WO2020050462A1 - Neuron-protective composition comprising cassia obtusifolia shoot-derived naphthopyrone derivative - Google Patents

Neuron-protective composition comprising cassia obtusifolia shoot-derived naphthopyrone derivative Download PDF

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WO2020050462A1
WO2020050462A1 PCT/KR2019/000399 KR2019000399W WO2020050462A1 WO 2020050462 A1 WO2020050462 A1 WO 2020050462A1 KR 2019000399 W KR2019000399 W KR 2019000399W WO 2020050462 A1 WO2020050462 A1 WO 2020050462A1
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formula
naphthopyrone
extract
composition
group
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PCT/KR2019/000399
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French (fr)
Korean (ko)
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권학철
박진수
권재영
황호성
이정환
이재욱
김동회
김형석
장성율
정상훈
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한국과학기술연구원
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Priority to CN201980072747.9A priority Critical patent/CN112996799A/en
Publication of WO2020050462A1 publication Critical patent/WO2020050462A1/en

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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/105—Plant extracts, their artificial duplicates or their derivatives
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C49/00—Ketones; Ketenes; Dimeric ketenes; Ketonic chelates
    • C07C49/587—Unsaturated compounds containing a keto groups being part of a ring
    • C07C49/613—Unsaturated compounds containing a keto groups being part of a ring polycyclic
    • C07C49/617—Unsaturated compounds containing a keto groups being part of a ring polycyclic a keto group being part of a condensed ring system
    • C07C49/643—Unsaturated compounds containing a keto groups being part of a ring polycyclic a keto group being part of a condensed ring system having three rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
    • C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
    • C07D311/78—Ring systems having three or more relevant rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H1/00—Processes for the preparation of sugar derivatives
    • C07H1/06—Separation; Purification
    • C07H1/08—Separation; Purification from natural products
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
    • C07H15/26—Acyclic or carbocyclic radicals, substituted by hetero rings
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0618—Cells of the nervous system
    • C12N5/0621—Eye cells, e.g. cornea, iris pigmented cells
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2300/00—Processes
    • A23V2300/14—Extraction
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2501/00—Active agents used in cell culture processes, e.g. differentation
    • C12N2501/999—Small molecules not provided for elsewhere

Definitions

  • the present specification relates to a composition
  • a composition comprising a naphthopyrone derivative and a shoot sprout extract comprising the same as an active ingredient.
  • the hippocampus is a structure of the medial temporal lobe of the brain and plays an important role in the cognitive function of humans and animals.
  • the hippocampus is known to be vulnerable to physiological and oxidative stress stimuli and to be a central tissue for cognitive impairment caused by stress. Stress can cause hippocampal structure and brain cell production, synaptic plasticity, and behavioral changes related to the hippocampus (Eunjoo Kim, Journal of the Korean Psychological Association: Cognitive and Biology, 2012, 24, 65-88).
  • ischemic nerve damage When oxidative stress in neurons is induced in the hippocampus, pituitary gland, striatum, black matter, the whole cortex, or the hypothalamus, neuronal cell death increases and neurons and growth factors decrease, resulting in amyotrophic lateral sclerosis (ALS), Parkinson's disease, or brain It is known to cause acute or chronic neurological diseases such as ischemic nerve damage (Rahman, T. et al. Adv. Biosci. Biotechnol. 2012, 3, 997-1019).
  • the number of patients with retinal disease has increased by about 8% per year from about 830,000 in 2010 to about 12.5 million in 2015.
  • the retina is the thinner nerve film inside the eye that is likened to the film of a camera.
  • Over 100 million photoreceptor cells (light-sensing cells) and over 1 million optic nerve cells are present in the retina, converting light into electrical signals and transmitting the image of the object through the nerve to the brain. If the nerves in the retina are damaged or nerve function is abnormal, vision and vision problems may occur.
  • Representative retinal diseases include diabetic retinopathy, age-related macular degeneration, retinal pigmentary degeneration, retinal detachment and retinal vascular obstruction, including decreased vision, blurred vision and blindness.
  • Glaucoma is a typical eye disease in which retinal optic nerve damage occurs, and the optic nerve is damaged due to increased intraocular pressure, ischemia and oxidative stress (Kim, NY et al. J. Korean Opthalmol. Soc. 2015, 56, 70-79; Kang, JH et al. J. Korean Ophthalmol. Soc. 2003, 44, 965-970; Lee, SM et al. J. Korean Ophthalmol. Soc. 2002, 43, 2577-2584).
  • Glutamate is an excitatory neurotransmitter that plays an important role in the central nervous system of vertebrates. Glutamate is involved in the expression of various physiological functions by acting on quisqualate receptors, NMDA (N-methyl-D-aspartate) receptors, and Kainite receptors, which are concentrated in the cerebellar amygdala and hippocampus, which are known to be involved in memory and learning in brain tissues. .
  • quisqualate receptors quisqualate receptors
  • NMDA N-methyl-D-aspartate
  • Kainite receptors which are concentrated in the cerebellar amygdala and hippocampus, which are known to be involved in memory and learning in brain tissues.
  • glutamate increases in the outside of neurons and the concentration of glutamate outside the cell increases rapidly, it can act as an oxidative neurotoxic substance (Hyun-Jeong Kim et al., J. Life Sci. 2009, 19, 963-967).
  • Excessive glutamate is known to be associated with various acute neurological disorders, including anemia, oxygen deficiency, hypoglycemia, trauma, and several chronic degenerative neurological disorders. Glutamate in the eye has been implicated in acute disorders and death of retinal ganglion cells (Otori, Y. et al. Invest. Ophthalmol. Vis. Sci. 1998, 39, 972-981). It is known that overproduction of reactive oxygen species (ROS) is stimulated by the production of a large amount of glutamate and presynaptic glutamate receptor activation (Tarasenko A. et al. Neurochem. Int. 2012, 61, 1044 -1051).
  • ROS reactive oxygen species
  • ROS Reactive Oxygen Species
  • ROS Since ROS is chemically unstable and highly reactive, it can react with lipids, nucleic acids, and proteins in vivo to cause DNA damage, increase the concentration of free calcium and iron in the cell, and damage the ion transport system of the biofilm (Kiselyov, K. et al. Cell Calcium. 2016, 60, 108-114). In addition, ROS is also known to induce light-induced damage to the optic nerve by generating light (Masuda, T. et al. Oxid.Med. Cell. Longevity. 2017; article ID 9208489, 14 pages).
  • the deficiency has traditionally been used for eye health and is known to have excellent antioxidant activity. It is a mature seed of the herbaceous plant belonging to the legume (Cassia obtusifolia L.) or Ginjiang Namcha (Cassia tora L.). It is cultivated in all parts of Korea and glossed in a bow-shaped pod that is about 10 cm after the leaves are cut. Seeds containing a single line (Yen, G.-C. et al. J. Agric. Food Chem. 1998, 46, 820-824). The extract of terminator has been studied and reported for improving diabetes, dyslipidemia, liver protection, antibacterial, and hypotensive effects (Dong, X. et al. Mol. Med. Rep. 2017, 16, 2331-2346).
  • the shoot sprout extract produced a new antioxidant component compared to the extract from the shooter, increasing the antioxidant active ingredient and significantly increasing it.
  • the present inventors have confirmed that the antioxidant activity and neuronal cell protective action, and naphtopyron component including the novel compound 7-hydroxymusininyl- lubrofusarin-8'-O-glucoside isolated from the shoot sprout extract.
  • the present invention has been completed by confirming that they exhibit the effect of protecting retinal neurons and hippocampal neurons from glutamate induced oxidative stress.
  • Patent Document 1 KR 10-1503429 B1
  • Patent Document 2 KR 10-2010-0082054 A
  • Patent Document 3 KR 10-0877371 B1
  • Patent Document 4 KR 10-1807367 B1
  • Patent Document 5 KR 10-2016-0058613 A
  • Non-Patent Document 1 Jung, H. A. Journal of Ethnopharmacology, 2016, vol 191, 152-160; Inhibitory activities of major anthraquinones and other constituents from Cassia obtusifolia against ⁇ -secretase and cholinesterases.
  • Non-patent document 2 Shrestha, S. et al. Archives Pharmacal Research, 2018, online publication https://doi.org/10.1007/s12272-018-1044-0; Two new naphthalenic lactone glycosides from Cassia obtusifolia L. seeds.
  • Another object of the present specification is to provide a composition that exhibits an effect of protecting neuronal cell damage from oxidative stress or inhibiting neuronal cell death.
  • Still another object of the present specification is to provide a composition that exhibits a prophylactic or therapeutic effect of a neurological damaging disease caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • Another object of the present specification is to provide a composition that exhibits an antioxidant effect.
  • the present invention provides a naphthopyrone derivative of Formula 1, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof.
  • the present invention provides one or more naphthopyrone derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvents thereof selected from the group consisting of naphthopyrone derivatives of the following Chemical Formulas 1 to 5 Provides a method of manufacturing the cargo.
  • the present invention is one or more naphthopyron derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or the like selected from the group consisting of naphthopyrone derivatives of Formula 1 to Formula 5.
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or One or more selected from the group consisting of solvates, Cassia obtusifolia L. or Cassia tora L. bud extract, or a fraction of the bud containing it as an active ingredient, comprising retinal nerve cells or hippocampal nerve cells.
  • a composition for the prevention or treatment of a neuro-damaging disease caused by damage or death is provided.
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or It provides at least one selected from the group consisting of solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extract, or a bud fraction containing the same as an active ingredient, provides an antioxidant composition.
  • the composition according to an aspect of the present invention has an antioxidant effect and an effect of protecting nerve cells from oxidative stress or inhibiting apoptosis, and in particular, damage or death of retinal nerve cells or hippocampal nerve cells due to glutamate toxicity. It has an inhibitory effect. Therefore, the composition according to one aspect of the present invention can be used for treating or preventing vision loss and decline and eye diseases caused by optic nerve damage by protecting retinal nerve cells, and memory loss and learning due to brain nerve damage by protecting hippocampal nerve cells. It can be used for the treatment or prevention of deterioration, development and deterioration of depressive disorders and neurodegenerative diseases, and also for eye health such as improvement of memory and learning ability, stress relief, and protection of optic nerve and decreased vision. It can be used as a bar, pharmaceutical or food composition.
  • FIG. 1 is a diagram showing the free radical scavenging activity of DPPH ( ⁇ , ⁇ -diphenyl- ⁇ -picrylhydrazyl) of the Cassia tora extract (ST) and Cassia tora sprout extract (STS).
  • DPPH ⁇ , ⁇ -diphenyl- ⁇ -picrylhydrazyl
  • FIG. 2 is a diagram showing ABTS online antioxidant HPLC chromatograms of Cassia tora extract (ST) and Cassia tora sprout extract (STS).
  • the chromatogram of the blue line indicated in the upward direction is a diagram showing the components detected at 254 nm of ultraviolet (UV), and the chromatogram of the red line indicated in the downward direction reacts with the ABTS reagent to UV / VIS (ultraviolet / visible light) 734 nm
  • UV / VIS ultraviolet ultraviolet / visible light
  • FIG. 3 is a diagram showing ABTS online antioxidant HPLC chromatograms of bearer bud extracts (STS-C, STS-385, STS-465, STS-645 and STS-780) grown under various light conditions.
  • the chromatogram indicated in the upward direction is a diagram showing the components detected at ultraviolet (UV) 254 nm
  • the chromatogram indicated in the downward direction reacts with ABTS reagent to react with UV / VIS ( UV / Visible light)
  • UV / VIS UV / Visible light
  • the sample of (A) is a seedling sprout extract grown for 2 weeks under light-shielding conditions (STS of Example 1)
  • the sample of (B) is a seedling sprout extract grown for 2 weeks under normal light conditions (STS of Example 2)
  • the samples of C) and (C) were cultivated for 2 weeks under 385 nm LED illumination (STS-385 of Example 2), and the samples of (D) were cultivated for 2 weeks under 465 nm LED illumination
  • the extract (STS-465 of Example 2), the sample of (E) was cultivated for 2 weeks under 645 nm LED illumination, and the bud extract (STS-645 of Example 2), the sample of (F) was 780 nm LED illumination It was a cultivated shoot extract (STS-780 of Example 2) grown for 2 weeks under.
  • Figure 4 is an HPLC chromatogram showing the peaks of the main components separated from the bud extract (STS) of Example 1, which is an embodiment of the present invention.
  • Figure 5 is a comparative example 1 of the present invention extract (ST), cultivated bud extracts grown in light-shielding conditions (STS of Example 1) and cultivated bud extracts grown under various light conditions (STS, STS of Example 2) -C, STS-385, STS-465, STS-645 and STS-780) are graphs showing the efficacy of protecting damage to retinal progenitor cells (R28) caused by glutamate toxicity.
  • Figure 6 is a retinal progenitor cell induced by glutamate toxicity by concentration of each compound (Compounds 1 to 5 of Examples 4 and 5, and Compounds A to Compound X of Comparative Example 2) isolated from the seedling sprout extract (STS) (R28) It is a graph showing the efficacy of protecting damage.
  • Figure 7 is a comparative example 1 of the present invention extracts (ST), the seedling sprout extract grown under light-shielding conditions (STS of Example 1) and the seedling sprout extracts grown under various light conditions (STS, STS of Example 2) -C, STS-385, STS-465, STS-645 and STS-780) are graphs showing the efficacy of protecting hippocampal nerve cell (HT-22) damage caused by glutamate toxicity.
  • STS of Example 1 the seedling sprout extract grown under light-shielding conditions
  • STS, STS of Example 2 the seedling sprout extracts grown under various light conditions
  • STS-385, STS-465, STS-645 and STS-780 are graphs showing the efficacy of protecting hippocampal nerve cell (HT-22) damage caused by glutamate toxicity.
  • the present invention may relate to a naphthopyrone derivative of Formula 1, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof.
  • the naphthopyrone derivative of Formula 1 may be 7-hydroxymusizinyl-rubrofusarin-8'-O-glucoside (7-hydroxymusizinyl-rubrofusarin-8'-O-glucopyranoside).
  • pharmaceutically acceptable means the approval of a government or equivalent regulatory body to use in animals, more specifically in humans, by avoiding significant toxic effects when used in conventional medical dosages. It is meant to be recognized or approved, or recognized as listed in a pharmacopeia or other general pharmacopeia.
  • salts means salts according to one aspect of the invention that are pharmaceutically acceptable and have the desired pharmacological activity of the parent compound.
  • the salt is formed from (1) an inorganic acid such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, or the like; Or acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3- (4-hydroxybenzoyl) Benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethane-disulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenes
  • optical isomers eg, essentially pure enantiomers, essentially pure diastereomers or mixtures thereof
  • conformational isomers conformation isomers (i.e., isomers differing only in that angle of one or more chemical bonds), positional isomers (especially tautomers) or geometric isomers (e.g., cis-trans isomers) do.
  • “essentially pure”, when used in connection with, for example, enantiomers or diastereomers, contains at least about 90%, preferably at least about 95% of specific compounds that may exemplify enantiomers or diastereomers. , More preferably at least about 97% or at least about 98%, even more preferably at least about 99%, even more preferably at least about 99.5% (w / w).
  • hydrate refers to a compound to which water is bound, and is a broad concept including an inclusion compound having no chemical bonding force between water and the compound.
  • solvate refers to a higher order compound formed between molecules or ions of a solute and molecules or ions of a solvent.
  • the present invention provides one or more naphthopyrone derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvents thereof selected from the group consisting of naphthopyrone derivatives of the following Chemical Formulas 1 to 5
  • a method for preparing a cargo comprising the naphthopyron derivative, its stereoisomer, pharmaceutically acceptable salt thereof, hydrate thereof, or solvate thereof from Sprout of Cassia obtusifolia L. or Cassia tora L.
  • a method for preparing a naphthopyron derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof comprising the step of separating one or more selected from the group.
  • the compound of formula 1 may be 7-hydroxymushizinyl-rubbrofusarin-8'-O-glucoside
  • the compound of formula 2 is isotolactone ( isotoralactone)
  • the compound of formula 3 may be toralactone (toralactone)
  • the compound of formula 4 may be torosacrysone (torosachrysone)
  • the The compound of Formula 5 may be rubrofusarin.
  • the sprout may be grown by short-term germination of seeds, germinated seeds indoors, or may be a sprout that is generally sold.
  • the sprouted or sprouted plant of the above-described manufacturing method was seeded from the seeded plant ( Cassia obtusifolia L.) or Ginseng tea ( Cassia tora L. or Senna tora ), and then grown or grown naturally for 2 to 31 days.
  • It may be a sprout or a young plant, specifically 2 days or more, 3 days or more, 4 days or more, 5 days or more, 6 days or more, 7 days or more, 8 days or more, 9 days or more, 10 days or more, 11 days or more, 12 days or more, 13 days or more, 14 days or more, 15 days or more, 16 days or more, 17 days or more, 18 days or more, 19 days or more, 20 days or more, 21 days or more, 22 days or more, 23 days or more, 24 days More than 25 days, more than 26 days, more than 27 days, more than 28 days, more than 29 days or more than 30 days Naturally grown or grown buds or young plants, 31 days or less, 30 days or less, 29 days or less, 28 days or less, 27 days or less, 26 days or less, 25 days or less, 24 days or less, 23 days or less, 22 days or less, 21 days or less, 20 days or less, 19 days or less, 18 days or less, 1 7 days or less, 16 days or less, 15 days or less, 14 days or less,
  • the term ( Cassia obtusifolia L. or Cassia tora L.) refers to the herbaceous genus of dicotyledonous plant family Rosaceae, and the term of the term (Cassia obtusifolia L.) or Ginseng tea (Cassia tora L. or Senna) tora) as mature seeds, the extract of terminator is known to improve diabetes, improve dyslipidemia, protect liver, antibacterial, and lower blood pressure (Dong, X. et al. Mol. Med. Rep. 2017, 16, 2331-2346) , It is not known about the efficacy of improving or inhibiting the damage of retinal nerve cells or hippocampal nerve cells due to oxidative stress or drug toxicity, especially glutamate.
  • the germinating bud or young plant may be all or part of the germinating bud or young plant.
  • the part may be an outpost or above ground or underground.
  • the ground portion may be a stem, a leaf, a flower, or a combination thereof.
  • the basement part may be a root.
  • the plant may be naturally grown or artificially cultivated.
  • the bud of the present invention can be easily grown indoors and can be grown and used for a short period of time within several weeks without supplying additional nutrients, so it has the advantage of great industrial utility.
  • the shoot sprout extract also includes a crude extract (crude extract) or an additional fraction (fractionation) of the extract.
  • the seedling sprout extract may be a crude extract, fraction, or a combination thereof.
  • the crude extract refers to the one obtained by contacting the sprouted bud with the extraction solvent.
  • the fraction refers to the separation of a substance containing specific components with respect to the crude extract.
  • the extract, or a fraction thereof may be an extract of a shoot sprout plant, a fraction thereof, or a small fraction of each of the compositions of the present invention, or as a mixture thereof.
  • the small fraction may be obtained by passing an ultrafiltration membrane having a cut-off value, and may be obtained by column chromatography or solvent fractionation.
  • the sprout bud extract or fraction may contain one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of the formula (1) to (5).
  • the separation may be by filtration, dipping, centrifugation, solvent fractionation, chromatography or a combination thereof.
  • the chromatography is prepared for separation according to various conditions, i.e., size, charge, hydrophobicity or affinity, ion exchange chromatography, affinity chromatography, size exclusion chromatography, HPLC, high-speed chromatography, column chromatography, reverse phase Column chromatography or combinations thereof.
  • the extraction may include incubating the plant in a solvent for a period of time.
  • the extraction may be performed with or without stirring, or may include heating.
  • the incubation may be performed at room temperature to reflux temperature with or without stirring.
  • Incubation temperature may be appropriately selected depending on the solvent selected. For example, the temperature may be from room temperature to reflux temperature, from 30 ° C. to reflux temperature, and from 40 ° C. to reflux temperature.
  • the heating may include heating to 50 ° C., 60 ° C., 70 ° C., 80 ° C., or reflux temperature.
  • the heating may be to 50 °C to reflux temperature, 60 °C to reflux temperature, 70 °C to reflux temperature, 80 °C to reflux temperature, or to the reflux temperature.
  • the extraction time may vary depending on the temperature selected, for example 1 hour to 2 months, for example 1 hour to 1 month, 1 hour to 15 days, 1 hour to 10 days, 1 hour to 5 days, 1 hour to 3 days. , 1 hour to 2 days, 1 hour to 1 day, 5 hours to 1 month, 5 hours to 15 days, 5 hours to 10 days, 5 hours to 5 days, 5 hours to 3 days, 5 hours to 2 days, 5 Hour to 1 day, 10 hours to 1 month, 10 hours to 15 days, 10 hours to 10 days, 10 hours to 5 days, 10 hours to 3 days, or 10 hours to 2 days.
  • the extraction may be to extract the plant under reflux in a solvent.
  • the solvent may have a volume of 1 times, 2 times, 5 times, 10 times, or 15 times or more with respect to the weight of the plant.
  • the solvent may have a volume of 1 to 15 times, 2 to 15 times, 5 to 15 times, 10 times to 15 times, or about 15 times the weight of the plant.
  • the plant may be dried in shade or shading facility or warm air or drying device.
  • the extraction method conventional methods in the art such as filtration, hot water extraction, dipping extraction, cold dipping, microwave extraction, reflux cooling extraction, pressure extraction, subcritical extraction, supercritical extraction, and ultrasonic extraction may be used.
  • Immersion extraction may be to be immersed at warm or room temperature, it may be one to five times to extract.
  • the seedling sprout plant may be in contact with the extraction solvent of 0.1 to 10 times or 1 to 6 times.
  • Cold needle extraction temperature may be 20 °C to 40 °C.
  • the warm needle or heat extraction temperature may be 40 ° C to 100 ° C.
  • Cold needle extraction time may be 24 hours to 120 hours, the temperature of the hot or heated extraction may be 0.5 hours to 48 hours.
  • the extraction may also include removing the solvent from the extract obtained by known methods such as evaporation or reduced pressure concentration.
  • the extraction may also include preparing a dry extract by drying the obtained extract, such as lyophilization.
  • the decompression concentration may be to use a vacuum decompression concentrator or a vacuum rotary evaporator.
  • the drying may be drying under reduced pressure, vacuum drying, boiling drying, spray drying or freeze drying.
  • the manufacturing method may further include the step of extracting the missing sprout as a solvent selected from the group consisting of water, C1 to C6 alcohol, and a mixed solvent thereof.
  • the alcohol may be C1 to C3 alcohol, C1 to C4, C1 to C5, or C1 to C6 alcohol.
  • the alcohol may be a primary alcohol.
  • the alcohol of C1 to C6 may be methanol, ethanol, propanol, isopropanol, butanol or a mixture thereof.
  • the preparation method may further comprise the step of fractionating to at least one selected from the group consisting of water, ethyl acetate, hexane, methylene chloride, chloroform, methanol, ethanol, acetone, and a mixed solvent thereof.
  • the preparation method is based on the total weight of the extract obtained through the extraction of the shoots sprouts one or more naphthopyron derivatives selected from the group consisting of the naphthopyrone derivatives of Formula 1 to Formula 5, stereoisomers thereof, Pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, which may comprise the step of preparing a shoot sprout fraction comprising 1 to 20% by weight, specifically 1% by weight, 2% by weight, At least 3 wt%, at least 4 wt%, at least 5 wt%, at least 6 wt%, at least 7 wt%, at least 8 wt%, at least 9 wt%, at least 10 wt%, at least 11 wt%, at least 12 wt%, It may be to include a process for producing a fraction of the shoots containing at least 13% by weight, at least 14% by weight, at least 15% by weight, at least 16% by weight, at least 17% by weight, at least 18% by weight or at least
  • One or more naphthopyron derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof selected from the group consisting of naphthopyrone derivatives of Formulas 1 to 5 may be May be included in the range of 1 to 50% by weight, specifically, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight, 6% by weight, 7% by weight % By weight, 8% by weight, 9% by weight, 10% by weight, 11% by weight, 12% by weight, 13% by weight, 14% by weight, 15% by weight, 16% by weight, 17% by weight At least 18% by weight at least 19% by weight at least 20% by weight at least 21% by weight at least 22% by weight at least 23% by weight at least 24% by weight at least 25% by weight at least 26% by weight.
  • % By weight, 28% by weight, 29% by weight, 30% by weight, 31% by weight, 32% by weight At least 33 wt%, at least 34 wt%, at least 35 wt%, at least 36 wt%, at least 37 wt%, at least 38 wt%, at least 39 wt%, at least 40 wt%, at least 41 wt%, 42 wt% At least 43 wt%, at least 44 wt%, at least 45 wt%, at least 46 wt%, at least 47 wt%, at least 48 wt% or at least 49 wt%, and at most 50 wt%, at most 49 wt%, 48 wt% or less, 47 wt% or less, 46 wt% or less, 45 wt% or less, 44 wt% or less, 43 wt% or less, 42 wt% or less, 41 wt% or less, 40 wt% or less, 39 wt
  • the present invention provides at least one naphthopyrone derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvent thereof.
  • the one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 are from the naphthopyrone derivatives of Formula 1 and the naphthopyrone derivatives of Formulas 2 to 5 It may be one or more selected naphthopyrone derivatives.
  • Descriptions of the naphthopyrone derivatives of Formulas 1 to 5, pharmaceutically acceptable salts, stereoisomers, hydrates, solvates, modified buds, modified bud extracts or fractions are as described above.
