KR100204500B1 - Use of parthenolide as therpeutics for septicemia and preparation method thereof - Google Patents

Abstract

The present invention relates to a novel use of a parthenolide compound as a synthetic inhibitor of tumor necrosis factor-alpha (TNF-α), nitric oxide (NO) and prostaglandin E ₂ (PGE ₂ ), and a method for preparing the same. will be.

More specifically, the present invention utilizes a parthenolide, a sesquiterpene lactone compound, as a inhibitor of the synthesis of tumor necrosis factor-alpha, nitric oxide, and prostaglandin E ₂ , and a parthenolide as a magnolia. grandiflora L.) is a novel method of extraction, the parthenolide of the present invention is an immune disease caused by lipopolysaccharide (LPS), a cell wall component of gram-negative bacteria, such as sepsis (sepsis. endotoxemia) It can be widely used as a therapeutic agent.

Description

A novel use of a parthenolide compound as an agent for the treatment of sepsis and a method of preparing the same.

In the present invention, a parthenolide compound of Formula 1 is used for tumor necrosis factor-α (TNF-α), nitric oxide (NO) and prostaglandin E ₂ (prostaglandin E ₂ , PGE). _It relates to a new use used as a synthetic inhibitor of ₂ ) and a preparation method thereof.

More specifically, the present invention utilizes a parthenolide, a sesquiterpene lactone compound, as a inhibitor of the synthesis of tumor necrosis factor-alpha, nitric oxide, and prostaglandin E ₂ , and a parthenolide as a magnolia. grandiflora L.) is a novel method of extraction, the parthenolide of the present invention is an immune disease caused by lipopolysaccharide (LPS), a cell wall component of gram-negative bacteria, such as sepsis (sepsis. endotoxemia) It can be widely used as a therapeutic agent.

Sepsis (sotopisemia) is an inflammatory response caused by excessive activation of the body's immune system by acting as a toxin of lipopolysaccharide (LPS), a component of the cell wall, when pathogenic Gram-negative bacteria are infected with a living body. Patients admitted to the hospital intensive care unit are the leading cause of death and are a very serious disease with a mortality rate of more than 30%. In addition, sepsis has been a very important problem for the treatment of the pathogens with increasing resistance to various antibiotics, but no suitable treatment has been developed.

Specifically, the sepsis may be caused by the lipopolysaccharide (LPS) excessively advancing the immune system in vivo, thereby leading to tumor necrosis factor-α (TNF-α) and interleukin-1 (TNF-α) in immune cells such as macrophages. It is caused by the production of inflammation-causing substances such as interleukine-1, interleukin-6 prostaglandin, leucotriene and nitric oxide (NO). The toxicity of lipopolysaccharide in vivo It is known that it is due to each of the above-mentioned proinflammatory factors or their synergistic action.

As seen above, tumor necrosis factor-alpha in sepsis is produced from macrophages or fibroblasts activated in vivo, and treated with BCG (Bacillus Calmette-Guerin) or Ropopolysaccharide (LPS). Can be obtained from serum. This tumor necrosis factor-alpha has been reported to have functions such as killing cells in vivo, inducing the production of prostaglandins, and inducing the production of cytokines such as interleukin-1 and interleukin-6. . Therefore, it is known to be directly or indirectly related to various immune diseases and particularly diseases such as arthritis, autoimmune diseases, and sepsis, which are of particular interest. In particular, recent studies have shown that mice with damaged tumor necrosis factor-alpha receptor genes are resistant to lethal toxicity by lipopolysaccharide. In one case, there was a prophylactic effect on the lethal toxicity (Beutler, B., Science, 229, 869 m 1985; Pfeffer, K. et al., Cell, 73, 457, 2993).

