ring structure, and 159.1 ppm of carbon is carbon to which -OH group is bonded. Methylene carbon was observed at 39.5 ppm and O-methyl carbon was observed at 53.8 ppm, respectively. On the other hand, two carbonyl carbons were observed at 170.6 and 171.6 ppm, and three quaternary carbons were observed at 140.1 (C-2), 128.8 (C-3) and 86.8 (C-4) ppm. It was found to form a γ-butyrolactone ring with the carbonyl carbon of (C-1). ¹ H NMR and ¹³ C NMR spectral data of this compound 161A are shown in Table 2.

¹ H NMR and ¹³ C NMR spectral data of 161A C δ _C (ppm) δ _H (ppm) One 170.6 C = O 2 140.1 C 3 128.8 C 4 86.8 C 5 39.5 CH ₂ 3.41 ( d , 14.5), 3.46 ( d , 14.5) 6 125.1 7 132.3 CH 6.42 ( d , 2.0) 8 128.4 9 154.9 10 115.0 CH 6.50 ( d , 8.0) 11 129.7 CH 6.54 ( d , 2.0, 8.0) 12 28.6 CH ₂ 3.07 ( dd , 6.5), 3.09 ( dd , 6.5) 13 123.5 CH 5.07 ( t , 6.5) 14 132.9 C 15 25.9 CH ₃ 1.66 ( s ) 16 17.7 CH ₃ 1.57 ( s ) 17 123.3 C 18 130.3 CH 7.60 ( d , 9.0) 19 116.6 CH 6.88 ( d , 9.0) 20 159.1 C 21 116.6 CH 6.88 ( d , 9.0) 22 130.3 CH 7.60 ( d , 9.0) 23 171.6 C OMe 53.8 CH ₃ 3.76 ( s )

The complete chemical structure of the 161A compound was determined by interpretation of the HMBC spectrum. The HMBC spectrum and the 1H-13C long-range correlation shown as a result of the analysis are shown in FIG. 4. Analysis of the HMBC spectra showed that coupling of C-8 and C-9 and C-7 to H-12 from methyl protons of H-12 of the isoprenyl group was observed. And it was found. On the other hand, the C-6, C-7 and C-11 carbons of the benzene ring to which isoprenyl group is bound from the methylne proton of H-5, and conversely, the long- Range correlation was observed to confirm that the methylene group was bound to C-6 of the benzene ring. In addition, long-range coupling from protons of H-5 to quaternary carbons, 86.8 ppm (C-4), 128.8 ppm (C-3) and 171.6 ppm carbonyl carbon (C-23), was observed. Long-range coupling from the protons of the methyl group to the carbonyl carbons of C-23 was observed, respectively. Therefore, it was found that C-5 (39.5 ppm) and methoxy carbonyl group were bound to C-4 (86.8 ppm), the γ-position of the γ-butyrolactone ring. The long-range coupling of C-3 (128.8 ppm) of the β-position of the γ-butyrolactone ring from H-18 and H-22 on the 1,4-disubstituted benzene ring results in a long-range coupling. It was confirmed that it is bound to C-3).

From the above results, the structure of the 161A compound was α-oxo-β- ( p -hydroxyphenyl) -γ- ( p- hydroxy- m- 3,3-dimethylallylbenzyl) -γmethox-ycarbony- as shown in the following general formula [I]. γ-butyrolactone was determined.

Example 4 Chemical Structure of Inventive Compound 161B

In order to elucidate the chemical structure of compound 161B, ¹ H NMR, ¹³ C NMR and HMBC spectra were measured.

¹ H NMR and ¹³ C NMR spectra and analysis results of the 161B compound are shown in FIGS. 5A and 5B and Table 3.

¹ H NMR, ¹³ C NMR Spectrum Data of 161B Compound C δ _C (ppm) δ _H (ppm) One 170.2 C = O 2 139.6 C 3 129.2 C 4 86.7 C 5 39.5 CH ₂ 3.47 ( s ) 6 125.2 C 7 132.5 CH 6.64 ( d , 8.5) 8 115.5 C 6.52 ( d , 8.0) 9 157.4 C 10 115.5 CH 6.52 ( d , 8.0) 11 132.5 CH 6.64 ( d , 8.5) 12 123.0 C 13 130.3 CH 7.60 ( d , 9.0) 14 116.6 CH 6.87 ( d , 9.0) 15 130.3 CH 7.60 ( d , 9.0) 16 116.6 CH 6.87 ( d , 9.0) 17 130.3 CH 7.60 ( d , 9.0) 18 171.4 C OMe 53.9 CH ₃ 3.77 ( s )

