Patent application title: Method for Preparation of Extract from Angelica Gigas Nakai and Composition Containing the Extract
Sang Jae Park (Yongin City, KR)
Won Cheol Choi (Incheon City, KR)
Sung Pil Kwon (Seoul, KR)
AZI Co., Ltd.
IPC8 Class: AA61K3623FI
Class name: Drug, bio-affecting and body treating compositions plant material or plant extract of undetermined constitution as active ingredient (e.g., herbal remedy, herbal extract, powder, oil, etc.)
Publication date: 2011-01-06
Patent application number: 20110003017
Patent application title: Method for Preparation of Extract from Angelica Gigas Nakai and Composition Containing the Extract
Sang Jae Park
Won Cheol Choi
Sung Pil Kwon
ANDRUS, SCEALES, STARKE & SAWALL, LLP
Origin: MILWAUKEE, WI US
IPC8 Class: AA61K3623FI
Publication date: 01/06/2011
Patent application number: 20110003017
Disclosed is Angelica gigas Nakai extract powder prepared by a process
including: (a) preparing 85-100% (v/v) pure ethanol and adjusting a pH
value of the ethanol to pH 7-9 with an alkaline solution; (b) adding
3-10× of the prepared ethanol to the total weight of Angelica gigas
Nakai herb, which is naturally grown or harvested; (c) conducting
extraction of the same at 30-75° C. for 3-8 hrs; (d) measuring a
pH value of the treated mixture at 15 m intervals and adjusting the same
to pH 7-9 with the alkaline solution during the extraction, while
stopping the pH control at 1 hr prior to completion of the extraction;
(e) filtering and concentrating the obtained extract to 3× the net
weight of Angelica gigas Nakai; (f) cooling and settling the treated
mixture at 10-30° C. for 1-24 hrs to remove a settled precipitate;
(g) adding an excipient such as microcrystalline cellulose, dextrin, etc.
to a concentrate remaining after removal of the precipitate; and (g)
drying and pulverizing the resulting product.
1. A composition including Angelica gigas Nakai extract wherein decursin,
decursinol angelate and decursinol are contained in relative ratios of 50
to 200:1 in terms of (decursin+decursinol angelate) to decursinol.
2. A method for the preparation of Angelica gigas Nakai extract as set forth in claim 1, including: preparing 85 to 100% (v/v) ethanol as solvents and adjusting the pH value of the ethanol to pH 7 to 9 with an alkaline solution; adding 3 to 10 times of the prepared ethanol to the total weight of Angelica gigas Nakai herb in a dried state; elevating a temperature of the mixture and conducting extraction of the same for 3 to 8 hours; measuring a pH value of the treated mixture at 15 minute intervals and adjusting the same to pH 7 to 9 with the alkaline solution during the extraction; concentrating the obtained extract and cooling to remove a precipitates; adding an excipient to the concentrate remaining after removal of the precipitates; and drying and pulverizing the resulting product.
3. A composition with analgesic efficacies, including the composition as set forth in claim 1 as a main component.
4. A composition with antioxidant effects, including the composition as set forth in claim 1 as a main component.
5. The method according to claim 2, wherein the alkaline solution comprises at least one selected from a group consisting of sodium hydroxide, potassium hydroxide, magnesium hydroxide and calcium hydroxide.
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to a method for preparation of an extract of Angelica gigas Nakai and a composition containing the extract prepared by the same, more particularly, to a method for preparation of an ethanol extract of Angelica gigas Nakai, which contains decursin, decursinol, and decursinol angelate in a relative ratio of 50 to 200:1 in terms of (decursin+decursinol angelate) to decursinol known as active ingredients, so as to accomplish considerably improved antioxidant and analgesic effects.