In addition, prostaglandins induced by lipopolysaccharide are a kind of hormone derived from arachidonic acid and have various physiological activities, including prostaglandin E ₂ (PGE ₂ ), prostaglandin 2α (PGF 2α), and prostaglandin. I ₂ (PGI ₂ ). In fact, nonsteroidal anti-inflammatory drugs such as aspirin and indomethacin are known to exhibit potent anti-inflammatory effects by inhibiting PGHS (prostaglandin H ₂ synthase, also known as cyclooxygenase), which acts on the synthesis of these prostaglandins. In addition, since aspirin and indomethacin have excellent effects on arthritis and the like, it can be expected that a compound that inhibits the synthesis of prostaglandin may have effects such as anti-inflammatory and arthritis treatment other than the above action.

On the other hand, recent studies have shown that PGHS enzymes that act as rate-limiting enzymes during the synthesis of prostaglandins are expressed by mitogens such as PGHS-1 enzymes and lipopolysaccharides and cytokines such as lipopolysaccharides. It has been reported to be divided into two types of PGHS-2 enzymes that are induced. In addition, the aspirin is known to exhibit a pharmacological effect by acting on the PGHS-2 enzyme, which is expressed by a specific factor, and inhibits its activity.

Nitric oxide is also a substance produced by endothelial cells or macrophages, which also causes the toxicity of lipopolysaccharides. Nitric oxide exhibits a variety of physiological activities, including killing cells or killing bacteria in vivo, especially the maintenance of homeostasis of blood pressure during vascular endothelial relaxation. . Therefore, blood pressure drop, the main symptom of toxicity caused by lipopolysaccharide, is known to be due to excess nitric oxide synthesis induced by lipopolysaccharide (Kilbourn, RG et al., Proc. Natl. Acad. Sci USA, 87, 3629, 1990).

Therefore, the compounds inhibiting the synthesis of tumor necrosis factor-alpha, prostaglandin and nitric oxide are considered to be excellent in the treatment of sepsis, an immune disease (Beutler, B., Science, 229, 869). , 1985; Pfeffer, K. et al., Cell, 73, 457, 1993; Kilbourn, RG et al., PNAS, USA, 87, 3629, 1990; Novogrodsky, A. et. Al., Science, 264, 1319 , 1996).

The therapeutic agents for sepsis studied so far include monoclonal antibodies against lipid A, a cell wall component of Gram-negative bacteria, antibodies against tumor necrosis factor-alpha (TNF-α), and interleukin-1 receptor (IL-1 receptor). ), But the specific clinical efficacy has not yet been demonstrated (Stone, R. Science, 264, 365, 1994). Recently, tyrpostin derivative AG126, an inhibitor acting on protein tyrosine kinase, inhibits lethal toxicity induced by lipopolysaccharide in mice, and the anti-septic effect of the derivative AG126 has a tumor necrosis factor-alpha ( TNF-α) has been reported to be associated with the effect of inhibiting the synthesis (Novogrodsky, A, et al., Science, 264, 1319, 1994).

Sesquiterpene lactone is a compound widely distributed in plants and has been isolated from the extract of Tanacetum parthenium L., especially known as the summer feverfew, which has the effect of treating migraine headaches. It is known to be the main active ingredient to have (Johnson, ES et al., British Med. J., 291, 569, 1985). In addition, sesquiterpene lactone compounds have been reported to have a variety of bioactivity, including showing cell reading in cancer cell lines (Satter, EA et al., J. Nat. Prod., 59, 403, 1996; Picman, A, K., Biochemical systematics and ecology, 14, 255-281, 1986). Recently, it has also been reported that summer chrysanthemum extracts containing patenolide have no effect on edema and arthritis caused by carrageenan (Pattrick, MR et al., Annals Rheumatic Diseases, 48, 547, 1989). However, there have been no reports of the fact that the patenolide compound is effective in treating sepsis caused by lipopolysaccharide and the like (sepsis.endotoxemia).

The present inventors studied extracts of various plants to develop a therapeutic agent for treating sepsis. As a result, nitric oxide, tumor induced by lipopolysaccharide in the leaves and stems of Magnolia grandiflora L. belonging to the Magnolia family Find and isolate active substances that inhibit the synthesis of necrosis factor-alpha and prostaglandin E ₂ . Purifying, identifying its chemical structure and confirming it with a previously reported sesquiterpene lactone compound, patenolide, provides a novel use of magnolia extract and patenolide compound as a treatment for sepsis to complete the present invention It was.

