KR100328478B1 - Sin extract and anti-asthmatic lignan compounds isolated from shin - Google Patents

Abstract

The present invention relates to a lignan compound separated from the god Shin ( Magnoliae flos ) extract and god Shin having an asthma efficacy, the god of the present invention obtained by extracting the god Shin with an organic solvent and separated from it Lignan compounds may be usefully used as anti-asthma agonists, including asthma prophylactic, therapeutic and therapeutic aids.

[Formula 1]

[Formula 2]

[Formula 3]

Description

Sinyi Extract and Anti-asthma Efficacy and Lignan Compounds Isolated from Sinui

The present invention relates to a god sensitizing ( Manoliae flos ) extract and a lignan compound isolated from the god yishin , and more specifically to the god yi extract obtained by extracting the god yi with an organic solvent and the following formulas 1 to 4 Lignan compounds, and their use as anti-asthma agonists.

Formula 1

Formula 2

Formula 3

Formula 4

There God (Magnoliae flos) has a spicy taste to dry the Chinese god (Magnolia fargesii Cheng, M. biondii Pamp .) Fiji buds do the (hwaroe) belonging to the Magnolia (Magnoliaceae) (Namba, T., Coloured Illustration of Wakan-Yaku , Hoikusha Publishing, Osaka, Vol. II , 127-129, 1980). It has been used to treat headaches, malignant non-mucosal ulcers, sinusitis and toothache, and has been used for some fragrances (Tang, W and G. Eisenbrand, Chinese Drugs of Plant Origin: Chemistry, Pharmacology and Use in Traditional and Morden Medicine , Springer-Verlag, Berlin, 639-646, 1992).

To date, various kinds of essential oils, lignans, sesquiterpenes, etc. have been separated from the gods, and they have been reported to exhibit calcium (Ca ²⁺ ) antagonistic activity, platelet activating factor (PAF) antagonistic activity and antihistamine activity (Jung, KY, et al. , Arch. Pharm. Res. , 20 , 363-367, 1997; Chen, CC, et al. , Planta Med., 54 , 438-440, 1988; Pan, JX, et al. , Phytochemistry , 26 , 1377-1379, 1987; Nagashima, S., et al. , J. Takeda Res. Lab., 40 , 27-36, 1981).

On the other hand, asthma is secreted by a variety of interleukins sequentially from various immune cells present in vivo by an allergen inhaled into the airways, whereby leukotriene (LTC ₄ ) is released from mast cells. , Prostaglandins (PGD ₂ ), histamines, etc. . This phenomenon, if symptoms are sensitized by the immunogen again after the symptoms recover, the bronchial hyperresponsiveness reoccurs, resulting in a stronger and more persistent bronchial contraction.

For the prevention and treatment of such asthma symptoms, bronchial relaxants such as beta-2 agonists (beta2-agonist), inhibitors of the production of various inflammatory mediators or antagonists have been developed, and various enzymes involved in the metabolism of the inflammatory mediators. Inhibitors have been developed, and immunomodulators, cytokine inhibitors and cell adhesion blockers have been developed from various angles.

In the case of acute bronchial contraction, many asthma patients use short-acting beta2 agonists and periodically inhale corticosteroids for proper and sustained asthma prevention. However, continuous inhalation of steroids causes various side effects. As a remedy, a method of topically using a long-acting beta2 agonist and a corticosteroid in combination has been introduced. Recently, the development of anti-asthma drugs using new bronchial relaxants and inhibitors of the production of inflammatory mediators or metabolic inhibitors focused on various inflammatory reactions involved in asthma.

Accordingly, the present inventors have identified a substance having an effect of inhibiting airway smooth muscle contraction of an animal sensitized by a kidney in order to develop a therapeutic agent or a preventive agent for bronchial contraction reaction caused by asthma from traditional natural products, and from this, four kinds of lignan The compounds were isolated and their structure examined by spectroscopic method, and the present invention was completed by confirming that the compounds, mixtures thereof, and god extracts containing them have anti-asthmatic efficacy.

It is an object of the present invention to provide a god extract having anti-asthmatic efficacy and a lignan compound isolated from the god.

1 is a result of sensitizing guinea pigs with ovalbumin, and then extracting airway sections and re-exposing to ovalbumin (1 µg / ml) to measure the effect of inhibiting airway contraction by god extract.

Figure 2 is the result of measuring the concentration of the airway extract to prevent airway contraction,

--- ● --- Chromoglycate Sodium

--- □ --- God Extract

3 is a result showing the airway contraction preventive effect of the lignan compound isolated from the gods,

4 is a result of recognizing the airway contraction preventive effect of the lignan compound isolated from the kidney in vivo ( in vivo ) test.