Analysis of the ¹ H NMR spectrum of Compound 161B showed only six signals. The aromatic protons of 6.52 ( d , 8.0 Hz), 6.64 ( d , 8.5 Hz), 6.87 ( d , 9.0 Hz) and 7.60 ppm ( d , 9.0 Hz) in the aromatic region were used as their integrals. From the above results, it was found that two protons overlap each other, which is almost identical to the chemical shift and coupling constant of the protons forming the 1,4-disubstituted benzene ring structure in the 161A compound. In addition, the proton spectrum of the compound showed methyl proton (3.47 ppm) and O-methyl group (3.77 ppm) that appeared in 161A compound. However, the isoprenyl group that was present in the 161A compound was not present in the 161B compound. Thus, this compound was assumed to be a deisoprenylated compound of 161A compound.

^{In the 13} C NMR spectrum, methyl, methine and methylene carbon, which formed isoprenyl group, were not observed in 161A compound. On the other hand, aromatic carbons contain two overlapping 132.5 (C-7, C-11), 115.5 ppm (C-8, C-10) and 157.4 (C-9) and 125.2 ppm (C-6) carbons. Forming a 1,4-disubstituted benzene ring of which is also two overlapping carbons: 130.3 ppm (C-13, C-17), 116.6 ppm (C-14, C-16) and 123.0 ppm (C -12) and 159.3 ppm (C-15) of carbon constituted another 1,4-disubstituted benzene ring. From chemical shifts of C-9 and C-15, it can be seen that -OH groups are bonded to these carbons. Other carbons were similar to those observed for the 161A compound, with methylene carbon at 39.5 ppm, O-methyl carbon at 53.9 ppm, and methoxy carbonyl carbonyl carbon (C-18) at 171.4 ppm, respectively. In addition, carbons forming the γ-butyrolactone ring were observed at 170.2 (C-1), 139.6 (C-2), 129.2 (C-3) and 86.7 ppm (C-4), respectively. The ¹ H- ¹³ C long-range correlation shown in the HMBC spectrum analysis of the 161B compound is shown in FIG. 6B. As a result, it was almost identical to the 161A compound except that the correlation of the isoprenyl group shown in 161A was not shown. Therefore, the chemical structure of the 161B compound was determined as α-oxo-β- ( p- hydroxyphenyl) -γ- ( p- hydroxybenzyl) -γ-methoxycarbony-γ-butyrolactone as shown in the following general formula [II].

Example 4 Chemical Structure of Inventive Compound 161C

The chemical structure of compound 161C was analyzed by analysis of ¹ H NMR, ¹³ C NMR and HMBC spectra.

¹ H NMR and ¹³ C NMR spectra and analysis results of the 161C compound are shown in FIGS. 7A and 7B and Table 4.

¹ H NMR, ¹³ C NMR Spectrum Data of 161C Compound C δ _C δ _H One C 176.8 2 C 94.4 3 C 179.2 4 C 147.6 5 CH 104.1 6.14 ( s ) 6 C 125.8 7 CH 129.9 7.50 ( dd , 8.5, 3.0) 8 CH 115.9 6.72 ( d , 8.5) 9 C 156.0 10 C 128.7 11 CH 132.8 7.43 ( d , 3.0) 12 CH ₂ 29.5 3.30 ( d , 8.0) 13 CH 124.8 5.34 ( m ) 14 132.9 14'-Me CH ₃ CH ₃ 17.926.0 1.71 ( s ) 1.74 ( s ) 6 ' C 125.8 7 ' CH 129.3 7.72 ( dd , 8.5, 3.0) 8' CH 115.3 6.70 ( d , 8.5) 9 ' C 152.8 10 ' C 128.3 11 ' CH 132.0 7.83 ( d , 3.0) 12 ' CH ₂ 29.4 3.30 ( d , 1.5) 13 ' CH 124.8 5.34 ( m ) 14 ' 132.9 14'-Me CH ₃ CH ₃ 17.926.0 1.71 ( s ) 1.74 ( s )

The ¹ H NMR spectrum of Compound 161B showed four methyl groups, two each at 1.71 (6H) and 1.74 ppm (6H), and 3.30 ppm (4H, d , 8.0 Hz), 5.34 ppm (2H) , proton signals of m) were observed. It is assumed that they constitute two 3,3-dimethylallyl groups. In addition, the six protons (6.70-7.83ppm) found in the aromatic region were estimated to form two 1,2,4-tri-substituted benzene rings. In addition, a disconnected methine peak was observed at 6.14 ppm.

