JP4922551B2 - Maple syrup fortified with bioactive phenolic compounds - Google Patents

Description

  The present invention is present in a maple family plant and has physiological activities such as alpha glucosidase inhibitory activity, active oxygen scavenging ability (SOD activity) or growth inhibition activity of human promyelocytic leukemia cells (HL-60), diabetes, The present invention relates to phenolic compounds useful for the prevention and treatment of lifestyle-related diseases such as obesity and cancer, and foods containing these compounds.

  In recent years, diseases such as diabetes, obesity, and cancer are also called lifestyle-related diseases, and improvement of lifestyle habits such as improving eating habits and lack of exercise and avoiding excessive stress are the first prevention and treatment. It is said to be one step. The effect of active oxygen has been pointed out on the aging phenomenon, and the degree of progression is said to be greatly influenced by lifestyle habits. It is thought that the influence of eating habits is the largest among lifestyles, and it is fundamental to take a diet that is nutritionally balanced, but recently it has been further promoted to prevent lifestyle-related diseases. The development of health foods that allow intake of therapeutically effective ingredients in daily eating habits has also been actively conducted. If such a physiologically active ingredient is found in natural food ingredients and can be taken naturally in the daily diet, it is the most desirable food form for the prevention and treatment of lifestyle-related diseases. Can be.

  For this purpose, the first step is to find effective ingredients for prevention and treatment of lifestyle-related diseases from natural materials. Conventionally, as an example of finding such an active ingredient from among plant components, a phenolic compound called desmanthin-1 was extracted from a genus plant Myrecia multiflora (Lam.) DC, which has an aldose reductase activity. Therefore, it has been reported that it has an effect on antidiabetic properties (see Non-Patent Document 1).

  On the other hand, it has been proposed by the present inventors to enhance nutrition by improving vitamins in maple syrup (see Patent Document 1). Maple syrup is made by collecting sap from maple trees that grow in the continental region from Quebec, Canada to Ohio, USA, for example, from late February to around April, and heating and concentrating them. As a natural sweetener with a safe and favorable taste, it has been used for hundreds of years worldwide.

As for the components of maple syrup, monosaccharides and oligosaccharides as saccharides, as well as various minerals, vitamins, and the like have been investigated, and there have been few reports on research on components other than these. Recently, it has been proposed to fortify vitamin C as in Patent Document 1, but in addition to this, the chemical study of the components of maple plants and the biochemical and drug related to the health promotion effect of maple syrup The physical examination has not been sufficiently conducted to date.
WO 03/070021 A1 Chem. Pharm. Bull. 46 (1) 113-119 (1998)

  The desire to prevent and treat lifestyle-related diseases such as diabetes, obesity, and cancer and the suppression of aging phenomena through daily diets is higher than ever. For this purpose, it is important to obtain these ingredients having physiological activity effective for prevention and treatment from natural food materials. Maple syrup, on the other hand, is a traditional sweetener with a history of hundreds of years. Although it is widely used around the world, its effectiveness and uses other than sweeteners have not been developed yet. It is.

  Therefore, the object of the present invention is to conduct chemical and pharmacological research on the components of the maple family plant to obtain medicinal resources, in particular, components effective for the prevention and treatment of lifestyle-related diseases and aging, and then the active components It is to develop a food containing sucrose so that it can be easily consumed in daily eating habits.

  Under such circumstances, the present inventors have focused on a maple family plant as a natural material and have started searching for and using an active ingredient contained therein. That is, component extraction is performed using each part of a maple plant as a sample, each fraction is subjected to biochemical evaluation, an effective fraction is separated, further purification is performed, a compound is isolated, and its chemical structure As a result, the effectiveness of prevention and treatment of lifestyle-related diseases was examined. As a result, the effects of some phenolic compounds were recognized, and the present invention was completed through further research.

That is, the present invention provides the following phenolic compounds and foods containing them.
1) A component of a maple family plant, characterized by having at least one physiological activity of alpha glucosidase inhibitory active oxygen scavenging ability (SOD activity) and growth inhibitory activity of human promyelocytic leukemia cells (HL-60). Phenolic compounds.

2) A food comprising the phenolic compound described in 1) above as an active ingredient.
3) A maple syrup obtained by strengthening the phenolic compound described in 1) above.
The significance of the three physiological activities of the phenolic compound of the present invention and the measuring methods thereof are as follows.