2. Description of the Related Art
Angelica gigas Nakai is well known as a nontoxic medical herb and is widely used as a folk medicine. Oriental herbal medicine is typically used in an extract form prepared by hot water extraction and medical efficacies thereof have been studied and disclosed in prior documents including, for example: S. M. Kim et al., Journal of the Korean Society of Food Science and Nutrition, 1998, reporting antioxidant effects of herbal medicine to lipids; S. A. Kang et al., The Korean Society of Community Nutrition [Poster SESSION] p4-6, 2004 (J. Korean Soc. Food Sci. Nutr., 33(7), 1112-8, 2004) reporting radical removal and antioxidant effects of a methanol extract and a hot water extract; S. Y. Lee et al., Food Sci Biotechnol., 12(5), 491-6, 2003 describing various ingredients contained in Angelica gigas Nakai extract such as decursin, decursinol, decursionl angelate, essential oils, and the like. Moreover, K. S. Ahn et al., reported at the 1996 Spring Conference of the Society, P89, that the medial efficacy, that is, cell toxicity of decursinol angelate and decursin contained in Angelica gigas Nakai extract is based on activation of protein kinase C; S. H. Lee et al. described anti-cancer efficacy of the above ingredients (Arch. Pharm. Res. 26(9), 727-30, 2003); Y. W. Lee et al. disclosed the medical efficacy in preventing platelet coagulation and blood coagulation attained by the above ingredients as well as nodakenin (Arch. Pharm. Res., 26(9), 723-6, 2003); K. W. Yoon et al. reported a methanol extract with anti-microbial activity prepared using extracted fractions of Angelica gigas Nakai and Angelica acutilobae in an organic solvent (Korean J. Plant Res., 17(3) 278-82, 2004); S. H. Lee et al. suggested a methanol extract with improved anti-microbial activity (Arch. Pharm. Res., 26(6), 449-52, 2003); J. B. Hwang et al. disclosed that Angelica gigas extract contains 18.1 wt. % protein, 8.9 wt. % lipid, 57 wt. % carbohydrate, etc. through analysis of chemical composition thereof (Korean J. Food Sci. Technol., 29(6), 1113-8, 1997); B. S, Noh et al. reported results of analysis for flavor constituents of Angelica gigas using a surface acoustic wave sensor (Korean J. Food Sci. Technol., 35(1), 144-8, 2003); M. K. Cho et al. reported results of a study for composition of volatile compounds and constitutional ingredients in Angelica gigas (Korean J. Medicinal Crop Sci., 11(5), 352-7, 2003); J. J. Kwak et al. reported results of analysis for volatile flavor compounds of oleoresin in Angelica gigas (J. Korean Soc. Tobacco Sci., 20(2), 210-7, 1998), and so forth.
As to preparation of Angelica gigas Nakai extract, any organic solvent or water is mostly used and, in particular, studies for identification of constituents and/or observation of efficacies thereof have usually employed organic solvent extraction and/or fractions thereof. S. Y. Lee et al. proposed a micro-wave process for extraction of active ingredients from Angelica gigas (J. Korean Soc. Food Sci. Nutr., 29(3), 442-7, 2000), and M. S. Lee et al. disclosed results of a study for production of Angelica gigas extract using hot water and alcohol (Korean J. Food & Nutr., 14(6), 543-7, 2001).
As for Angelica gigas Nakai, it is known that carbohydrate content is relatively high to about 50% by weight of the extract thereof (Korean J. Food Sci. Technol., 29(6), 1113-8, 1997). However, contents of mono-saccharides and/or di-saccharides such as glucose, fructose, sucrose, etc. are very low, down to around 0.5 wt. %. Therefore, it is expected that sugar ingredients in Angelica gigas Nakai are polymeric substances. Based on this fact, Y. S. Park et al. disclosed results of a study for fractionation of stem cells of angelan as a polysaccharide from Angelica gigas (Korean J. Medicinal Crop Sci., 10(4), 2002).
Since Angelica gigas Nakai as a traditional herbal medicine was used for a long time, a number of technologies patented or described in pending applications for commercially available use thereof have been suggested.
