KR19990048807A - Extraction and Separation Methods of Natural Antioxidant Components from Bark of Oak Tree - Google Patents

Abstract

The present invention relates to a natural antioxidant component extracted from bark of an oak bark tree, and more particularly, to extract the bark of an oak bark tree, which is discarded in the existing wood industry, with a mixed solution of methanol and water, and then washed with heptane and diethyl ether. It relates to a natural antioxidant component excellent in the antioxidant effect obtained by fractionation.

Description

Extraction and Separation Methods of Natural Antioxidant Components from Bark of Oak Tree

The present invention relates to a natural antioxidant component extracted from bark of an oak bark tree, and more particularly, to extract the bark of an oak bark tree, which is discarded in the existing wood industry, with a mixed solution of methanol and water, and then washed with heptane and diethyl ether. It relates to a natural antioxidant component excellent in the antioxidant effect obtained by fractionation.

In general, antioxidants are known to be very beneficial to the human body. However, conventional synthetic antioxidants are toxic, unlike natural products, and because they are additives that are artificially added to human intakes such as foods, they are problematic in nature. Efforts have been made to obtain non-toxic natural antioxidants.

For example, the extract obtained by extracting degreasing rice bran with a mixture of water and ethanol is filtered and concentrated, and then an antioxidant is prepared [Domestic Patent Publication No. 95-10536] and green tea leaves are extracted with calcium carbonate-added water. After adsorbing to an adsorption resin, eluting with an organic solvent, converting it into ethyl acetate, and concentrating and drying to obtain a catechin powder, a method of preparing a natural antioxidant (Korean Patent Publication No. 93-5694) and the like are known. However, these methods are difficult to industrialize because the use of relatively rare materials such as rice bran and green tea has a problem in supply and demand of raw materials in mass production.

In addition, there is also a method of extracting and concentrating ethanol from turmeric, hawthorn, wormwood, donkey, larvae, gojija, and sanjoin to obtain an antioxidant substance [Korean Patent Publication No. 96-3620], which is a relatively simple extraction method. Although it is possible to obtain a good natural antioxidant, in this case, too, since the material used is a relatively expensive herbal medicine, there is a problem in mass production as in the above case.

On the contrary, in recent years, extracts have been obtained at room temperature or warm conditions using water or some organic solvents alone or in a mixture of bark or leaves of pineaceae plants, and the water and fat-soluble solvents are added to the extracts to give an aqueous solution layer. Separation using HPLC and then recrystallization to obtain an antioxidant furanone material or a natural antioxidant containing the same (Korean Patent Publication No. 95-29281) has been disclosed. However, this method can be said that industrial production is possible in terms of supply and demand by using pine and bark as raw materials.However, since HPLC is used to separate materials, it takes a lot of time and money, and separates single materials. Attempts have been made to increase the antioxidant activity, but the use of such a single substance is significantly lower in yield compared to the use of a few ingredients, so that the overall economic efficiency has not been improved despite the use of cheap bark raw materials. There is an urgent need for improvement in the separation method.

Therefore, there is a limit to producing a competitive product that can replace the existing synthetic antioxidants or antioxidants produced by conventional methods.

In addition, there are known methods for separating natural antioxidants from various plant materials [US Patent Nos. 5,427,806, 5,209,870, 5,102,659, 5,026,550, 4,923,679]. As it is a foreign kind, there is a problem in supply and demand of raw materials in domestic production, and the separation method is also focused on the separation of too pure material, which has a fundamental problem of costly and time-consuming separation.

As such, conventional methods of extracting and separating natural antioxidant components can be summarized as extracting and concentrating antioxidant components using water or an organic solvent, and separating the components having excellent antioxidant effects using a column or HPLC. Can be. However, the conventional method of separating using a column or HPLC in this way not only takes a long time but also consumes a lot of reagents consumed in the separation, resulting in high cost due to high separation costs. In addition, by limiting the extraction raw materials of conventional natural antioxidant components to edible or medicinal plants, or by using a separation method to obtain too pure substances even if using cheap materials, the raw material cost is high, and by using rare materials that are difficult to obtain in some parts As a material and separation method adopted in the prior art, such as supply and demand of raw materials, it was difficult to mass-produce and industrialize natural antioxidant components.

Acanthopanaxs sessiliflorus used in the present invention is a deciduous shrub with flowers blooming in August to September, the fruit ripening in October. Since ancient times, ogalpi trees have been used for medicinal, edible and ornamental purposes, and in Chinese medicine, they are used for the purpose of 지 志, soothing, 심 心, 中心, 中風, and tonic. come. Efforts have recently been made to utilize extracts of the ogalpi trees. For example, reports on the anticomplement activity of polysaccharides derived from Acanthopanaxs enticosus [Korean Journal of Agricultural Chemistry, 1992, 35:42] and the effect on the fat accumulation of hot water extract of Ogalpi [Korean Journal of Nutrition and Nutrition, 1992, 21] : 9] and the like, and related patents are related to the manufacturing method and the anti-inflammatory effect of dieterpen and its derivatives isolated from the extracts of the bark of bark and bark roots. ], A beverage production method using the extract of the roots and fruit of the organza (Korean Patent Publication No. 94-618), and a method for purifying saponin from the extract of the organza [Korean Patent Publication No. 89-1097].

