KR20160115039A - A method for removing toxic substance from an extract of Asiasarum root or rhizome and the preparation comprising the purified extract as an active ingredient - Google Patents

Abstract

The present invention relates to a producing method of an Asiasarum root and rhizome extract removing toxic substances and herbal preparation comprising the extract as an active ingredient. The producing method obtaining the Asiasarum root and rhizome extract removing toxic substances comprises: a first step of obtaining a first Asiasarum root and rhizome extract; a second step of obtaining an alkalized suspension; a third step of a second extract; and fourth step of depressurizing and concentrating the second extraction.

Description

[0001] The present invention relates to a method for producing a sesquicentin extract which is free from toxic components and a preparation containing the extract as an active ingredient,

The present invention relates to a method for preparing a sesquiterp extract and a preparation containing the extract as an active ingredient.

[Literature 1] Encyclopedia of Korean Culture, encykorea.aks.ac.kr, Central Research Institute of Korean Studies

[Literature 2] Oriental medicine, pp. 347-348, 1986, Korean Pharmacists Association

[Literature 3] Korean Pharmacopoeia 10th Revision, 2012, Food and Drug Administration

[Document 4] Herbal medicine, Dongmyungsa, pp 151-153, 2006,

[Literature 5] Zhao et al., 2008, v. 15, pp. 741-748, Phytomedicine

The present invention relates to a method for preparing a sesquiterp extract and a herbal preparation containing the extract as an active ingredient.

Socheongryongtang is one of the most commonly used herbal remedies for the treatment of coughing, whooping cough, bronchitis, bronchial asthma and allergic rhinitis. There are eight kinds of Sukyongrytang: mahwang (麻黄), 藥药 (藥 药), Omija 五 (半 夏), Ssin, 건강 (枝) It is made up of herbal medicine. In general, about 6g of saccharin, health, stamina and licorice is used in about 4g of saccharin, peony, omija, and bananas, but it depends on the symptoms and condition of the patient. encykorea.aks.ac.kr, Central Research Institute for Korean Studies, Korean Pharmacology, pp.347-348, 1986, Korean Pharmacists Association)

The double seshin is the dried root of the bamboo shoots, and the name seshin is named because it has a very thin and very spicy taste. In the 10th amendment of the Korean Pharmacopoeia, Asiasarum root and rhizome were deposited in the tribal Asiasarum There are heterotropoides . mandshuricum or Seoul Pedestrian Pool Asiasarum I have sieboldii . It is stipulated that roots and rootstocks of seoulense i (rats and Aristolochiaceae) should contain more than 0.6 mL of essential oil per 30 g. In China, Asian History Room hetero teuropoyi Death (Asiasarum heterotropoides var. Mandshuricum), Asian History Room Bar Seoul City boldi Lawrence (Asiasarum I have sieboldii . seoulense), Asian history room When boldi (Asiasarum sieboldii ). (Korean Pharmacopoeia 10th Revision, 2012, Korea Food and Drug Administration, Korea Food and Drug Administration), pp 151-153, 2006,

Ssein shows fever, tremor, sedation, local anesthesia, antiinflammatory, immunosuppressive action, and it has kinking, vasodilation, smooth muscle relaxation, lipid metabolism enhancement, and blood sugar elevation. In addition, cecin has antibacterial activity, antiallergic action, antitumor action, and cholesterol lowering action. Sethin contains 2-3% essential oil, and its components are methyluegenol, asarylketone, cineol, safrole, limonene, eucarvone (azulene), elemicin, kakuol, 1.8-cineol and γ-asarone. In addition to essential oils, components such as N-isobutyl-2,4,8,10-dodecatetraenamide, pellitorin, asarinin, xanthoxylol, sesamin, aristolochic acid (AA), aristolactam 3 and higenamine are known. (Genetic Medicine, Tongmyungsa, pp. 151-153, 2006, Compilation Committee of Korean Medical Science Textbook)

Among the components of acinic acid, aristolochic acid (hereinafter referred to as AA) of the following formula 1 is known as a renal toxic substance. Especially in Belgium and Hong Kong, nephrotoxicity due to AA has been shown to limit the use of herbal medicines containing AA. The AA in Seesin is especially abundant in the top part of Seesin. Seeds contain 0.93 ~ 2.16 ug / g in the roots of Korean tall grass and 6.14 ~ 11.91 ug / g in the ground and 0.19 ug / g AA (Zhao et al., 2008, v. 15, pp. 741-748, Phytomedicine).