  Alpha glucosidase inhibitory activity is an action that suppresses the action of alpha glucosidase, a digestive enzyme that breaks down disaccharides into glucose, and suppresses absorption of carbohydrates taken by meals from the small intestine, resulting in post-meal blood glucose Is expected to have an effect of suppressing the level of diabetes, and is expected to have an effect of preventing obesity from the prevention and treatment effects of diabetes, as well as the low intake of fructose. The alpha glucosidase inhibitory activity can be measured according to the method of A. Dahlqvist [Anal. Biochem. 7, 18-25 (1964)].

  An active oxygen scavenging enzyme (SOD active enzyme) is produced in animals and plants and has a function of removing active oxygen (superoxide) with a metal-containing enzyme. Typical diseases caused by active oxygen include cancer, leukemia, cerebral blood flow, adult diseases such as cerebral infarction and myocardial infarction, and aging. Substances with active oxygen scavenging ability are expected to have preventive and therapeutic effects. The SOD activity can be measured according to the method of M.W.Sutherland et al. [Free Rad. Res., 27,283 (1997)].

  The growth inhibitory action on human promyelocytic leukemia cells is based on the growth of human leukemia cell cultures and is used to evaluate the antitumor activity. The inhibitory substance is effective in preventing cancer and inhibiting cancer cell growth. Be expected. The growth inhibitory activity of human promyelocytic leukemia cells (HL-60) can be measured according to the method of P.R.Twentyman et al. [Cancer, 56, 279-285 (1987)].

  The phenolic compound of the present invention is present in a maple family plant which is a source of maple syrup that has been edible for many years. 60) It has at least one kind of physiological activity of the growth inhibitory activity. Therefore, based on each of these active effects, the prophylactic / therapeutic effect on the above-mentioned diseases is sufficiently expected. In addition, by including the phenolic compound in various foods, it can be safely ingested through daily eating habits, and a health promotion effect can be expected. The phenolic compound of the present invention is particularly useful for the prevention / treatment of diseases based on alpha glucosidase inhibitory activity and active oxygen scavenging ability among the above three physiological activities.

  The phenolic compound of the present invention can be isolated and purified from a maple plant and has at least one activity of alpha glucosidase inhibitory activity, reactive oxygen scavenging activity and growth inhibition activity of human promyelocytic leukemia cells (HL-60). The compound which has is included. Representative phenolic compounds (compounds 1 to 11) of the present invention are exemplified below.

In the above formula, Me represents a methyl group, Glc represents a glucose group, Rha represents a rhamnose group, and galloyl represents a galloyl group.
As exemplified above, the phenolic compound of the present invention has a phenol skeleton in the chemical structural formula and a plurality of hydroxy groups in the molecule, as long as the hydroxy group has the physiological activity described above. May be substituted. The compound of the present invention includes a phenol skeleton including a compound having only a phenol skeleton such as compound 3, for example, a benzofuran derivative (compounds 2 and 4), a coumarin derivative (compounds 5, 6, 7, and 8), Each derivative constituting a flavone derivative (compounds 9, 10, 11) or a lignan derivative (compound 1) is included.

Examples of the substituent of the hydroxy group include a lower alkyl group (such as methyl) or a glycosyl group (such as glucose and rhamnose).
Next, as a maple family plant referred to in the present invention, red maple (Acer rubrum), silver maple (Silver Maple: Acer saccharinum), manitoba maple (Manitoba Maple; Acer negrundo), black maple (Black Maple; Acer nigrum), Norway Maple (Acer platanoides), Sycamore Maple (Acer pseudoplatanus), Canyon Maple (Acer grandidentatum) or Bigleaf Maple (Acer macreophyllum). Among these, the maple family plants that use the sap to produce maple syrup include silver maple and black maple, which are useful as a source of the phenolic compound of the present invention.

  The phenolic compounds of the present invention are obtained from sites such as leaves, xylem, sap, fruits or roots of maple plants. In order to obtain the phenolic compound of the present invention from such a site, plant component separation and purification means can be appropriately combined. For example, when a silver maple xylem is used as a material, it is immersed and extracted at room temperature for about 45 days using an alcohol solvent such as ethanol, and the soluble part is collected and concentrated to obtain the phenolic compound of the present invention. A crude extract containing can be obtained. As a solvent used when obtaining a crude extract, solvents such as water, alcohol, acetone, and ethyl acetate can be used alone or in combination. The extraction temperature can be selected as desired under temperature conditions such as room temperature, ice-cooling such as 0 ° C., or near the boiling point of the solvent.