However, to date, no extracts of oak bark have been studied or used for antioxidant purposes.

Therefore, the present invention is to use a bark of bark, a cheap material that is discarded in the timber industry, in order to improve various problems in the extraction and separation method of the antioxidant component of the prior art, unlike the conventional method of simple extraction and separation method The purpose of the present invention is to produce and use the natural antioxidant component which is low in manufacturing cost compared to the existing antioxidants and exhibits at least two times the antioxidant effect.

1 is a process diagram schematically illustrating a process of extracting an antioxidant component from the bark of an oak bark according to the present invention,

FIG. 2 is a process diagram schematically illustrating a method of separating an antioxidant component from a bark extract obtained through the process of FIG. 1 through a solvent fractionation process.

3 shows the IR spectrum for the diethyl ether fraction.

The present invention is characterized by a natural antioxidant component obtained by pulverizing bark of bark, extracted with a mixed solution of water and methanol, washed with heptane and fractionated with diethyl ether.

Referring to the present invention in more detail as follows.

The present invention extracts antioxidant components from the bark of the ogopi tree, which is discarded as waste in the timber industry, with a mixture of water and organic solvent, and simply washes and fractionates them by polarity using several organic solvents. To get.

As used in the present invention, for example, one or two or more selected from among the gangolpipi, wanggaopipi, island ogalpi, talugalpi, Seoul ogalpi, Jirisan ogalpi, etc. can be suitably used. Wastes produced in the wood industry can also be used. They are crushed to a size of 3 mm or less to make them suitable for extraction, such as to facilitate solvent penetration.

In the present invention, the organic solvent used in the initial solvent extraction and mixed with water is suitable for methanol, the mixing ratio of methanol: water mixed with a volume ratio of 5: 1 to 1: 5 is used to complete the active ingredient Good in terms of extraction,

On the other hand, the present invention is an important feature to greatly improve the extraction effect by the method of washing the extract sequentially with different organic solvents for the separation of effective antioxidants, solvent extraction is degreased with, for example, heptane The antioxidant components can then be effectively separated by washing with diethyl ether. At this time, degreasing with heptane is for removing fat-soluble components, and for effective degreasing, it is preferable to use after dissolving the extract before use of heptane in methanol.

The extraction and separation method according to the present invention will be described in more detail in a series of processes with reference to FIGS. 1 and 2 as follows.

As shown in FIG. 1, the extract obtained from the bark is pulverized to a size of 3 mm or less in diameter to facilitate penetration of the solvent, and then the raw material is methanol: water of 5: 1 to 1: 5. It is immersed in the mixed solution mixed by volume ratio, left for several days, and then filtered to obtain an extract. Extraction at this time can be carried out repeatedly several times to increase the yield of the extract. The extracts thus obtained are combined and concentrated to about 1/10 of the original volume using a reduced pressure concentrator. In this state, mostly water is present.

The concentrated extract is washed with a process as shown in FIG. 2. First, methanol is added to completely dissolve all components, and then heptane is added to perform degreasing through layer separation, which is also repeated several times. Then, diethyl ether was added to the concentrated heptane washed solution, followed by washing several times with fractions to remove nonpolar components. The diethyl ether fraction thus obtained is concentrated and spray-dried to complete the commercialization of the natural antioxidant component.

In the washing fraction process according to the present invention, in addition to diethyl ether, ethyl acetate, alcohols and the like may also be used as the solvent for fractionation, but the experimental results of the present invention showed that the extract fractionated with diethyl ether after washing with heptane showed the best antioxidant activity.

As a result of the antioxidant activity measurement using DPPH for the antioxidant natural extract prepared according to the present invention, the ED50% value was found to be 3 μl / ml or less, and the total phenol content was obtained at 65% or more.

The present invention will be described in more detail based on the following examples, but the present invention is not limited thereto.

Example

After the bark of the oak bark was pulverized to a size of 3 mm or less in diameter, the pulverized sample was immersed in a 10-fold methanol: water (3: 1, v / v) mixed solution, left for 2-3 days, and then filtered and extracted. Gathered bay. In order to increase the yield of the extract was again replaced with a new methanol: water mixture solution, and the extraction was left for 2 to 3 more days, and then filtered again to obtain an extract. The obtained extracts were combined and concentrated to about 1/10 of the original volume using a vacuum concentrator. In this state, mostly water is present.

The concentrated extract was added with the same amount of methanol to completely dissolve all the components, and then shaken vigorously by adding the same amount of heptane, followed by standing. Thereafter, when the layer was formed, degreasing was performed by removing the heptane layer, and the above procedure was repeated twice (preparation of heptane fraction). An equal amount of diethyl ether (Et ₂ O) was poured into the concentrated solution of heptane washing, and washing fractions were carried out three times in the same manner as in the case of heptane to remove nonpolar components (Et ₂ O fraction preparation). For a more thorough wash fraction, the same amount of ethyl acetate (EA) having a polarity similar to diethyl ether was poured into the concentrate and washed three times in the same manner as above (EA fraction preparation). Each fraction thus obtained and the methanol / aqueous layer remaining after fractionation were concentrated and dried using a spray dryer to powder.