Structure of Aristolochic acid (AA)

However, none of the above documents discloses or teaches a preparation method for completely removing the arrythroloquinic acid (AA) kidney toxic component contained in the sesquicent extract.

Accordingly, the inventors of the present invention have studied a manufacturing method for producing a safe herbal preparation free from nephrotoxicity by completely removing the arrytholic acid (AA) kidney toxic component contained in the sesquicent extract. As a result, (AA) kidney toxic components were almost completely eliminated by HPLC. The extracts of sesquicin prepared by this manufacturing process were confirmed by HPLC. The present invention has been completed upon confirming that the present invention can be effectively used for the preparation of a safe herbal medicine agent free from side effects due to sezin.

It is an object of the present invention to provide a method for producing a sesquicin extract which removes aristolechinic acid (AA) which is a kidney toxicity inducing component of acorn extract.

In addition, the present invention provides a herbal medicine or a pharmaceutical composition containing the sesquicin extract prepared by the above production method.

In order to achieve the above object, the present invention provides an organic solvent selected from water, a lower alcohol having 1 to 4 carbon atoms such as methanol, ethanol, propanol, and butanol, or a mixed solvent thereof, Is added in a mixed solvent of water and ethanol, more preferably 30 to 80% ethanol, at a temperature of 10 to 100 캜, preferably 20 to 90 캜 for 30 minutes to 72 hours, preferably 6 to 48 hours A first step of obtaining a sesquist first extract by performing an extraction method, preferably a reflux extraction method, selected from a combination of a cold extraction method, an ultrasonic extraction method, a reflux cooling extraction method, a hot water extraction method, a room temperature extraction method, a reflux extraction method or a combination of at least one of these extraction methods; A second step of adding an alkaline solution such as ammonia water to the first extract of the first step to obtain an alkalized suspension; The alkalized suspension of the second step is subjected to a resin treatment method using ion exchange resin column chromatography or a solvent extraction method using a non-polar solvent such as hexane, chloroform, ethyl acetate, or diethyl ether to obtain a third extract step; A fourth step of concentrating the second extraction liquid in the third step under reduced pressure; And a method for producing a sesquicin extract in which a toxic component including a process is removed.

In the third step of the manufacturing process, the alkali solution is characterized by containing an alkalinizing solution such as ammonia water, NaOH, KOH, and Ca (OH) 2.

In the third step of the manufacturing process, the alkalized suspension is characterized by a pH ranging from 7 to 12, preferably from 8 to 10.

In the fourth step of the production process, the ion exchange resin column chromatography is carried out by a chromatography method using a cation exchange resin, an anion exchange resin, or a synthetic adsorbent, preferably a chromatography method using an anion exchange resin, For example, ToyoPearl QEA (available from Tosoh), Q Sepharose FF (GE < (R) >), (Available from Healthcare) or Fractogel EMD, Fractogel TMAE or Fractogel HICAP (available from Merck KGaA, Darmstadt Germany), and even more preferably SA21A.

In the fourth step of the above process, the non-polar solvent is preferably ethyl acetate or diethyl ether, more preferably diethyl ether.

It was confirmed by HPLC that arsistolechinic acid (AA), which is a kidney toxic component, was almost completely removed from the sesquicin extract prepared through the manufacturing process of the present invention. The extract of Sekisin is applied to various herbal preparations including Sekin, and thus it can be usefully used for the production of safe herbal preparations without side effects due to Sekin.

Accordingly, the present invention provides a pharmaceutical preparation or a pharmaceutical composition containing a cedar leaf extract, wherein the toxic component produced by the method is removed.

As used herein, " pharmaceutical preparations containing a sesquiter extract with the removal of a toxic component " include, but are not limited to, Socheongryangtang, MaengpuBujiaShinTang, GumiGanghongTang, .