  • the composition may be a composition for protecting neurons or inhibiting neuronal death, and specifically, may be a composition for protecting neurons or inhibiting neuronal death from oxidative stress induced by metabolic toxicity, neurotoxicity, chemical causes, and the like.
  • the oxidative stress may be caused by glutamate, glutamate toxicity, or glutamate neurotoxicity, or calcium homeostatic dysregulation, mitochondrial dysfunction, excitatory cytotoxicity, similar to oxidative stress caused by glutamate neurotoxicity, Oxidative stress, such as depletion of nutritional factors.
  • the composition may reduce, inhibit, ameliorate, or prevent damage to retinal neurons or hippocampal neurons caused by glutamate, glutamate toxicity, or glutamate neurotoxicity, or resuscitate or kill dead retinal neurons or hippocampal neurons. It may be regenerative and protects nerve cells by regulating antioxidant activation or glutamate metabolism, which is a defense against oxidative stress caused by glutamate neurotoxicity, abnormalities in calcium homeostasis, mitochondrial dysfunction, excitatory cytotoxicity, depletion of nutritional factors, etc. Or inhibit neuronal cell death.
  • the extract of shoot sprouts has a DPPH free radical scavenging effect of 1.7 times or more superior to the extract of shoot shoots (Test Example 1), and the compounds of Formula 1 to Formula 5 corresponding to antioxidant components (naph Topirone derivative) has a high content and shows a better antioxidant effect (Test Example 2)
  • the composition of the present invention is an antioxidant action that is a defense mechanism against oxidative stress is activated to protect neurons or inhibit neuronal cell death Was confirmed to be excellent.
  • the composition may be treated before, concurrent with, or after development of neuronal damage or death.
  • the composition is a group consisting of at least one naphthopyrone derivative selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof.
  • sprout extract or a sprout sprout fraction comprising the same may comprise 0.001% to 80% by weight relative to the total weight of the composition, specifically 0.001% or more, 0.01% or more, 0.05% or more, 0.1% or more, 1% or more, 2% or more, 3% or more, 4% or more, 5% or more, 10% or more, 20 wt% or more, 30 wt% or more, 40 wt% or more, or 60 wt% or more, 80 wt% or less, 60 wt% or less, 40 wt% or less, 30 wt% or less, 20 wt% or less, 10 It may include less than 5% by weight, less than 5% by weight, less than 4% by weight, less than 3% by weight, less than 2% by weight, or less than 1% by weight.
  • the above-mentioned sprout extract or fraction is one or more naphthopyron derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or the like selected from the group consisting of naphthopyrone derivatives of the above Chemical Formulas 1 to 5.
  • Solvate may comprise from 1 to 50% by weight, based on the total weight of the shoot sprout extract or fraction, specifically, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight % By weight, 6% by weight, 7% by weight, 8% by weight, 9% by weight, 10% by weight, 11% by weight, 12% by weight, 13% by weight, 14% by weight, 15% by weight At least 16% by weight at least 17% by weight at least 18% by weight at least 19% by weight at least 20% by weight at least 21% by weight at least 22% by weight at least 23% by weight at least 24% by weight.
  • % By weight, 26% by weight, 27% by weight, 28% by weight, 29% by weight At least 30%, at least 31% by weight, at least 32% by weight, at least 33% by weight, at least 34% by weight, at least 35% by weight, at least 36% by weight, at least 37% by weight, at least 38% by weight, at 39% by weight At least 40% at least 41% at least 42% at least 43% at least 44% at least 44% at least 45% at least 46% at least 47% at least 48% or at least 49% It may include at least 50%, up to 49%, up to 48%, up to 47%, up to 47%, up to 46%, up to 45%, up to 44%, up to 43%, up to 42% Up to 41 wt%, up to 40 wt%, up to 39 wt%, up to 38 wt%, up to 37 wt%, up to 36 wt%, up to 35 wt%, up to 34 wt%, up to 33 wt
  • the nerve cell may be a central nerve, a peripheral nerve or an optic nerve associated with the brain hippocampus or the eye retina, and specifically, may be a retinal nerve cell or a hippocampal nerve cell.
  • the retinal nerve cells are rod cells, cone cells, bipolar cells, retinal amacrine cells, horizontal cells and retinal ganglion cells. It may be one or more cells selected from the group consisting of).
  • the rod cells and cone cells are nerve cells that sense light by sensing light, and the dipole cells, retinal amacrine cells, and horizontal cells are nerve cells that transmit visual information to the retinal ganglion cells, and the retinal ganglion cells are retinal ganglion cells. It is a nerve cell that delivers visual information to the brain.
  • R28 retinal progenitor cells or R28 retinal progenitor cells of the present invention express the traits of various constituent cells of the retina and survive even when transplanted into the retina, apoptosis occurs due to hypoxia or serum deficiency, It can also be useful for apoptosis and cytotoxicity studies that are linked to related glutamate or GABA receptor expression.
  • R28 cells can differentiate from retinal progenitor cells to retinal ganglion cells depending on the culture conditions, and can be used not only for overall cytological studies of the retina, but also for basic research of retinal ganglion cells (Jungil Lee, Jaewoo Kim, Korean Ophthalmology) Journal, 2009, 50, 919-922).
  • the hippocampal nerve cells are nerve cells of the hippocampus (hippocampus), which is a structure of the medial temporal lobe of the brain, and the hippocampus is vulnerable to physiological / oxidative stress stimulation and is known as a central tissue causing damage to cognitive function due to stress. May cause hippocampal structure, brain cell production, synaptic plasticity, and behavioral changes associated with hippocampus (Kim, Eun-ju, Korean Psychological Association: Cognitive and Biological, 2012, 24, 65-88).
  • HT-22 hippocampal neurons of one embodiment of the present invention can be used as an in vitro model system for studying neurotoxicity induced by oxidative stress due to sensitivity to glutamate (Liu, J. et al. Life Science , 2009, 84, 267-271; Kim Ji-hyun, Jeon Soon-sil, Korean Journal of Food Science and Nutrition, 2017, 46, 886-890).
  • the present invention provides at least one naphthopyrone derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvent thereof.
  • Injury of retinal nerve cells or hippocampal nerve cells comprising as an active ingredient at least one selected from the group consisting of cargo, Cassia obtusifolia L. or Cassia tora L. Or it may be directed to a composition for preventing or treating a neurologically damaging disease caused by death, or may be related to a composition for preventing or treating a neurologically damaging disease caused by glutamate neurotoxicity.
  • the at least one naphthopyrone derivative selected from the group consisting of naphthopyrone derivatives of Formulas 1 to 5 is selected from the group consisting of naphthopyrone derivatives of Formula 1 and naphthopyrone derivatives of Formulas 2 to 5 It may be one or more naphthopyrone derivatives.
  • the neurological damaging disease caused by damage or death of the retinal nerve cells or hippocampal nerve cells may be due to glutamate neurotoxicity, and the description of the retinal nerve cells and hippocampal nerve cells is as described above.
  • the neurological damaging disease may be at least one selected from the group consisting of diabetic retinopathy due to retinal nerve cell injury or death, macular degeneration, vision disorder due to retinal cell damage, retinal pigmentation, retinal detachment, retinal vessel occlusion and glaucoma.
  • the disease May be one or more selected from the group consisting of memory loss, loss of learning ability, depressive disorder, amyotrophic lateral sclerosis (Lou Gehrig's disease), Parkinson's disease and cerebral ischemic nerve injury due to hippocampal nerve cell injury or death,
  • the disease is not limited as long as the disease is caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • the glutamate neurotoxicity refers to toxicity caused by glutamate acting as an oxidative neurotoxic substance due to a rapid increase in glutamate concentration outside the neuron.
  • the glutamate neurotoxicity may be caused by glutamate introduced from outside of the subject, and the influx of glutamate may be ingested with food containing glutamate, therapeutic agent containing glutamate, preventive agent containing glutamate, antibiotic containing glutamate, and the like. Or by administration, but is not limited thereto.
  • One or more naphthopyron derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 may be separated or purified from the sprout ( Cassia obtusifolia L. or Cassia tora L.), specifically, the sprout
  • the ethanol extract may be separated or purified from the ethanol extract, and the ethanol extract from the shoots may be fractionated with ethyl acetate, and then the ethyl acetate fraction may be separated by chromatography, or the ethyl acetate fraction may be separated with a mixed solvent of hexane, methylene chloride and methanol. It may be fractionated and separated by chromatography.
  • the seedling sprout extract may be extracted as a solvent selected from the group consisting of water, C1 to C6 alcohol and a mixed solvent thereof.
  • the alcohol may be C1 to C3 alcohol, C1 to C4, C1 to C5, or C1 to C6 alcohol.
  • the alcohol may be a primary alcohol.
  • the alcohol of C1 to C6 may be methanol, ethanol, propanol, isopropanol, butanol or a mixture thereof.
  • the fraction may further include fractionating into one or more selected from the group consisting of water, ethyl acetate, hexane, methylene chloride, chloroform, methanol, ethanol, acetone, and a mixed solvent thereof.
  • the one or more naphthopyrone derivatives selected from the group consisting of naphthopyrone derivatives of the formula (1) to (5) is contained in a higher content in the shoot sprout extract than the extract of the fault as an antioxidant component (Test Example 2), the naphthopyron derivative is excellent in protecting the retinal nerve cells (R28) and hippocampal nerve cells (HT-22) damaged or killed by glutamate treatment (Test Examples 4 and 6).
  • the naphthopyrone derivatives of Formulas 1 to 5 are effective ingredients of the shoot sprout having the efficacy of preventing or treating neuronal damage diseases caused by glutamate neurotoxicity.
  • the present invention provides at least one naphthopyrone derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvent thereof. At least one selected from the group consisting of cargoes, Cassia obtusifolia L. or Cassia tora L. Sprout extracts comprising the same, or may be related to the composition for antioxidant, comprising a fraction of the sprouts containing the same as an active ingredient.
  • the at least one naphthopyrone derivative selected from the group consisting of naphthopyrone derivatives of Formulas 1 to 5 is selected from the group consisting of naphthopyrone derivatives of Formula 1 and naphthopyrone derivatives of Formulas 2 to 5 It may be one or more naphthopyrone derivatives.
  • the present invention in one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formula 1 to Formula 5 in the individual in need of neuronal cell protection from oxidative stress or neuronal cell death inhibition, the stereoscopic thereof One or more selected from the group consisting of isomers, pharmaceutically acceptable salts thereof, hydrates, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or missing bud fractions comprising the same It may be related to a method for protecting neurons from oxidative stress or inhibiting neuronal death. In one aspect of the present invention, administration of the method may be performed according to the administration method and administration dose described herein.
  • the present invention is selected from the group consisting of the naphthopyrone derivatives of Formula 1 to Formula 5 in the individual in need of prevention or treatment of neuro-injury disease caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • It may be related to a method for preventing or treating a neuro-damaging disease caused by damage or death of retinal nerve cells or hippocampal nerve cells, comprising administering a bud extract or a bud fraction containing a bud thereof.
  • administration of the method may be performed according to the administration method and administration dose described herein.
  • the present invention is a naphthopyrone derivative selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 in an individual in need of antioxidant, stereoisomers thereof, pharmaceutically acceptable salts thereof, and hydrates thereof , Or one or more selected from the group consisting of solvates thereof, and related to an antioxidant method comprising administering a bud extract comprising Cassia obtusifolia L. or Cassia tora L., or a bud bud fraction comprising the same. have.
  • administration of the method may be performed according to the administration method and administration dose described herein.
  • the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a pharmaceutical composition for protecting neurons from oxidative stress or inhibiting neuronal death.
  • One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
  • the present invention is a naphthopyrone derivative of Chemical Formulas 1 to 5 for preparing a pharmaceutical composition for the prevention or treatment of neuro-injury disease caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a pharmaceutical composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or the named bud fractions comprising the same.
  • the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a cosmetic composition for protecting neurons from oxidative stress or inhibiting neuronal death.
  • One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
  • the present invention is a naphthopyrone of Formula 1 to Formula 5 for preparing a cosmetic composition for improving, preventing or treating a neuro-damage disease caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • One or more naphthopyrone derivatives selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, one or more selected from the group consisting of, Cassia obtusifolia L. Or Cassia tora L.) bud extract, or the use of a named bud fraction comprising the same.
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a cosmetic composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or the named bud fractions comprising the same.
  • the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a food composition for protecting neurons from oxidative stress or inhibiting neuronal death.
  • One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
  • the present invention is a naphthopyrone of Formula 1 to Formula 5 for preparing a food composition for improving, preventing or treating a neuro-damaging disease caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • One or more naphthopyrone derivatives selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, one or more selected from the group consisting of, Cassia obtusifolia L. Or Cassia tora L.) bud extract, or the use of a named bud fraction comprising the same.
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a food composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or the named bud fractions comprising the same.
  • the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a quasi-drug composition for protecting neurons from oxidative stress or inhibiting neuronal death.
  • One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
  • the present invention is a naphthopyrone of Formula 1 to Formula 5 for preparing a quasi-drug composition for the improvement, prevention or treatment of a neuro-damage disease caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • One or more naphthopyrone derivatives selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, one or more selected from the group consisting of, Cassia obtusifolia L. Or Cassia tora L.) bud extract, or the use of a named bud fraction comprising the same.
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a quasi-drug composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or bear sprout fractions comprising them.
  • one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or solvents thereof
  • one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or solvents thereof
  • one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or solvents thereof
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for protecting neurons from oxidative stress or inhibiting the death of neurons. At least one selected from the group consisting of stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extract, or crystals comprising the same It may be related to the use of the sprout fraction.
  • the present invention is selected from the group consisting of naphthopyrone derivatives of Formula 1 to Formula 5 for preventing or treating neuro-injury diseases caused by damage or death of retinal nerve cells or hippocampal nerve cells.
  • the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for antioxidant, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, Or it may be related to the use of at least one selected from the group consisting of solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extract, or a named bud fraction comprising the same.
  • composition of the present invention may be a pharmaceutical composition, cosmetic composition or food composition.
  • compositions according to one aspect of the invention may be formulated in oral or parenteral dosage forms.
  • it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents and surfactants.
  • Solid form preparations for oral administration include tablets, pills, powders, granules, soft or hard capsules, and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose, or the like. Or lactose, gelatin, or the like is mixed.
  • lubricants such as magnesium stearate and talc are also used.
  • Liquid preparations for oral administration include suspending agents, intravenous solutions, emulsions, syrups, etc.
  • various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included.
  • Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, and suppositories.
  • a non-aqueous solvent and a suspension solvent propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate may be used.
  • As a base for suppositories witepsol, macrogol, tween 61, cacao butter, laurin butter, and glycerogelatin may be used.
  • compositions according to one aspect of the invention may be used in the form of their pharmaceutically acceptable salts, and may also be used alone or in combination with other pharmaceutically active compounds as well as in a suitable collection.
  • the salt is not particularly limited as long as it is pharmaceutically acceptable.
  • hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrofluoric acid, hydrobromic acid, formic acid acetic acid, tartaric acid, lactic acid, citric acid, fumaric acid, maleic acid, succinic acid, methanesulfonic acid , Benzene sulfonic acid, toluene sulfonic acid, naphthalene sulfonic acid and the like can be used.
  • Parenteral dosage forms may also be dermal application patches, ointments, creams, eye drops, sprays or injections.
  • the invention may be a method for protecting neurons in a subject comprising administering the pharmaceutical composition to the subject.
  • the subject may be a mammal, eg, a human, a cow, a horse, a pig, a dog, a sheep, a goat, or a cat, the mammal may be a human, and an effective dosage for a human body of a compound of the invention Depends on the age, weight, sex, dosage form, health condition and degree of disease of the patient.
  • a mammal eg, a human, a cow, a horse, a pig, a dog, a sheep, a goat, or a cat
  • an effective dosage for a human body of a compound of the invention Depends on the age, weight, sex, dosage form, health condition and degree of disease of the patient.
  • the administration may be administered in various formulations for oral administration or parenteral administration such as intravenous, intraperitoneal, intradermal, subcutaneous, epithelial or intramuscular administration, and when formulated, commonly used fillers, extenders, binders, wetting agents, shelf life It is prepared using diluents or excipients such as releases, surfactants and the like.
  • the administration can be administered by methods known in the art. Administration can be administered directly to the subject by any means, eg, by intravenous, intramuscular, oral, or subcutaneous administration.
  • the administration can be administered systemically or locally.
  • the administration may be locally administered to a site where neurons are present or expected to occur.
  • Food composition according to an aspect of the present invention may be a health functional food composition.
  • the food composition is not particularly limited in dosage form, but may be formulated in a form in which the concentrate or powder is ingested or ingested directly or diluted, for example, a liquid such as tablets, granules, powders, drinks, caramels , Gels, bars and the like.
  • the food composition of each formulation may be suitably selected by those skilled in the art according to the formulation or purpose of use in addition to the active ingredient, and a synergistic effect may occur when simultaneously applied with other raw materials.
  • the food composition may contain various flavors or natural carbohydrates as additional ingredients.
  • the natural carbohydrate may be glucose, monosaccharides such as fructose, maltose, disaccharides such as sucrose, and polysaccharides such as dextrin, cyclodextrin, sugar alcohols such as xylitol, sorbitol, and erythritol.
  • sweetener natural sweeteners such as taumatin and stevia extract, and synthetic sweeteners such as saccharin and aspartame can be used.
  • the ratio of the natural carbohydrate may be selected from 0.01 to 0.04 parts by weight, specifically about 0.02 to 0.03 parts by weight per 100 parts by weight of the composition.
  • the food composition may contain various nutrients, vitamins, electrolytes, flavors, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohols, carbonic acid. Carbonating agents and the like used in beverages.
  • the functional food of the present invention may contain a flesh for preparing natural fruit juice, fruit juice beverage and vegetable beverage. These ingredients can be used independently or in combination. The proportion of such additives is not so critical but is typically included in the range of 0 to about 20 parts by weight per 100 parts by weight of the composition herein.
  • the dosage determination of the active ingredient is within the level of those skilled in the art, the daily dosage of which is for example from 0.1 mg / kg / day to 5000 mg / kg / day, more specifically 50 mg / kg It may be / day to 500 mg / kg / day, but is not limited thereto, and may vary depending on various factors such as age, health condition, complications of the subject to be administered.
  • the food composition according to one aspect of the present invention includes, for example, health products including chewing gum, caramel products, candy, ice cream, confectionary, various food products such as soft drinks, mineral water, alcoholic beverages, vitamins and minerals, and the like. Functional foods.
  • composition of the present invention may be a quasi-drug composition.
  • the term 'out of quasi-drugs' refers to drugs based on the classification criteria set by the Ministry of Health and Welfare for articles that have less effect on the human body than drugs used for the treatment or prevention of diseases according to the Pharmaceutical Affairs Act. do. Therefore, it may include fiber and rubber products used for the treatment or prevention of human or animal diseases, mild or non- direct action on the human body, non-apparatus or machinery, and the like, or disinfectants and insecticides to prevent infectious diseases. Can be.
  • the quasi-drug composition of the present invention may be for the improvement, protection or treatment of damage to nerve cells by oxidative stress, may be for the prevention or improvement of neurological damage diseases caused by glutamate neurotoxicity.
  • the quasi-drug composition may further include an quasi-pharmaceutically acceptable excipient or carrier.
  • the quasi-drug composition may be formulated in the form of skin smear, cream, paste, eye drop, spray.
  • the dried plant was placed in an extraction container, 5.5 L of ethanol was added thereto, shaken at room temperature, stirred and extracted for 7 days, and the mixture was mixed with a Whatman paper filter paper having a film thickness of 0.34 mm and a diameter of 30.
  • the process of gravity filtration using a cm glass funnel was repeated twice ('extraction-filtration' twice) to obtain a filtered extract.
  • the filtered extract was placed in a reduced pressure concentrator and concentrated by evaporation of the solvent completely at 35 ° C. under reduced pressure to obtain 66.7 g (hereinafter, referred to as 'STS') of Clarified Sprout Extract (yield 9.55%).
  • the seedling sprouts were grown by varying the light wavelength conditions. Cultivated buds are grown in the same manner as in Example 1, but the selective light quality of the fluorescent light, LED 385 nm, 465 nm, 645 nm, 780 nm, respectively, in a large amount of wavelength, not light-shielding conditions at the time of growing the germination buds Irradiated with light, each was grown for 6 days.
  • each of the extracts of Cultivated Sprout was prepared in the same manner as in Example 1, and the extracts obtained by cultivating under fluorescent light conditions were 'STS-C' under LED light conditions of 385 nm.
  • 'STS-385' the extract obtained by cultivation under the light condition of LED 465 nm, 'STS-465', the extract obtained by cultivating under the light condition of LED 645 nm 'STS-645', LED 780 nm
  • the extract obtained by cultivation under light condition of 'STS-780' was called.
  • ethyl acetate fraction (STS- EA) 23.5 g were obtained.
  • the ethyl acetate fraction was mixed with methylene chloride (or ethyl acetate) alone or with a mixed solvent of n -hexane, methylene chloride (or ethyl acetate) and methanol (or ethanol) ( n -hexane: methylene chloride mixture ratio (volume ratio) of 1).
  • the ethyl acetate fraction (STS-EA) is contacted with the eight fraction solvents, stirred in celite to evaporate the solvent, and silica packed in a column having a diameter of 10 cm and a length of 20 cm. ) was developed in the resin. Chromatography eluting with each of the above solvents yielded 20 fractions (STS-EA-fraction 1 to STS-EA-fraction 20).
  • STS-EA-fraction 12 prepared in Example 3 (the second fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 10: 1)
  • preparative column chromatography was performed under the following separation method 1 to separate Compound 1 having the structure of Formula 1 at a retention time of about 221 minutes. (14.0 mg, 0.002% by weight of dry plant; 0.021% by weight of dry extract)
  • STS-EA-fraction 12 prepared in Example 3 (the second fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 10: 1)
  • preparative column chromatography was performed under the condition of separation method 1 of Example 4 to separate 1.2 mg of Compound 3 having the structure of Chemical Formula 3 at a retention time of about 24 minutes.
  • the STS-EA-fraction 8 of Example 3 (the first fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 50: 1) is concentrated under reduced pressure. Then, under the conditions of the following separation method 2, preparative column chromatography was performed to separate 7.5 mg of compound 2 having the structure of formula 2 at a retention time of about 36 minutes.
  • Example 3 STS-EA-fraction 13 (third fraction out of three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 10: 1) is concentrated under reduced pressure. Then, under the conditions of the following separation method 3, preparative column chromatography was performed to separate 0.6 mg of Compound 4 having the structure of Formula 4 between the retention time of about 71 minutes, and the Chemical Formula 5 at the retention time of about 120 minutes. 2.6 mg of Compound 5 having the structure of was isolated.
  • the STS-EA-fraction 8 of Example 3 was concentrated under reduced pressure, and preparative column chromatography was performed under the conditions of the separation method 2 of Example 4 to separate the components contained in the fraction. As a result, 7.2 mg of Compound C corresponding to Peak C of FIG. 4, 0.8 mg of Compound D corresponding to Peak D of FIG. 4, and 2.5 mg of Compound E corresponding to Peak E of FIG. 4 were isolated.
  • STS-EA-fraction 16 obtained in Example 3 (the third fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 5: 1)
  • silica gel column chromatography was performed under the following separation method 5 to fractionate into 14 small fractions.
  • the peak of FIG. 4 2.5 mg of compound M corresponding to M, 2.1 mg of compound N corresponding to peak N in FIG. 4, and 0.5 mg of compound O corresponding to peak O in FIG. 4 were separated.
  • STS-fraction 17 obtained in Example 3 (the first fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 3: 1) is concentrated under reduced pressure. Subsequently, the components containing the fractions were separated by HPLC separation under the conditions of Separation Method 9 below. As a result, 1.7 mg of Compound Q corresponding to the peak Q of FIG. 4 was isolated.
  • the STS-EA-fraction 18 obtained in Example 3 (the second fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 3: 1) is reduced.
  • silica gel column chromatography was performed under the conditions of the following separation method 10 to fractionate into 24 small fractions.
  • the small fractions 17 to 21 (the third and fourth fractions of the four small fractions in which the elution solvent in the separation method 10 is methylene chloride: methanol mixing ratio 3: 1 and the elution solvent is methylene chloride: methanol mixture ratio 2: 1
  • the second to fourth of 4 fractions were separated under the conditions of the following separation method 11 through HPLC separation to separate the components of the fraction.
  • 1.6 mg of Compound B corresponding to Peak B of FIG. 4 24.8 mg of Compound R corresponding to Peak R of FIG. 4, 5.5 mg of Compound S corresponding to Peak S of FIG. 4, and corresponding to Peak T of FIG. 4.
  • Compound U corresponding to peak U of FIG. 4 9.3 mg
  • peak X of FIG. 0.6 mg of the corresponding compound X was isolated.
  • HPLC peaks corresponding to all the substances (Compounds 1 to 5, and Compounds C to X) separated in Examples 4 and 5 and Comparative Example 2 are as indicated for each peak of the named bud extract HPLC chromatogram in FIG. 4.
  • a total of 29 substances, including naphthopyrone derivatives having the structures of Formulas 1 to 5 (Compounds 1 to 5), are components that can be separated from the extract of the resulting shoot sprout obtained in Example 1.
  • the 29 substances correspond to the main component of the shoot sprout extract.
  • the molecular weight of Compound 1 isolated in Example 4 was determined to be 664 by MS measurement using an Agilent 1100 Fast Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), ultraviolet (PerkinElmer 343 Polarimeter), infrared (Thermo Scientific).