Experimental Example

The extract of bark of bark tree prepared in Example was measured for antioxidant activity in order to compare the antioxidant capacity of the existing antioxidant (comparative example 1) and the extract of woody part of tree (comparative example 2), The results are shown in Table 1 below.

-How to measure antioxidant activity-

The antioxidant activity of the extract was measured by the following method using a spectrophotometer.

First, 2.96 mg of DPPH (1,1-diphenyl-2-picrylhydrazine) was dissolved in 50 ml of methanol to make a standard DPPH solution. Samples of fractions for which antioxidant activity was to be measured were dissolved in methanol, and seven samples were prepared such that the concentrations were 1.25, 2.5, 5.0, 10.0, 20.0, 40.0, and 80.0 μg / ml, respectively. Then, 1 ml of standard DPPH solution was added thereto, shaken and left for 30 minutes, and then absorbance was measured at 520 nm using a spectrophotometer. The result was obtained from the regression equation of absorbance at each concentration, and the concentration having the absorbance corresponding to half of the standard DPPH solution (hereinafter referred to as "ED50% value") was taken as the antioxidant capacity of the sample. ].

In general, the lower the value of ED50% is evaluated to have excellent antioxidant capacity.

division Antioxidant ED50% value (μg / ml) Existing Antioxidant (Comparative Example 1) α-tocopherol 6.53 Butylhydroxyanisole (BHA) 5.15 BHT (dibutylhydroxytoluene) 10.82 Oak Tree Wood Extract (Comparative Example 2) Heptane fraction 890.34 Et ₂ O Fraction 8.46 EA fraction 22.13 Methanol / water layer 72.40 Oak Tree Bark Extract (Example) Heptane fraction 160.51 Et ₂ O Fraction 2.76 EA fraction 44.15 Methanol / water layer 80.23

According to Table 1, the diethyl ether (Et ₂ O) fraction of the fraction of the bark extract of Agal bark shows the best antioxidant activity, which is 2.4 times improved compared to the tocopherol known as a natural antioxidant. Therefore, if you want to use the extract of the oakpi tree for antioxidant use, separate the skin of the bark as in the present experiment, extract with a mixture of water and methanol, concentrate and wash with heptane and extract only the active ingredient with diethyl ether. It can be seen that using as a natural antioxidant is the most effective extraction and separation method.

-Bark Extract Characteristics and Antioxidant Activity-

As a result of analysis of antioxidant properties of bark extracts, the total phenolic content and antioxidant activity of bark extracts were 0.93, indicating that total phenolic content was closely related to antioxidant activity.

As shown in Table 2, in the case of the diethyl ether (Et ₂ O) fraction having better antioxidant capacity than tocopherol, the total phenol content was analyzed to be about 68 ± 4%. Therefore, the ethyl ether (Et ₂ O) fraction in the bark extract of the Acanthopanax japonica can be found to have superior antioxidant capacity than natural tocopherol. In addition, the IR spectrum of the ethyl ether (Et ₂ O) fraction is shown in FIG. 3.

Total Phenolic Content (%) for Each Bark Extract Fraction division Heptane fraction Et ₂ O Fraction EA fraction Methanol / water layer Oak Tree Wood 2 ± 1% 56 ± 4% 42 ± 3% 18 ± 4% Oak Tree Bark 6 ± 3% 68 ± 5% 23 ± 5% 14 ± 3%

As described above, the present invention uses a cheap bark of bark as a raw material, unlike the prior art, the extraction and separation process is different, in particular through the diethyl ether fraction, the antioxidant extract is more than two times better than the conventional one It can be produced very economically.

Claims (7)

Extract the antioxidant components from the bark of the oak bark tree, which was pulverized, extracted with a mixed solution of water and methanol, concentrated, and then washed with heptane and fractionated with diethyl ether to obtain a natural extract. And how to separate. The method of claim 1, wherein the mixed solution of water and methanol is water: methanol in a ratio of 5: 1 to 1: 5 by weight. The method of extracting and separating antioxidant components from the bark of an oak bark according to claim 1, wherein the concentrate is sufficiently dissolved in methanol prior to degreasing to heptane. Antioxidant extract from bark of bark, characterized in that the extract is isolated according to claim 1. [6] The antioxidant extract from the bark of the bark of claim 4, wherein the tree is selected from one or two or more selected from thorn ogalpi, wangigao galpi, island ogalpi, hair ogalpi, seoul ogalpi, and zilic acid ogalpi. 5. The extract of claim 4, wherein the extract has an antioxidant activity as measured by DPPH and has an ED50% value of 3 µg / ml or less. 5. The extract of claim 4, wherein the extract has an average total phenol content of at least 65%.