The pharmaceutical composition containing the extract according to the present invention may be formulated in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols or the like, oral preparations such as external preparations, suppositories, Can be used. Examples of carriers, excipients and diluents that can be included in the composition containing the extract include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate , Cellulose, methylcellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, a diluent or excipient such as a filler, a weighting agent, a binder, a wetting agent, a releasing agent, a surfactant and the like is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose ), Lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories.

Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. As a suppository base, witepsol, macrogol, tween 61, cacao paper, laurin, glycerol gelatin and the like can be used.

The preferred dosage of the pharmaceutical composition of the present invention varies depending on the condition and the weight of the patient, the degree of disease, the drug form, the administration route and the period, but can be appropriately selected by those skilled in the art. However, for the desired effect, the pharmaceutical composition containing the extract of the present invention is preferably administered at 0.0001 to 100 mg / kg, preferably 0.001 to 100 mg / kg per day. The administration may be carried out once a day or divided into several times. The dose is not intended to limit the scope of the invention in any way.

The agent of the present invention can be administered to mammals such as rats, mice, livestock, humans, and the like in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injection.

The sesquicin extract prepared through the manufacturing process according to the present invention can be effectively used for the preparation of safe sesquicin containing herbal preparations which are free from side effects by virtually completely eliminating the kidney toxic ingredient aristololeucine (AA).

1 shows the results of HPLC chromatograms of the extracts of Reference Example 1 (A), Example 1 (B) and Example 2 (C).

Hereinafter, preferred embodiments of the present invention will be described in order to facilitate understanding of the present invention. However, the following examples are provided to further understand the present invention, and the present invention is not limited by the examples.

Reference Example 1: Preparation of unprocessed cedar leaf extract

200 mL of 70% ethanol was added to 10 g of Seisin (Kyungdong market) and refluxed for 3 hours. The extract was filtered and concentrated under reduced pressure using a vacuum condenser (EYELA, N-2100, JAPAN) to obtain 3.3 g of 70%

Example 1: Example of purification using resin treatment method (1)

The filtrate was filtered through a rotary evaporator (EYELA, N-2100, JAPAN) and concentrated under reduced pressure to remove the organic solvent. The filtrate was diluted to about 200 mL with water, Was added to adjust the pH to about 9. 20 g of a strongly basic anion exchange resin activated in cationic form was added thereto, and the mixture was slowly stirred for 1 hour. Then, the resin was filtered out and the solution was concentrated under reduced pressure at 40 ° C or lower to obtain 2.7 g of purified cetin extract.

Example 2: Purification Example using Solvent Extraction Method (1)

200 g of 70% ethanol was added to 10 g of sesquin, and the mixture was refluxed and vacuum-filtered. The extract was concentrated under reduced pressure using a vacuum condenser (EYELA, N-2100, JAPAN) to remove the organic solvent, Ammonia water was added to adjust the pH to about 9. This was transferred to a separatory funnel and repeatedly extracted with 200 mL of diethyl ether three times. The diethyl ether layer was combined, and the solvent was removed from the combined diethyl ether layer at 40 ° C or lower to obtain 1.1 g of purified sesqui extract.

Experimental Example 1: HPLC component analysis

High performance liquid chromatography was carried out under the same conditions as in Table 1 to confirm the content of aristolochic acid (AA) contained in the reference examples and examples obtained above. The results are shown in FIG.

High performance liquid chromatography conditions column Agilent Eclipse Plus C18 (250 * 4.6 mm, 5 [mu] m) Mobile phase Tilt elution 30% B (0 min) → 50% B (10 min) → 80% B (30 min) → 100% B (35 min). A: 0.2% formic acid aqueous solution; B: 0.2% formic acid methanol solution Detector wavelength 254 nm Mobile phase flow velocity Flow rate 1 mL / min detection UV 254 nm

As a result of the above experiment, as shown in Fig. 1, AA peak appeared at 29.9 minutes in the sesqui extract without any purification treatment in Reference Example 1, and NIDA (N-isobutyl-2,4,8 , 10-dodecatetraenamide) showed a strong peak at 35 min (chromatogram A). However, in the extracts of Examples 1 and 2 according to the present invention, as shown in Chromatograms B and C, it was confirmed that the active ingredient, NIDA, was not removed even though AA which is a toxic substance was effectively removed.