  The crude extract thus obtained was extracted with a mixed solvent of ethyl acetate and water, the ethyl acetate soluble part was fractionated by column chromatography, and each component was fractionated. A reactive oxygen scavenging ability (SOD activity), an alpha glucosidase inhibitory activity, and a growth inhibitory action on human promyelocytic leukemia cells are examined, and a fraction having at least one of these biological activities is separated.

For example, a crude extract of a silver maple xylem is extracted with a mixed solvent of ethyl acetate and water to obtain an ethyl acetate soluble part, which is then subjected to silica gel column chromatography to eliminate active oxygen in each fraction ( Using the SOD) action as an index, 12 active fractions are obtained. Subsequently, 8 compounds are isolated by performing re-column chromatography etc. These were recognized as phenolic compounds because of their physicochemical properties, and when their respective chemical structural formulas were determined from physicochemical studies, seven of them were known compounds and one was a new compound. When the SOD activity of each compound was examined, the activity was recognized in 7 types. It has never been reported that these compounds have been obtained from a maple family, and it is a new finding that these compounds have been found to have SOD activity. Among these, compounds 2, 5 and 6 have strong SOD activity exceeding that of ascorbic acid. On the other hand, the growth inhibitory action of these compounds on human promyelocytic leukemia cells (HL-60) is observed in a concentration-dependent manner in the range of 10 ⁻⁴ to 10 ⁻⁵ g / mL in compounds 6 and 7. It is done.

As a method for obtaining a compound having SOD activity from a maple plant, the present inventors separated active fractions and active compounds by a combination of normal phase, reverse phase column chromatography and a mixture of various solvents, and the like. It was the first time that they could be separated.
On the other hand, using the alpha glucosidase inhibitory activity, which is an evaluation system for anti-obesity and anti-diabetic prevention and treatment effects as an index, the extraction method for active ingredients in silver maple leaves is similarly extracted, separated, Isolation was performed to obtain three phenolic compounds. Of these three compounds, Compound 10 was found to have a strong alpha glucosidase inhibitory activity. This activity is more than 10 times as potent as 1-deoxynojirimycin, which is known to have this action. That is, the silver maple leaf itself or an extract thereof can be used for the prevention / treatment of anti-obesity and anti-diabetes.

  On the other hand, a component having alpha glucosidase activity can be obtained from maple syrup produced from sap such as silver maple. For example, Canada No.1 medium, which is representative of maple syrup, is adsorbed as a sample on column chromatography packed with porous filler Toyopearl HW-40, and then the column is eluted with water. Monosaccharides and some oligosaccharides are eluted in the fraction. A mixture of saccharide and phenolic compound is eluted in the fraction eluted with 5% methanol aqueous solution or 5% isopropyl alcohol aqueous solution. Subsequently, further elution with 50% methanol or 50% isopropyl alcohol gives a fraction of phenolic compounds. This phenolic compound fraction has a strong alpha glucosidase inhibitory activity. Moreover, alpha glucosidase activity is also observed in the elution fraction of the mixture of sugar and phenolic compound, but this activity is not observed in the sugar fraction. Since the alpha glucosidase inhibitory activity was observed in the group of phenolic compounds found from the leaves of maple, the anti-obesity and anti-diabetic effects of the anti-obesity and anti-diabetics as well as the components of the leaves, as will be apparent from Test Example 1 described later. There are compounds that can be expected to act, and these can be used for prevention and treatment.

  On the other hand, maple syrup produced late in the harvest tends to have a dark color and a savory taste, but the reason remains unclear. These products are referred to as Super Dark as a poor grade maple syrup, distinguished from those used as normal sweeteners. These quality products are used for flavoring tobacco rather than as sweeteners. For this Super Dark sample, the same active component separation experiment as that of the above sample was examined in comparison with the sample of the maple syrup Canadia No. 1 medium. % Methanol elution fractions were obtained as 0.9 g and 1.3 g, respectively, and contained more phenolic compounds than Canada No. 1 medium sample. This indicates that the Super Dark sample contains more bioactive components that are expected to have anti-obesity and anti-diabetic effects than the Canadia No. 1 medium sample.