It is an object of the present invention to provide a novel use of a setheter terpene lactone compound having a structure of the formula (1) as a therapeutic agent for sepsis caused by lipopolysaccharide as an immune disease.

Parthenolide compounds of the present invention can be used in the treatment of sepsis in combination with one or more selected from bronchodilators, antihistamines, anti-inflammatory analgesics and antibiotics.

The present invention also relates to the use of the parthenolide compound as a synthesis inhibitor of nitric oxide (NO).

It is also an object of the present invention to provide a use of a parthenolide compound as a tumor necrosis factor-alpha (TNF-α) synthesis inhibitor.

It is another object of the present invention to provide a use of a panenolide compound as a synthesis inhibitor of prostaglandin E ₂ (PGE ₂ ).

It is also an object of the present invention to provide a new method for obtaining a parthenolide compound from Magnolia grandiflora L. Specifically, the leaves and stems of the magnolia are extracted using lower alcohols, ethers, methylene chloride, ethyl acetate and mixed solvents thereof, and separated by silica gel column chromatography and recrystallization. Purify.

It is also an object of the present invention to provide a magnolia extract which can be used as a therapeutic agent for sepsis, including a parthenolide, and a preparation method thereof.

FIG. 1 shows the effect of inhibiting nitric oxide synthesis (NO) by treating the macrotene Raw 264.7 cell line of the present invention to a mouse macrophage Raw 264.7 cell line.

Figure 2 is to measure the effect of inhibiting the tumor necrosis factor (TNF-α) synthesis by treating the macrophage Raw 264.7 cell line of the present invention to the rat macrophage Raw 264.7 cell line.

FIG. 3 shows the effect of inhibiting prostaglandin E ₂ (PGE ₂ ) synthesis by treating the macrotene Raw 264.7 cell line of the present invention with a mouse macrophage Raw 264.7 cell line.

FIG. 4 shows the effect of inhibiting lethal toxicity induced by lipopolysaccharide (LPS) in experimental mice with the patenolide compound of the present invention.

The present invention relates to a nitric oxide NO, a tumor necrosis factor-α, a TNF-α in which a paresnolide, a sesquiterpene lactone compound, is involved in a cellular immune response. By using the activity of inhibiting the synthesis of) and prostaglandin E ₂ (prostaglandin PGE ₂ ), there is provided a new use of the parthenolide compound as a therapeutic agent for sepsis.

The present invention obtains the parthenolide compound from the leaves and stems of Magnolia grand iflora L. belonging to the Magnolia family. Magnolia is an evergreen tree native to North America, and is a common plant grown mainly in the southern part of Korea.

First, the present invention is to separate the extract from the leaves and stems of magnolia using a solvent, the activity of inhibiting the synthesis of nitric oxide, tumor necrosis factor-alpha and prostaglandin in cell lines stimulated with lipopolysaccharide Confirm that it is displayed.

The present invention isolates the active substance that inhibits the synthesis of nitric oxide, tumor necrosis factor-alpha and prostaglandin from the magnolia extract. Purification of the chemical structure confirmed that the active substance is a parthenolide compound represented by Chemical Formula 1, a sesquiterpene lactone compound that has been previously isolated from plants and animals.

Specifically, using the magnolia of the present invention to obtain the magnolia extract and to separate the parthenolide compound from it. For purification, the leaves and stems of magnolias are first extracted with lower alcohols. The alcohol used may be a lower alcohol having 1 to 4 carbon atoms. The extract is partitioned with water, methylene chloride and the like, concentrated, and then subjected to silica gel column chromatography several times.