--- ○ --- ketotifen

--- × --- Mixture of Lignan Compound

--- ● --- Pinoresinol Dimethyl Ether (Formula 1)

--- □ --- Magnoline (Formula 2)

In order to achieve the above object, the present invention provides a god extract having anti-asthma efficacy, a lignan compound represented by the formula (1), (2), (3) or (4) obtained from the god and an asthma agonist using the compound as an active ingredient do.

Hereinafter, the present invention will be described in detail.

First, the present invention provides a shinyi extract having anti-asthma effect obtained by extracting sinyi with lower alcohol or suspended in distilled water and extracted with an organic solvent in addition to the above process. As the organic solvent, hexane, ether, ethyl acetate, chloroform or alcohol may be used.

The shinyi extract may include one or more compounds selected from compounds of Formula 1, Formula 2, Formula 3, and Formula 4.

Formula 1

Formula 2

Formula 3

Formula 4

At this time, the compound of Formula 1 to Formula 4 is prepared by extracting sinyi with lower alcohol, suspended with distilled water, washed with hexane and extracted again with ethyl acetate to repeat the adsorption chromatography. Specifically, in the present invention, the extract is extracted with lower alcohol, filtered, concentrated under reduced pressure, suspended with distilled water, washed with hexane, extracted with ethyl acetate, and then repeatedly subjected to adsorption chromatography to formula (1), (2) and (3). And the compound of formula (4).

Preferably, the lower alcohol is methanol, and the adsorption chromatography is preferably silica gel column chromatography, reversed phase silica gel column chromatography, and preparative high performance liquid column chromatography. In this process, elution solvents are hexane-acetone, methanol- Preference is given to using water, a methanol-acetonitrile-water mixed solvent, or the like.

In addition, the present invention provides a pharmaceutical composition for an asthma agonist comprising at least one compound selected from Formula 1, Formula 2, Formula 3 and Formula 4 and derivatives thereof as an active ingredient. The anti-asthma agonist may be used as a preventive, therapeutic or therapeutic aid for asthma.

The pharmaceutical composition for an anti-asthmatic agonist of the present invention may be prepared in unit dose form or formulated in a multi-dose container by formulating with a pharmaceutically acceptable carrier or excipient.

The preparation form can then be in the form of solutions, suspensions or emulsions in oils or aqueous media, or in the form of extracts, powders, granules, tablets or capsules, and can include dispersants, suspensions or stabilizers.

The human dose of the active ingredient according to the present invention is appropriately selected depending on the absorption rate, inactivation rate and excretion rate of the active ingredient in the body, the age, sex and condition of the patient, the severity of the disease to be treated, but generally 15 to adult It is preferred to administer about 1500 mg, but about 100 to 800 mg. Therefore, the unit dosage form of the present invention is prepared to have an amount of 5 to 500 mg of the active substance of the present invention in consideration of the above-mentioned effective amount range. Preferably it is formulated to contain 50-300 mg. Such dosage unit dosage forms may be formulated using specialized dosing regimens as determined by the expert and by the needs of the individual to monitor or observe the administration of the medication as needed. It may be administered at regular time intervals, preferably from one to three times a day.

Toxicity experiments were conducted with respect to the pharmaceutical composition for an asthma agonist of the present invention. The mortality was measured by observing for 30 days after each administration of the pharmaceutical composition of the present invention to mice (10 per group). At this time, the pharmaceutical composition of the present invention was administered in two stages of 300 mg / kg, 100 mg / kg, and 50% lethal dose (LD ₅₀ ) was 300 mg / kg or more.

Hereinafter, the present invention will be described in detail with reference to Examples.

However, the following examples are illustrative of the present invention, and the content of the present invention is not limited by the examples.

<Example 1> Isolation of Lignan Compound from Xinyi

About 3 kg of the dried and ground crushed to a size of about 1 mm was extracted three times with 9 L of methanol at room temperature or 70 ℃, the extract was collected and concentrated under reduced pressure to 225g. After adding 1 L of water and 1 L of hexane and shaking, the shaking process of separating the hexane layer was repeated three times. Then, the hexane layers were collected and concentrated under reduced pressure to make 40 g. After adding 1 liter of ethyl acetate and shaking again, the process of separating the ethyl acetate layer was repeated three times, and the ethyl acetate layers were collected and concentrated under reduced pressure to 100 g.