  Therefore, the phenolic fraction can be separated using Super Dark and added to normal maple syrup to enhance and strengthen health promoting components such as anti-obesity and anti-diabetic effects. For example, 10 L of Maple syrup Super Dark is added to a Toyopearl HW-40 column (10 × 100 cm), and then the sugar fraction is sufficiently eluted and removed with 50 L of water, and then eluted with 50 L of 5% aqueous methanol solution. After removing the remaining sugar fraction, 13 g of the fraction eluted with 20 L of 50% aqueous methanol solution is collected. This brown fraction has alpha glucosidase inhibitory activity. The phenolic compound of the present invention can be strengthened by adding the brown fraction to ordinary maple syrup. For example, by adding 1 g of the brown fraction to 500 g of maple syrup Canada No. 1 medium, a maple syrup product in which the phenolic compound of the present invention is enhanced and strengthened can be produced. The addition amount of the phenolic compound of the present invention can be appropriately selected in consideration of taste and the like, but is generally about 0.01 to 20% by weight in maple syrup, preferably 0.1 to 5% by weight. %.

  Next, when the phenolic compound of the present invention is contained in various foods and ingested, the preventive / therapeutic effect of the corresponding disease is sufficiently expected based on the above physiological activity. In this case, you may add the phenolic compound of this invention to a foodstuff in the concentrated state, without isolating from the site | part of a maple, for example. For example, the above-mentioned phenolic compound in Super dark is added to maple syrup Canada No.1 medium without separation treatment to produce health foods having anti-obesity, anti-diabetes, anti-cancer, active oxygen scavenging, etc. can do. Although it does not specifically limit as the said target food, For example, syrups, drinks (coffee, wine etc.), confectionery (candy, cake, cookies, jelly, chocolate etc.), sweeteners, livestock foods (butter, Cheese)) or teas.

  As described above, the phenolic compound of the present invention can be safely ingested on a daily basis by incorporating it into various foods including the form of maple syrup. The phenolic compound of the present invention is generally taken in an amount of 0.1 to 200 mg, preferably 1 to 10 mg per day for an adult. What is necessary is just to make the said phenolic compound contain in target food so that it can ingest in this range.

Hereinafter, the present invention will be described more specifically with reference to examples.
Example 1
18 kg of ethyl alcohol was added to 3.7 kg of crushed xylem of silver maple (Acer saccharum), and the mixture was allowed to stand at room temperature for 45 days for extraction. Next, the ethyl alcohol-soluble part was filtered, and then the solvent was distilled off under reduced pressure to obtain a crude extract. This crude extract was extracted with a mixed solution of 9 L of ethyl acetate and 9 L of water, and fractionated into an ethyl acetate soluble part and a water soluble part, and the solvent was distilled off under reduced pressure for each. A soluble part (54.0 g) and a water-soluble part (140 g) were obtained.

The alpha glucosidase inhibitory activity of the obtained ethyl acetate soluble part showed strong activity (1 × 10 ⁻⁴ g / mL, 100%). Next, the ethyl acetate soluble part of the obtained strong active fraction was applied to a silica gel column chromatogram and eluted with a mixed solution of n-hexane-diisopropyl ether-methanol (25: 2: 0.1). The amount of isopropyl ether was increased, and column chromatography was performed with a mixed solution of n-hexane-diisopropyl ether-methanol (25: 8: 0.1) to obtain 12 fractions. The alpha glucosidase inhibitory activity of each fraction was measured at 1 × 10 ⁻⁴ g / mL, and the inhibitory activity was expressed in%. As a result, fractions 1-5 (57.7%), 6 (95.6%), 7 (97.4%), 8 (70.2%), 9 (93.1%), 10 (98.98). 9%), 11 (91.2%), and 12 (97.8%) showed activity. Of these, fractions 6, 7, 9 and 10 which showed strong activity were further purified by column chromatogram, and the following compounds were isolated.