Such patents and/or pending applications include, for example: Korean Patent No. 0,519,456 granted to the Korea Institute of Oriental Medicine, disclosing an extract for inhibition of side effects in cancer chemotherapy; Korean Patent No. 0,427,704 granted to GETWELL Bio Co., Ltd., describing a composition containing decursinol extracted from Angelica gigas Nakai for protection and treatment of gastric mucosal damage; Korean Patent No. 0,480,970 granted to Natural Endo Tech Co., Ltd., proposing an agonistic composition based on dioxins using Angelica gigas extract; Korean Patent No. 0,448,680 granted to Scigen Harvest Co., Ltd., describing a composition containing decursinol for treatment of hepatic damage such as hangover; Korean Patent Application No. 10-2003-0074646 filed by S. C. Lee, disclosing a composition containing Angelica gigas extract for prevention and treatment of dementia; Korean Patent No. 0,504,405 granted to S. Y. Chung, suggesting a decursin containing composition with inhibitory effects to nephrotoxicity; Korean Patent No. 0,441,644 granted to the Korea Research Institute of Bioscience and Biotechnology, describing a composition containing angelan as a polysaccharide from Angelica gigas Nakai for prevention and treatment of diabetes; Korean Patent Application No. 10-2000-0060196 filed by Scigenic Co., disclosing a composition containing decursinol for prevention and treatment of septic shock; Korean Patent No. 0,385,092 granted to INHA University, suggesting a process for preparation of active ingredients from Angelica gigas by culturing root cells of Angelica gigas Nakai; Korean Patent No. 0,252,194 granted to the Korea Institute of Science and Technology, disclosing separation of pectin based polysaccharides from Angelica gigas Nakai and use thereof for enhancement of immune effects; Korean Patent No. 0,187,881 granted to JEIL Pharmaceutical Co., Ltd., describing a process for preparation of an anticancer agent containing decursinol angelate; Korean Patent No. 0,059,142 granted to Korea Food Research Institute, suggesting a process for production of Angelica gigas based beverage; Korean Patent No. 0,132,609 granted to Rural Development Administration, proposing a process for production of granular tea based on Angelica gigas; Korean Patent No. 0,003,649 granted to H. J. Cho, suggesting an anti-tuberculosis agent extracted from Angelica gigas; Korean Patent No. 0,002,596 granted to J. Y. Wi, disclosing a method for preparation of Angelica gigas extract using petroleum ether and lead acetate; Korean Patent No. 0,003,195 granted to H. S. Kim, describing extraction of volatile compounds from Angelica gigas by pH control; Korean Patent Application No. 10-2004-0046403 filed by S. P. Han, disclosing a drug for treatment of urinary disturbance and a composition for treatment of oedema which both contain Angelica gigas extract; Korean Patent Application No. 10-2004-0036199 filed by MD Bio Alpha Co., Ltd., proposing a composition for treatment diabetics using Angelica gigas extract; Korean Patent Application No. 10-2005-0079103 filed by H. W. Kim, describing a blood vessel relaxant agent containing decursin and/or decursinol angelate; Korean Patent Application No. 10-2005-0087384 filed by STC Nara Co., Ltd., disclosing a composition for prevention or treatment of dementia and improvement of cognitive functions which contains Angelica gigas extract; Korean Patent Application No. 10-2004-0008975 filed by Elcomscience Co., Ltd., describing a complex extract composition containing Angelica gigas ingredients for improvement and activation of brain functions and cognitive functions; and Korean Patent No. 0,176,413 granted to Korea Institute of Science and Technology, suggesting use of decursin as an anticancer agent.
Furthermore, Korean Patent Nos. 0,775,741 and 0,749,233 granted to AZI Co., Ltd. proposes a process for preparation of Angelica gigas Nakai extract using alcohol fraction and, especially, described a composition containing decursion and decursinol in a ratio in contents of 200 to 400:1 as well as another composition including at least 30% of decursion and at least 0.25% of decursinol. Such technologies described above are substantially similar to one another in view of extraction of Angelica gigas Nakai using an ethanol based solvent, however, extraction methods are different in terms of detailed techniques to control compositional ratios of active ingredients and/or compositions obtained thereby have different constitutional ratios. The present invention is characterized by controlled pH values of an extraction solvent from pH 7 to 8 over an extraction period and, in addition, proposes a novel extracted product which includes decursinol angelate as well as decursin and decursinol so as to attain compositional ratios of to 200:1 for (decursin+decursinol angelate):decursinol. Therefore, it is believed that the present invention has novelty and an inventive step over conventional technologies. Of course, AZI's patent suggests an increase in content of decursinol by alkaline treatment. However, such increase of the decursinol content means further processing of the extract after extraction. Also, decursinol angelate is not contained in the product, thus exhibiting compositional ratios of the product which are clearly different from those of the present invention. Moreover, extraction conditions used in the present invention are distinguishable from conventional methods as follows:
1) The extraction is performed at PH 7 to 9 which was never disclosed in the prior art.
2) Compositional ratio for (decursin+decursinol angelate):decursinol in the extract ranges from 50 to 200:1 which was never proposed in the prior art.