In this case, a mixed solvent of dichloromethane and cyanide methane or a mixed solvent of nucleic acid and ethyl acetate is used as the developing solvent. Specifically, it is preferable to use a mixed solvent in the range of non-free 50: 1 to 3: 1 of dichloromethane and methane cyanide, and more preferably to use a mixed solvent while changing at a ratio of 10: 1 to 5: 1. Do. In addition, a mixed solvent having a ratio of nucleic acid: ethyl acetate in a ratio of 10: 1 to 2: 1 is also used, and a mixed solvent of 3: 1 is more preferable.

The active material is obtained through the above process, dissolved in an organic solvent such as ether, filtered, and recrystallized using ether and nucleic acid to recover the pure patenolide compound.

The physical and chemical properties of the parthenolide compound obtained in the present invention are examined by carbon and hydrogen nuclear magnetic resonance spectra, and it is confirmed that the present compound is the same as a known substance.

In order to confirm whether the patenolide compound can be used for the treatment of sepsis, the effect of the compound on the synthesis of nitric oxide, tumor necrosis factor-alpha, and prostaglandin E ₂ is described in the macrophage Raw 264.7 cell line of rats. Investigate using

The effect of inhibiting the synthesis of nitric oxide by the patenolide compound is lipopolysaccharide and the addition of patenolide to the cell culture to induce the synthesis of nitric oxide and the amount of nitric dioxide (NO ₂ ) produced therefrom Is determined by the method of Ding et al. (Ding A, H, et al., J. Immunol., 141, 2047, 1988). As a result, the parthenolide compound of the present invention was found to inhibit the nitric oxide synthesis by 73% or more at a concentration of 0.5 µg / ml or more (see FIG. 1).

The effect of inhibiting the synthesis of tumor necrosis factor-alpha by the parthenolide compound is the addition of lipopolysaccharide and parthenide to the cells, and then the cell cultures are recovered and the amount of tumor necrosis factor-alpha now present is L929 cells. It is measured and confirmed by the bioassay method (Aggarwal, BB et al., J. Biol. Chem., 260,2345,1985). Fibroblasts L929 cells are sensitive to tumor necrosis-alpha and useful for measuring the effect, and the survival rate of L929 cells is SRB (Skehan, PR et al., J. Natl. Cancer Inst., 82, 1107, 1990) Measure with As a result, the parthenolide compound of the present invention was found to inhibit the synthesis of tumor necrosis factor-alpha by more than 80% at 1 μg / ml concentration (see FIG. 2).

The parthenolide compound inhibits the synthesis of prostaglandin E ₂ by treating aspirin with each well for 3 hours to inactivate the enzyme PGHS-1 (aka cyclooxygenase-1) present in cells, lipopolysaccharide and Parthenolide is added to the cells, and the culture supernatant is recovered from each well, and quantified and confirmed by prostaglandin E ₂ by radioimmuno assay. As a result, the parthenolide was found to inhibit the synthesis of prostaglandin by 95% or more at a concentration of 5 µg / ml or more (see FIG. 3).

More specifically, to determine whether the patenolide compound of the present invention is effective in the treatment of sepsis, it is investigated whether lethal toxicity induced by lipopolysaccharide in experimental mice is inhibited by the treatment of the patenolide compound. .

As a result, in the control group administered only lipopolysaccharide, 87% of the experimental mice died within 3 days, and in the experimental group to which the patenolide was added 2 hours before the lipopolysaccharide was administered, the pateno was administered. It can be observed that the mortality rate is drastically lowered depending on the amount of ride. Therefore, it can be seen that the parthenolide of the present invention has an excellent effect of significantly lowering the lethal toxicity by lipopolysaccharide and protecting the living body against it (see FIG. 4).

Through the above results, it is confirmed that the magnolia extract and the parthenolide compound of the present invention can be usefully used as inhibitors of the synthesis of nitric oxide, tumor necrosis factor-alpha and prostaglandin E ₂ . Therefore, the pharmaceutical composition comprising the magnolia extract or the parthenolide compound as an active ingredient may be usefully used as a therapeutic agent for sepsis.