Isolation and qualification of compound 7:
Fraction 6 (3.14 g) was subjected to silica gel column chromatography with an elution solvent of diisopropyl ether-methanol (20: 1) to isolate Compound 7 (157.8 mg). From amorphous powder, FAB-MS: m / z: 191 [M (C ₁₀ H ₈ O ₄ ) -H], this product is a compound already isolated by T. Morinaga et al., Scopoletin [Chem. Pharm. Bull., 51, 62-67 (1989)].

Isolation and qualification of compound 2:
Fraction 7 (2.64 g) was subjected to silica gel column chromatogram with diisopropyl ether-methanol-water (25: 2: 0.1) solvent to isolate compound 2 (14.6 mg). Amorphous powder, [α] _D -8.53 (C = 6.7, methanol), FAB-MS: m / z: 359 [M (C ₂₀ H ₂₄ O ₆ ) -H]. It was found to be identical to the compound [Phytochemistry, 28, 3483-3485, (1989)] already isolated by Miyase et al.

Isolation and qualification of compounds 5 and 8:
Fraction 9 (2.25 g) was subjected to silica gel column chromatogram with diisopropyl ether-methanol-water (25: 3: 0.1) solvent to isolate compound 5 (85.0 mg). From amorphous powder, [α] _D −8.0 (C = 0.2, methanol: pyridine = 1: 1), FAB-MS: m / z: 415 [M (C ₂₁ H ₂₀ O ₉ ) —H] This product was found to be identical to the compound clemiscosin C [Phytochemistry, 27,636-638, (1998)] already isolated by ABRay et al.

Compound 8 (4.3 mg) was then isolated. From amorphous powder, [α] _D -14.9 (C = 0.3, methanol), FAB-MS: m / z: 401 [M (C ₂₀ H ₁₈ O ₉ ) -H], this product is obtained from XFCheng et al. Was identified as 5'-dimethylaquillochin [Fitoterapia, 71, 341-342 (2000)] already isolated.
Isolation and qualification of compounds 5 and 6:
Fraction 10 (4.99 g) was subjected to silica gel column chromatogram with diisopropyl ether-methanol-ethyl acetate-water (6: 2: 4: 1) solvent, and then HPLC (ODS) to 30-60% aqueous methanol. And compound 5 (37.3 mg) was isolated. Compound 6 (13.4 mg) was then isolated. From amorphous powder, [α] _D −4.42 (C = 0.5, methanol), FAB-MS: m / z: 415 [M (C ₂₁ H ₂₀ O ₉ ) -H], this product is ABRay et al. Was identified as clemiscosin D [Phytochemistry, 27,636-638, (1998)].

Isolation and qualification of compounds 1, 3 and 4:
Fraction 10 (4.99 g) was subjected to silica gel column chromatogram with diisopropyl ether-methanol-ethyl acetate-water (6: 2: 4: 1) solvent, followed by LH-20 column chromatography and further HPLC (ODS) chromatography. Compounds 1, 3 and 4 were isolated respectively.

Compound 1 (22.0 mg) was first isolated. Compound 1 includes amorphous powder, [α] _D -40.1 (C = 1.70, methanol), HR-FAB-MS: m / z: 549.2006 [MH] ⁻ (calcd for C ₂₇ H ₃₃ O ₁₂ ), FT-IR (film): 3400, 1760, 1050 cm ⁻¹ , and from the analysis results of various spectra, it was determined that the compound was a novel compound having the above chemical structural formula. Compound 3 (38.7 mg) was then isolated. Compound 3 is amorphous powder, [α] _D -65.7 (C = 1.0, methanol: pyridine = 1: 1), FAB-MS: m / z: 345 [M (C ₁₅ H ₂₂ O ₉ ) − H], this product was found to be identical to koaburside (Phytochemistry, 45, 149-157, (1997), a compound isolated by H. Achenbach et al., Followed by compound 4 (138.6 mg). This compound was isolated from amorphous powder, [α] _D -184.7 (C = 2.0, methanol), FAB-MS: m / z: 505 [M (C ₂₆ H ₃₄ O ₁₀ ) —H. From this result, this product was identified as identical to icariside E4 [Phytochemistry, 28, 3483-3485, (1989)], a compound already isolated by T.Miyase et al.

Example 2:
In Example 1, the active oxygen scavenging ability (SOD activity) of each compound isolated from the xylem of maple (Acer saccharum) was measured by the method of MW Sutherland et al. [Free Rad. Res., 27,283 (1997)]. Is shown in Table 1.