The present invention relates to a method for preparation of Angelica gigas extract with a high content of decursinol having high pharmacological activity. A number of methods for preparation of Angelica gigas extract are currently known. For example, a pressing process includes compressing freshly harvested Angelica gigas herb having high water content without drying the same. Although the pressing process requires simple equipment and operation, production yield is relatively low. Another method is steam distillation including extraction at 100quadrature or more, by which heat sensitive ingredients are susceptible to decomposition and/or degradation by heat. Use of an organic solvent may extract active ingredients very efficiently, however, a great amount of solvent is required for extraction and energy consumption is considerably increased during removal of the solvent. A supercritical fluid extraction process has advantages of a high extraction rate and reduced consumption of solvent, however, encounters problems in scale-up of equipment and requires expensive capital costs. In addition, an extraction process using microwaves is known. However, such a process also requires expensive capital costs and has problems such as scale-up of equipments and/or considerably increased energy consumption.
Current methods for preparation of Angelica gigas extract generally use water, ethanol, ether or hexane as an extraction solvents and the extract obtained thereby is mostly blended with other herbal extracts to form a simple mixture without requiring an alternative process for purification thereof. As for the Angelica gigas extract, numerous studies and investigations have been conducted into processes for separation of polysaccharides from the extract and/or extraction of active ingredients such as decursin, decursinol, decursinol algelate, etc. However, there has yet to be proposed a process for preparation of Angelica gigas extract with an improved content of decursinol.
Even though results of a study for analgesic effects of decursinol as an active ingredient contained in Angelica gigas Nakai were reported in Life Science 73 (2003), 471-485, this study was only conducted for a single substance, that is, decursinol. On the other hand, a composition containing decursin, decursinol angelate and decursinol according to the present invention exhibits more noticeably enhanced analgesic effects and may favorably be used in a variety of industrial applications.
SUMMARY OF THE INVENTION
Therefore, the present invention has been made in view of the above problems, and it is an object of the present invention to provide a process for preparation of an extract from Angelica gigas Nakai with improved performances wherein the extract contains decursin, decursinol angelate and decursinol in relative compositional ratios of 50 to 200:1 in terms of (decursin+decursinol angelate) to decursinol and, in addition, a composition including the extract.
As for Angelica gigas extract typically prepared by any conventional method, the ratio of decursin to decursinol in the extract is generally known to range from 20 to 400:1. In such a case, adding an excessive amount of sodium hydroxide to the extract maintains the extract in the range of pH 9 to 14 in order to accelerate hydrolysis of decursin thereof. However, this is a very severe and undesirable process for the extract and hydrolysis of numerous natural substances occurring under strong alkaline conditions is duly expected.
Accordingly, the present invention is directed to development of a process for preparation of an extract containing functional components with specified compositional ratios thereof under mild conditions instead of severe conditions described above, as well as a composition including the extract prepared by the above process. In addition, another object of the present invention is to provide a composition with superior analgesic effects over those of pure decursinol.
In order to achieve the above objects of the present invention, there is provided an extract powder prepared from Angelica gigas Nakai while maintaining an extraction solvent at pH 7 to 8, wherein the extract powder contains decursin, decursinol angelate and decursinol and a content of decursinol is at least 0.1 wt. % so as to attain a specified compositional ratio of 50 to 200:1 in terms of (decursin+decursinol angelate) to decursinol. The present invention also provides a process for preparation of the extract powder described above under mild conditions. A composition containing decursin and decursinol angelate is produced using the above prepared extract powder, exhibiting improved analgesic effects over those of pure decursinol.
As described above, the present invention is characterized in that a favorably mild process is conducted to produce Angelica gigas Nakai extract in place of a conventional strong alkaline treatment, so as to obtain a natural extract-like product while considerably enhancing performance thereof. Therefore, the present invention has merits of preventing hydrolysis of natural substances due to a severe alkaline treatment and producing an extract composition with excellent antioxidant and analgesic effects.
BRIEF DESCRIPTION OF THE DRAWINGS
The above and other objects, features and other advantages of the present invention will be more clearly understood from the following detailed description taken in conjunction with the accompanying drawings, in which:
FIG. 1 illustrates a chemical structure of decursinol;
FIG. 2 illustrates a chemical structure of decursin;
FIG. 3 illustrates a chemical structure of decursinol angelate;
FIG. 4 illustrates analysis results of Angelica gigas Nakai extract according to the present invention by gas chromatography; and
FIG. 5 illustrates results of analgesic effects of the extract according to the present invention through a writhing test in mice.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
Hereinafter, exemplary embodiments of the present invention will be described in detail with reference to the following examples as well as the accompanying drawings.