The ethyl acetate layer was separated by silica gel column chromatography (eluent: hexane-acetone = 99: 1 → 1:99) into 19 fractions, of which part 11 (1.6 g) was again reversed-phase silica gel column chromatography. (Eluent: methanol-water = 3: 2), silica gel column chromatography (eluent: hexane-acetone = 2: 1) and preparative high-performance column chromatography (eluent: methanol-acetonitrile-water = 6 : 7: 12) was repeated to give the compound of formula 1 [pinoresinol dimethyl ether], the compound of formula 2 [magnolin], the compound of formula 3 [lilorresinol-B ether ( liroresinol-B dimethyl ether)] and a compound of formula 4 [(+)-epimagnoline A ((+)-epimagnolin A)].

Example 2 Anti-asthma Efficacy Test of Sinyi Extract

A male guinea pig (male, 8 weeks) was injected subcutaneously with ovalbumin and alumina gel (10 ㎍ + alumina gel 1 mg, dissolved in 0.5 ml saline), followed by sensitization two weeks later. It was. After 1-2 weeks, sensitized guinea pigs were sacrificed by cervical dislocation, followed by airway extraction to make spiral sections in cold PBS-buffered saline (pH 7.4), followed by KRB (Krebs- Ringer bicarbonate was measured in the tissue chamber through which the buffer solution was perfused to the isometric force transducer to measure the degree of contraction. After restoring a resting tension of 0.5 g, the response to the drug was measured. After checking the state of the tissue in response to histamine, the sensitized airway sections were exposed to ovalbumin (1 μg / ml) to measure the effect of inhibiting contraction by the extract of god. The results are shown in FIG. 1 .

As shown in Figure 1 , when exposed to ovalbumin in the state where the god is not extract, the degree of tissue contraction is large, whereas when exposed to the ovalbumin in the state where the extract is added, the degree of tissue contraction is low, The extract was confirmed to have an effect of inhibiting the contraction of airway sections sensitized to ovalbumin.

In addition, in order to determine whether the effect of the god extract on the airway contraction of the extract is concentration-dependent, airway slices were previously exposed to various concentrations of god extract or sodium chromoglycate, a control drug, for 5 minutes, followed by ovalbumin. The degree of contraction of the airway sections was measured by administration. As shown in Figure 2 , according to the concentration of the extract of the god Shin, the prevention of airway contraction increased, unlike the chromoglycate sodium used as a control drug, Sinyi extract showed a 100% airway contraction preventive effect, IC ₅₀ value 0.1mg It was about / ml.

As a result of the above experiment, it was shown that the extract of Sinyi effectively blocks airway contraction caused by sensitizing antigen.

Example 3 Anti-asthma Efficacy Test of Lignan Compound Isolated from Divine Extract

The anti-airway contraction effect of the lignan compound isolated from the extract was determined in the same manner as in Example 2 was determined to have anti-asthmatic efficacy. As shown in FIG. 3 , the lignan compound of the present invention isolated from the god extract was found to have an increased potency for airway contraction compared to the god extract.

〈 Example 4> In vivo of the lignan compound isolated from god extract ( in vivo Anti-asthma efficacy test

The prevention of airway contraction of the compound of the present invention isolated from the god was examined by an in vivo test. First, sensitized guinea pigs were anesthetized and a tubule was inserted into the airway and connected to a mechanical respirator and a pressure transducer using a 3-way cock. Airway resistance was measured by the pressure difference in expiration-inspiration. Once the exhalation-intake pressure differential was stable, ovalbumin was dissolved in water at 1 mg / ml and inhaled with a nebulizer and the increase in pressure difference thereby was recorded in a polygraph. Drug treatment was intraperitoneally injected 10 minutes prior to ovalbumin inhalation, and ketotifen was used as a positive control drug, and the results are shown in FIG. 4 .

As shown in FIG. 4 , the compound of Formula 2 exhibited the efficacy and potency corresponding to ketotifen, and the compound of Formula 1 also exhibited the corresponding effect, indicating that the compound of the present invention has excellent airway contraction prevention effect. Could.

As discussed above, the extract of the present invention and the lignan compound isolated therefrom are very excellent in anti-asthma efficacy and can be usefully used as a preventive, therapeutic and therapeutic aid for asthma.

Claims (5)

After extracting Sinyi with methanol, the extract was concentrated under reduced pressure, shaked with addition of water and hexane, the hexane layer was separated and concentrated under reduced pressure, shaken with addition of ethyl acetate, and the ethyl acetate layer was separated and concentrated under reduced pressure. Dragon God Extract. An asthma treatment agent comprising the lignan compound represented by the formula (1) as an active ingredient. Formula 1 An asthma treatment agent comprising the lignan compound represented by the formula (2) as an active ingredient. Formula 2 An asthma treatment agent comprising the lignan compound represented by the formula (3) as an active ingredient. Formula 3 An asthma treatment agent comprising the lignan compound represented by the formula (4) as an active ingredient. Formula 4