As apparent from Table 1, compounds 1, 2, 3, 4, 5 and 6 showed SOD activity, and in particular, the activities of compounds 2, 5, and 6 exceeded those of vitamin C.
Example 3:
To 320 g of silver maple (Acer saccharum) leaves, 2 L of ethyl alcohol was added, and the mixture was allowed to stand at room temperature for 45 days for extraction. The ethyl alcohol-soluble part was filtered, and then the solvent was distilled off under reduced pressure to obtain a crude extract. The crude extract was extracted with a mixed solution of ethyl acetate and water, and separated into an ethyl acetate soluble part and a water soluble part, and the solvent was distilled off under reduced pressure to dissolve the ethyl acetate soluble part. Part (13.5 g) and a water-soluble part were obtained.

The alpha glucosidase inhibitory activity of the obtained ethyl acetate soluble part showed strong activity (1 × 10 ⁻⁴ g / mL, 100%). Next, the ethyl acetate soluble part of the strong active fraction obtained was applied to a silica gel column chromatogram and eluted with a mixed solution of n-hexane-ethyl acetate (1: 1). The column chromatography was performed until the mixture was eluted and the mixed solution of n-hexane-ethyl acetate (0:10) was eluted to obtain 6 fractions. The alpha glucosidase inhibitory activity of each fraction was measured at 1 × 10 ⁻⁴ g / ml, and the inhibitory activity was expressed in%. As a result, fractions 1 (67.4%), 2 (99.3%), 3 (62.0%), 4 (93.5), 5 (80.8%), 6 (94.3) Indicated. Fraction 4 which showed strong activity was further purified by column chromatogram, and the following compounds were isolated together with compound 1-8.

Isolation and qualification of compounds 9, 10 and 11:
Fraction 4 is applied to a silica gel column chromatogram, column chromatography is performed with an elution solvent of n-hexane-ethyl acetate (4: 6), and then eluted with 55% methyl alcohol using an HPLC (ODS) column. The fractions were further rechromatographed to isolate compounds 9 (71.3 mg), 10 (9.22 mg) and 11 (5.02 mg) from the 45% methyl alcohol elution fraction. Compound 9 is an amorphous powder, [α] _D -70.5 (C = 3.6, pyridine), FAB-MS: m / z: 447 [M (C ₂₀ H ₂₀ O ₁₁ ) -H]. Are compounds already isolated by K. Hyong Ja et al. [J. Nat. Prod., 61,145-148, (1998)]. Compound 10 was obtained from amorphous powder, [α] _D -5.9 (C = 0.9, pyridine), FAB-MS: m / z: 599 [M (C ₂₈ H ₂₄ O ₁₅ ) -H]. Was identified to be identical to the compound [Chem. & Pharm. Bull., 47, 1484-1486 (1999)] already isolated by P. Si-Hyung et al. From Compound 11, amorphous powder, [α] _D -127.2 (C = 0.4, pyridine), FAB-MS: m / z: 431 [M (C ₂₁ H ₂₀ O ₁₀ ) -H] K. Hyong Ja et al. Have been identified as the same compound [J. Nat. Prod., 61, 145-148, (1998)] already isolated.

Example 4
The alpha glucosidase inhibitory activity of each compound (1-11) isolated from xylem and leaves of maple (Acer saccharum), which was carried out in Examples 1 and 3, was determined by the method of A. Dahlqvist [Anal. Biochem. , 7, 18-25 (1964)]. Each activity of Compound 1-11 was expressed as IC ₅₀ (μm).

  The results were as follows: Compound 1 (> 100), Compound 2 (79.3), Compound 3 (> 100), Compound 4 (not identified), Compound 5 (> 100), Compound 6 (not identified), Compound 7 ( 14.8), Compound 8 (not identified), Compound 9 (not identified), Compound 10 (3.6) and Compound 11 (not identified). Compound 10 was found to have 10 times or more stronger activity than 1-deoxynijrimycyn (51.0) of the comparative control compound.