In general, it is difficult to increase the content of decursinol in an extract which is prepared from Angelica gigas Nakai, since the decursinol content in this herb is very low. The present inventors found that hydrolysis of decursin and decursinol angelate, contents of which are high in Angelica gigas Nakai, under alkaline condition could produces decursinol. Therefore, the present invention proposes a process for preparation of Angelica gigas Nakai extract with increased content of decursinol, wherein Angelica gigas Nakai is subjected to extraction using alcohol while adding an alkaline compounds such as sodium hydroxide, potassium hydroxide, calcium hydroxide, calcium oxide, etc. to the solvent in order to maintain the extract solution at pH 7 to 9 until 1 hour before completing the extraction. It is known that decursin is a structural isomer with decursinol angelate as illustrated in FIGS. 1 and 3, respectively. On the other hand, decursinol has a chemical structure shown in FIG. 2. Decursin (or decursinol angelate) is a compound obtained by esterification of a hydroxyl group (--OH) of decursinol. Such an ester is decomposed into an alcohol (--OH form) and an acetic acid (--COOH form) by hydrolysis thereof under alkaline conditions. The present inventors applied such principles in production of Angelica gigas extract so as to accomplish a process for preparation of Angelica gigas Nakai extract with increased content of decursinol. Korean Patent No. 0,749,233 described an extraction process involving use of a solvent with controlled pH 9 to 14 by an alkaline substance such as sodium hydroxide. However, sodium hydroxide is a strong alkali to induce a condition for simultaneous hydrolysis of multiple ingredients in a natural substance, causing a significant variation in compositional ratio of the extract. Conversely, the present invention continuously maintains the pH value of the extract in the range of pH 7 to 9 while conducting the extraction so that a possibility of undesirable hydrolysis may be considerably reduced.
According to the present invention, a process for preparation of Angelica gigas Nakai extract powder includes: preparing 85 to 100% (v/v) ethanol and adjusting a pH value of the ethanol to pH 7 to 9 with an alkaline solution; adding 3 to 10 times of the prepared ethanol to the total weight of Angelica gigas Nakai herb in a dried state; conducting extraction of the same at 30 to 75quadrature for 3 to 8 hours; measuring a pH value of the treated mixture at 15 minute intervals and adjusting the same to pH 7 to 9 with the alkaline solution during the extraction, while stopping the pH control at 1 hour before completing the extraction; filtering and concentrating the obtained extract to 2.5 to 3.5 fold of the net weight of Angelica gigas Nakai; cooling and settling the treated mixture at 10 to 30quadrature for 1 to 24 hours to remove a settled precipitate; adding an excipient such as microcrystalline cellulose, dextrin, etc. to a concentrate remaining after removal of the precipitate; and drying and pulverizing the resulting product. The present invention also provides a composition including the extract powder prepared as described above.
The ethanol with a purity of 80 to 100% (v/v) used herein may maximize an extraction efficiency of key compounds in the Angelica gigas Nakai herb. The pH value of the extraction solvent is preferably in the range of pH 7 to 9. If the pH value exceeds pH 9, alkalinity of the solvent is too high, causing significant hydrolysis of extract ingredients. In fact, it was observed that the extract solution became yellow when the pH value of the solution was above pH 9. Therefore, the pH value of the extraction solvent is adjusted using a sodium hydroxide solution to maintain pH 7 to 9 at 15 minute intervals during the extraction. The reason for pH control in such a way described above is that a content of decursinol may be increased and significant hydrolysis may be prevented by adjusting the pH value at 15 minute intervals, since an acidic substance is excessively extracted while continuing the extraction to decrease the pH value of the extract solution. In the present invention, this pH control may be particularly achieved for main ingredients of Angelica gigas Nakai extract which are decursinol derivatives. The 15 minute interval in pH control is preferably defined on the ground that the pH value of the extraction solvent containing excess ethanol is difficult to measure using any typical pH electrode. The pH control may be conducted during the extraction or until 1 hour prior to completion of extraction. More preferably, the pH control is conducted while extracting for at least 2 hours, and then, the extraction may be continued for another 1 hour without the pH control. The obtained extract solution may have a pH value in the range of 4.6 to 5.4. In order to adjust the pH value of the extract solution, an alkaline solution is added. Such alkaline solution may comprise sodium hydroxide or calcium hydroxide in water and, in addition, magnesium hydroxide or barium hydroxide may be used as an alkali agent. Among these, sodium hydroxide is the most efficient and readily available compound suitable to apply Angelica gigas extract in manufacturing food products. The extraction time may range from 3 to 8 hours in consideration of common extraction yield and/or generation of decursinol. If the extraction time is less than 3 hours, the extraction yield and the generation of decursinol are drastically reduced. On the other hand, when the extraction time exceeds 8 hours, there is no further increase in extraction yield and decursin is excessively decomposed to generate too much decursinol and/or induce further decomposition thereof, causing a decrease in both contents of decursin and decursinol.