^{In the 13} C NMR spectrum, methyl, methine, and methylene carbon, which are considered to form two isoprenyl groups, were almost overlapped with each other. In other words, two methyl carbones of 17.4 and 26.0 ppm each, and a mathine carbone of 124.8 ppm were overlapped, and methylene carbon was almost overlapped at 29.4 and 29.5 ppm, respectively. On the other hand, 12 aromatic carbons between 115.3 and 156.0 ppm were estimated to form two isoprenyl groups and hydroxy-dimethylallylphenyl groups, respectively. In addition, carbonyl carbon at 176.8 ppm, mathine carbon at 104.1 ppm, and three quaternary carbons at 94.4, 147.6 and 179.2 ppm were observed.

Complete chemical structure determination of the 161C compound was performed by analysis of the HMBC spectrum, and the ¹ H- ¹³ C long-range correlation shown in the HMBC spectrum analysis is shown in FIG. 8. As a result, the long-range coupling between isoprenyl group and phenyl group as shown in 161A was observed. In other words, the two isoprenyl groups are each bonded to C-10 and C-10 'of the phenyl ring. On the other hand, the long-range correlation from the methine protons of H-5 to the C-7 and C-11 carbons of the benzene ring to which isoprenyl group is bound, and conversely from the C-7 and C-11 carbons to H-5 protons This observation confirmed that the methylene group is bonded to C-6 of the benzene ring. In addition, long-range coupling was observed from the protons of H-5 to 147.6 ppm (C-4) and 179.2 ppm (C-3), the quaternary carbon, indicating that C-4 (147.6) is the γ position of the C-5 lactone ring. ppm). On the other hand, another hydroxy-dimethylallylphenyl group, at 7.83 (H-11 ') and 7.72 ppm (H-7'), was at 94.4 ppm (C-2) at the α position of the lactone ring from the long-range coupling, namely C-2. It can be seen that the combination. Of the four quaternary carbons, 176.8 ppm of carbon is carbonyl carbon and the carbon observed at 179.2 ppm of low magnetic field is interpreted as having an OH group. From the above results, the structure of the 161C compound was determined as shown in the following general formula [III].

Example 5 Analysis of Lipid Peroxidation Inhibitory Activity of Compounds 161A and 161B of the Invention

Lipid peroxidation inhibitory activity using the rat liver microsomes of Compounds 161A and 161B was measured by thiobabituric acid (TBA) method. In this method, oxygen in the reaction medium is reduced by divalent iron to produce superoxide radicals, and the microsomes of rat liver added as lipid sources by the radical chain reaction thereof are peroxidated. This method is to measure lipid peroxidation by reacting malondialdehyde (MDA) with TBA. As shown in Figure 9 161A and 161B compounds showed superior lipid peroxidation inhibitory activity compared to vitamin E, a representative antioxidant used as a control. That is, the concentration of the active substance (IC ₅₀ ) required to inhibit lipid peroxidation of rat liver microsomes by 50% was 4.3 ㎍ / ml for vitamin E, whereas 161A was only 0.12 ㎍ / ml and 161B was only 1 ㎍ / ml. Therefore, the lipid peroxidation inhibitory activity of the rat liver microsomes of 161A and 161B compounds was 40 and 4 times stronger than vitamin E, respectively. These results demonstrate that 161A and 161B compounds have potent lipid peroxidation inhibitory activity, which shows the necessity of reviewing their pharmacological activity in the future.

As is apparent from the above examples, the present invention is a fungus Aspergillus sp. Butyrolactone system, which shows the strong free radical scavenging activity of F80161, which newly separates and accumulates in vivo from it, destroys cells and removes reactive oxygen species that cause fatal oxygen toxicity to the biological defense system. There is an excellent effect of providing compounds 161A, 161B, 161C. In addition, as a result of testing the lipid peroxidation inhibitory activity of the butyrolactone compound 161A, 161B, 161C of the present invention by using the rat liver microsome TBH method, it showed very excellent lipid peroxidation activity and prevented aging by active oxygen It is a very useful invention in the pharmaceutical manufacturing industry and cosmetic manufacturing industry because it shows an excellent effect to apply to medicines, cosmetics, etc.

Claims (5)

Butyllactone compound 161A having a lipid peroxidation inhibitory activity or an active oxygen scavenging activity represented by the following general formula [I]. Butyrolactone compound 161B having a lipid peroxidation inhibitory activity or an active oxygen scavenging activity represented by the following general formula [II]. Butyllactone compound 161C having a lipid peroxidation inhibitory activity or an active oxygen scavenging activity represented by the following general formula [III]. Aspergillus sp. F80161 strain (KCTC 0895BP), which produces the compound of claims 1 to 3. A lipid peroxidation inhibitory activity or an active oxygen scavenging activity composition comprising the compound of any one of claims 1 to 3 as an active ingredient.