  • Nicolet iS50 1 H and 13 C NMR spectrum analysis using non-photoluminescence (PerkinElmer Lambda 35) spectroscopy and nuclear magnetic resonance (Bruker 400 MHz, 100 MHz NMR) to obtain the structure of the formula
  • Eggplants were determined with 7-hydroxymusgininyl-rubrufusarin-8'-0-glucoside, a naphthopyrone derivative.
  • Compound 1 is a novel compound that has not been reported to date and has been isolated as one of the main components only in the shoot buds, which was not found or contained in very small amounts in the shooter or fully grown shooter plants.
  • the molecular weight of Compound 2 isolated in Example 5 was determined to be 272 by MS measurement using an Agilent 1100 High-Speed Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR). 1 H NMR spectra were used to estimate isotoractone, a naphthopyrone derivative having the structure shown in Formula 2 below. In addition, the structure of the NMR data was compared with that of the existing literature (Kitanaka, S. et al. Phytochemistry, 1981, 20, 1951-1953).
  • the molecular weight of Compound 3 isolated in Example 5 was determined to be 272 by MS measurement using an Agilent 1100 Fast Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR).
  • the structure was estimated by 1 H NMR spectral analysis as toralactone, a naphthopyrone derivative having the structure shown in Chemical Formula 3 below.
  • the structure of the NMR data was compared with that of the existing literature (Newman, AG et al. J Am Chem Soc 2016, 138, 4219-4228).
  • the molecular weight of Compound 4 isolated in Example 5 was determined to be 288 by MS measurement using an Agilent 1100 Fast Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR).
  • the structure was estimated by 1 H NMR spectral analysis as torosachrysone, a naphthopyron derivative having the structure shown in Chemical Formula 4 below.
  • the structure of the NMR data was compared with that of the existing literature (Gill, M. et al. Aust J Chem 2000, 53, 213-220).
  • the molecular weight of Compound 5 isolated in Example 5 was determined to be 288 by MS measurement using an Agilent 1100 High-Speed Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR). Through 1 H NMR spectroscopic analysis, the structure was estimated to be rubrofusarin, a naphthopyron derivative having the structure shown in Chemical Formula 5 below. In addition, the structure of NMR data was compared with that of the existing literature (Alemayehu, G. et al. Phytochemistry, 1993, 32, 1273-1277).
  • Dissolver extract (ST) prepared in Comparative Example 1 and the crystallized sprout extract (STS) prepared in Example 1 was dissolved in an aqueous solution of alcohol at the same concentration, according to the following analysis conditions ABTS online antioxidant high performance liquid chromatography (HPLC ) Was performed.
  • ABTS composition water containing 0.08 mM ABTS and 0.12 mM potassium persulfate
  • Detector detection wavelength ultraviolet 254 nm; 734 nm
  • Cassiae extract (ST) and Cassiae sprout extract (STS) showed very different antioxidant components.
  • the components of the Cassia vulgaris extract (ST) and Cassia vulgaris extract (STS) were found to be composed of flavonoids and naphthalene derivatives having sulfated activity 15 minutes before the retention time. At the same time as the contents of the above components were increased, naphthalene and anthraquinone derivatives having antioxidant activities were additionally observed after 15 minutes.
  • Figure 2 (A) and (B) is a result of performing the ABTS online antioxidant HPLC of the deficiency and the germinated bud extract, ultraviolet (UV) detector 254 nm (chromatic gram of the blue line indicated in the upward direction), 734 nm (downward) Chromatogram obtained by detecting wavelengths of red lines indicated by?).
  • UV detector 254 nm chromatic gram of the blue line indicated in the upward direction
  • 734 nm downward Chromatogram obtained by detecting wavelengths of red lines indicated by?
  • Clarified sprout extract prepared in Example 1 and Clarified sprout extract grown under various light conditions prepared in Example 2 (STS-C, STS-385, STS-465, STS-645 and STS-780) was dissolved in an aqueous alcohol solution, and subjected to ABTS online antioxidant high performance liquid chromatography (HPLC) according to the same analysis conditions as in Test Example 1.
  • HPLC high performance liquid chromatography
  • A, B, C, D, E, F of Figure 3 are ABTS online antioxidants of the six species of Sprout bud extracts STS, STS-C, STS-385, STS-465, STS-645 and STS-780, respectively.
  • the degree of antioxidant activity was determined by the area of the peaks and peaks of the UV detector 254 nm (showing peaks of the components absorbing UV 254 nm) and 734 nm wavelengths (antioxidative action against ABTS). It is a chromatogram obtained by detecting each).
  • Test Example 3 Example 7 kinds of 1 and 2 Fault Of sprout extract Retinal progenitor cells (R28) Protective effect
  • R28 cells at 37 ° C. in Dulbecco's modified eagle's medium (DMEM) / low glucose medium containing 10% fetal bovine serum (FBS), 100 U / mL penicillin, and 100 ⁇ g / mL streptomycin. After incubation under 5% CO 2 conditions in an incubator, passaged with 0.05% trypsin every 2 days, inoculated at a density of 1 ⁇ 10 4 in 96 plates and cultured for 24 hours. The 7 extracts were treated to the cultured R28 cells at the concentrations of (A) to (C) of FIG.
  • DMEM Dulbecco's modified eagle's medium
  • FBS fetal bovine serum
  • FIG 5 (A) is a light extract of Example 2, the extract of the crystallized shooter (ST) of the Comparative Example 1, ST extract of Example 1, at three concentrations from the oxidative stress caused by glutamate This is the result of the cell survival rate of the R28 cell protection effect of the five species of sprout sprouts cultivated differently (STS-C, STS-385, STS-465, STS-645, STS-780).
  • Figure 5 (B) and (C) are R28 cells of the clarifier extract (ST) of the Comparative Example 1 and the sprout bud extract (STS) of Example 1 from the oxidative stress caused by glutamate at each of five concentrations, respectively Protective efficacy is shown as cell viability.
  • Example 5 and 6 Fault Of compounds 1 to 5 isolated from sprout extract Retinal progenitor cells (R28) Confirmation of protective efficacy
  • Compounds 1 to 5 exhibited an effect of protecting R28 cells from neurotoxicity induced by glutamate, and the compounds have a relatively high content in the Sprout Sprout Extract (STS) or the Sprout Sprout Extract. Since the compounds are mainly present in (STS), it was confirmed that the R28 cell protective effect of the shoot sprout extract (STS) is superior to that of the shooter extract (ST).
  • Test Example 5 Example 7 kinds of 1 and 2 Fault Protective Effect of Sprout Extract on Hippocampal Neuronal Cells (HT-22)
  • HT-22 cells are 10% fetal bovine serum (FBS), 100 U / mL penicillin, and 100 ⁇ g / mL streptomycin containing DMEM (Dulbecco's modified eagle's medium), 5% in a 37.5 ° C incubator in low glucose medium. Incubated under CO 2 conditions, subcultured with 0.05% trypsin every 2 days, inoculated at a density of 3 ⁇ 10 3 in 96 plates and cultured for 24 hours. The 7 extracts were treated to the cultured HT-22 cells at the concentrations of (A) to (C) of FIG. 7, and 2 hours later, 5 mM glutamate was added and cultured for 22 hours. Then, 10 ⁇ L EZ-cytox was treated and maintained for 2 hours to measure cell viability, and UV absorbance was measured at 450 nm.
  • FBS fetal bovine serum
  • penicillin 100 U / mL penicillin
  • streptomycin containing DMEM Dulbecco
  • Figure 7 (A) is the light extract of Example 2, the extract of the glenja extract of Comparative Example 1 (ST), the shoot sprout of Example 1 (STS) at three concentrations from the oxidative stress caused by glutamate HT-22 cell protection effect of the five species of sprout sprouts (STS-C, STS-385, STS-465, STS-645, STS-780) cultivated differently to the cell survival rate.
  • Figure 7 (B) and (C) is HT-22 of the deficiency extract (ST) of the Comparative Example 1 and the deficiency sprout extract (STS) of Example 1 from the oxidative stress caused by glutamate at five concentrations It is the result of cell protection effect by cell viability.
  • HT-22 cell viability was measured in the same manner as in Test Example 5, Comparative Example 1, Example 1, Example 2 extract gyeongjab (ST), the shoot sprout extract (STS, STS-C, STS-385, STS-465 , STS-645 and STS-780) instead of compounds 1 to 5 of Examples 5 and 6 were treated with 50 ⁇ M, 16.6 ⁇ M, 5.55 ⁇ M, and the results are shown in FIG. 8.
  • Table 1 The physiological activity of each component evaluated in Test Example 1, Test Example 4 and Test Example 6 is shown in Table 1 below.
  • Table 1 below relates to the physiological activity of the components isolated from the bud extract (STS) of Example 1.

Abstract

The present invention relates to a naphthopyrone derivative and a composition comprising, as an active ingredient, a Cassia obtusifolia shoot extract including the naphthopyrone derivative. A composition according to an aspect of the present invention has an antioxidative effect and an effect of protecting neurons from oxidative stress or inhibiting the death of neurons, and in particular, has an effect of inhibiting damage or death of retinal neurons or hippocampal neurons due to glutamate toxicity, and thus the composition can be used as a pharmaceutical or food composition for the treatment or prevention of eye diseases and deterioration and loss of vision caused by optic nerve injury, and for the treatment or prevention memory loss, decline in learning ability, onset and worsening of depressive disorders, and degenerative neurological diseases caused by brain nerve damage.

Description

결명 새싹 유래 나프토파이론 유도체를 포함하는 신경세포 보호용 조성물Neuroprotective composition containing naphthopyrone derivative derived from bud
본 명세서는 나프토파이론 유도체 및 이를 포함하는 결명 새싹 추출물을 유효성분으로 포함하는 조성물에 관한 것이다.The present specification relates to a composition comprising a naphthopyrone derivative and a shoot sprout extract comprising the same as an active ingredient.
뇌 해마(hippocampus)는 뇌 내측 측두엽의 한 구조체로써, 인간과 동물의 인지적 기능에 중요한 역할을 한다. 해마는 생리적/산화적 스트레스 자극에 취약하고 스트레스로 인한 인지기능의 손상을 일으키는데 중심 조직으로 알려져 있다. 스트레스는 해마의 구조와 뇌세포 생성, 시냅스 가소성 및 해마와 관련된 행동학적 변화를 일으킬 수 있다(김은주, 한국심리학회지: 인지 및 생물, 2012, 24, 65-88). 신경세포에서의 산화적 스트레스가 해마, 뇌하수체, 선조체, 흑질, 전뇌피질 또는 시상하부에서 유발되면 신경세포사멸이 증가하고 뉴런 및 성장인자를 감소시켜 근위축성 측삭 경화증(루게릭병), 파킨슨병 또는 뇌허혈성 신경손상 등의 급성 또는 만성 신경질환을 초래하는 것으로 알려져 있다(Rahman, T. et al. Adv. Biosci. Biotechnol. 2012, 3, 997-1019).The hippocampus is a structure of the medial temporal lobe of the brain and plays an important role in the cognitive function of humans and animals. The hippocampus is known to be vulnerable to physiological and oxidative stress stimuli and to be a central tissue for cognitive impairment caused by stress. Stress can cause hippocampal structure and brain cell production, synaptic plasticity, and behavioral changes related to the hippocampus (Eunjoo Kim, Journal of the Korean Psychological Association: Cognitive and Biology, 2012, 24, 65-88). When oxidative stress in neurons is induced in the hippocampus, pituitary gland, striatum, black matter, the whole cortex, or the hypothalamus, neuronal cell death increases and neurons and growth factors decrease, resulting in amyotrophic lateral sclerosis (ALS), Parkinson's disease, or brain It is known to cause acute or chronic neurological diseases such as ischemic nerve damage (Rahman, T. et al. Adv. Biosci. Biotechnol. 2012, 3, 997-1019).
국민건강보험공단 건강보험 자료에 의하면 망막질환 환자는 2010년 약 83만명에서 2015년에 약 125만명으로 연평균 약 8% 증가해 왔다. 망막은 카메라의 필름에 비유되는 눈 내부의 얇은 신경막이다. 망막에는 일억개 이상의 광수용체 세포(빛 감지 세포) 및 백만개 이상의 시신경 세포가 존재하여 빛을 전기적 신호로 바꾸고 신경을 통해 눈으로 보는 사물의 상을 뇌로 전달한다. 이러한 망막의 신경이 손상되거나 신경 기능에 이상이 생기면 시력과 시야에 문제가 생긴다. 대표적인 망막질환은 시력 저하, 시야 방해 및 실명을 포함하여 당뇨병성 망막병증, 연령 관련 황반변성, 망막색소변성, 망막박리 및 망막혈관폐쇄 등이 있다. 녹내장은 망막의 시신경 손상이 발생하는 대표적인 안질환으로, 안압상승, 허혈 및 산화스트레스 등의 이유로 시신경이 손상된다(Kim, N. Y. et al. J. Korean Opthalmol. Soc. 2015, 56, 70-79; Kang, J. H. et al. J. Korean Ophthalmol. Soc. 2003, 44, 965-970; Lee, S. M. et al. J. Korean Ophthalmol. Soc. 2002, 43, 2577-2584).According to the National Health Insurance Corporation's health insurance data, the number of patients with retinal disease has increased by about 8% per year from about 830,000 in 2010 to about 12.5 million in 2015. The retina is the thinner nerve film inside the eye that is likened to the film of a camera. Over 100 million photoreceptor cells (light-sensing cells) and over 1 million optic nerve cells are present in the retina, converting light into electrical signals and transmitting the image of the object through the nerve to the brain. If the nerves in the retina are damaged or nerve function is abnormal, vision and vision problems may occur. Representative retinal diseases include diabetic retinopathy, age-related macular degeneration, retinal pigmentary degeneration, retinal detachment and retinal vascular obstruction, including decreased vision, blurred vision and blindness. Glaucoma is a typical eye disease in which retinal optic nerve damage occurs, and the optic nerve is damaged due to increased intraocular pressure, ischemia and oxidative stress (Kim, NY et al. J. Korean Opthalmol. Soc. 2015, 56, 70-79; Kang, JH et al. J. Korean Ophthalmol. Soc. 2003, 44, 965-970; Lee, SM et al. J. Korean Ophthalmol. Soc. 2002, 43, 2577-2584).
글루타메이트는 척추동물의 중추신경계에서 중요한 역할을 하는 흥분성 신경전달물질이다. 글루타메이트는 뇌 조직 중에서 기억력과 학습능력에 관여한다고 알려진 소뇌편도와 해마에 집중적으로 분포되어 있는 quisqualate 수용체, NMDA (N-methyl-D-aspartate) 수용체 및 Kainite 수용체에 작용하여 다양한 생리작용 발현에 관여한다. 반면, 글루타메이트가 신경세포 내에서 밖으로 유리가 증가되어 세포 밖의 글루타메이트 농도가 급속히 증가하게 되면 산화적 신경독성 물질로 작용할 수 있다(김현정 등, J. Life Sci. 2009, 19, 963-967). 과도한 글루타메이트(Glutamate)는 빈혈, 산소결핍, 저혈당, 트라우마, 여러 만성 퇴행성 신경질환을 포함한 다양한 급성 신경학적 장애와 관련되어 있다고 알려져 있다. 눈에서 글루타메이트는 망막 신경절세포의 급성적인 장애와 사멸에 관련되어 있다(Otori, Y. et al. Invest. Ophthalmol. Vis. Sci. 1998, 39, 972-981). 글루타메이트가 다량 생성되어 시냅스전(presynaptic) 글루타메이트 수용체가 활성화되면서 활성산소종 (Reactive Oxygen Species, ROS)의 과생성이 자극되는 것으로 알려져 있다 (Tarasenko A. et al. Neurochem. Int. 2012, 61, 1044-1051). 또한, 글루타메이트에 반응하여 발휘되는 산화작용은 NMDA 수용체의 활성화에 의해 매개되며, 칼슘이온 의존성을 나타내는 것으로 알려져 있다. 이러한 과도한 NMDA 수용체 활성화와 칼슘이온의 미토콘드리아로의 흡수(uptake)는 ROS 생성으로 이어지는 것으로 보고되었다(Reynolds I. J. et al. J. Neurosci. 1995, 15, 3318-3327). 활성 산소종(Reactive Oxygen Species, ROS)은 생물체 세포 내의 미토콘드리아에서 호흡과 면역 반응에 의한 산소의 산화·환원을 통해 지속적으로 생성된다. ROS는 화학적으로 불안정하고 반응성이 높아 생체내의 지질, 핵산, 단백질 등과 반응하여 DNA 손상, 세포내 유리 칼슘과 철 농도 증가, 생체막의 이온 수송계 손상 등을 일으킬 수 있다(Kiselyov, K. et al. Cell Calcium. 2016, 60, 108-114). 또한, ROS는 빛에 의해 생성이 유도되기도 하여 시신경의 광산화 손상을 유도하는 것으로 알려져 있다(Masuda, T. et al. Oxid. Med. Cell. Longevity. 2017; article ID 9208489, 14 pages).Glutamate is an excitatory neurotransmitter that plays an important role in the central nervous system of vertebrates. Glutamate is involved in the expression of various physiological functions by acting on quisqualate receptors, NMDA (N-methyl-D-aspartate) receptors, and Kainite receptors, which are concentrated in the cerebellar amygdala and hippocampus, which are known to be involved in memory and learning in brain tissues. . On the other hand, when glutamate increases in the outside of neurons and the concentration of glutamate outside the cell increases rapidly, it can act as an oxidative neurotoxic substance (Hyun-Jeong Kim et al., J. Life Sci. 2009, 19, 963-967). Excessive glutamate is known to be associated with various acute neurological disorders, including anemia, oxygen deficiency, hypoglycemia, trauma, and several chronic degenerative neurological disorders. Glutamate in the eye has been implicated in acute disorders and death of retinal ganglion cells (Otori, Y. et al. Invest. Ophthalmol. Vis. Sci. 1998, 39, 972-981). It is known that overproduction of reactive oxygen species (ROS) is stimulated by the production of a large amount of glutamate and presynaptic glutamate receptor activation (Tarasenko A. et al. Neurochem. Int. 2012, 61, 1044 -1051). In addition, oxidative action exerted in response to glutamate is mediated by activation of the NMDA receptor, and is known to exhibit calcium ion dependence. This excessive NMDA receptor activation and uptake of calcium ions into the mitochondria have been reported to lead to ROS production (Reynolds I. J. et al. J. Neurosci. 1995, 15, 3318-3327). Reactive Oxygen Species (ROS) are continuously produced in the mitochondria in living cells through the oxidation and reduction of oxygen by respiratory and immune responses. Since ROS is chemically unstable and highly reactive, it can react with lipids, nucleic acids, and proteins in vivo to cause DNA damage, increase the concentration of free calcium and iron in the cell, and damage the ion transport system of the biofilm (Kiselyov, K. et al. Cell Calcium. 2016, 60, 108-114). In addition, ROS is also known to induce light-induced damage to the optic nerve by generating light (Masuda, T. et al. Oxid.Med. Cell. Longevity. 2017; article ID 9208489, 14 pages).
결명자는 전통적으로 눈 건강을 위해 활용되어 왔으며 우수한 항산화 작용을 가진다고 알려졌다. 결명자는 콩과에 속하는 초본인 결명(Cassia obtusifolia L.) 또는 긴강남차 (Cassia tora L.)의 성숙한 종자로서 우리나라 각지에서 재배되며, 잎이 진 뒤에 약 10 cm정도 되는 활모양의 꼬투리 속에 윤기가 나는 종자가 한 줄로 들어 있다(Yen, G.-C. et al. J. Agric. Food Chem. 1998, 46, 820-824). 결명자 추출물은 당뇨병 개선, 이상지질혈증 개선, 간보호, 항균, 혈압저하 효능이 연구·보고되었다(Dong, X. et al. Mol. Med. Rep. 2017, 16, 2331-2346).The deficiency has traditionally been used for eye health and is known to have excellent antioxidant activity. It is a mature seed of the herbaceous plant belonging to the legume (Cassia obtusifolia L.) or Ginjiang Namcha (Cassia tora L.). It is cultivated in all parts of Korea and glossed in a bow-shaped pod that is about 10 cm after the leaves are cut. Seeds containing a single line (Yen, G.-C. et al. J. Agric. Food Chem. 1998, 46, 820-824). The extract of terminator has been studied and reported for improving diabetes, dyslipidemia, liver protection, antibacterial, and hypotensive effects (Dong, X. et al. Mol. Med. Rep. 2017, 16, 2331-2346).
그러나, 결명 새싹 추출물 또는 이로부터 유래한 나프로파이론 유도체인 7-하이드록시무시지닐-루브로푸사린-8'-O-글루코사이드 (7-hydroxymusizinyl-rubrofusarin-8'-O-glucopyranoside), 이소토라락톤 (isotoralactone), 토라락톤 (toralactone), 토로사크리손 (torosachrysone) 및 루브로푸사린 (rubrofusarin)의 글루타메이트 유도 신경독성에 대한 신경보호작용은 보고된 바 없으며, 결명 새싹 추출물을 활용한 항산화 및 신경보호용 조성물에 대한 기술은 알려진 바 없다.However, 7-hydroxymusinginyl-rubrofusarin-8'-O-glucoside, isotora, which is a bud extract or napropyrone derivative derived therefrom No neuroprotective action of glutamate-induced neurotoxicity of isotoralactone, toralactone, torosachrysone and torochrysone and rubrofusarin has been reported. There is no known technique for protective compositions.
또한, 결명 새싹 추출물을 활용한 건강 기능성 및 약효성분 연구개발 사례는 보고된 바 없는바, 본 발명자들은 결명 새싹 추출물이 결명자 추출물에 비해 새로운 항산화 성분이 생성되어 항산화 활성 성분이 증가되고 이를 통해 현저히 증가된 항산화 작용과 신경세포 보호 작용을 한다는 사실을 본 발명자들이 확인하였으며, 결명 새싹 추출물로부터 분리한 신규 화합물 7-하이드록시무시지닐-루브로푸사린-8'-O-글루코사이드를 포함한 나프토파이론 성분들이 글루타메이트 유도 산화적 스트레스로부터 망막신경세포 및 해마신경세포를 보호하는 효능을 나타낸다는 사실을 확인함으로써 본 발명을 완성하였다.In addition, there have been no cases of research and development of health functional and medicinal ingredients using the shoot sprout extract, and the present inventors found that the shoot sprout extract produced a new antioxidant component compared to the extract from the shooter, increasing the antioxidant active ingredient and significantly increasing it. The present inventors have confirmed that the antioxidant activity and neuronal cell protective action, and naphtopyron component including the novel compound 7-hydroxymusininyl- lubrofusarin-8'-O-glucoside isolated from the shoot sprout extract The present invention has been completed by confirming that they exhibit the effect of protecting retinal neurons and hippocampal neurons from glutamate induced oxidative stress.
[선행기술문헌][Advanced technical literature]
[특허문헌][Patent Document]
(특허문헌 1) KR 10-1503429 B1(Patent Document 1) KR 10-1503429 B1
(특허문헌 2) KR 10-2010-0082054 A(Patent Document 2) KR 10-2010-0082054 A
(특허문헌 3) KR 10-0877371 B1(Patent Document 3) KR 10-0877371 B1
(특허문헌 4) KR 10-1807367 B1(Patent Document 4) KR 10-1807367 B1
(특허문헌 5) KR 10-2016-0058613 A(Patent Document 5) KR 10-2016-0058613 A
[비특허문헌][Non-patent literature]
(비특허문헌 1) Jung, H. A. Journal of Ethnopharmacology, 2016, vol 191, 152-160; Inhibitory activities of major anthraquinones and other constituents from Cassia obtusifolia against β-secretase and cholinesterases. (Non-Patent Document 1) Jung, H. A. Journal of Ethnopharmacology, 2016, vol 191, 152-160; Inhibitory activities of major anthraquinones and other constituents from Cassia obtusifolia against β-secretase and cholinesterases.
(비특허문헌 2) Shrestha, S. et al. Archives Pharmacal Research, 2018, online publication https://doi.org/10.1007/s12272-018-1044-0; Two new naphthalenic lactone glycosides from Cassia obtusifolia L. seeds. (Non-patent document 2) Shrestha, S. et al. Archives Pharmacal Research, 2018, online publication https://doi.org/10.1007/s12272-018-1044-0; Two new naphthalenic lactone glycosides from Cassia obtusifolia L. seeds.
본 명세서의 목적은 하기 화학식 1의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물을 제공하는 것이다.It is an object of the present specification to provide a naphthopyrone derivative of Formula 1, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof.
[화학식 1][Formula 1]
Figure PCTKR2019000399-appb-img-000001
Figure PCTKR2019000399-appb-img-000001
본 명세서의 다른 목적은 산화적 스트레스로부터의 신경세포 손상을 보호하거나 신경세포 사멸을 억제하는 효과를 나타내는 조성물을 제공하는 것이다. Another object of the present specification is to provide a composition that exhibits an effect of protecting neuronal cell damage from oxidative stress or inhibiting neuronal cell death.
본 명세서의 또 다른 목적은 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료 효과를 나타내는 조성물을 제공하는 것이다. Still another object of the present specification is to provide a composition that exhibits a prophylactic or therapeutic effect of a neurological damaging disease caused by damage or death of retinal nerve cells or hippocampal nerve cells.
본 명세서의 또 다른 목적은 항산화 효과를 나타내는 조성물을 제공하는 것이다. Another object of the present specification is to provide a composition that exhibits an antioxidant effect.