Pharmaceutical compositions comprising the patenolide compounds of the present invention may be administered orally or parenterally for the treatment of sepsis. In addition, it is more preferable to administer the parthenolide compound of the present invention in admixture with a pharmaceutically acceptable excipient such as a bronchodilator, an antihistamine, an antibiotic and an anti-inflammatory drug.

Specifically, the parthenolide compound of the present invention is preferably administered 2-3 times a day by 1-10mg per 1kg of body weight, but in acute diseases such as sepsis, 10-100mg per 1kg of body weight is administered 2-3 times a day. desirable.

The present invention will be described in detail by the following examples.

The following examples illustrate the present invention in detail, and the content of the present invention is not limited by the examples.

Example 1

[Isolation and Purification of Parthenolide Compounds from Magnolias]

The leaves and stems of magnolias (1.5Kg) were obtained and dried well. Then, they were crushed and extracted three times with 10 L of methanol at room temperature. The methanol extract thus obtained was concentrated and partitioned between water and methylene chloride, and the methylene chloride layer was concentrated under reduced pressure to obtain 65 g of a concentrated extract. Silica gel column chromatography is performed using the extract (65 g), wherein a mixed solvent of dichloromethane and cyanide methane is used as a developing solvent. Specifically, the ratio of dichloromethane and cyanide methane is 10: 1. A mixed solvent in a ratio of 5: 1 was used [CH ₂ Cl ₂ : CH ₃ CN (10: 1) → CH ₂ Cl ₂ : CH ₃ CN (5: 1)).

Through the above procedure, the fraction having the highest activity inhibiting the synthesis of nitric oxad was obtained and concentrated to obtain an extract residue (5 g). Silica gel chromatography was performed again using the fraction thus obtained, wherein a mixed solvent of nucleic acid and ethyl acetate was used as a developing solvent, and specifically, a mixed solvent having a ratio of 3: 1 of nucleic acid: ethyl acetate was used.

The active ingredient was concentrated and concentrated by evaporation. As a result, 1.5 g of the active substance was obtained, dissolved in 100 ml of ether, filtered, and nucleic acid was added to the filtrate to obtain 770 mg of the active substance. The precipitate thus obtained was recrystallized again using ether and nucleic acid, and as a result, 450 mg of pure patenolide compound was recovered.

Example 2

[Physical and Chemical Properties of Parthenolide Compounds from Magnolias]

Separation from Magnolia Extract obtained in Example 1. The physical and chemical properties of the purified sesquiterpene lactone compound, patenolide, were as follows.

1) Appearance of material: white plate crystal

2) Molecular Weight: 248

3) Molecular formula: C ₁₅ H ₂₀ O ₃

4) Melting Point: 113-115 ℃

5) Hydrogen Nuclear Magnetic Resonance Spectrum

The result is as follows.

δ: 1.31 (3H, s, H-15), 1.72 (3H, s, H-10), 2.78 (1H, d, J = 9 Hz, H-5), 3.86 (1H, t, J = 8 Hz, H -6), 5.21 (1H, m, H-1), 5.62 (1H, d, J = 3 Hz, H-13), 6.33 (1H, d, J = 3 Hz, H-13)

6) Carbon nuclear magnetic resonance spectrum

The result is as follows.

δ: 125.2 (CH, C-1), 24.1 (CH ₂ , C-2), 36.3 (CH ₂ , C-3), 61.5 (C, C-4), 66.3 (CH, C-5), 82.4 (CH, C-6), 47.6 (CH, C-7), 30.6 (CH ₂ , C-8), 41.4 (CH ₂ , C-9), 134.6 (C, C-10), 139.2 (C, C-11), 169.2 (C = 0, C-12), 121.1 (= CH ₂ , C-13), 16.9 (CH3, C-14), 17.2 (CH ₃ , C-15)

7) Chemical formula

Example 3

[Inhibition of Synthesis of Nitric Oxide by Parthenolide]

In order to investigate the degree of inhibition of nitric oxide synthesis by the patenolide compound, a mouse macrophage Raw 264.7 cell line was used.