A ratio in contents of (decursin+decursinol angelate) to decursinol contained in Angelica gigas extract prepared by the present invention may range from 50 to 200:1, compared to 400 to 1,000:1 for an extract obtained by any conventional process (extracting without pH control). Consequently, the content of decursinol may be increased to the maximum 50 fold over the prior art. Angelica gigas Nakai extract with increased content of decursinol is subjected to measurement of solid content therein, followed by addition of microcrystalline cellulose or dextrin as an excipient and pulverization thereof. The resultant extract powder substantially has at least 0.1 wt. % of decursinol as well as a compositional ratio of 50 to 200:1 in terms of (decursin+decursinol angelate) to decursin.
Concentrating the obtained extract may result in a concentrate with 2.5 to 3.5 times the weight of Angelica gigas Nakai. The concentrate undergoes cooling and settling at 10 to 30quadrature for 1 to 24 hours to remove a settled precipitate, so as to remove polysaccharides excessively extracted from Angelica gigas Nakai. As a result, excess polysaccharide and/or oligosaccharide extracts may be removed from the resulting product.
In order to prepare Angelica gigas Nakai extract with a high content of decursinol, an extraction process using hydrolysis of decursin under alkaline conditions may be proposed, wherein an alkali agent is introduced to a resultant solution after completing the extraction instead of a treated solution during the extraction, and then, the mixture is subjected to hydrolysis while elevating a temperature thereof.
A diluted alcohol was used as an extraction solvent in the present invention. However, water may also be used in preparation of Angelica gigas extract. Even when water is used as the extraction solvent, an alkali agent may be added and the resultant extract may be substantially identical to that obtained using alcohol as the extraction solvent. However, since volatile compounds contained in Angelica gigas Nakai herb are generally water insoluble, water extraction may cause a decrease in extraction yield and exhibit inferior flavor quality in inherent Angelica gigas than an alcohol extraction.
Functional efficacies of Angelica gigas Nakai extract according to the present invention, in particular, antioxidant effects and analgesic effects were determined as follows.
The antioxidant effects were determined using 1,1-diphenyl-2-picryl hydrazyl (DPPH). Analysis of the extract using DPPH is performed by the following method.
DPPH having radicals is mostly used for determining radical scavenging performance. DPPH was purchased from Sigma. The following procedures were carried out:
1) 2 ml of 0.1 mM DHHP (Sigma, D-9132) in ethanol was added to 2 ml of a sample dissolved in ethanol;
2) After mixing the solution for 10 minutes, the mixture was left in a dark place;
3) Absorbance of the resultant product at 520 nm was measured;
4) Calculation of antioxidant efficacy (electron donating ability),
EDA (electron donating ability)=(Cabs-Sabs)/Cabs*100
wherein Cabs: absorbance of a negative control (which contains ethanol only instead of a sample), Sabs: absorbance of a sample, and Positive control: Vitamin E (Fluka, 95240) as a sample.
The analgesic effects were determined by the following procedures.
As described in a prior document, Life Science 73 (2003), 471-485, the analgesic effects were determined by writhing comparison using acetic acid. In a writhing test, after injecting 1% acetic acid to a mouse, the mouse was placed in a cage with dimensions of 20 cm (diameter)×20 cm (height). After 30 minutes, writhing numbers were measured and compared.
Hereinafter, preferred examples of the present invention will be described in detail. However, these examples are given for the purpose of illustration and are not intended to limit the invention.