상기 목적을 달성하기 위하여 본 발명은 일 측면에 있어서, 하기 화학식 1의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물을 제공한다.In order to achieve the above object, in one aspect, the present invention provides a naphthopyrone derivative of Formula 1, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof.
[화학식 1][Formula 1]
Figure PCTKR2019000399-appb-img-000002
Figure PCTKR2019000399-appb-img-000002
다른 측면에 있어서, 본 발명은 하기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물의 제조방법을 제공한다.In another aspect, the present invention provides one or more naphthopyrone derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvents thereof selected from the group consisting of naphthopyrone derivatives of the following Chemical Formulas 1 to 5 Provides a method of manufacturing the cargo.
[화학식 1][Formula 1]
Figure PCTKR2019000399-appb-img-000003
Figure PCTKR2019000399-appb-img-000003
[화학식 2][Formula 2]
Figure PCTKR2019000399-appb-img-000004
Figure PCTKR2019000399-appb-img-000004
[화학식 3][Formula 3]
Figure PCTKR2019000399-appb-img-000005
Figure PCTKR2019000399-appb-img-000005
[화학식 4][Formula 4]
Figure PCTKR2019000399-appb-img-000006
Figure PCTKR2019000399-appb-img-000006
[화학식 5][Formula 5]
Figure PCTKR2019000399-appb-img-000007
Figure PCTKR2019000399-appb-img-000007
또 다른 측면에 있어서, 본 발명은 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 산화적 스트레스로부터의 신경세포 보호용 또는 신경세포 사멸 억제용 조성물을 제공한다.In another aspect, the present invention is one or more naphthopyron derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or the like selected from the group consisting of naphthopyrone derivatives of Formula 1 to Formula 5. At least one selected from the group consisting of solvates, Cassia obtusifolia L. or Cassia tora L. Sprout extract, or a deficiency sprout fraction comprising the same for the protection of neurons from oxidative stress, comprising as an active ingredient Or it provides a composition for inhibiting neuronal cell death.
또 다른 측면에 있어서, 본 발명은 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료용 조성물을 제공한다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or One or more selected from the group consisting of solvates, Cassia obtusifolia L. or Cassia tora L. bud extract, or a fraction of the bud containing it as an active ingredient, comprising retinal nerve cells or hippocampal nerve cells Provided is a composition for the prevention or treatment of a neuro-damaging disease caused by damage or death.
또 다른 측면에 있어서, 본 발명은 상기 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 항산화용 조성물을 제공한다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or It provides at least one selected from the group consisting of solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extract, or a bud fraction containing the same as an active ingredient, provides an antioxidant composition.
본 발명의 일 측면에 따른 조성물은 항산화 효과, 및 산화적 스트레스로부터 신경세포를 보호하거나 신경세포 사멸을 억제하는 효과가 있으며, 특히, 글루타메이트 독성에 의한 망막신경세포 또는 해마신경세포의 손상 또는 사멸을 억제하는 효과가 있다. 따라서, 본 발명의 일 측면에 따른 조성물은 망막신경세포를 보호하여 시신경 손상에 따른 시력저하와 감퇴 및 안질환 치료 또는 예방에 사용될 수 있으며, 해마신경세포를 보호하여 뇌신경 손상에 따른 기억력 감퇴, 학습능력 감퇴, 우울장애 발생 및 악화와 퇴행성 뇌신경 질환의 치료 또는 예방에 사용될 수 있고, 나아가 기억력과 학습능력 개선, 스트레스 완화 등의 정신건강과 시신경 보호와 시력 저하 개선 등의 눈 건강을 위해 사용될 수 있는바, 약학 또는 식품 조성물로서 사용될 수 있다. The composition according to an aspect of the present invention has an antioxidant effect and an effect of protecting nerve cells from oxidative stress or inhibiting apoptosis, and in particular, damage or death of retinal nerve cells or hippocampal nerve cells due to glutamate toxicity. It has an inhibitory effect. Therefore, the composition according to one aspect of the present invention can be used for treating or preventing vision loss and decline and eye diseases caused by optic nerve damage by protecting retinal nerve cells, and memory loss and learning due to brain nerve damage by protecting hippocampal nerve cells. It can be used for the treatment or prevention of deterioration, development and deterioration of depressive disorders and neurodegenerative diseases, and also for eye health such as improvement of memory and learning ability, stress relief, and protection of optic nerve and decreased vision. It can be used as a bar, pharmaceutical or food composition.
도 1는 결명자 추출물(ST)과 결명 새싹 추출물(STS)의 DPPH(α,α-diphenyl-β-picrylhydrazyl) 자유라디컬 소거 활성을 나타낸 도이다. 1 is a diagram showing the free radical scavenging activity of DPPH (α, α-diphenyl-β-picrylhydrazyl) of the Cassia tora extract (ST) and Cassia tora sprout extract (STS).
도 2은 결명자 추출물(ST)와 결명 새싹 추출물(STS)의 ABTS 온라인 항산화 HPLC 크로마토그램을 나타낸 도이다. 윗쪽 방향으로 표기된 파란색 선의 크로마토그램은 자외선(UV) 254 nm 에서 검출되는 성분들을 나타낸 도이고, 아랫쪽 방향으로 표기된 붉은색 선의 크로마토그램은 ABTS 시약과 반응하여 UV/VIS(자외선/가시광선) 734 nm에서 검출되는 항산화 성분들을 나타낸 도이다.FIG. 2 is a diagram showing ABTS online antioxidant HPLC chromatograms of Cassia tora extract (ST) and Cassia tora sprout extract (STS). The chromatogram of the blue line indicated in the upward direction is a diagram showing the components detected at 254 nm of ultraviolet (UV), and the chromatogram of the red line indicated in the downward direction reacts with the ABTS reagent to UV / VIS (ultraviolet / visible light) 734 nm It is a diagram showing the antioxidant components detected in.
도 3은 다양한 빛 조건 하에서 재배된 결명 새싹 추출물(STS-C, STS-385, STS-465, STS-645 및 STS-780)의 ABTS 온라인 항산화 HPLC 크로마토그램을 나타낸 도이다. (A) 내지 (F)의 각 크로마토그램에서 윗쪽 방향으로 표기된 크로마토그램은 자외선(UV) 254 nm 에서 검출되는 성분들을 나타낸 도이고, 아랫쪽 방향으로 표기된 크로마토그램은 ABTS 시약과 반응하여 UV/VIS(자외선/가시광선) 734 nm에서 검출되는 항산화 성분들을 나타낸 도이다. (A)의 시료는 차광 조건에서 2주 동안 재배한 결명 새싹 추출물(실시예 1의 STS), (B)의 시료는 일반 빛 조건에서 2주 동안 재배한 결명 새싹 추출물(실시예 2의 STS-C), (C)의 시료는 385 nm LED 조명하에서 2주 동안 재배한 결명 새싹 추출물(실시예 2의 STS-385), (D)의 시료는 465 nm LED 조명하에서 2주 동안 재배한 결명 새싹 추출물(실시예 2의 STS-465), (E)의 시료는 645 nm LED 조명하에서 2주 동안 재배한 결명 새싹 추출물(실시예 2의 STS-645), (F)의 시료는 780 nm LED 조명하에서 2주 동안 재배한 결명 새싹 추출물(실시예 2의 STS-780)이다. FIG. 3 is a diagram showing ABTS online antioxidant HPLC chromatograms of bearer bud extracts (STS-C, STS-385, STS-465, STS-645 and STS-780) grown under various light conditions. In each chromatogram of (A) to (F), the chromatogram indicated in the upward direction is a diagram showing the components detected at ultraviolet (UV) 254 nm, and the chromatogram indicated in the downward direction reacts with ABTS reagent to react with UV / VIS ( UV / Visible light) It is a diagram showing antioxidant components detected at 734 nm. The sample of (A) is a seedling sprout extract grown for 2 weeks under light-shielding conditions (STS of Example 1), and the sample of (B) is a seedling sprout extract grown for 2 weeks under normal light conditions (STS of Example 2) The samples of C) and (C) were cultivated for 2 weeks under 385 nm LED illumination (STS-385 of Example 2), and the samples of (D) were cultivated for 2 weeks under 465 nm LED illumination The extract (STS-465 of Example 2), the sample of (E) was cultivated for 2 weeks under 645 nm LED illumination, and the bud extract (STS-645 of Example 2), the sample of (F) was 780 nm LED illumination It was a cultivated shoot extract (STS-780 of Example 2) grown for 2 weeks under.
도 4는 본 발명의 일 실시예인 실시예 1의 결명 새싹 추출물(STS)로부터 분리된 주요 성분들의 피크들을 표기한 HPLC 크로마토그램이다. Figure 4 is an HPLC chromatogram showing the peaks of the main components separated from the bud extract (STS) of Example 1, which is an embodiment of the present invention.
도 5는 본 발명의 비교예 1인 결명자 추출물(ST), 차광 조건에서 재배한 결명 새싹 추출물(실시예 1의 STS) 및 다양한 빛 조건하에서 재배된 결명 새싹 추출물들(실시예 2의 STS, STS-C, STS-385, STS-465, STS-645 및 STS-780)이 글루타메이트(glutamate) 독성에 의해 유발된 망막전구세포(R28) 손상을 보호하는 효능을 나타낸 그래프이다. Figure 5 is a comparative example 1 of the present invention extract (ST), cultivated bud extracts grown in light-shielding conditions (STS of Example 1) and cultivated bud extracts grown under various light conditions (STS, STS of Example 2) -C, STS-385, STS-465, STS-645 and STS-780) are graphs showing the efficacy of protecting damage to retinal progenitor cells (R28) caused by glutamate toxicity.
도 6은 결명 새싹 추출물(STS)로부터 분리한 각 화합물(실시예 4 및 5의 화합물 1 내지 화합물 5, 및 비교예 2의 화합물 A 내지 화합물 X)의 농도별 글루타메이트 독성에 의해 유발된 망막전구세포(R28) 손상을 보호하는 효능을 나타낸 그래프이다. Figure 6 is a retinal progenitor cell induced by glutamate toxicity by concentration of each compound (Compounds 1 to 5 of Examples 4 and 5, and Compounds A to Compound X of Comparative Example 2) isolated from the seedling sprout extract (STS) (R28) It is a graph showing the efficacy of protecting damage.
도 7은 본 발명의 비교예 1인 결명자 추출물(ST), 차광 조건에서 재배한 결명 새싹 추출물(실시예 1의 STS) 및 다양한 빛 조건하에서 재배된 결명 새싹 추출물들(실시예 2의 STS, STS-C, STS-385, STS-465, STS-645 및 STS-780)이 글루타메이트(glutamate) 독성에 의해 유발된 해마신경세포(HT-22) 손상을 보호하는 효능을 나타낸 그래프이다. Figure 7 is a comparative example 1 of the present invention extracts (ST), the seedling sprout extract grown under light-shielding conditions (STS of Example 1) and the seedling sprout extracts grown under various light conditions (STS, STS of Example 2) -C, STS-385, STS-465, STS-645 and STS-780) are graphs showing the efficacy of protecting hippocampal nerve cell (HT-22) damage caused by glutamate toxicity.
도 8은 결명 새싹 추출물(STS)로부터 분리한 각 화합물(실시예 4 및 5의 화합물 1 내지 화합물 5, 및 비교예 2의 화합물 A 내지 화합물 X)의 농도별 글루타메이트 독성에 의해 유발된 해마신경세포(HT-22) 손상을 보호하는 효능을 나타낸 그래프이다. 8 is a hippocampal neuron induced by glutamate toxicity by concentration of each compound (Compounds 1 to 5 of Examples 4 and 5, and Compounds A to Compound X of Comparative Example 2) isolated from the seedling sprout extract (STS). (HT-22) It is a graph showing the efficacy of protecting damage.
본 발명은 일 측면에 있어서, 하기 화학식 1의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물에 관한 것일 수 있다.In one aspect, the present invention may relate to a naphthopyrone derivative of Formula 1, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof.
[화학식 1][Formula 1]
Figure PCTKR2019000399-appb-img-000008
Figure PCTKR2019000399-appb-img-000008
상기 화학식 1 의 나프토파이론 유도체는 7-하이드록시무시지닐-루브로푸사린-8'-O-글루코사이드(7-hydroxymusizinyl-rubrofusarin-8'-O-glucopyranoside)일 수 있다.The naphthopyrone derivative of Formula 1 may be 7-hydroxymusizinyl-rubrofusarin-8'-O-glucoside (7-hydroxymusizinyl-rubrofusarin-8'-O-glucopyranoside).
본 명세서에서 "약학적으로 허용 가능"이란 통상의 의약적 복용량(medicinal dosage)으로 이용할 때 상당한 독성 효과를 피함으로써, 동물, 더 구체적으로는 인간에게 사용할 수 있다는 정부 또는 이에 준하는 규제 기관의 승인을 받을 수 있거나 승인 받거나, 또는 약전에 열거되거나 기타 일반적인 약전에 기재된 것으로 인지되는 것을 의미한다.As used herein, "pharmaceutically acceptable" means the approval of a government or equivalent regulatory body to use in animals, more specifically in humans, by avoiding significant toxic effects when used in conventional medical dosages. It is meant to be recognized or approved, or recognized as listed in a pharmacopeia or other general pharmacopeia.
본 명세서에서 "약학적으로 허용 가능한 염"은 약학적으로 허용 가능하고 모 화합물(parent compound)의 바람직한 약리 활성을 갖는 본 발명의 일측면에 따른 염을 의미한다. 상기 염은 (1) 염산, 브롬화수소산, 황산, 질산, 인산 등과 같은 무기산으로 형성되거나; 또는 아세트산, 프로파이온산, 헥사노산, 시클로펜테인프로피온산, 글라이콜산, 피루브산, 락트산, 말론산, 숙신산, 말산, 말레산, 푸마르산, 타르타르산, 시트르산, 벤조산, 3-(4-히드록시벤조일) 벤조산, 신남산, 만델산, 메테인설폰산, 에테인설폰산, 1,2-에테인-디설폰산, 2-히드록시에테인설폰산, 벤젠설폰산, 4-클로로벤젠설폰산, 2-나프탈렌설폰산, 4-톨루엔설폰산, 캄퍼설폰산, 4-메틸바이시클로 [2,2,2]-oct-2-엔-1-카르복실산, 글루코헵톤산, 3-페닐프로파이온산, 트리메틸아세트산, tert-부틸아세트산, 라우릴 황산, 글루콘산, 글루탐산, 히드록시나프토산, 살리실산, 스테아르산, 뮤콘산과 같은 유기산으로 형성되는 산 부가염(acid addition salt); 또는 (2) 모 화합물에 존재하는 산성 프로톤이 치환될 때 형성되는 염을 포함할 수 있다.As used herein, "pharmaceutically acceptable salts" means salts according to one aspect of the invention that are pharmaceutically acceptable and have the desired pharmacological activity of the parent compound. The salt is formed from (1) an inorganic acid such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, or the like; Or acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3- (4-hydroxybenzoyl) Benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethane-disulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo [2,2,2] -oct-2-ene-1-carboxylic acid, glucoheptonic acid, 3-phenylpropionic acid, trimethylacetic acid, tert Acid addition salts formed with organic acids such as butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid; Or (2) salts formed when the acidic protons present in the parent compound are substituted.
본 명세서에서 "이성질체"는 특히 광학 이성질체(optical isomers)(예를 들면, 본래 순수한 거울상 이성질체(essentially pure enantiomers), 본래 순수한 부분 입체 이성질체(essentially pure diastereomers) 또는 이들의 혼합물)뿐만 아니라, 형태 이성질체(conformation isomers)(즉, 하나 이상의 화학 결합의 그 각도만 다른 이성질체), 위치 이성질체(position isomers)(특히, 호변이성체(tautomers)) 또는 기하 이성질체(geometric isomers)(예컨대, 시스-트랜스 이성질체)를 포함한다.“Isomers” as used herein, in particular optical isomers (eg, essentially pure enantiomers, essentially pure diastereomers or mixtures thereof), as well as conformational isomers ( conformation isomers (i.e., isomers differing only in that angle of one or more chemical bonds), positional isomers (especially tautomers) or geometric isomers (e.g., cis-trans isomers) do.
본 명세서에서 "본래 순수(essentially pure)"란, 예컨대 거울상 이성질체 또는 부분 이성질체와 관련하여 사용한 경우, 거울상 이성질체 또는 부분 이성질체를 예로 들 수 있는 구체적인 화합물이 약 90% 이상, 바람직하게는 약 95% 이상, 보다 바람직하게는 약 97% 이상 또는 약 98% 이상, 보다 더 바람직하게는 약 99% 이상, 보다 더욱 더 바람직하게는 약 99.5% 이상(w/w) 존재하는 것을 의미한다.As used herein, “essentially pure”, when used in connection with, for example, enantiomers or diastereomers, contains at least about 90%, preferably at least about 95% of specific compounds that may exemplify enantiomers or diastereomers. , More preferably at least about 97% or at least about 98%, even more preferably at least about 99%, even more preferably at least about 99.5% (w / w).
본 명세서에서 "수화물(hydrate)"은 물이 결합되어 있는 화합물을 의미하며, 물과 화합물 사이에 화학적인 결합력이 없는 내포 화합물을 포함하는 광범위한 개념이다.As used herein, "hydrate" refers to a compound to which water is bound, and is a broad concept including an inclusion compound having no chemical bonding force between water and the compound.
본 명세서에서 "용매화물"은 용질의 분자나 이온과 용매의 분자나 이온 사이에 생긴 고차의 화합물을 의미한다.As used herein, "solvate" refers to a higher order compound formed between molecules or ions of a solute and molecules or ions of a solvent.
본 발명은 일 측면에 있어서, 하기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물의 제조방법으로서, 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹(sprout)으로부터 상기 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상을 분리하는 과정을 포함하는, 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물의 제조방법에 관한 것일 수 있다.In one aspect, the present invention provides one or more naphthopyrone derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvents thereof selected from the group consisting of naphthopyrone derivatives of the following Chemical Formulas 1 to 5 A method for preparing a cargo, comprising the naphthopyron derivative, its stereoisomer, pharmaceutically acceptable salt thereof, hydrate thereof, or solvate thereof from Sprout of Cassia obtusifolia L. or Cassia tora L. And a method for preparing a naphthopyron derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof, comprising the step of separating one or more selected from the group.
[화학식 1][Formula 1]
Figure PCTKR2019000399-appb-img-000009
Figure PCTKR2019000399-appb-img-000009
[화학식 2][Formula 2]
Figure PCTKR2019000399-appb-img-000010
Figure PCTKR2019000399-appb-img-000010
[화학식 3][Formula 3]
Figure PCTKR2019000399-appb-img-000011
Figure PCTKR2019000399-appb-img-000011
[화학식 4][Formula 4]
Figure PCTKR2019000399-appb-img-000012
Figure PCTKR2019000399-appb-img-000012
[화학식 5][Formula 5]
Figure PCTKR2019000399-appb-img-000013
Figure PCTKR2019000399-appb-img-000013
상기 화학식 1의 화합물(나프토파이론 유도체)은 7-하이드록시무시지닐-루브로푸사린-8'-O-글루코사이드일 수 있고, 상기 화학식 2 의 화합물(나프토파이론 유도체)은 이소토라락톤(isotoralactone)일 수 있고, 상기 화학식 3 의 화합물(나프토파이론 유도체)은 토라락톤(toralactone)일 수 있고, 상기 화학식 4 의 화합물(나프토파이론 유도체)은 토로사크리손(torosachrysone)일 수 있고, 상기 화학식 5 의 화합물(나프토파이론 유도체)은 루브로푸사린(rubrofusarin)일 수 있다.The compound of formula 1 (naphthopyrone derivative) may be 7-hydroxymushizinyl-rubbrofusarin-8'-O-glucoside, and the compound of formula 2 (naphthopyrone derivative) is isotolactone ( isotoralactone), the compound of formula 3 (naphthopyrone derivative) may be toralactone (toralactone), the compound of formula 4 (naphthopyrone derivative) may be torosacrysone (torosachrysone), the The compound of Formula 5 (naphthopyrone derivative) may be rubrofusarin.
상기 새싹은 씨앗을 발아시켜 단기간에 재배한 것으로, 실내에서 종자를 발아시켜 재배한 것일 수 있고, 또는 일반적으로 판매되는 새싹인 것일 수 있다.The sprout may be grown by short-term germination of seeds, germinated seeds indoors, or may be a sprout that is generally sold.
상기 제조방법의 결명 새싹(sprout) 또는 결명 새싹 식물체는 결명( Cassia obtusifolia L.) 또는 긴강남차 ( Cassia tora L. 또는 Senna tora)의 종자가 발아하여 2일 내지 31일동안 자연 생장하였거나 재배한 새싹 또는 어린 식물체일 수 있으며, 구체적으로 2일 이상, 3일 이상, 4일 이상, 5일 이상, 6일 이상, 7일 이상, 8일 이상, 9일 이상, 10일 이상, 11일 이상, 12일 이상, 13일 이상, 14일 이상, 15일 이상, 16일 이상, 17일 이상, 18일 이상, 19일 이상, 20일 이상, 21일 이상, 22일 이상, 23일 이상, 24일 이상, 25일 이상, 26일 이상, 27일 이상, 28일 이상, 29일 이상 또는 30일 이상 자연 생장하였거나 재배한 새싹 또는 어린 식물체일 수 있고, 31일 이하, 30일 이하, 29일 이하, 28일 이하, 27일 이하, 26일 이하, 25일 이하, 24일 이하, 23일 이하, 22일 이하, 21일 이하, 20일 이하, 19일 이하, 18일 이하, 17일 이하, 16일 이하, 15일 이하, 14일 이하, 13일 이하, 12일 이하, 11일 이하, 10일 이하, 9일 이하, 8일 이하, 7일 이하, 6일 이하, 5일 이하, 4일 이하 또는 3일 이하 자연 생장하였거나 재배한 새싹 또는 어린 식물체일 수 있고, 보다 구체적으로 2일 내지 21일까지의 기간 중 선택된 기간 동안 자연 생장하였거나 재배한 새싹 또는 어린 식물체일 수 있다.The sprouted or sprouted plant of the above-described manufacturing method was seeded from the seeded plant ( Cassia obtusifolia L.) or Ginseng tea ( Cassia tora L. or Senna tora ), and then grown or grown naturally for 2 to 31 days. It may be a sprout or a young plant, specifically 2 days or more, 3 days or more, 4 days or more, 5 days or more, 6 days or more, 7 days or more, 8 days or more, 9 days or more, 10 days or more, 11 days or more, 12 days or more, 13 days or more, 14 days or more, 15 days or more, 16 days or more, 17 days or more, 18 days or more, 19 days or more, 20 days or more, 21 days or more, 22 days or more, 23 days or more, 24 days More than 25 days, more than 26 days, more than 27 days, more than 28 days, more than 29 days or more than 30 days Naturally grown or grown buds or young plants, 31 days or less, 30 days or less, 29 days or less, 28 days or less, 27 days or less, 26 days or less, 25 days or less, 24 days or less, 23 days or less, 22 days or less, 21 days or less, 20 days or less, 19 days or less, 18 days or less, 1 7 days or less, 16 days or less, 15 days or less, 14 days or less, 13 days or less, 12 days or less, 11 days or less, 10 days or less, 9 days or less, 8 days or less, 7 days or less, 6 days or less, 5 days Hereinafter, it may be a young plant or a young plant that has been naturally grown or cultivated for up to 4 days or less, and more specifically, may be a sprout or a young plant that has been naturally grown or cultivated for a selected period of the period from 2 days to 21 days.
상기 결명( Cassia obtusifolia L. 또는 Cassia tora L.)은 속씨식물문 쌍떡잎식물강 장미목 콩과의 초본을 의미하며, 결명자는 상기 결명(Cassia obtusifolia L.) 또는 긴강남차(Cassia tora L. 또는 Senna tora)의 성숙한 종자로서 결명자 추출물은 당뇨병 개선, 이상지질혈증 개선, 간보호, 항균, 혈압저하 효능이 알려져 있으나(Dong, X. et al. Mol. Med. Rep. 2017, 16, 2331-2346), 산화적 스트레스 또는 약물 독성, 특히 글루타메이트로 인한 망막신경세포 또는 해마신경세포의 손상을 개선하거나 사멸을 억제하는 효능에 대하여는 알려진 바가 없다.The term ( Cassia obtusifolia L. or Cassia tora L.) refers to the herbaceous genus of dicotyledonous plant family Rosaceae, and the term of the term (Cassia obtusifolia L.) or Ginseng tea (Cassia tora L. or Senna) tora) as mature seeds, the extract of terminator is known to improve diabetes, improve dyslipidemia, protect liver, antibacterial, and lower blood pressure (Dong, X. et al. Mol. Med. Rep. 2017, 16, 2331-2346) , It is not known about the efficacy of improving or inhibiting the damage of retinal nerve cells or hippocampal nerve cells due to oxidative stress or drug toxicity, especially glutamate.
상기 결명 새싹 또는 어린 식물체는 결명 새싹 또는 어린 식물체의 전체 또는 부분일 수 있다. 상기 부분은 전초 또는 지상부 또는 지하부일 수 있다. 상기 지상부는 줄기, 잎, 꽃 또는 이들의 조합일 수 있다. 상기 지하부는 뿌리일 수 있다. 상기 식물체는 자연적으로 생장하였거나 또는 인위적으로 재배된 것일 수 있다. 본 발명의 결명 새싹은 실내에서 용이하게 재배할 수 있으며 별도의 영양분 공급 없이 수 주 이내의 단기간에 재배하여 사용할 수 있으므로 산업적 활용성이 크다는 장점을 가진다.The germinating bud or young plant may be all or part of the germinating bud or young plant. The part may be an outpost or above ground or underground. The ground portion may be a stem, a leaf, a flower, or a combination thereof. The basement part may be a root. The plant may be naturally grown or artificially cultivated. The bud of the present invention can be easily grown indoors and can be grown and used for a short period of time within several weeks without supplying additional nutrients, so it has the advantage of great industrial utility.