The raw 264.7 cell line was suspended using Dulbecco Modified Eagle's Medium (DMEM) medium, and then 1 × 10 cells were added to each well in a 96-well plate and cultured at 37 ° C. and 5% CO ₂ to attach the cells to each well. After 3 hours, each well was added to the final concentration of lipopolysaccharide to 1 µg / ml, and the parthenolide was added to 25 µg, 5 µg, 0.5 µg, 0.05 µg, 0.005 µg per ml of medium. . Rifu polysaccharide and parthenolide were added to the medium, followed by incubation for 20 hours, and the amount of nitrite (NO ₂ ) generated from the nitric oxide induced in the culture was measured by ding or the like.

Specifically, 100 μl of the cell culture solution and the same volume of grease reagent [1% sulfanilamide dissolved in 5% phosphoric acid and 0.1% naphtylethylene diamine Reagent mixed with an aqueous solution of hydrochloric acid at a ratio of 1: 1] was added to a 96-well plate and left for 10 minutes, and then absorbance was measured at 550 nm. The amount of nitric dioxide in each sample was quantified using a standard curve of sodium nitrite (NaNO ₂ ).

As a result, the parthenolide compound of the present invention was found to inhibit the nitric oxide synthesis by 73% or more at a concentration of 0.5 µg / ml or more (see FIG. 1).

Example 4

[Suppression of Synthesis of Tumor Necrosis Factor-alpha by Parthenolide]

In the present invention, to examine the degree of inhibition of tumor necrosis factor-alpha synthesis by the parthenolide compound, a mouse macrophage Raw 264.7 cell line was used as in Example 3.

First, Raw264.7 cells were suspended in DMEM medium, added to a concentration of 0.5 x 10 cells / ml in a 24-well plate, and cultured at 37 ° C and 5% CO ₂ to attach the cells to each well. After 3 hours, each well was added to the final concentration of lipopolysaccharide to 1 μg / ml, and the parthenide was added to 10 μg, 5 μg, 1 μg, 0.5 μg, 0.1 μg, 0.01 μg per 1 ml of medium. It was added as possible. After culturing for 20 hours, the cell culture was recovered, centrifuged, and dialyzed, and filtered using a 0.2 μm membrane filter. The amount of each tumor necrosis factor-alpha in the filtrate was measured by a bioassay method using L929 cells.

Specifically, L929 cells, which are fibroblasts sensitive to tumor necrosis factor-alpha, are suspended in RPMI medium containing 50% fetal bovine serum (FBS) to give 2 × 10 ⁴ cells per well in 96-well plates. Incubated at 37 ° C., 5% CO ₂ for 24 hours. Subsequently, remove the medium, add Actinomycin-D (manufactured by Sigma) and the above-described Raw 264.7 cell culture solution to 50 µl per well, and add fetal bovine serum (FBS) to a final concentration of 5%, followed by 37 ° C and 5% CO _2. Incubated for 24 hours at. The survival rate of L929 cells was measured by SRB method.

As a result, the parthenolide compound of the present invention was found to inhibit tumor necrosis factor-alpha synthesis by more than 80% at 1 μg / ml concentration (see FIG. 2).

Example 5

[Inhibition of Synthesis of Prostaglandin E ₂ by Parthenolide]

In order to investigate the degree of inhibition of prostaglandin synthesis by the patenolide compound as shown in Example 3, the macrophage Raw 264.7 cell line of the mouse was suspended in DMEM medium in a concentration of 0.5 × 10 ⁶ cells / ml in a 24-well plate. Cells were added and cultured at 37 ° C., 5% CO ₂ . After incubation for 20 hours, each well was treated with 500 μm of aspirin for 3 hours to inactivate the enzyme PGHS-1 present in the cells.

After washing these cells several times, the lipopolysaccharide was added to a final concentration of 1 μg / ml and the parthenolide was 10 μg, 3 μg, 1 μg, 0.3 μg, 0.1 μg, 0.01 μg per ml of medium. And incubated in a CO ₂ incubator. After 8 hours, the whole culture supernatant (1 ml) was recovered from each well, and prostaglandin E ₂ was quantified by radioimmuno assay.