Example 5
Alpha glucosidase inhibitory activity was measured using Canada No1 medium as a sample of maple syrup. Dilute 500 mL of the sample to 1 L with water, apply it to a HW-40 column (100 mL), and then elute with 500 mL of water, then elute with 300 mL of 5% isopropyl alcohol and 500 mL of 50% isopropyl alcohol for 2 minutes. Each was concentrated under reduced pressure to obtain a crude fraction, a 5% isopropyl alcohol fraction (30.5 mg), and a 50% isopropyl alcohol fraction (60 mg). The alpha glucosidase inhibitory activity (1 × 10 ⁻³ g / mL) of each fraction was 0.09 and 0.67, respectively. Strong activity was observed in the 5% isopropyl alcohol fraction.

Example 6
Isolation of alpha glucosidase inhibitory activity using Super dark as a sample of maple syrup. Dilute 500 mL of the sample to 1 L with water, apply it to a HW-40 column (100 mL), elute with 500 mL of water, elute with 300 mL of 5% isopropyl alcohol and 500 mL of 50% isopropyl alcohol for 2 minutes. Each was concentrated under reduced pressure to obtain a crude fraction, a 5% isopropyl alcohol fraction (34 mg), and a 50% isopropyl alcohol fraction (55 mg). The alpha glucosidase inhibitory activity (1 × 10 ⁻³ g / mL) of each fraction was 0.15 and 1.70, respectively. Strong activity was observed in the 5% isopropyl alcohol fraction.

Example 7
1 g of the 5% isopropyl alcohol fraction obtained in Example 5 was added to 500 mL of Canada No1 medium of maple syrup, and the mixture was stirred to obtain a syrup in which the component of alpha glucosidase inhibitory activity was enhanced.
Example 8
0.3g of compound 10 was added to 500mL of maple syrup Canada No1 medium, and the syrup which strengthened alpha glucosidase inhibitory activity was obtained.

Example 9
1 mL, 2 mL, 3 mL, 4 mL, and 5 mL of Canada No.3 Dark, a dark quality maple syrup, was added to each normal Canadian maple syrup, Canada.
By adding to 9 mL, 8 mL, 7 mL, 6 mL, and 5 mL of No. 1 Medium, the maple component containing the phenolic compound of the present invention is 10%, 20%, respectively, compared to normal maple syrup. 30%, 40% and 50% fortified maple syrup was obtained. When the fortified maple syrup was evaluated with a focus on the taste and the degree of coloring, no problem was found in quality.

Test example 1
The results of HPLC analysis of the phenolic compound of the present invention are shown below.
HPLC analysis conditions:
Column Unsion UK-Phenyl 100 x 4.6 mm (Imtakt)
CH ₃ CN: 10% AcOH = 15: 85 Flow rate: 1 mL / min Detection: UV 254 nm
Table 2 shows the retention times of compounds 1-8.

  Canada medium No. 1 5% isopropyl alcohol elution and Super dark 5% isopropyl alcohol elution both contained the above-mentioned compounds, and also showed 4 or 5 compound peaks by 10 minutes retention time. Overall, there is no significant difference between Canada medium No. 1 elution with 5% isopropyl alcohol and Super dark 5% isopropyl alcohol. In addition, both 50% isopropyl alcohol elution and Super dark 50% isopropyl alcohol elution of Canada medium No. 1 contain the above-mentioned compounds, and there is no significant difference between the two components. Compared with the 5% elution, the retention time was slow, and it was confirmed that the contents of Compound 2 (18.0 minutes), Compound 6 (28.5 minutes) and Compound (35.0 minutes) were both high.

  The phenolic compound of the present invention exists in a maple family plant and has physiological activities such as alpha glucosidase inhibitory activity, active oxygen scavenging ability (SOD activity), or growth inhibition activity of human promyelocytic leukemia cells (HL-60). For example, health can be promoted safely by containing it in food and taking it on a daily basis.

Claims (3)

A component of Aceraceae Plants, have at least one biological activity of alpha-glucosidase inhibitory activity and scavenging activity (SOD activity), whose chemical structure is the following compound 1,2,3,4,5 and 10

(Wherein Me is a methyl group, Glc is a glucose group, Rha is a rhamnose group, and galloyl is a galloyl group) syrup. In strengthening the phenolic compound, a fraction containing the phenolic compound and having at least one physiological activity of alpha glucosidase inhibitory activity and active oxygen scavenging ability (SOD activity) is isolated from a maple family plant. The maple syrup according to claim 1, which is used for strengthening. The maple syrup according to claim 1 or 2, wherein the maple plant material is maple syrup graded as Super Dark.