상기 결명 새싹 추출물은 조추출물(crude extract) 또는 추출물을 추가적으로 분획(fractionation)한 분획물도 포함한다. 구체적으로, 상기 결명 새싹 추출물은 결명 새싹의 조추출물, 분획물, 또는 그 조합일 수 있다. 상기 조추출물은 결명 새싹을 추출 용매와 접촉시켜 얻어지는 것을 말한다. 상기 분획물은 상기 조추출물에 대하여 특정 성분들을 포함하는 물질을 분리한 것을 말한다. 상기 추출물, 또는 그의 분획물은 본 발명의 조성물 중에 결명 새싹 식물체의 추출물, 그의 분획물, 또는 소분획물이 각각 또는 그 혼합물로서 포함된 것일 수 있다. 상기 소분획물은 컷-오프 값을 갖는 한외 여과막을 통과시켜 얻은 것일 수 있으며, 컬럼크로마토그래피 또는 용매분획을 통해 수득한 것일 수 있다. 상기 결명 새싹 추출물 또는 분획물은 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체를 함유하는 것일 수 있다. The shoot sprout extract also includes a crude extract (crude extract) or an additional fraction (fractionation) of the extract. Specifically, the seedling sprout extract may be a crude extract, fraction, or a combination thereof. The crude extract refers to the one obtained by contacting the sprouted bud with the extraction solvent. The fraction refers to the separation of a substance containing specific components with respect to the crude extract. The extract, or a fraction thereof, may be an extract of a shoot sprout plant, a fraction thereof, or a small fraction of each of the compositions of the present invention, or as a mixture thereof. The small fraction may be obtained by passing an ultrafiltration membrane having a cut-off value, and may be obtained by column chromatography or solvent fractionation. The sprout bud extract or fraction may contain one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of the formula (1) to (5).
상기 분리는 여과, 침지, 원심 분리, 용매분획, 크로마토그래피 또는 이들의 조합에 의한 것일 수 있다. 상기 크로마토그래피는 다양한 조건 즉, 크기, 전하, 소수성 또는 친화성에 따른 분리를 위해 제작된 것으로서 이온교환 크로마토그래피, 친화성 크로마토그래피, 크기배제 크로마토그래피, HPLC, 고속유체 크로마토그래피, 컬럼 크로마토그래피, 역상 컬럼 크로마토그래피 또는 그 조합일 수 있다. The separation may be by filtration, dipping, centrifugation, solvent fractionation, chromatography or a combination thereof. The chromatography is prepared for separation according to various conditions, i.e., size, charge, hydrophobicity or affinity, ion exchange chromatography, affinity chromatography, size exclusion chromatography, HPLC, high-speed chromatography, column chromatography, reverse phase Column chromatography or combinations thereof.
상기 추출은 상기 식물체를 용매 중에 일정 시간 동안 인큐베이션하는 것을 포함할 수 있다. 상기 추출은 교반 또는 교반 없이 수행되는 것일 수 있고, 또는 가열하는 것을 포함할 수 있다. 상기 인큐베이션은 실온 내지 환류 온도에서 교반 또는 교반 없이 수행되는 것일 수 있다. 인큐베이션 온도는 선택되는 용매에 따라 적절하게 선택될 수 있다. 예를 들면, 실온 내지 환류 온도, 30℃ 내지 환류 온도, 40℃ 내지 환류 온도일 수 있다. 상기 가열은 50℃, 60℃, 70℃, 80℃, 또는 환류 온도까지 가열하는 것을 포함할 수 있다. 상기 가열은 50℃ 내지 환류 온도, 60℃ 내지 환류 온도, 70℃ 내지 환류 온도, 80℃ 내지 환류 온도, 또는 환류 온도까지 가열하는 것일 수 있다. 상기 추출 시간은 선택된 온도에 따라 달라질 수 있는데 1시간 내지 2개월, 예를 들면, 1시간 내지 1개월, 1시간 내지 15일, 1시간 내지 10일, 1시간 내지 5일, 1시간 내지 3일, 1시간 내지 2일, 1시간 내지 1일, 5시간 내지 1개월, 5시간 내지 15일, 5시간 내지 10일, 5시간 내지 5일, 5시간 내지 3일, 5시간 내지 2일, 5시간 내지 1일, 10시간 내지 1개월, 10시간 내지 15일, 10시간 내지 10일, 10시간 내지 5일, 10시간 내지 3일, 또는 10시간 내지 2일일 수 있다. 상기 추출은 상기 식물체를 용매 중에서 환류 추출하는 것일 수 있다. 상기 용매는 상기 식물체 중량에 대하여 1배, 2배, 5배, 10배 또는 15배 이상의 부피를 갖는 것일 수 있다. 상기 용매는 상기 식물체 중량에 대하여 1배 내지 15배, 2배 내지 15배, 5배 내지 15배, 10배 내지 15배 또는 약 15배의 부피를 갖는 것일 수 있다. 상기 식물체는 음지 또는 차광설비 또는 온풍 또는 건조장치에 건조한 것일 수 있다.The extraction may include incubating the plant in a solvent for a period of time. The extraction may be performed with or without stirring, or may include heating. The incubation may be performed at room temperature to reflux temperature with or without stirring. Incubation temperature may be appropriately selected depending on the solvent selected. For example, the temperature may be from room temperature to reflux temperature, from 30 ° C. to reflux temperature, and from 40 ° C. to reflux temperature. The heating may include heating to 50 ° C., 60 ° C., 70 ° C., 80 ° C., or reflux temperature. The heating may be to 50 ℃ to reflux temperature, 60 ℃ to reflux temperature, 70 ℃ to reflux temperature, 80 ℃ to reflux temperature, or to the reflux temperature. The extraction time may vary depending on the temperature selected, for example 1 hour to 2 months, for example 1 hour to 1 month, 1 hour to 15 days, 1 hour to 10 days, 1 hour to 5 days, 1 hour to 3 days. , 1 hour to 2 days, 1 hour to 1 day, 5 hours to 1 month, 5 hours to 15 days, 5 hours to 10 days, 5 hours to 5 days, 5 hours to 3 days, 5 hours to 2 days, 5 Hour to 1 day, 10 hours to 1 month, 10 hours to 15 days, 10 hours to 10 days, 10 hours to 5 days, 10 hours to 3 days, or 10 hours to 2 days. The extraction may be to extract the plant under reflux in a solvent. The solvent may have a volume of 1 times, 2 times, 5 times, 10 times, or 15 times or more with respect to the weight of the plant. The solvent may have a volume of 1 to 15 times, 2 to 15 times, 5 to 15 times, 10 times to 15 times, or about 15 times the weight of the plant. The plant may be dried in shade or shading facility or warm air or drying device.
상기 추출 방법으로는 여과법, 열수 추출, 침지 추출, 냉침법, 마이크로웨이브 추출, 환류냉각 추출, 가압추출, 아임계추출, 초임계추출 및 초음파추출 등 당업계의 통상적인 방법을 이용할 수 있다. 예를 들면, 침지 추출 방법일 수 있다. 침지 추출은 온침 또는 상온에서 침지하는 것일 수 있고, 1회 내지 5회 추출하는 것일 수 있다. 상기 결명 새싹 식물체는 0.1배 내지 10배 또는 1배 내지 6배의 추출 용매에 접촉되는 것일 수 있다. 냉침 추출온도는 20℃ 내지 40℃인 것일 수 있다. 온침 또는 가열 추출온도는 40℃ 내지 100℃일 수 있다. 냉침 추출시간은 24시간 내지 120시간인 것일 수 있으며, 온침 또는 가열 추출온도는 0.5시간 내지 48시간인 것일 수 있다.As the extraction method, conventional methods in the art such as filtration, hot water extraction, dipping extraction, cold dipping, microwave extraction, reflux cooling extraction, pressure extraction, subcritical extraction, supercritical extraction, and ultrasonic extraction may be used. For example, it may be a dipping extraction method. Immersion extraction may be to be immersed at warm or room temperature, it may be one to five times to extract. The seedling sprout plant may be in contact with the extraction solvent of 0.1 to 10 times or 1 to 6 times. Cold needle extraction temperature may be 20 ℃ to 40 ℃. The warm needle or heat extraction temperature may be 40 ° C to 100 ° C. Cold needle extraction time may be 24 hours to 120 hours, the temperature of the hot or heated extraction may be 0.5 hours to 48 hours.
상기 추출은 또한 얻어진 추출액으로부터 증발 또는 감압 농축과 같은 알려진 방법에 의하여 용매를 제거하는 것을 포함할 수 있다. 상기 추출은 또한 얻어진 추출물을 동결건조와 같은 건조에 의하여 건조 추출물을 제조하는 것을 포함할 수 있다. 상기 감압농축은 진공 감압농축기 또는 진공 회전증발기를 이용하는 것일 수 있다. 또한, 건조는 감압건조, 진공건조, 비등건조, 분무건조 또는 동결건조하는 것일 수 있다.The extraction may also include removing the solvent from the extract obtained by known methods such as evaporation or reduced pressure concentration. The extraction may also include preparing a dry extract by drying the obtained extract, such as lyophilization. The decompression concentration may be to use a vacuum decompression concentrator or a vacuum rotary evaporator. In addition, the drying may be drying under reduced pressure, vacuum drying, boiling drying, spray drying or freeze drying.
상기 제조방법은 상기 결명 새싹을 물, C1 내지 C6의 알코올 및 이들의 혼합 용매로 이루어진 군으로부터 선택된 용매로 하여 추출하는 과정을 더 포함하는 일 수 있다. 상기 알코올은 C1 내지 C3의 알코올, C1 내지 C4, C1 내지 C5 또는 C1 내지 C6의 알코올일 수 있다. 상기 알코올은 일차 알콜일 수 있다. 상기 C1 내지 C6의 알코올은 메탄올, 에탄올, 프로판올, 이소프로판올, 부탄올 또는 이들의 혼합물인 것일 수 있다.The manufacturing method may further include the step of extracting the missing sprout as a solvent selected from the group consisting of water, C1 to C6 alcohol, and a mixed solvent thereof. The alcohol may be C1 to C3 alcohol, C1 to C4, C1 to C5, or C1 to C6 alcohol. The alcohol may be a primary alcohol. The alcohol of C1 to C6 may be methanol, ethanol, propanol, isopropanol, butanol or a mixture thereof.
또한, 상기 제조방법은 물, 에틸아세테이트, 헥산, 염화메틸렌, 클로로포름, 메탄올, 에탄올, 아세톤, 및 이들의 혼합 용매로 이루어진 군으로부터 선택된 하나 이상으로 분획하는 과정을 더 포함하는 것일 수 있다. 상기 제조방법은 상기 결명 새싹을 추출하는 과정을 통해 수득된 추출물의 총 중량을 기준으로 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물이 1 내지 20 중량%로 포함된 결명 새싹 분획물을 제조하는 과정을 포함하는 것일 수 있고, 구체적으로 1 중량% 이상, 2 중량% 이상, 3 중량% 이상, 4 중량% 이상, 5 중량% 이상, 6 중량% 이상, 7 중량% 이상, 8 중량% 이상, 9 중량% 이상, 10 중량% 이상, 11 중량% 이상, 12 중량% 이상, 13 중량% 이상, 14 중량% 이상, 15 중량% 이상, 16 중량% 이상, 17 중량% 이상, 18 중량% 이상 또는 19 중량% 이상 포함된 결명 새싹 분획물을 제조하는 과정을 포함하는 것일 수 있고, 20 중량% 이하, 19 중량% 이하, 18 중량% 이하, 17 중량% 이하, 16 중량% 이하, 15 중량% 이하, 14 중량% 이하, 13 중량% 이하, 12 중량% 이하, 11 중량% 이하, 10 중량% 이하, 9 중량% 이하, 8 중량% 이하, 7 중량% 이하, 6 중량% 이하, 5 중량% 이하, 4 중량% 이하, 3 중량% 이하 또는 2 중량% 이하 포함된 결명 새싹 분획물을 제조하는 과정을 포함하는 것일 수 있다. In addition, the preparation method may further comprise the step of fractionating to at least one selected from the group consisting of water, ethyl acetate, hexane, methylene chloride, chloroform, methanol, ethanol, acetone, and a mixed solvent thereof. The preparation method is based on the total weight of the extract obtained through the extraction of the shoots sprouts one or more naphthopyron derivatives selected from the group consisting of the naphthopyrone derivatives of Formula 1 to Formula 5, stereoisomers thereof, Pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, which may comprise the step of preparing a shoot sprout fraction comprising 1 to 20% by weight, specifically 1% by weight, 2% by weight, At least 3 wt%, at least 4 wt%, at least 5 wt%, at least 6 wt%, at least 7 wt%, at least 8 wt%, at least 9 wt%, at least 10 wt%, at least 11 wt%, at least 12 wt%, It may be to include a process for producing a fraction of the shoots containing at least 13% by weight, at least 14% by weight, at least 15% by weight, at least 16% by weight, at least 17% by weight, at least 18% by weight or at least 19% by weight, 20 wt% or less , 19 wt% or less, 18 wt% or less, 17 wt% or less, 16 wt% or less, 15 wt% or less, 14 wt% or less, 13 wt% or less, 12 wt% or less, 11 wt% or less, 10 wt% or less A process for preparing a fraction seedling fraction containing up to 9% by weight, up to 8% by weight, up to 7% by weight, up to 6% by weight, up to 5% by weight, up to 4% by weight, up to 3% by weight or up to 2% by weight It may be to include.
상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물은 결명 새싹 추출물 또는 분획물의 총 중량을 기준으로 1 내지 50 중량%로 포함될 수 있고, 구체적으로, 1 중량% 이상, 2 중량% 이상, 3 중량% 이상, 4 중량% 이상, 5 중량% 이상, 6 중량% 이상, 7 중량% 이상, 8 중량% 이상, 9 중량% 이상, 10 중량% 이상, 11 중량% 이상, 12 중량% 이상, 13 중량% 이상, 14 중량% 이상, 15 중량% 이상, 16 중량% 이상, 17 중량% 이상, 18 중량% 이상, 19 중량% 이상, 20 중량% 이상, 21 중량% 이상, 22 중량% 이상, 23 중량% 이상, 24 중량% 이상, 25 중량% 이상, 26 중량% 이상, 27 중량% 이상, 28 중량% 이상, 29 중량% 이상, 30 중량% 이상, 31 중량% 이상, 32 중량% 이상, 33 중량% 이상, 34 중량% 이상, 35 중량% 이상, 36 중량% 이상, 37 중량% 이상, 38 중량% 이상, 39 중량% 이상, 40 중량% 이상, 41 중량% 이상, 42 중량% 이상, 43 중량% 이상, 44 중량% 이상, 45 중량% 이상, 46 중량% 이상, 47 중량% 이상, 48 중량% 이상 또는 49 중량% 이상 포함될 수 있고, 50 중량% 이하, 49 중량% 이하, 48 중량% 이하, 47 중량% 이하, 46 중량% 이하, 45 중량% 이하, 44 중량% 이하, 43 중량% 이하, 42 중량% 이하, 41 중량% 이하, 40 중량% 이하, 39 중량% 이하, 38 중량% 이하, 37 중량% 이하, 36 중량% 이하, 35 중량% 이하, 34 중량% 이하, 33 중량% 이하, 32 중량% 이하, 31 중량% 이하, 30 중량% 이하, 29 중량% 이하, 28 중량% 이하, 27 중량% 이하, 26 중량% 이하, 25 중량% 이하, 24 중량% 이하, 23 중량% 이하, 22 중량% 이하, 21 중량% 이하, 20 중량% 이하, 19 중량% 이하, 18 중량% 이하, 17 중량% 이하, 16 중량% 이하, 15 중량% 이하, 14 중량% 이하, 13 중량% 이하, 12 중량% 이하, 11 중량% 이하, 10 중량% 이하, 9 중량% 이하, 8 중량% 이하, 7 중량% 이하, 6 중량% 이하, 5 중량% 이하, 4 중량% 이하, 3 중량% 이하 또는 2 중량% 이하 포함될 수 있다.One or more naphthopyron derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof selected from the group consisting of naphthopyrone derivatives of Formulas 1 to 5 may be May be included in the range of 1 to 50% by weight, specifically, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight, 6% by weight, 7% by weight % By weight, 8% by weight, 9% by weight, 10% by weight, 11% by weight, 12% by weight, 13% by weight, 14% by weight, 15% by weight, 16% by weight, 17% by weight At least 18% by weight at least 19% by weight at least 20% by weight at least 21% by weight at least 22% by weight at least 23% by weight at least 24% by weight at least 25% by weight at least 26% by weight. % By weight, 28% by weight, 29% by weight, 30% by weight, 31% by weight, 32% by weight At least 33 wt%, at least 34 wt%, at least 35 wt%, at least 36 wt%, at least 37 wt%, at least 38 wt%, at least 39 wt%, at least 40 wt%, at least 41 wt%, 42 wt% At least 43 wt%, at least 44 wt%, at least 45 wt%, at least 46 wt%, at least 47 wt%, at least 48 wt% or at least 49 wt%, and at most 50 wt%, at most 49 wt%, 48 wt% or less, 47 wt% or less, 46 wt% or less, 45 wt% or less, 44 wt% or less, 43 wt% or less, 42 wt% or less, 41 wt% or less, 40 wt% or less, 39 wt% or less, 38 wt% or less, 37 wt% or less, 36 wt% or less, 35 wt% or less, 34 wt% or less, 33 wt% or less, 32 wt% or less, 31 wt% or less, 30 wt% or less, 29 wt% or less, 28 wt% or less, 27 wt% or less, 26 wt% or less, 25 wt% or less, 24 wt% or less, 23 wt% or less, 22 wt% or less, 21 wt% or less, 20 wt% or less, 19 wt% or less, 18 wt% or less, 17 wt% or less , 16% or less, 15% or less, 14% or less, 13% or less, 12% or less, 11% or less, 10% or less, 9% or less, 8% or less, 7% or less , 6 wt% or less, 5 wt% or less, 4 wt% or less, 3 wt% or less, or 2 wt% or less.
본 발명은 일 측면에 있어서, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 산화적 스트레스로부터의 신경세포 보호용 또는 신경세포 사멸 억제용 조성물에 관한 것일 수 있다. 상기 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체는 상기 화학식 1의 나프토파이론 유도체, 및 상기 화학식 2 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체일 수 있다. 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체, 약학적으로 허용가능한 염, 입체이성질체, 수화물, 용매화물, 결명 새싹, 결명 새싹 추출물 또는 분획물에 관한 설명은 상술한 바와 같다.In one aspect, the present invention provides at least one naphthopyrone derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvent thereof. At least one selected from the group consisting of cargoes, including Cassia obtusifolia L. or Cassia tora L. sprout sprout extract, or Cassia sprout bud fraction comprising the same as an active ingredient, for protecting nerve cells from oxidative stress or It may be directed to a composition for inhibiting neuronal cell death. The one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 are from the naphthopyrone derivatives of Formula 1 and the naphthopyrone derivatives of Formulas 2 to 5 It may be one or more selected naphthopyrone derivatives. Descriptions of the naphthopyrone derivatives of Formulas 1 to 5, pharmaceutically acceptable salts, stereoisomers, hydrates, solvates, modified buds, modified bud extracts or fractions are as described above.
상기 조성물은 신경세포 보호용 또는 신경세포 사멸 억제용 조성물일 수 있으며, 구체적으로 대사성 독성, 신경 독성, 화학적 원인 등으로 유도되는 산화적 스트레스로부터의 신경세포 보호용 또는 신경세포 사멸 억제용 조성물일 수 있다. 상기 산화적 스트레스는 글루타메이트(glutamate), 글루타메이트 독성 또는 글루타메이트 신경독성에 의해 유발된 것일 수 있으며, 또는 글루타메이트 신경독성에 의해 유발된 산화적 스트레스와 유사한 칼슘 항상성 조절 이상, 미토콘드리아 기능 이상, 흥분세포 독성, 영양 인자 고갈 등에 의한 산화적 스트레스일 수 있다. 구체적으로, 상기 조성물은 글루타메이트, 글루타메이트 독성 또는 글루타메이트 신경독성에 의해 유발된 망막신경세포 또는 해마신경세포의 손상을 감소, 억제, 개선, 또는 예방하거나, 사멸된 망막신경세포 또는 해마신경세포를 소생 또는 재생하는 것일 수 있으며, 글루타메이트 신경독성, 칼슘 항상성 조절 이상, 미토콘드리아 기능 이상, 흥분세포 독성, 영양 인자 고갈 등에 의해 유발된 산화적 스트레스에 대한 방어 기작인 항산화 활성화 또는 글루타메이트 대사 조절 등에 의해 신경세포를 보호하거나 신경세포 사멸을 억제하는 것일 수 있다.The composition may be a composition for protecting neurons or inhibiting neuronal death, and specifically, may be a composition for protecting neurons or inhibiting neuronal death from oxidative stress induced by metabolic toxicity, neurotoxicity, chemical causes, and the like. The oxidative stress may be caused by glutamate, glutamate toxicity, or glutamate neurotoxicity, or calcium homeostatic dysregulation, mitochondrial dysfunction, excitatory cytotoxicity, similar to oxidative stress caused by glutamate neurotoxicity, Oxidative stress, such as depletion of nutritional factors. Specifically, the composition may reduce, inhibit, ameliorate, or prevent damage to retinal neurons or hippocampal neurons caused by glutamate, glutamate toxicity, or glutamate neurotoxicity, or resuscitate or kill dead retinal neurons or hippocampal neurons. It may be regenerative and protects nerve cells by regulating antioxidant activation or glutamate metabolism, which is a defense against oxidative stress caused by glutamate neurotoxicity, abnormalities in calcium homeostasis, mitochondrial dysfunction, excitatory cytotoxicity, depletion of nutritional factors, etc. Or inhibit neuronal cell death.
본 발명의 일 실시예에 따르면, 결명 새싹 추출물은 결명자 추출물에 비하여 DPPH 자유라디컬 소거 효능이 1.7배 이상 우수하였으며(시험예 1), 항산화 성분에 해당하는 상기 화학식 1 내지 화학식 5의 화합물(나프토파이론 유도체)의 함량이 높아 보다 우수한 항산화 효과를 나타내어(시험예 2), 본 발명의 조성물은 산화적 스트레스에 대한 방어 기작인 항산화 작용이 활성화되어 신경세포를 보호하거나 신경세포 사멸을 억제하는 효과가 우수함을 확인하였다.According to an embodiment of the present invention, the extract of shoot sprouts has a DPPH free radical scavenging effect of 1.7 times or more superior to the extract of shoot shoots (Test Example 1), and the compounds of Formula 1 to Formula 5 corresponding to antioxidant components (naph Topirone derivative) has a high content and shows a better antioxidant effect (Test Example 2), the composition of the present invention is an antioxidant action that is a defense mechanism against oxidative stress is activated to protect neurons or inhibit neuronal cell death Was confirmed to be excellent.
상기 조성물은 신경세포 손상 또는 사멸 발생 전, 발생과 동시, 또는 발생 후에 처리하는 것일 수 있다. 상기 조성물은 상기 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 조성물 총 중량에 대하여 0.001 중량% 내지 80 중량% 포함할 수 있고, 구체적으로 0.001 중량% 이상, 0.01 중량% 이상, 0.05 중량% 이상, 0.1 중량% 이상, 1 중량% 이상, 2 중량% 이상, 3 중량% 이상, 4 중량% 이상, 5 중량% 이상, 10 중량% 이상, 20 중량% 이상, 30 중량% 이상, 40 중량% 이상 또는 60 중량% 이상 포함할 수 있고, 80 중량% 이하, 60 중량% 이하, 40 중량% 이하, 30 중량% 이하, 20 중량% 이하, 10 중량% 이하, 5 중량% 이하, 4 중량% 이하, 3 중량% 이하, 2 중량% 이하 또는 1 중량% 이하 포함할 수 있다.The composition may be treated before, concurrent with, or after development of neuronal damage or death. The composition is a group consisting of at least one naphthopyrone derivative selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof. One or more selected from, Cassia obtusifolia L. or Cassia tora L. sprout extract, or a sprout sprout fraction comprising the same may comprise 0.001% to 80% by weight relative to the total weight of the composition, specifically 0.001% or more, 0.01% or more, 0.05% or more, 0.1% or more, 1% or more, 2% or more, 3% or more, 4% or more, 5% or more, 10% or more, 20 wt% or more, 30 wt% or more, 40 wt% or more, or 60 wt% or more, 80 wt% or less, 60 wt% or less, 40 wt% or less, 30 wt% or less, 20 wt% or less, 10 It may include less than 5% by weight, less than 5% by weight, less than 4% by weight, less than 3% by weight, less than 2% by weight, or less than 1% by weight.