Specifically, 200 μl of the cell culture solution was taken and diluted in 300 μl of PBS-gel (phosphate buffer solution containing 0.1% gelatin), followed by 100 μl of radiolabeled prostaglandin [ ³ H] -PGE ₂ solution. 200 μl each of prostaglandin E ₂ -antiserum was added and mixed well, which was left at 4 ° C. for 12 hours. During this process, prostaglandin E ₂ present in the sample binds to prostaglandin E ₂ -antiserum competitively with [ ³ H] -PGE ₂ . After leaving for 12 hours, free [ ³ H] -PGE ₂ and prostaglandin E _{2 which} are not bound are removed by adding activated carbon and standing at 4 ° C. for 15 minutes.

[ ³ H] -PGE ₂ and prostaglandin E ₂ adsorbed on the activated carbon are removed by centrifugation, and liquid scintillation (Liquid) is used to detect radioactivity present in the complex of [ ³ H] -PGE ₂ and the antibody in the supernatant. It was measured using a scintillation counter. In addition, the amount of prostaglandin E ₂ was quantified in the measured value.

As a result, patorelide was shown to inhibit the synthesis of prostaglandin by 95% or more at a concentration of 5 µg / ml or more (see FIG. 3).

Example 6

[Effect of Parthenolide to Inhibit Lethal Toxicity of Lipopolysaccharide]

As shown in the above examples, the parthenolide compound strongly inhibits the synthesis of nitric oxide, tumor necrosis factor-alpha, and prostaglandin E ₂ in the macrophage Raw 264.7 cell line of rats. To determine whether it was effective in the treatment of sepsis, it was examined whether lethal toxicity induced by lipopolysaccharide in experimental mice was inhibited by parthenolide.

Specific pathogen free ICR mice (females, 22-27 g, 6 weeks old) were divided into four experimental groups, a comparison group and a parthenide-administered group. In comparison group, only 200% of 30% PEG 400 was injected intraperitoneally, and in the parthenolide group, the parthenolide was dissolved in 30% PEG 400 at 1 mg / ml, 0.5 mg / ml and 0.25 ml / ml. Parthenolide was administered to the abdominal cavity. Two hours after the administration of the Parthenolide, lipopolysaccharide (Sigma: E. coli 055: B5 LPS) dissolved in physiological saline was administered intraperitoneally at a dose of 3 mg / ml and the mice were treated for 6 days. Lethality was observed.

As a result, in the control group administered only lipopolysaccharide, 87% of the experimental mice died within 3 days, but in the experimental group to which the patenolide was added 2 hours before the lipopolysaccharide was administered, the dose of 1 mg / ml In mortality, mortality was lowered to 27% and mortality was reduced to 47% and 67% at 0.5 and 0.25 mg / mol doses, respectively. Therefore, the parthenolide of the present invention was found to significantly lower the lethal toxicity caused by lipopolysaccharide and protect the living body against it (see FIG. 4).

The Magnolia extract and the Parthenolide compound of the present invention effectively inhibit the synthesis of nitric oxide, tumor necrosis factor-alpha and prostaglandin E ₂ involved in the biological immune response, as shown in the above Examples, pharmaceutical composition comprising the same Can be widely used as an excellent treatment to overcome sepsis, which has been difficult to treat so far.

In addition, the parthenolide compounds of the present invention are easily separated from magnolias. Since it can be purified and already associated with arthritis, inflammation, headache, etc., and can be used as a therapeutic agent for various diseases in addition to the treatment for sepsis, the manufacturing method of the present invention can be said to be very economically useful.

Claims (3)

Sepsis therapeutic agent containing the parthenolide compound of formula (1) as an active ingredient. Inhibitor of synthesis of nitrile oxide comprising a parthenolide compound of formula (1). A method of preparing a parthenolide compound represented by Chemical Formula 1 by extracting magnolia (Magnolis grandiflora L.) with a lower alcohol and then performing silica gel column chromatography and recrystallization using an organic solvent.