Preparation of an Extract from Angelica gigas Nakai Using Alcohol
10 kg of air dried Angelica gigas Nakai herb was purchased at a marketplace named "Kyung-dong market," in Korea. 1 kg of the herb was used. 5 L of 93% (v/v) ethanol was prepared by diluting alcohol, followed by adding 15 wt. % of sodium hydroxide solution thereto so as to prepare an ethanol solvent with pH 8.5. The herb was added to the prepared solvent and subjected to extraction at 65quadrature. Sampling the extract at 15-minute intervals, the sample underwent pH measurement using a pH measuring tape, followed by adjusting the pH value to 8.0. 4 hours after the extraction, the pH adjustment was terminated and additional extraction was conducted for 1 hour. After the extraction, a filtrate was separated and concentrated to a volume of 3 L. Settling the concentrate at 10quadrature for 10 hours, a settled precipitate was filtered and removed to yield a solution which in turn was again subjected to concentration. Adding 100 g of microcrystalline cellulose as an excipient to the resultant concentrate, the mixture was completely dried to yield 198 g of light yellow powder as a final product, which is Angelica gigas Nakai extract powder.
Contents of decursinol, decursin and decursinol angelate in the extract were analyzed by gas chromatography and the results are shown in FIG. 4. From FIG. 4, it can be seen that a peak at 17.533 minutes exhibited decursinol. Similarly, a peak at 24.054 minutes and another peak at 25.718 minutes showed decursinol angelate and decursin, respectively. From quantification results thereof, it was determined that a relative content ratio of (decursin+decursinol angelate) to decursinol was 110:1 and a content of decursinol was 0.15 wt.%.
Preparation of an Extract from Angelica gigas Nakai Using Water
Adding 100 g of water to Angelica gigas Nakai herb prepared as described in Example 1, the mixture was subjected to extraction for 3 hours. The obtained concentrate was dried and 21 g of the dried product was added to 21 g of microcrystalline cellulose. Adding a small amount of water re-dissolved the mixture. The re-dissolved solution was dried to yield 41 g of powder as a final product, which is Angelica gigas Nakai extract powder.
The extract was analyzed by gas chromatography and the results show that a relative content ratio of (decursin+decursinol angelate) to decursinol was 825:1 and a content of decursinol was 0.0083 wt.%.
From the above results in Examples 1 and 2, it is found that Angelica gigas Nakai extract obtained using water in Example 2 has decursinol content 18 times that of the extract obtained using alcohol in Example 1.
Comparison of Antioxidant Effects
Antioxidant effects were compared between both extract powders obtained in Examples 1 and 2. Using DPPH analyzed and quantified EDA to determine the antioxidant effects of the extract powder. IC50 means a concentration required for scavenging 50% of radicals and it is determined that the antioxidant effects are enhanced as IC50 value is lowered. It was demonstrated that the extract prepared according to the present invention has antioxidant effects 10 times better than those of an extract obtained using water as a control.
TABLE-US-00001 TABLE 1 Comparison of antioxidant effects of Angelica gigas Nakai extracts IC50 (mg/L) Vitamin E 15 Angelica gigas Nakai 230 extract obtained using water Extract of the present 22 invention
Comparison of Analgesic Effects
Analgesic effects were compared between the extract obtained according to the present invention in Example 1 and pure decursinol. The analgesic effects were determined and results thereof are shown in FIG. 5. Referring to FIG. 5, `Control` means a test group without treatment, `Decursinol 20` means a 20 mg/Kg decursinol treatment group, and `DNS` is a group administered with an extract of the present invention containing 20 m/Kg in terms of decursinol. Assuming decursinol contents are equal, it can be identified that additional administration of decursin and decursinol angelate improved the analgesic effects. From results of the test shown in FIG. 5, it was demonstrated that the inventive extract exhibited analgesic effects more than 2.7 times better than those of the others.
As is apparent from the above detailed description, the extract of the present invention exhibits excellent analgesic and antioxidant effects, thereby being effectively utilized in a variety of applications including foods, cosmetics, and natural medicines for human health.
Although the preferred embodiment of the present invention has been disclosed for illustrative purposes, those skilled in the art will appreciate that various modifications, additions and substitutions are possible, without departing from the scope and spirit of the invention as disclosed in the appended claims.
Patent applications by Sung Pil Kwon, Seoul KR
Patent applications by AZI Co., Ltd.
Patent applications in class PLANT MATERIAL OR PLANT EXTRACT OF UNDETERMINED CONSTITUTION AS ACTIVE INGREDIENT (E.G., HERBAL REMEDY, HERBAL EXTRACT, POWDER, OIL, ETC.)
Patent applications in all subclasses PLANT MATERIAL OR PLANT EXTRACT OF UNDETERMINED CONSTITUTION AS ACTIVE INGREDIENT (E.G., HERBAL REMEDY, HERBAL EXTRACT, POWDER, OIL, ETC.)