또한, 상기 결명 새싹 추출물 또는 분획물은 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물을 결명 새싹 추출물 또는 분획물의 총 중량을 기준으로 1 내지 50 중량%로 포함할 수 있고, 구체적으로, 1 중량% 이상, 2 중량% 이상, 3 중량% 이상, 4 중량% 이상, 5 중량% 이상, 6 중량% 이상, 7 중량% 이상, 8 중량% 이상, 9 중량% 이상, 10 중량% 이상, 11 중량% 이상, 12 중량% 이상, 13 중량% 이상, 14 중량% 이상, 15 중량% 이상, 16 중량% 이상, 17 중량% 이상, 18 중량% 이상, 19 중량% 이상, 20 중량% 이상, 21 중량% 이상, 22 중량% 이상, 23 중량% 이상, 24 중량% 이상, 25 중량% 이상, 26 중량% 이상, 27 중량% 이상, 28 중량% 이상, 29 중량% 이상, 30 중량% 이상, 31 중량% 이상, 32 중량% 이상, 33 중량% 이상, 34 중량% 이상, 35 중량% 이상, 36 중량% 이상, 37 중량% 이상, 38 중량% 이상, 39 중량% 이상, 40 중량% 이상, 41 중량% 이상, 42 중량% 이상, 43 중량% 이상, 44 중량% 이상, 45 중량% 이상, 46 중량% 이상, 47 중량% 이상, 48 중량% 이상 또는 49 중량% 이상 포함할 수 있고, 50 중량% 이하, 49 중량% 이하, 48 중량% 이하, 47 중량% 이하, 46 중량% 이하, 45 중량% 이하, 44 중량% 이하, 43 중량% 이하, 42 중량% 이하, 41 중량% 이하, 40 중량% 이하, 39 중량% 이하, 38 중량% 이하, 37 중량% 이하, 36 중량% 이하, 35 중량% 이하, 34 중량% 이하, 33 중량% 이하, 32 중량% 이하, 31 중량% 이하, 30 중량% 이하, 29 중량% 이하, 28 중량% 이하, 27 중량% 이하, 26 중량% 이하, 25 중량% 이하, 24 중량% 이하, 23 중량% 이하, 22 중량% 이하, 21 중량% 이하, 20 중량% 이하, 19 중량% 이하, 18 중량% 이하, 17 중량% 이하, 16 중량% 이하, 15 중량% 이하, 14 중량% 이하, 13 중량% 이하, 12 중량% 이하, 11 중량% 이하, 10 중량% 이하, 9 중량% 이하, 8 중량% 이하, 7 중량% 이하, 6 중량% 이하, 5 중량% 이하, 4 중량% 이하, 3 중량% 이하 또는 2 중량% 이하 포함할 수 있다.In addition, the above-mentioned sprout extract or fraction is one or more naphthopyron derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or the like selected from the group consisting of naphthopyrone derivatives of the above Chemical Formulas 1 to 5. Solvate may comprise from 1 to 50% by weight, based on the total weight of the shoot sprout extract or fraction, specifically, 1% by weight, 2% by weight, 3% by weight, 4% by weight, 5% by weight % By weight, 6% by weight, 7% by weight, 8% by weight, 9% by weight, 10% by weight, 11% by weight, 12% by weight, 13% by weight, 14% by weight, 15% by weight At least 16% by weight at least 17% by weight at least 18% by weight at least 19% by weight at least 20% by weight at least 21% by weight at least 22% by weight at least 23% by weight at least 24% by weight. % By weight, 26% by weight, 27% by weight, 28% by weight, 29% by weight At least 30%, at least 31% by weight, at least 32% by weight, at least 33% by weight, at least 34% by weight, at least 35% by weight, at least 36% by weight, at least 37% by weight, at least 38% by weight, at 39% by weight At least 40% at least 41% at least 42% at least 43% at least 44% at least 44% at least 45% at least 46% at least 47% at least 48% or at least 49% It may include at least 50%, up to 49%, up to 48%, up to 47%, up to 47%, up to 46%, up to 45%, up to 44%, up to 43%, up to 42% Up to 41 wt%, up to 40 wt%, up to 39 wt%, up to 38 wt%, up to 37 wt%, up to 36 wt%, up to 35 wt%, up to 34 wt%, up to 33 wt%, up to 32 wt% Or less, 31 wt% or less, 30 wt% or less, 29 wt% or less, 28 wt% or less, 27 wt% or less, 26 wt% or less, 25 wt% or less, 24 wt% or less, 23 wt% or less, 22 wt% Or less, 21% by weight or less, 20% by weight Lower, 19% by weight, 18% by weight, 17% by weight, 16% by weight, 15% by weight, 14% by weight, 13% by weight, 12% by weight, 11% by weight, 10% by weight Or less, 9 wt% or less, 8 wt% or less, 7 wt% or less, 6 wt% or less, 5 wt% or less, 4 wt% or less, 3 wt% or less, or 2 wt% or less.
상기 신경세포는 뇌 해마 또는 눈 망막과 연관된 중추 신경, 말초 신경 또는 시신경일 수 있으며, 구체적으로 망막신경세포 또는 해마신경세포일 수 있다. The nerve cell may be a central nerve, a peripheral nerve or an optic nerve associated with the brain hippocampus or the eye retina, and specifically, may be a retinal nerve cell or a hippocampal nerve cell.
상기 망막신경세포는 막대세포(rod cell), 원뿔세포(cone cell), 쌍극세포(bipolar cell), 망막 아마크린 세포(retinal amacrine cell), 수평세포(horizontal cell) 및 망막 신경절세포(retinal ganglion cell)로 이루어진 군으로부터 선택된 하나 이상의 세포일 수 있다. 상기 막대세포 및 원뿔세포는 빛을 감지하여 사물을 인식하는 신경세포이고, 상기 쌍극세포, 망막 아마크린 세포 및 수평세포는 시각정보를 망막신경절세포로 보내는 신경세포이며, 상기 망막 신경절세포는 망막의 시각 정보를 뇌에까지 전달하는 신경세포이다. 본 발명의 일 실시예의 R28 망막전구세포 또는 R28 망막신경전구세포는 망막의 각종 구성세포의 형질을 발현하며 망막에 이식해도 생존하는 세포로, 저산소증이나 혈청의 결핍에 대해 세포사멸이 일어나고, 녹내장과도 관련된 글루타메이트나 GABA 수용체 발현과 연결되는 세포사멸과 세포독성 연구에 유용하게 사용될 수 있다. 또한, R28 세포는 배양조건에 따라 망막전구세포에서 망막신경절세포로도 분화할 수 있으며, 망막의 전반적인 세포학적 연구뿐 만 아니라 망막 신경절세포의 기초연구에 활용될 수 있다(이정일, 김재우, 대한안과학회지, 2009, 50, 919-922). The retinal nerve cells are rod cells, cone cells, bipolar cells, retinal amacrine cells, horizontal cells and retinal ganglion cells. It may be one or more cells selected from the group consisting of). The rod cells and cone cells are nerve cells that sense light by sensing light, and the dipole cells, retinal amacrine cells, and horizontal cells are nerve cells that transmit visual information to the retinal ganglion cells, and the retinal ganglion cells are retinal ganglion cells. It is a nerve cell that delivers visual information to the brain. R28 retinal progenitor cells or R28 retinal progenitor cells of the present invention express the traits of various constituent cells of the retina and survive even when transplanted into the retina, apoptosis occurs due to hypoxia or serum deficiency, It can also be useful for apoptosis and cytotoxicity studies that are linked to related glutamate or GABA receptor expression. In addition, R28 cells can differentiate from retinal progenitor cells to retinal ganglion cells depending on the culture conditions, and can be used not only for overall cytological studies of the retina, but also for basic research of retinal ganglion cells (Jungil Lee, Jaewoo Kim, Korean Ophthalmology) Journal, 2009, 50, 919-922).
상기 해마신경세포는 뇌 내측 측두엽의 한 구조체인 해마(hippocampus)의 신경세포로서, 해마는 생리적/산화적 스트레스 자극에 취약하고 스트레스로 인한 인지기능의 손상을 일으키는데 중심 조직으로 알려져 있으며, 상기 스트레스는 해마의 구조와 뇌세포 생성, 시냅스 가소성 및 해마와 관련된 행동학적 변화를 일으킬 수 있다(김은주, 한국심리학회지: 인지 및 생물, 2012, 24, 65-88). 본 발명의 일 실시예의 HT-22 해마신경세포는 글루타메이트(glutamate)에 민감하여 산화적 스트레스로 유발되는 신경독성을 연구하기 위한 in vitro 모델 시스템으로 사용될 수 있다(Liu, J. et al. Life Science, 2009, 84, 267-271; 김지현, 전순실, 한국식품영양과학회지, 2017, 46, 886-890).The hippocampal nerve cells are nerve cells of the hippocampus (hippocampus), which is a structure of the medial temporal lobe of the brain, and the hippocampus is vulnerable to physiological / oxidative stress stimulation and is known as a central tissue causing damage to cognitive function due to stress. May cause hippocampal structure, brain cell production, synaptic plasticity, and behavioral changes associated with hippocampus (Kim, Eun-ju, Korean Psychological Association: Cognitive and Biological, 2012, 24, 65-88). HT-22 hippocampal neurons of one embodiment of the present invention can be used as an in vitro model system for studying neurotoxicity induced by oxidative stress due to sensitivity to glutamate (Liu, J. et al. Life Science , 2009, 84, 267-271; Kim Ji-hyun, Jeon Soon-sil, Korean Journal of Food Science and Nutrition, 2017, 46, 886-890).
본 발명의 일 실시예에 따르면, 글루타메이트 처리에 의하여 산화적 스트레스가 유발된 R28 세포에 결명 새싹 추출물, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체 각각을 처리 시, 글루타메이트에 의해 감소된 세포생존율이 대조군에 상응하는 정도로 상승하였으며, 이러한 망막세포 보호 효능은 비교예인 결명자 추출물보다 우수하였는바, 본 발명의 조성물은 산화적 스트레스에 의한 망막신경세포의 손상을 보호하거나 망막신경세포 사멸을 억제하는 효과가 우수함을 확인하였다(시험예 3 및 4).According to one embodiment of the present invention, the treatment of the scavenged sprout extract, each of the naphthopyrone derivatives of Formulas 1 to 5 above, to R28 cells induced by oxidative stress by glutamate treatment, reduced cell viability by glutamate. Elevated to the control group, and the retinal cell protection effect was superior to that of the comparative extracts bar, the composition of the present invention has the effect of protecting the retinal nerve cells damage caused by oxidative stress or inhibit retinal neuronal cell death It confirmed that it was excellent (Test Examples 3 and 4).
또한, 본 발명의 일 실시예에 따르면, 글루타메이트 처리에 의하여 산화적 스트레스가 유발된 HT-22 세포에 결명 새싹 추출물, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체 각각을 처리 시, 글루타메이트에 의해 감소된 세포생존율이 대조군에 상응하는 정도로 상승하였으며, 이러한 해마신경세포 보호 효능은 비교예인 결명자 추출물보다 우수하였는바, 본 발명의 조성물은 산화적 스트레스에 의한 해마신경세포의 손상을 보호하거나 해마신경세포 사멸을 억제하는 효과가 우수함을 확인하였다(시험예 5 및 6).In addition, according to an embodiment of the present invention, upon treatment with each of the naphthopyrone derivatives of Formula 1 to Formula 5, which is a modified bud extract, in HT-22 cells in which oxidative stress is induced by glutamate treatment, is reduced by glutamate. The increased cell viability was increased to a level corresponding to that of the control group, and the effect of protecting the hippocampal neurons was superior to that of the extract of the comparative example. It was confirmed that the effect of inhibiting is excellent (Test Examples 5 and 6).
본 발명은 일 측면에 있어서, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료용 조성물에 관한 것일 수 있고, 또는 글루타메이트 신경독성에 의해 유발되는 신경 손상성 질환의 예방 또는 치료용 조성물에 관한 것일 수 있다. 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체는 상기 화학식 1의 나프토파이론 유도체, 및 상기 화학식 2 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체일 수 있다. 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체, 약학적으로 허용가능한 염, 입체이성질체, 수화물, 용매화물, 결명 새싹, 결명 새싹 추출물 또는 분획물, 함량에 관한 설명은 상술한 바와 같다.In one aspect, the present invention provides at least one naphthopyrone derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvent thereof. Injury of retinal nerve cells or hippocampal nerve cells, comprising as an active ingredient at least one selected from the group consisting of cargo, Cassia obtusifolia L. or Cassia tora L. Or it may be directed to a composition for preventing or treating a neurologically damaging disease caused by death, or may be related to a composition for preventing or treating a neurologically damaging disease caused by glutamate neurotoxicity. The at least one naphthopyrone derivative selected from the group consisting of naphthopyrone derivatives of Formulas 1 to 5 is selected from the group consisting of naphthopyrone derivatives of Formula 1 and naphthopyrone derivatives of Formulas 2 to 5 It may be one or more naphthopyrone derivatives. The naphthopyron derivatives of the formulas (1) to (5), pharmaceutically acceptable salts, stereoisomers, hydrates, solvates, germinated buds, germinated bud extracts or fractions, as described above.
상기 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환은 글루타메이트 신경독성에 의한 것일 수 있고, 상기 망막신경세포 및 해마신경세포에 관한 설명은 상술한 바와 같다. 상기 신경 손상성 질환은 망막신경세포 손상 또는 사멸에 의한 당뇨병성 망막병증, 황반변성, 망막세포 손상에 의한 시력장애, 망막색소변성, 망막박리, 망막혈관폐쇄 및 녹내장으로 이루어진 군으로부터 선택된 하나 이상일 수 있고, 해마신경세포 손상 또는 사멸에 의한 기억력 감퇴, 학습능력 감퇴, 우울장애, 근위축성 측삭 경화증(루게릭병), 파킨슨병 및 뇌허혈성 신경손상으로 이루어진 군으로부터 선택된 하나 이상일 수 있으나, 글루타메이트 신경독성에 의해 유발되는 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의한 질환이라면 제한되지 않는다.The neurological damaging disease caused by damage or death of the retinal nerve cells or hippocampal nerve cells may be due to glutamate neurotoxicity, and the description of the retinal nerve cells and hippocampal nerve cells is as described above. The neurological damaging disease may be at least one selected from the group consisting of diabetic retinopathy due to retinal nerve cell injury or death, macular degeneration, vision disorder due to retinal cell damage, retinal pigmentation, retinal detachment, retinal vessel occlusion and glaucoma. May be one or more selected from the group consisting of memory loss, loss of learning ability, depressive disorder, amyotrophic lateral sclerosis (Lou Gehrig's disease), Parkinson's disease and cerebral ischemic nerve injury due to hippocampal nerve cell injury or death, The disease is not limited as long as the disease is caused by damage or death of retinal nerve cells or hippocampal nerve cells.
상기 글루타메이트 신경독성은 신경세포 밖의 글루타메이트 농도가 급속히 증가하게 되어 글루타메이트가 산화적 신경독성 물질로 작용함으로써 나타나는 독성을 의미한다. 상기 글루타메이트 신경독성은 개체 외부에서 유입된 글루타메이트에 의해 유발된 것일 수 있고, 상기 글루타메이트의 유입은 글루타메이트가 함유된 식품, 글루타메이트가 함유된 치료제, 글루타메이트가 함유된 예방제, 글루타메이트가 함유된 항생제 등의 섭취 또는 투여에 의할 수 있으나, 이에 제한되는 것은 아니다.The glutamate neurotoxicity refers to toxicity caused by glutamate acting as an oxidative neurotoxic substance due to a rapid increase in glutamate concentration outside the neuron. The glutamate neurotoxicity may be caused by glutamate introduced from outside of the subject, and the influx of glutamate may be ingested with food containing glutamate, therapeutic agent containing glutamate, preventive agent containing glutamate, antibiotic containing glutamate, and the like. Or by administration, but is not limited thereto.
상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹으로부터 분리 또는 정제된 것일 수 있고, 구체적으로 결명 새싹 에탄올 추출물로부터 분리 또는 정제된 것일 수 있으며, 결명 새싹 에탄올 추출물을 에틸아세테이트로 분획한 뒤, 상기 에틸아세테이트 분획물을 크로마토그래피로 분리하거나 또는 상기 에틸아세테이트 분획물을 다시 헥산, 염화메틸렌 및 메탄올의 혼합 용매로 분획하고 이를 크로마토그래피로 분리한 것일 수 있다.One or more naphthopyron derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 may be separated or purified from the sprout ( Cassia obtusifolia L. or Cassia tora L.), specifically, the sprout The ethanol extract may be separated or purified from the ethanol extract, and the ethanol extract from the shoots may be fractionated with ethyl acetate, and then the ethyl acetate fraction may be separated by chromatography, or the ethyl acetate fraction may be separated with a mixed solvent of hexane, methylene chloride and methanol. It may be fractionated and separated by chromatography.
상기 결명 새싹 추출물은 결명 새싹을 물, C1 내지 C6의 알코올 및 이들의 혼합 용매로 이루어진 군으로부터 선택된 용매로 하여 추출한 것일 수 있다. 상기 알코올은 C1 내지 C3의 알코올, C1 내지 C4, C1 내지 C5 또는 C1 내지 C6의 알코올일 수 있다. 상기 알코올은 일차 알콜일 수 있다. 상기 C1 내지 C6의 알코올은 메탄올, 에탄올, 프로판올, 이소프로판올, 부탄올 또는 이들의 혼합물인 것일 수 있다.The seedling sprout extract may be extracted as a solvent selected from the group consisting of water, C1 to C6 alcohol and a mixed solvent thereof. The alcohol may be C1 to C3 alcohol, C1 to C4, C1 to C5, or C1 to C6 alcohol. The alcohol may be a primary alcohol. The alcohol of C1 to C6 may be methanol, ethanol, propanol, isopropanol, butanol or a mixture thereof.
상기 분획물은 물, 에틸아세테이트, 헥산, 염화메틸렌, 클로로포름, 메탄올, 에탄올, 아세톤, 및 이들의 혼합 용매로 이루어진 군으로부터 선택된 하나 이상으로 분획하는 과정을 더 포함하는 것일 수 있다.The fraction may further include fractionating into one or more selected from the group consisting of water, ethyl acetate, hexane, methylene chloride, chloroform, methanol, ethanol, acetone, and a mixed solvent thereof.
본 발명의 일 실시예에 따르면, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체는 항산화 성분으로서 결명자 추출물에 비해 결명 새싹 추출물에 보다 높은 함량으로 포함되어 있으며(시험예 2), 상기 나프토파이론 유도체는 글루타메이트 처리에 의해 손상 내지 사멸된 망막신경세포(R28) 및 해마신경세포(HT-22)을 보호하는 효능이 우수한바(시험예 4 및 6), 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체가 글루타메이트 신경독성에 의해 유발되는 신경손상성 질환을 예방 또는 치료하는 효능을 가지는 결명 새싹의 유효성분임을 확인하였다.According to one embodiment of the present invention, the one or more naphthopyrone derivatives selected from the group consisting of naphthopyrone derivatives of the formula (1) to (5) is contained in a higher content in the shoot sprout extract than the extract of the fault as an antioxidant component (Test Example 2), the naphthopyron derivative is excellent in protecting the retinal nerve cells (R28) and hippocampal nerve cells (HT-22) damaged or killed by glutamate treatment (Test Examples 4 and 6). , It was confirmed that the naphthopyrone derivatives of Formulas 1 to 5 are effective ingredients of the shoot sprout having the efficacy of preventing or treating neuronal damage diseases caused by glutamate neurotoxicity.
본 발명은 일 측면에 있어서, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 항산화용 조성물에 관한 것일 수 있다. 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체는 상기 화학식 1의 나프토파이론 유도체, 및 상기 화학식 2 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체일 수 있다. 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체, 약학적으로 허용가능한 염, 이성질체, 수화물, 용매화물, 결명 새싹, 결명 새싹 추출물 또는 분획물, 함량에 관한 설명은 상술한 바와 같다.In one aspect, the present invention provides at least one naphthopyrone derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvent thereof. At least one selected from the group consisting of cargoes, Cassia obtusifolia L. or Cassia tora L. Sprout extracts comprising the same, or may be related to the composition for antioxidant, comprising a fraction of the sprouts containing the same as an active ingredient. The at least one naphthopyrone derivative selected from the group consisting of naphthopyrone derivatives of Formulas 1 to 5 is selected from the group consisting of naphthopyrone derivatives of Formula 1 and naphthopyrone derivatives of Formulas 2 to 5 It may be one or more naphthopyrone derivatives. The naphthopyron derivatives of the formulas (1) to (5), pharmaceutically acceptable salts, isomers, hydrates, solvates, unidentified sprouts, unidentified sprout extracts or fractions, as described above.
본 발명은 다른 관점에서, 산화적 스트레스로부터의 신경세포 보호 또는 신경세포 사멸 억제가 필요한 개체에 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 투여하는 것을 포함하는 산화적 스트레스로부터의 신경세포 보호 또는 신경세포 사멸 억제 방법에 관한 것일 수 있다. 본 발명의 일 관점에서, 상기 방법의 투여는 본 명세서에 기재된 투여 방법 및 투여 용량에 따라 수행될 수 있다.In another aspect, the present invention, in one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formula 1 to Formula 5 in the individual in need of neuronal cell protection from oxidative stress or neuronal cell death inhibition, the stereoscopic thereof One or more selected from the group consisting of isomers, pharmaceutically acceptable salts thereof, hydrates, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or missing bud fractions comprising the same It may be related to a method for protecting neurons from oxidative stress or inhibiting neuronal death. In one aspect of the present invention, administration of the method may be performed according to the administration method and administration dose described herein.
본 발명은 다른 관점에서, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료가 필요한 개체에 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 투여하는 것을 포함하는 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료 방법에 관한 것일 수 있다. 본 발명의 일 관점에서, 상기 방법의 투여는 본 명세서에 기재된 투여 방법 및 투여 용량에 따라 수행될 수 있다.In another aspect, the present invention is selected from the group consisting of the naphthopyrone derivatives of Formula 1 to Formula 5 in the individual in need of prevention or treatment of neuro-injury disease caused by damage or death of retinal nerve cells or hippocampal nerve cells. One or more selected from the group consisting of one or more naphthopyrone derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates, or solvates thereof, and a name comprising the same ( Cassia obtusifolia L. or Cassia tora L.) It may be related to a method for preventing or treating a neuro-damaging disease caused by damage or death of retinal nerve cells or hippocampal nerve cells, comprising administering a bud extract or a bud fraction containing a bud thereof. In one aspect of the present invention, administration of the method may be performed according to the administration method and administration dose described herein.
본 발명은 다른 관점에서, 항산화가 필요한 개체에 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 투여하는 것을 포함하는 항산화 방법에 관한 것일 수 있다. 본 발명의 일 관점에서, 상기 방법의 투여는 본 명세서에 기재된 투여 방법 및 투여 용량에 따라 수행될 수 있다.In another aspect, the present invention is a naphthopyrone derivative selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 in an individual in need of antioxidant, stereoisomers thereof, pharmaceutically acceptable salts thereof, and hydrates thereof , Or one or more selected from the group consisting of solvates thereof, and related to an antioxidant method comprising administering a bud extract comprising Cassia obtusifolia L. or Cassia tora L., or a bud bud fraction comprising the same. have. In one aspect of the present invention, administration of the method may be performed according to the administration method and administration dose described herein.
본 발명은 또 다른 관점에서, 산화적 스트레스로부터의 신경세포 보호 또는 신경세포 사멸 억제용 약학 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a pharmaceutical composition for protecting neurons from oxidative stress or inhibiting neuronal death. One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
본 발명은 또 다른 관점에서, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료용 약학 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is a naphthopyrone derivative of Chemical Formulas 1 to 5 for preparing a pharmaceutical composition for the prevention or treatment of neuro-injury disease caused by damage or death of retinal nerve cells or hippocampal nerve cells. One or more selected from the group consisting of one or more naphthopyrone derivatives selected from the group consisting of, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates, or solvates thereof, and crystals comprising them ( Cassia obtusifolia L. or Cassia tora L.) bud extract, or a named bud fraction comprising the same.
본 발명은 또 다른 관점에서, 항산화용 약학 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a pharmaceutical composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or the named bud fractions comprising the same.
본 발명은 또 다른 관점에서, 산화적 스트레스로부터의 신경세포 보호 또는 신경세포 사멸 억제용 화장료 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a cosmetic composition for protecting neurons from oxidative stress or inhibiting neuronal death. One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
본 발명은 또 다른 관점에서, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 개선, 예방 또는 치료용 화장료 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is a naphthopyrone of Formula 1 to Formula 5 for preparing a cosmetic composition for improving, preventing or treating a neuro-damage disease caused by damage or death of retinal nerve cells or hippocampal nerve cells. One or more naphthopyrone derivatives selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, one or more selected from the group consisting of, Cassia obtusifolia L. Or Cassia tora L.) bud extract, or the use of a named bud fraction comprising the same.
본 발명은 또 다른 관점에서, 항산화용 화장료 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a cosmetic composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or the named bud fractions comprising the same.
본 발명은 또 다른 관점에서, 산화적 스트레스로부터의 신경세포 보호 또는 신경세포 사멸 억제용 식품 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a food composition for protecting neurons from oxidative stress or inhibiting neuronal death. One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
본 발명은 또 다른 관점에서, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 개선, 예방 또는 치료용 식품 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is a naphthopyrone of Formula 1 to Formula 5 for preparing a food composition for improving, preventing or treating a neuro-damaging disease caused by damage or death of retinal nerve cells or hippocampal nerve cells. One or more naphthopyrone derivatives selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, one or more selected from the group consisting of, Cassia obtusifolia L. Or Cassia tora L.) bud extract, or the use of a named bud fraction comprising the same.
본 발명은 또 다른 관점에서, 항산화용 식품 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a food composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or the named bud fractions comprising the same.