Such sesquin can be safely used without fear of the renal toxicity of AA. Especially, when using the sesquin according to the present invention, it can be safely applied without toxicity.

Formulation example  1. Herbal medicine containing sesquist

The preparation examples of the composition containing the purified cetin extract described above (Table 1) will be described below, but the present invention is not intended to be limited thereto but is specifically described.

Prescription Furtherance Indications Sochyong-tang 6 g of each, 黃 마 마 黃 白 白 白 白 白 白 건강 건강 건강 건강 건강 건강 건강 건강 건강 건강 각 ((((((((((((((((,炒 甘草) 4g each Cough and pertussis due to colds, bronchitis Ma Zhang Fujushushin Tang 8g each of mahwang and seshin, 4g of forbuja Dyspnea, Gumi Fortress 6 g each of ganghwam and windbreak, 4 g each of ganghwa, white bamboo, 黄 ·, raw yellow bamboo, 甘草, Each 2g Tetanus, flu Tenshin Mint, 160 grams, molds, 80 grams, 40 grams of ganghwamun, white bamboo, 甘 草, 20 grams of spicy, 30 grams of windshield Migraine (migraine headache), nasal congestion, ominous wind, dizziness A horny bath 2.8g each, 熟地黄, 芎궁 ·, ginseng, 백 령 白 백 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 슬 2 ㎝ of licorice, 甘 草 1.2g of each of the broiler meat, 膝,), 秦 세, 신 세 세, Arthritis, neuralgia Inspiration Omi Gangshin Bokryeong 8, Licorice, Omiza, Health, Seishin 6 Body cold, runny nose

Claims (9)

An organic solvent selected from the group consisting of water, methanol, ethanol, propanol, butanol or a mixed solvent thereof is added to the dried cedar and the mixture is heated at a temperature of 10 to 100 캜 for 30 minutes to 72 hours by a cold extraction method, A first step of obtaining a sesquist primary extract by performing an extraction method selected from an extraction method, a room temperature extraction method, a reflux extraction method, or a combination of at least one of these extraction methods; A second step of adding an alkali solution to the first extract of the first step to obtain an alkalized suspension; A third step of obtaining a secondary extract by subjecting the alkalized suspension of the second step to a resin treatment method using ion exchange resin column chromatography or a solvent extraction method using a non-polar solvent; A fourth step of concentrating the second extraction liquid in the third step under reduced pressure; A process for producing a sesquicin extract from which a toxic component containing a process is removed. The method according to claim 1,
Wherein in the first step of the manufacturing process, the organic solvent is a mixed solvent of water and ethanol. The method according to claim 1,
Wherein in the second step of the manufacturing process, the alkali solution is ammonia water, NaOH, KOH, or a Ca (OH) 2 alkali solution. The method according to claim 1,
Wherein in the second step of the manufacturing process, the alkali solution is in the range of pH 6-12. The method according to claim 1,
Wherein the ion exchange resin column chromatography method is a chromatography method using a cation exchange resin, an anion exchange resin, or a synthetic adsorbent in the third step of the manufacturing process. 6. The method of claim 5,
Wherein the ion exchange resin column chromatography method is a chromatography method using an anion exchange resin. The method according to claim 1,
Wherein in the third step of the manufacturing process, the nonpolar solvent is hexane, chloroform, ethyl acetate, or diethyl ether. A pharmaceutical preparation comprising a sesamin extract obtained by removing the toxic components produced by the manufacturing method of claim 1. [Claim 9] The pharmaceutical preparation according to claim 8, wherein the toxic component is removed from the pharmaceutical preparation, wherein the pharmaceutical preparation comprises sesquiterpin extract is selected from the group consisting of Sochungryongtang, Mawangbuja Sechinhang, Gumi Ganghongtang,

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