본 발명은 또 다른 관점에서, 산화적 스트레스로부터의 신경세포 보호 또는 신경세포 사멸 억제용 의약외품 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrones selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a quasi-drug composition for protecting neurons from oxidative stress or inhibiting neuronal death. One or more selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or combinations thereof This may be related to the use of the fraction of the sprouted shoots.
본 발명은 또 다른 관점에서, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 개선, 예방 또는 치료용 의약외품 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is a naphthopyrone of Formula 1 to Formula 5 for preparing a quasi-drug composition for the improvement, prevention or treatment of a neuro-damage disease caused by damage or death of retinal nerve cells or hippocampal nerve cells. One or more naphthopyrone derivatives selected from the group consisting of derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof, one or more selected from the group consisting of, Cassia obtusifolia L. Or Cassia tora L.) bud extract, or the use of a named bud fraction comprising the same.
본 발명은 또 다른 관점에서, 항산화용 의약외품 조성물을 제조하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for preparing a quasi-drug composition for antioxidant, stereoisomers thereof, and pharmaceutically acceptable thereof It may be related to the use of at least one selected from the group consisting of salts, hydrates thereof, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extracts, or bear sprout fractions comprising them.
본 발명은 또 다른 관점에서, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 산화적 스트레스로부터의 신경세포 보호 또는 신경세포 사멸 억제 용도에 관한 것일 수 있다.In another aspect, the present invention, one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or solvents thereof One or more selected from the group consisting of cargo, Cassia obtusifolia L. or Cassia tora L. bud extract, or a bud fraction comprising the same, for protecting neurons from oxidative stress or inhibiting neuronal death It may be about
본 발명은 또 다른 관점에서, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료 용도에 관한 것일 수 있다.In another aspect, the present invention, one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or solvents thereof One or more selected from the group consisting of cargo, Cassia obtusifolia L. or Cassia tora L. bud extract, or a fraction of the resulting bud containing it, caused by damage or death of retinal nerve cells or hippocampal nerve cells It may be related to the use of a prophylactic or therapeutic neurodegenerative disease.
본 발명은 또 다른 관점에서, 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 항산화 용도에 관한 것일 수 있다.In another aspect, the present invention, one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts, hydrates thereof, or solvents thereof One or more selected from the group consisting of cargo, Cassia obtusifolia L. or Cassia tora L. sprout extract, or antioxidant sprout fractions comprising the same.
본 발명은 또 다른 관점에서, 산화적 스트레스로부터 신경세포를 보호하거나 또는 신경세포의 사멸을 억제하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for protecting neurons from oxidative stress or inhibiting the death of neurons. At least one selected from the group consisting of stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates, or solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extract, or crystals comprising the same It may be related to the use of the sprout fraction.
본 발명은 또 다른 관점에서, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환을 예방 또는 치료하기 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is selected from the group consisting of naphthopyrone derivatives of Formula 1 to Formula 5 for preventing or treating neuro-injury diseases caused by damage or death of retinal nerve cells or hippocampal nerve cells. One or more selected from the group consisting of at least one naphthopyrone derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof, a Cassia obtusifolia L. or Cassia tora L. bud containing it It may be related to the use of an extract, or a fraction of a named sprout comprising it.
본 발명은 또 다른 관점에서, 항산화를 위한 상기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물의 용도에 관한 것일 수 있다.In another aspect, the present invention is one or more naphthopyrone derivatives selected from the group consisting of the naphthopyrone derivatives of Formulas 1 to 5 for antioxidant, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, Or it may be related to the use of at least one selected from the group consisting of solvates thereof, Cassia obtusifolia L. or Cassia tora L. bud extract, or a named bud fraction comprising the same.
본 발명의 조성물은 약학 조성물, 화장료 조성물 또는 식품 조성물일 수 있다.The composition of the present invention may be a pharmaceutical composition, cosmetic composition or food composition.
본 발명의 일 측면에 따른 약학 조성물은 경구 또는 비경구 투여 형태로 제형화될 수 있다. 제제화할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 조제된다. 경구투여를 위한 고형제제에는 정제, 환제, 산제, 과립제, 연질 또는 경질 캡슐제 등이 포함되며, 이러한 고형제제는 하나 이상의 화합물에 적어도 하나 이상의 부형제 예를 들면, 전분, 탄산칼슘, 수크로오스(sucrose) 또는 락토오스(lactose), 젤라틴 등을 섞어 조제된다. 또한 단순한 부형제 이외에 스테아린산 마그네슘, 탈크 등과 같은 윤활제들도 사용된다. 경구투여를 위한 액상제제로는 현탁제, 내용액제, 유제, 시럽제 등이 해당되는데 흔히 사용되는 단순 희석제인 물, 리퀴드 파라핀 이외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다. 비경구투여를 위한 제제에는 멸균된 수용액, 비수성용제, 현탁제, 유제, 동결건조제제, 좌제가 포함된다. 비수성용제, 현탁용제로는 프로필렌글리콜(propylene glycol), 폴리에틸렌 글리콜, 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테르 등이 사용될 수 있다. 좌제의 기제로는 위텝솔(witepsol), 마크로골, 트윈(tween) 61, 카카오지, 라우린지, 글리세로젤라틴 등이 사용될 수 있다.Pharmaceutical compositions according to one aspect of the invention may be formulated in oral or parenteral dosage forms. In the case of formulation, it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents and surfactants. Solid form preparations for oral administration include tablets, pills, powders, granules, soft or hard capsules, and the like, which may contain at least one excipient such as starch, calcium carbonate, sucrose, or the like. Or lactose, gelatin, or the like is mixed. Also, in addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral administration include suspending agents, intravenous solutions, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used as diluents, various excipients such as wetting agents, sweeteners, fragrances, and preservatives may be included. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, and suppositories. As a non-aqueous solvent and a suspension solvent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate may be used. As a base for suppositories, witepsol, macrogol, tween 61, cacao butter, laurin butter, and glycerogelatin may be used.
본 발명의 일 측면에 따른 조성물의 약학적 투여 형태는 이들의 약학적으로 허용 가능한 염의 형태로도 사용될 수 있고, 또한 단독으로 또는 타 약학적 활성 화합물과 결합뿐만 아니라 적당한 집합으로 사용될 수 있다. 상기 염으로는 약학적으로 허용되는 것이면 특별히 한정되지 않으며, 예를 들어 염산, 황산, 질산, 인산, 불화수소산, 브롬화수소산, 포름산 아세트산, 타르타르산, 젖산, 시트르산, 푸마르산, 말레산, 숙신산, 메탄술폰산, 벤젠술폰산, 톨루엔술폰산, 나프탈렌술폰산 등을 사용할 수 있다. 또한, 비경구 투여 제형은 피부적용 패치, 연고, 크림, 점안제, 분무제 또는 주사제일 수 있다.Pharmaceutical dosage forms of the compositions according to one aspect of the invention may be used in the form of their pharmaceutically acceptable salts, and may also be used alone or in combination with other pharmaceutically active compounds as well as in a suitable collection. The salt is not particularly limited as long as it is pharmaceutically acceptable. For example, hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrofluoric acid, hydrobromic acid, formic acid acetic acid, tartaric acid, lactic acid, citric acid, fumaric acid, maleic acid, succinic acid, methanesulfonic acid , Benzene sulfonic acid, toluene sulfonic acid, naphthalene sulfonic acid and the like can be used. Parenteral dosage forms may also be dermal application patches, ointments, creams, eye drops, sprays or injections.
다른 측면에서, 본 발명은 상기 약학 조성물을 개체에 투여하는 단계를 포함하는 개체의 신경세포 보호하기 위한 방법일 수 있다.In another aspect, the invention may be a method for protecting neurons in a subject comprising administering the pharmaceutical composition to the subject.
상기 개체는 포유동물, 예를 들면, 사람, 소, 말, 돼지, 개, 양, 염소, 또는 고양이일 수 있으며, 상기 포유동물은 인간일 수 있으며, 본 발명의 화합물의 인체에 대한 효과적인 투여량은 환자의 나이, 몸무게, 성별, 투여형태, 건강 상태 및 질환 정도에 따라 달라질 수 있다.The subject may be a mammal, eg, a human, a cow, a horse, a pig, a dog, a sheep, a goat, or a cat, the mammal may be a human, and an effective dosage for a human body of a compound of the invention Depends on the age, weight, sex, dosage form, health condition and degree of disease of the patient.
상기 투여는 경구 투여 또는 정맥, 복강, 피내, 피하, 상피 또는 근육투여 등과 같은 비경구 투여를 위해 여러 가지 제형으로 투여될 수 있으며, 제제화할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 제조된다.The administration may be administered in various formulations for oral administration or parenteral administration such as intravenous, intraperitoneal, intradermal, subcutaneous, epithelial or intramuscular administration, and when formulated, commonly used fillers, extenders, binders, wetting agents, shelf life It is prepared using diluents or excipients such as releases, surfactants and the like.
상기 투여는 당업계에 알려진 방법에 의하여 투여될 수 있다. 투여는 예를 들면, 정맥내, 근육내, 경구, 또는 피하 투여와 같은 경로로, 임의의 수단에 의하여 개체로 직접적으로 투여될 수 있다. 상기 투여는 전신적으로 또는 국부적으로 투여될 수 있다. 상기 투여는 신경세포가 존재하거나 발생이 예상되는 부위에 국소적으로 투여하는 것일 수 있다.The administration can be administered by methods known in the art. Administration can be administered directly to the subject by any means, eg, by intravenous, intramuscular, oral, or subcutaneous administration. The administration can be administered systemically or locally. The administration may be locally administered to a site where neurons are present or expected to occur.
본 발명의 일 측면에 따른 식품 조성물은 건강기능식품 조성물일 수 있다.Food composition according to an aspect of the present invention may be a health functional food composition.
상기 식품 조성물은 제형은 특별히 한정되지 않으나, 농축액 또는 분말을 직접 또는 희석하여 섭취하거나 경구로 섭취하는 형태로 제형화될 수 있으며, 예를 들어, 정제, 과립제, 분말제, 드링크제와 같은 액제, 캐러멜, 겔, 바 등으로 제형화될 수 있다. 각 제형의 식품 조성물은 유효 성분 이외에 해당 분야에서 통상적으로 사용되는 성분들을 제형 또는 사용 목적에 따라 당업자가 어려움 없이 적의 선정하여 배합할 수 있으며, 다른 원료와 동시에 적용할 경우 상승 효과가 일어날 수 있다. 구체적으로, 상기 식품 조성물은 여러 가지 향미제 또는 천연 탄수화물 등을 추가 성분으로서 함유할 수 있다. 상기 천연 탄수화물은 포도당, 과당과 같은 모노사카리드, 말토스, 슈크로스와 같은 디사카리드, 및 덱스트린, 사이클로덱스트린과 같은 폴리사카리드, 자일리톨, 소르비톨, 에리트리톨 등의 당알콜일 수 있다. 감미제로서는 타우마틴, 스테비아 추출물과 같은 천연 감미제나, 사카린, 아스파르탐과 같은 합성 감미제 등을 사용할 수 있다. 상기 천연 탄수화물의 비율은 상기 조성물 100 중량부당 0.01 ~ 0.04 중량부, 구체적으로는 약 0.02 ~ 0.03 중량부 범위에서 선택할 수 있다. 또한, 상기 식품 조성물은 여러 가지 영양제, 비타민, 전해질, 풍미제, 착색제, 펙트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알콜, 탄산 음료에 사용되는 탄산화제 등을 함유할 수 있다. 그밖에 본 발명의 기능성 식품은 천연 과일쥬스, 과일쥬스 음료 및 야채 음료의 제조를 위한 과육을 함유할 수 있다. 이러한 성분은 독립적으로 또는 조합하여 사용할 수 있다. 이러한 첨가제의 비율은 그렇게 중요하진 않지만 본 명세서의 조성물 100 중량부 당 0 내지 약 20 중량부의 범위에서 포함되는 것이 일반적이다.The food composition is not particularly limited in dosage form, but may be formulated in a form in which the concentrate or powder is ingested or ingested directly or diluted, for example, a liquid such as tablets, granules, powders, drinks, caramels , Gels, bars and the like. In addition to the active ingredient, the food composition of each formulation may be suitably selected by those skilled in the art according to the formulation or purpose of use in addition to the active ingredient, and a synergistic effect may occur when simultaneously applied with other raw materials. Specifically, the food composition may contain various flavors or natural carbohydrates as additional ingredients. The natural carbohydrate may be glucose, monosaccharides such as fructose, maltose, disaccharides such as sucrose, and polysaccharides such as dextrin, cyclodextrin, sugar alcohols such as xylitol, sorbitol, and erythritol. As the sweetener, natural sweeteners such as taumatin and stevia extract, and synthetic sweeteners such as saccharin and aspartame can be used. The ratio of the natural carbohydrate may be selected from 0.01 to 0.04 parts by weight, specifically about 0.02 to 0.03 parts by weight per 100 parts by weight of the composition. In addition, the food composition may contain various nutrients, vitamins, electrolytes, flavors, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH regulators, stabilizers, preservatives, glycerin, alcohols, carbonic acid. Carbonating agents and the like used in beverages. In addition, the functional food of the present invention may contain a flesh for preparing natural fruit juice, fruit juice beverage and vegetable beverage. These ingredients can be used independently or in combination. The proportion of such additives is not so critical but is typically included in the range of 0 to about 20 parts by weight per 100 parts by weight of the composition herein.
상기 식품 조성물에 있어서, 상기 유효 성분의 투여량 결정은 당업자의 수준 내에 있으며, 이의 1일 투여 용량은 예를 들어 0.1mg/kg/일 내지 5000mg/kg/일, 보다 구체적으로는 50 mg/kg/일 내지 500 mg/kg/일이 될 수 있으나, 이에 제한되지 않으며, 투여하고자 하는 대상의 연령, 건강 상태, 합병증 등 다양한 요인에 따라 달라질 수 있다.In the food composition, the dosage determination of the active ingredient is within the level of those skilled in the art, the daily dosage of which is for example from 0.1 mg / kg / day to 5000 mg / kg / day, more specifically 50 mg / kg It may be / day to 500 mg / kg / day, but is not limited thereto, and may vary depending on various factors such as age, health condition, complications of the subject to be administered.
본 발명의 일 측면에 따른 식품 조성물은, 예를 들어, 츄잉껌, 캐러멜 제품, 캔디류, 빙과류, 과자류 등의 각종 식품류, 청량 음료, 미네랄 워터, 알코올 음료 등의 음료 제품, 비타민이나 미네랄 등을 포함한 건강기능성 식품류일 수 있다.The food composition according to one aspect of the present invention includes, for example, health products including chewing gum, caramel products, candy, ice cream, confectionary, various food products such as soft drinks, mineral water, alcoholic beverages, vitamins and minerals, and the like. Functional foods.
본 발명의 조성물은 의약외품 조성물일 수 있다. The composition of the present invention may be a quasi-drug composition.
상기 '의약외품'이란 약사법에 따라 의약품의 용도로 사용되는 물품을 제외한 것으로서, 질병을 치료하거나 예방하기 위해 쓰는 의약품보다 인체에 대한 작용이 경미한 물품에 대해 보건복지부가 따로 정한 분류 기준에 의한 약품을 의미한다. 따라서 인간 또는 동물의 질병 치료나 예방에 쓰이는 섬유ㆍ고무 제품, 인체에 대한 작용이 경미하거나 직접 작용하지 않으며, 기구 또는 기계가 아닌 것과 이와 유사한 것, 또는 감염병을 막기 위한 살균ㆍ살충제 등을 포함할 수 있다. 본 발명의 상기 의약외품 조성물은 산화적 스트레스에 의한 신경세포의 손상 개선, 보호 또는 치료를 위한 것일 수 있고, 글루타메이트 신경독성에 의해 유발되는 신경 손상성 질환의 예방 또는 개선을 위한 것일 수 있다.The term 'out of quasi-drugs' refers to drugs based on the classification criteria set by the Ministry of Health and Welfare for articles that have less effect on the human body than drugs used for the treatment or prevention of diseases according to the Pharmaceutical Affairs Act. do. Therefore, it may include fiber and rubber products used for the treatment or prevention of human or animal diseases, mild or non- direct action on the human body, non-apparatus or machinery, and the like, or disinfectants and insecticides to prevent infectious diseases. Can be. The quasi-drug composition of the present invention may be for the improvement, protection or treatment of damage to nerve cells by oxidative stress, may be for the prevention or improvement of neurological damage diseases caused by glutamate neurotoxicity.
상기 의약외품 조성물은 의약외품적으로 허용가능한 부형제 또는 담체를 더 포함할 수 있다.The quasi-drug composition may further include an quasi-pharmaceutically acceptable excipient or carrier.
상기 의약외품 조성물은 피부도말제, 크림, 파스, 점안제, 분무제 형태로 제형화될 수 있다.The quasi-drug composition may be formulated in the form of skin smear, cream, paste, eye drop, spray.
이하 본 발명을 실시예 및 시험예를 통하여 보다 상세하게 설명한다. 그러나, 이들 실시예 및 시험예는 본 발명을 예시적으로 설명하기 위한 것으로 본 발명의 범위가 이들 실시예 및 시험예에 한정되는 것은 아니다.Hereinafter, the present invention will be described in more detail with reference to Examples and Test Examples. However, these Examples and Test Examples are intended to illustrate the present invention by way of example, but the scope of the present invention is not limited to these Examples and Test Examples.
[[ 실시예Example 1]  One] 결명Fault 새싹 추출물의 제조 Preparation of Sprout Extract
소독되지 않은 국내산 결명자( Cassia obtusifolia L.)를 구입하여(㈜두손애약초, 2016년 1월) 표백분(CaOCl 2) 20 mg/mL 농도로 15분간 소독한 후 물로 충분히 세척하고 정제수에 4시간 동안 침지한 후, 차광 조건에서 6일간 재배하였다. 상기 재배를 통해 수득한 결명 새싹을 32 ℃에서 22시간 동안 온풍(38°C) 건조하고 분쇄하여 698.4 g의 건조된 식물체를 얻었다. 그 후, 상기 건조된 식물체를 추출 용기에 넣은 다음, 에탄올 5.5 L를 가한 후 상온에서 흔들어서 교반하여 7일 동안 추출하고 상기 혼합물을 0.34 mm의 필름 두께를 가지는 와트만(whatman) 종이 여과지와 직경 30 cm 유리 깔대기를 사용하여 중력 여과하는 과정을 2회 반복(‘추출-여과' 2 회)하여, 여과된 추출액을 수득하였다. 상기 여과된 추출액을 감압 농축기에 넣고 감압 하에 35 ℃에서 용매를 완전히 증발시켜 농축하여 결명 새싹 추출물 66.7 g(이하 ‘STS’라고 함)을 수득하였다(수율 9.55%).By purchasing undisinfected domestic Cassia obtusifolia L. (Dooson Aecho Herbs Co., Ltd., January 2016), bleached powder (CaOCl 2 ) was sterilized at a concentration of 20 mg / mL for 15 minutes, washed thoroughly with water, and purified water for 4 hours. After immersion, it was cultivated for 6 days under light-shielding conditions. The resulting buds obtained through the above cultivation were dried and crushed at 32 ° C. for 22 hours in a warm air (38 ° C.) to obtain 698.4 g of dried plants. Then, the dried plant was placed in an extraction container, 5.5 L of ethanol was added thereto, shaken at room temperature, stirred and extracted for 7 days, and the mixture was mixed with a Whatman paper filter paper having a film thickness of 0.34 mm and a diameter of 30. The process of gravity filtration using a cm glass funnel was repeated twice ('extraction-filtration' twice) to obtain a filtered extract. The filtered extract was placed in a reduced pressure concentrator and concentrated by evaporation of the solvent completely at 35 ° C. under reduced pressure to obtain 66.7 g (hereinafter, referred to as 'STS') of Clarified Sprout Extract (yield 9.55%).
[[ 비교예Comparative example 1] 결명자 추출물의 제조 1] Preparation of Clarifier Extract
소독되지 않은 국내산 결명자( Cassia obtusifolia L.)를 구입하여(㈜두손애약초, 2016년 1월) 분쇄한 후 15 mL 추출 용기에 넣은 다음, 에탄올 11 mL을 가한 후 상온에 흔들어 교반하여 7일 추출하고 이 혼합물을 0.34 mm의 필름 두께를 가지는 와트만(whatman) 종이 여과지와 직경 10 cm 유리 깔대기를 사용하여 중력 여과하는 과정을 실시하여('추출-여과' 2 회), 여과된 추출액을 수득하였다(이하 ‘ST’라고 함). 상기 여과된 추출액을 감압 농축기에 넣고 감압 하에 35 ℃에서 용매를 완전히 증발시켜 농축하였다. (수율 3.34%)Purchase undisinfected domestic Cassia obtusifolia ( Cassia obtusifolia L.) ( Dusson Aecho Herbs Co., Ltd., January 2016), pulverize, place in a 15 mL extraction container, add 11 mL of ethanol, shake at room temperature and stir to extract for 7 days And the mixture was subjected to gravity filtration using a Whatman paper filter paper having a film thickness of 0.34 mm and a glass funnel of 10 cm in diameter ('extraction-filtration' twice) to obtain a filtered extract. (Hereinafter referred to as 'ST'). The filtered extract was placed in a reduced pressure concentrator, and concentrated under reduced pressure by evaporating the solvent completely at 35 ° C. (Yield 3.34%)
[[ 실시예Example 2] 다양한 빛  2] various light 조건 하에서Under conditions 재배된  Cultivated 결명Fault 새싹 추출물의 제조 Preparation of Sprout Extract
상기 실시예 1의 차광 조건에서 재배한 결명 새싹 추출물(STS)의 성분과 특정 빛 조건에서 재배한 결명 새싹 추출물의 성분을 비교 분석하기 위하여 빛의 파장 조건을 다르게 하여 결명 새싹을 재배하였다. 상기 실시예 1과 동일한 방법으로 결명 새싹을 재배하되, 결명 새싹 재배 시의 빛 조건을 차광 조건이 아닌, 형광등, LED 385 nm, 465 nm, 645 nm, 780 nm을 각각 다량 파장으로 하는 선택적 광질의 빛을 조사하여 각각 6일간 재배하였다. 상기 재배를 통해 수득한 5 종의 새싹들을 이용하여 상기 실시예 1과 동일한 방법으로 각각 결명 새싹 추출물을 제조하였으며, 형광등 조건에서 재배하여 얻은 추출물을 'STS-C', LED 385 nm의 빛 조건에서 재배하여 얻은 추출물을 'STS-385', LED 465 nm의 빛 조건에서 재배하여 얻은 추출물을 'STS-465', LED 645 nm의 빛 조건에서 재배하여 얻은 추출물을 'STS-645', LED 780 nm의 빛 조건에서 재배하여 얻은 추출물을 'STS-780'이라 하였다.In order to compare and analyze the components of the seedling sprout extract (STS) grown under the light shielding conditions of Example 1 and the components of the seedling sprout extract grown under specific light conditions, the seedling sprouts were grown by varying the light wavelength conditions. Cultivated buds are grown in the same manner as in Example 1, but the selective light quality of the fluorescent light, LED 385 nm, 465 nm, 645 nm, 780 nm, respectively, in a large amount of wavelength, not light-shielding conditions at the time of growing the germination buds Irradiated with light, each was grown for 6 days. Using the five kinds of sprouts obtained through the above cultivation, each of the extracts of Cultivated Sprout was prepared in the same manner as in Example 1, and the extracts obtained by cultivating under fluorescent light conditions were 'STS-C' under LED light conditions of 385 nm. 'STS-385', the extract obtained by cultivation under the light condition of LED 465 nm, 'STS-465', the extract obtained by cultivating under the light condition of LED 645 nm 'STS-645', LED 780 nm The extract obtained by cultivation under light condition of 'STS-780' was called.
[[ 실시예Example 3]  3] 결명Fault 새싹 추출물( Sprout Extract ( STSSTS )의 )of 분획물Fraction 제조 Produce
상기 실시예 1의 결명 새싹 추출물(STS) 66.7 g을 물 600 mL에 현탁시킨 후, 에틸아세테이트 1.8 L를 가하고 상온에서 흔들어 교반하여 2시간씩 4회에 걸쳐 분획을 실시하여 에틸아세테이트 분획물(STS-EA) 23.5 g을 수득하였다. 상기 에틸아세테이트 분획물을 염화메틸렌(또는 에틸아세테이트) 단독 또는 n-헥산, 염화메틸렌(또는 에틸아세테이트), 메탄올(또는 에탄올)의 혼합 용매( n-헥산 : 염화메틸렌의 혼합비율(부피비)이 1:1, 염화메틸렌 : 메탄올의 혼합비율(부피비)이 200:1, 50:1, 10:1, 5:1, 3:1 및 1:1) 2.4L씩을 분획용매로 사용하여 상기 STS-EA의 분획물, 즉 결명 새싹 추출물(STS)의 분획물을 제조하였다.Suspend 66.7 g of the Clarified Sprout Extract of Example 1 in 600 mL of water, 1.8 L of ethyl acetate was added thereto, and the mixture was stirred at room temperature and stirred four times for 2 hours to obtain ethyl acetate fraction (STS- EA) 23.5 g were obtained. The ethyl acetate fraction was mixed with methylene chloride (or ethyl acetate) alone or with a mixed solvent of n -hexane, methylene chloride (or ethyl acetate) and methanol (or ethanol) ( n -hexane: methylene chloride mixture ratio (volume ratio) of 1). 1, methylene chloride: methanol mixing ratio (volume ratio) of 200: 1, 50: 1, 10: 1, 5: 1, 3: 1 and 1: 1) by using 2.4L each fraction of the STS-EA Fractions, ie fractions of Clarified Sprout Extract (STS), were prepared.
구체적으로, 상기 에틸아세테이트 분획물(STS-EA)을 상기 8종의 분획용매에 접촉시켜 셀라이트(celite)에 교반하여 용매를 증발시키고, 직경 10 cm 및 길이 20 cm의 컬럼에 충진된 실리카(silica) 수지에 전개하였다. 상기 각 용매로 용출하는 크로마토그래피를 수행하여 20종의 분획물(STS-EA-분획 1 내지 STS-EA-분획 20)을 수득하였다.Specifically, the ethyl acetate fraction (STS-EA) is contacted with the eight fraction solvents, stirred in celite to evaporate the solvent, and silica packed in a column having a diameter of 10 cm and a length of 20 cm. ) Was developed in the resin. Chromatography eluting with each of the above solvents yielded 20 fractions (STS-EA-fraction 1 to STS-EA-fraction 20).
[[ 실시예Example 4]  4] 결명Fault 새싹 추출물로부터 화합물 1의 분리 Isolation of Compound 1 from Sprout Extract
상기 실시예 3에서 제조한 STS-EA-분획 12(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 10:1인 분획 3개 중 2번째 분획)을 감압 농축한 후, 하기 분리 방법 1의 조건 하에서 분취용 컬럼크로마토그래피를 수행하여 머무름시간 약 221분에서 상기 화학식 1의 구조를 가지는 화합물 1을 분리하였다. (14.0 mg, 건조 식물체 무게 대비 0.002%; 건조 추출물 무게 대비 0.021%) STS-EA-fraction 12 prepared in Example 3 (the second fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 10: 1) After concentration under reduced pressure, preparative column chromatography was performed under the following separation method 1 to separate Compound 1 having the structure of Formula 1 at a retention time of about 221 minutes. (14.0 mg, 0.002% by weight of dry plant; 0.021% by weight of dry extract)
분리 방법 1Separation Method 1
사용장비: YMC LC-Forte REquipment: YMC LC-Forte R
컬럼: Waters Delta-Pak C-18 RP HPLC 컬럼 (30.0x300 ㎜, 15 ㎛)Column: Waters Delta-Pak C-18 RP HPLC Columns (30.0x300 mm, 15 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 40:60으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 40:60
0분에서 60분 (a)용매의 비율을 40%에서 100%로 증가               0 min to 60 min (a) Increase solvent to 40% to 100%
이동상 유속: 10.0 mL/min Mobile phase flow rate: 10.0 mL / min
검출기: 자외선 254 nm; ELSDDetector: ultraviolet 254 nm; ELSD
[[ 실시예Example 5]  5] 결명Fault 새싹 추출물로부터 화합물 2 내지 5의 분리 Isolation of Compounds 2 to 5 from Sprout Extract
(1) 화합물 3의 분리 (1) Isolation of Compound 3
상기 실시예 3에서 제조한 STS-EA-분획 12(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 10:1인 분획 3개 중 2번째 분획)을 감압 농축한 후, 상기 실시예 4의 분리 방법 1의 조건 하에서 분취용 컬럼크로마토그래피를 수행하여 머무름시간 약 24분에서 상기 화학식 3의 구조를 가지는 1.2 mg의 화합물 3을 분리하였다. STS-EA-fraction 12 prepared in Example 3 (the second fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 10: 1) After concentration under reduced pressure, preparative column chromatography was performed under the condition of separation method 1 of Example 4 to separate 1.2 mg of Compound 3 having the structure of Chemical Formula 3 at a retention time of about 24 minutes.
(2) 화합물 2의 분리 (2) Isolation of Compound 2
상기 실시예 3의 STS-EA-분획 8(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 50:1인 분획 3개 중 1번째 분획)을 감압 농축한 후, 하기 분리 방법 2의 조건 하에서 분취용 컬럼크로마토그래피를 수행하여 머무름시간 약 36분에서 상기 화학식 2의 구조를 가지는 7.5 mg의 화합물 2를 분리하였다. The STS-EA-fraction 8 of Example 3 (the first fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 50: 1) is concentrated under reduced pressure. Then, under the conditions of the following separation method 2, preparative column chromatography was performed to separate 7.5 mg of compound 2 having the structure of formula 2 at a retention time of about 36 minutes.
분리 방법 2 Removal method 2
사용장비: YMC LC-Forte REquipment: YMC LC-Forte R
컬럼: Waters Delta-Pak C-18 RP HPLC 컬럼 (30.0x300 ㎜, 15 ㎛)Column: Waters Delta-Pak C-18 RP HPLC Columns (30.0x300 mm, 15 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 60:40으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 60:40
0분에서 60분 (a)용매의 비율을 60%에서 100%로 증가               0 min to 60 min (a) Increase solvent to 60% to 100%
이동상 유속: 10.0 mL/min Mobile phase flow rate: 10.0 mL / min
검출기: 자외선 254 nm; ELSDDetector: ultraviolet 254 nm; ELSD
(3) 화합물 4 및 5의 분리 (3) Separation of Compounds 4 and 5
상기 실시예 3에서 STS-EA-분획 13(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 10:1인 분획 3개 중 3번째 분획)을 감압 농축한 후, 하기 분리 방법 3의 조건 하에서 분취용 컬럼크로마토그래피를 수행하여 머무름시간 약 71분 사이에서 상기 화학식 4의 구조를 가지는 0.6 mg의 화합물 4를 분리하였고, 머무름시간 약 120분에서 상기 화학식 5의 구조를 가지는 2.6 mg의 화합물 5를 분리하였다. In Example 3, STS-EA-fraction 13 (third fraction out of three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 10: 1) is concentrated under reduced pressure. Then, under the conditions of the following separation method 3, preparative column chromatography was performed to separate 0.6 mg of Compound 4 having the structure of Formula 4 between the retention time of about 71 minutes, and the Chemical Formula 5 at the retention time of about 120 minutes. 2.6 mg of Compound 5 having the structure of was isolated.
분리 방법 3Removal method 3
사용장비: JASCO LC-2000PLUSEquipment: JASCO LC-2000PLUS
컬럼: Phenomenex Luna C-18 RP HPLC 컬럼 (21.2x250 ㎜, 10 ㎛)Column: Phenomenex Luna C-18 RP HPLC Columns (21.2 × 250 mm, 10 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 10:90으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 10:90
0분에서 60분 (a)용매의 비율을 10%에서 50%로 증가               0 min to 60 min (a) Increase solvent to 10% to 50%
60분에서 80분 (a)용매의 비율을 50%에서 51%로 증가               60 minutes to 80 minutes (a) Increase solvent to 50% to 51%
80분에서 140분 (a)용매 비율을 51%에서 58%로 증가               80 minutes to 140 minutes (a) Increased solvent ratio from 51% to 58%
140분에서 180분 (a)용매 비율을 58%에서 100%로 증가               140 minutes to 180 minutes (a) Increased solvent ratio from 58% to 100%
이동상 유속: 8 mL/minMobile phase flow rate: 8 mL / min
검출기: 자외선 254 nmDetector: UV 254 nm
[[ 비교예Comparative example 2]  2] 결명Fault 새싹 추출물로부터 화합물 A 내지 X의 분리 Isolation of Compounds A to X from Sprout Extract
(1) 화합물 F, G 및 H의 분리 (1) Isolation of Compounds F, G, and H
상기 실시예 3의 STS-EA-분획 12을 감압 농축한 후, 상기 실시예 4의 분리 방법 1의 조건 하에서 분취용 컬럼크로마토그래피를 수행하여 상기 분획물에 대한 성분 분리를 수행하였다. 그 결과, 도 4의 피크 F에 해당하는 화합물 F 0.8 mg, 도 4의 피크 G에 해당하는 화합물 G 0.9 mg, 도 4의 피크 H에 해당하는 화합물 H 0.5 mg을 분리하였다.After concentrating the STS-EA-fraction 12 of Example 3 under reduced pressure, preparative column chromatography was performed under the conditions of the separation method 1 of Example 4 to perform component separation on the fraction. As a result, 0.8 mg of compound F corresponding to peak F of FIG. 4, 0.9 mg of compound G corresponding to peak G of FIG. 4, and 0.5 mg of compound H corresponding to peak H of FIG. 4 were separated.
(2) 화합물 C, D 및 E의 분리 (2) Isolation of Compounds C, D, and E
상기 실시예 3의 STS-EA-분획 8을 감압 농축한 후, 상기 실시예 4의 분리 방법 2의 조건 하에서 분취용 컬럼크로마토그래피를 수행하여 상기 분획물에 함유된 성분들을 분리하였다. 그 결과, 도 4의 피크 C에 해당하는 화합물 C 7.2 mg, 도 4의 피크 D에 해당하는 화합물 D 0.8 mg, 도 4의 피크 E에 해당하는 화합물 E 2.5 mg을 분리하였다.The STS-EA-fraction 8 of Example 3 was concentrated under reduced pressure, and preparative column chromatography was performed under the conditions of the separation method 2 of Example 4 to separate the components contained in the fraction. As a result, 7.2 mg of Compound C corresponding to Peak C of FIG. 4, 0.8 mg of Compound D corresponding to Peak D of FIG. 4, and 2.5 mg of Compound E corresponding to Peak E of FIG. 4 were isolated.
(3) 화합물 I의 분리 (3) Isolation of Compound I
상기 실시예 3에서 수득한 STS-EA-분획 15(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 5:1인 분획 3개 중 2번째 분획)를 감압 농축한 후, 하기 분리 방법 4의 조건 하에서 HPLC 분리법을 통하여 상기 분획물의 함유 성분을 분리하였다. 그 결과, 도 4의 피크 I에 해당하는 화합물 I 0.7 mg을 분리하였다.STS-EA-fraction 15 obtained in Example 3 (the second fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 5: 1) After concentration under reduced pressure, the components containing the fractions were separated by HPLC separation under the conditions of separation method 4 below. As a result, 0.7 mg of compound I corresponding to peak I of FIG. 4 was isolated.
분리 방법 4 Removal method 4
사용장비: JASCO LC-2000PLUSEquipment: JASCO LC-2000PLUS
컬럼: Phenomenex Luna C-18 RP HPLC 컬럼 (21.2x250 ㎜, 10 ㎛)Column: Phenomenex Luna C-18 RP HPLC Columns (21.2 × 250 mm, 10 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 30:70으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 30:70
0분에서 240분 (a) 용매의 비율을 30%로 유지               0 to 240 minutes (a) Keep the solvent at 30%
이동상 유속: 8 mL/minMobile phase flow rate: 8 mL / min
검출기: 자외선 254 nmDetector: UV 254 nm
(4) 화합물 J, K, L, M, N, O, A 및 P의 분리 (4) Separation of Compounds J, K, L, M, N, O, A and P
상기 실시예 3에서 수득한 STS-EA-분획 16(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 5:1인 분획 3개 중 3번째 분획)을 감압 농축한 후, 하기 분리 방법 5의 조건 하에서 실리카겔 컬럼 크로마토그래피를 실시하여 14개의 소분획물로 분획하였다. STS-EA-fraction 16 obtained in Example 3 (the third fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 5: 1) After concentration under reduced pressure, silica gel column chromatography was performed under the following separation method 5 to fractionate into 14 small fractions.
분리 방법 5 Removal method 5
사용장비: 실리카 플래시 컬럼Equipment used: silica flash column
컬럼: 유리 컬럼 (25.0x200 ㎜)Column: glass column (25.0x200 mm)
용매: (a) 메틸렌 클로라이드 (염화메틸렌)Solvent: (a) Methylene chloride (methylene chloride)
(b) 메탄올      (b) methanol
이동상 조성: (a)용매:(b)용매 부피비 20:1으로 용출 시작Mobile phase composition: (a) Solvent: (b) Elution starts at 20: 1 solvent volume ratio
(a)용매:(b)용매 부피비 10:1, 7:1, 4:1, 2:1, 1:1로 변화            (a) Solvent: (b) Solvent volume ratio 10: 1, 7: 1, 4: 1, 2: 1, 1: 1
단계별 용매량: 300 mLSolvent Steps: 300 mL
상기 소분획물 3(상기 분리 방법 5에서의 용출 용매가 염화메틸렌: 메탄올 혼합비율 10:1인 소분획 3개 중 두번째 분획)으로부터 하기 분리 방법 6의 조건 하에서 HPLC 분리법을 통하여 도 4의 피크 J에 해당하는 화합물 J 1.8 mg, 도 4의 피크 K에 해당하는 화합물 K 1.9 mg, 도 4의 피크 L에 해당하는 화합물 L 1.0 mg을 분리하였다. From the small fraction 3 (the second fraction of the three small fractions in which the elution solvent in the separation method 5 is a methylene chloride: methanol mixture ratio of 10: 1) to the peak J in FIG. 4 through the HPLC separation method under the conditions of the separation method 6 below. The compound J 1.8 mg, the compound K 1.9 mg corresponding to the peak K in FIG. 4, and the compound L 1.0 mg corresponding to the peak L in FIG. 4 were separated.
분리 방법 6 Removal method 6
사용장비: Gilson 321 HPLCEquipment: Gilson 321 HPLC
컬럼: Phenomenex Luna C-18 RP HPLC 컬럼 (21.2x250 ㎜, 10 ㎛)Column: Phenomenex Luna C-18 RP HPLC Columns (21.2 × 250 mm, 10 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 20:80으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 20:80
0분에서 80분 (a)용매의 비율을 20%에서 40%로 증가               0 to 80 minutes (a) Increase solvent to 20% to 40%
80분에서 120분 (a)용매의 비율을 40%에서 100%로 증가               80 to 120 minutes (a) Increasing the proportion of solvent from 40% to 100%
이동상 유속: 8 mL/minMobile phase flow rate: 8 mL / min
검출기: 자외선 254 nmDetector: UV 254 nm
또한, 상기 소분획물 4(상기 분리 방법 5에서의 용출 용매가 염화메틸렌: 메탄올 혼합비율 10:1인 소분획 3개 중 세번째 분획)로부터 하기 분리 방법 7의 조건 하에서 HPLC 분리법을 통하여 도 4의 피크 M에 해당하는 화합물 M 2.5 mg, 도 4의 피크 N에 해당하는 화합물 N 2.1 mg, 도 4의 피크 O에 해당하는 화합물 O 0.5 mg을 분리하였다.In addition, from the small fraction 4 (the third fraction of the three small fractions in which the elution solvent in the separation method 5 is a methylene chloride: methanol mixture ratio of 10: 1) under the conditions of the separation method 7 below, the peak of FIG. 4 2.5 mg of compound M corresponding to M, 2.1 mg of compound N corresponding to peak N in FIG. 4, and 0.5 mg of compound O corresponding to peak O in FIG. 4 were separated.
분리 방법 7Removal method 7
사용장비: Gilson 321 HPLCEquipment: Gilson 321 HPLC
컬럼: Phenomenex Luna C-18 RP HPLC 컬럼 (21.2x250 ㎜, 10 ㎛)Column: Phenomenex Luna C-18 RP HPLC Columns (21.2 × 250 mm, 10 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 20:80으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 20:80
0분에서 100분 (a)용매의 비율을 20%에서 80%로 증가               0 to 100 minutes (a) Increase solvent to 20% to 80%
100분에서 160분 (a)용매의 비율을 80%에서 100%로 증가               100 minutes to 160 minutes (a) Increase solvent to 80% to 100%
이동상 유속: 8 mL/minMobile phase flow rate: 8 mL / min
검출기: 자외선 254 nmDetector: UV 254 nm
상기 소분획물 6(상기 분리 방법 5에서의 용출 용매가 염화메틸렌: 메탄올 혼합비율 7:1인 소분획 3개 중 두번째 분획)로부터 하기 분리 방법 8의 조건하에서 HPLC 분리법을 통하여 도 4의 피크 A에 해당하는 화합물 A 2.0 mg을 분리하였다. From the small fraction 6 (the second fraction of the three small fractions in which the elution solvent in the separation method 5 is methylene chloride: methanol mixture ratio 7: 1) to the peak A in FIG. 4 by HPLC separation under the conditions of the separation method 8 below. 2.0 mg of the corresponding Compound A was isolated.
또한, 상기 소분획물 7(상기 분리 방법 5에서의 용출 용매가 염화메틸렌: 메탄올 혼합비율 7:1인 소분획 3개 중 세번째 분획)로부터 하기 분리 방법 8의 조건 하에서 HPLC 분리법을 통하여 도 4의 피크 P에 해당하는 화합물 P 4.1 mg을 분리하였다.In addition, from the small fraction 7 (the third fraction of the three small fractions in which the elution solvent in the separation method 5 is methylene chloride: methanol mixture ratio 7: 1) is the peak of FIG. 4 through the HPLC separation method under the conditions of the separation method 8 below. 4.1 mg of the compound P corresponding to P was isolated.
분리 방법 8 Removal method 8
사용장비: Gilson 321 HPLCEquipment: Gilson 321 HPLC
컬럼: Phenomenex Luna C-18 RP HPLC 컬럼 (21.2x250 ㎜, 10 ㎛)Column: Phenomenex Luna C-18 RP HPLC Columns (21.2 × 250 mm, 10 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 17:83으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 17:83
0분에서 50분 (a)용매의 비율을 17%로 유지               0 to 50 minutes (a) Maintain solvent at 17%
50분에서 70분 (a)용매의 비율을 17%에서 25%로 증가               50 minutes to 70 minutes (a) Increase solvent to 17% to 25%
70분에서 150분 (a)용매의 비율을 25%에서 30%로 증가               70 minutes to 150 minutes (a) Increase solvent to 25% to 30%
150분에서 180분 (a)용매의 비율을 30%에서 100%로 증가               150 minutes to 180 minutes (a) Increase solvent to 30% to 100%
이동상 유속: 6.5 mL/minMobile phase flow rate: 6.5 mL / min
검출기: 자외선 254 nmDetector: UV 254 nm
(5) 화합물 Q의 분리 (5) Isolation of Compound Q
상기 실시예 3에서 수득한 STS-분획 17(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 3:1인 분획 3개 중 첫번째 분획)을 감압 농축한 후, 하기 분리 방법 9의 조건 하에서 HPLC 분리법을 통하여 상기 분획물의 함유 성분을 분리하였다. 그 결과, 도 4의 피크 Q에 해당하는 화합물 Q 1.7 mg을 분리하였다.STS-fraction 17 obtained in Example 3 (the first fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 3: 1) is concentrated under reduced pressure. Subsequently, the components containing the fractions were separated by HPLC separation under the conditions of Separation Method 9 below. As a result, 1.7 mg of Compound Q corresponding to the peak Q of FIG. 4 was isolated.
분리 방법 9Removal Method 9
사용장비: JASCO LC-2000PLUSEquipment: JASCO LC-2000PLUS
컬럼: Phenomenex Luna C-18 RP HPLC 컬럼 (21.2x250 ㎜, 10 ㎛)Column: Phenomenex Luna C-18 RP HPLC Columns (21.2 × 250 mm, 10 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 10:90으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 10:90
0분에서 40분 (a)용매 비율을 10%로 유지            0 to 40 minutes (a) Keep the solvent ratio at 10%
40분에서 70분 (a)용매 비율을 10%에서 18%로 증가            40 minutes to 70 minutes (a) Increased solvent ratio from 10% to 18%
70분에서 180분 (a)용매의 비율을 18%에서 22%로 증가            70 minutes to 180 minutes (a) Increase solvent to 18% to 22%
180분에서 260분 (a)용매의 비율을 22%에서 100%로 증가            180 minutes to 260 minutes (a) Increase solvent to 22% to 100%
이동상 유속: 6.5 mL/minMobile phase flow rate: 6.5 mL / min
검출기: 자외선 254 nmDetector: UV 254 nm
(6) 화합물 B, R, S, T, U, V, W 및 X의 분리 (6) Isolation of Compounds B, R, S, T, U, V, W and X
상기 실시예 3에서 수득한 STS-EA-분획 18(실시예 3에서의 용출 용매가 염화메틸렌(또는 에틸아세테이트) : 메탄올 (또는 에탄올) 혼합비율 3:1인 분획 3개 중 두번째 분획)을 감압 농축한 후, 하기 분리 방법 10의 조건 하에서 실리카겔 컬럼 크로마토그래피를 실시하여 24개의 소분획물로 분획하였다.The STS-EA-fraction 18 obtained in Example 3 (the second fraction of the three fractions in which the elution solvent in Example 3 is methylene chloride (or ethyl acetate): methanol (or ethanol) mixing ratio 3: 1) is reduced. After concentration, silica gel column chromatography was performed under the conditions of the following separation method 10 to fractionate into 24 small fractions.
분리 방법 10 Removal Method 10
사용장비: 실리카 플래시 컬럼Equipment used: silica flash column
컬럼: 유리 컬럼 (25.0x200 ㎜)Column: glass column (25.0x200 mm)
용매: (a) 메틸렌 클로라이드 (염화메틸렌)Solvent: (a) Methylene chloride (methylene chloride)
(b) 메탄올      (b) methanol
이동상 조성: (a)용매:(b)용매 부피비 20:1으로 용출 시작Mobile phase composition: (a) Solvent: (b) Elution starts at 20: 1 solvent volume ratio
(a)용매:(b)용매 부피비 10:1, 7:1, 5:1, 3:1, 2:1, 1:1로 변화            (a) Solvent: (b) Solvent volume ratio 10: 1, 7: 1, 5: 1, 3: 1, 2: 1, 1: 1
단계별 용매량: 400 mLSolvent Steps: 400 mL
10:1 (소분획물 3,4,5,6), 7:1 (소분획물 7,8,9,10), 5:1(소분획물 11,12,13,14), 3:1(소분획물 15,16,17,18), 2:1(소분획물 19,20,21,22)10: 1 ( small fraction 3,4,5,6), 7: 1 (small fraction 7,8,9,10), 5: 1 (small fraction 11,12,13,14), 3: 1 ( small Fractions 15,16,17,18), 2: 1 (small fractions 19,20,21,22)
상기 소분획물 17 내지 21(상기 분리 방법 10에서의 용출 용매가 염화메틸렌: 메탄올 혼합비율 3:1인 소분획 4개 중 세번째와 네번째 분획 및 용출 용매가 염화메틸렌: 메탄올 혼합비율 2:1인 소분획 4개 중 두번째 내지 네번째 분획)을 하기 분리 방법 11의 조건 하에서 HPLC 분리법을 통하여 상기 분획물의 함유 성분을 분리하였다. 그 결과, 도 4의 피크 B에 해당하는 화합물 B 1.6 mg, 도 4의 피크 R에 해당하는 화합물 R 24.8 mg, 도 4의 피크 S에 해당하는 화합물 S 5.5 mg, 도 4의 피크 T에 해당하는 화합물 T 2.3 mg, 도 4의 피크 U에 해당하는 화합물 U 9.3 mg, 도 4의 피크 V에 해당하는 화합물 V 6.4 mg, 도 4의 피크 W에 해당하는 화합물 W 4.1 mg, 도 4의 피크 X에 해당하는 화합물 X 0.6 mg을 분리하였다.The small fractions 17 to 21 (the third and fourth fractions of the four small fractions in which the elution solvent in the separation method 10 is methylene chloride: methanol mixing ratio 3: 1 and the elution solvent is methylene chloride: methanol mixture ratio 2: 1 The second to fourth of 4 fractions) were separated under the conditions of the following separation method 11 through HPLC separation to separate the components of the fraction. As a result, 1.6 mg of Compound B corresponding to Peak B of FIG. 4, 24.8 mg of Compound R corresponding to Peak R of FIG. 4, 5.5 mg of Compound S corresponding to Peak S of FIG. 4, and corresponding to Peak T of FIG. 4. Compound T 2.3 mg, Compound U corresponding to peak U of FIG. 4 9.3 mg, Compound V corresponding to peak V of FIG. 4 6.4 mg, Compound W corresponding to peak W of FIG. 4 4.1 mg, peak X of FIG. 0.6 mg of the corresponding compound X was isolated.
분리 방법 11Removal Method 11
사용장비: JASCO LC-2000PLUSEquipment: JASCO LC-2000PLUS
컬럼: Phenomenex Luna C-18 RP HPLC 컬럼 (21.2x250 ㎜, 10 ㎛)Column: Phenomenex Luna C-18 RP HPLC Columns (21.2 × 250 mm, 10 μm)
용매: (a) 0.02 % TFA를 함유한 아세토니트릴Solvent: (a) Acetonitrile with 0.02% TFA
(b) 0.02 % TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분에서 (a)용매:(b)용매 부피비 10:90으로 용출 시작Mobile phase composition: elution starts at 0 min with (a) solvent: (b) solvent volume ratio 10:90
0분에서 40분 (a)용매 비율을 10%로 유지             0 to 40 minutes (a) Keep the solvent ratio at 10%
40분에서 70분 (a)용매 비율을 10%에서 18%로 증가             40 minutes to 70 minutes (a) Increased solvent ratio from 10% to 18%
70분에서 180분 (a)용매의 비율을 18%에서 22%로 증가             70 minutes to 180 minutes (a) Increase solvent to 18% to 22%
180분에서 260분 (a)용매의 비율을 22%에서 100%로 증가               180 minutes to 260 minutes (a) Increase solvent to 22% to 100%
이동상 유속: 6.5 mL/minMobile phase flow rate: 6.5 mL / min
상기 실시예 4 및 5와 비교예 2에서 분리된 모든 물질(화합물 1 내지 5, 및 화합물 C 내지 X)에 대응하는 HPLC 피크는 도 4의 결명 새싹 추출물 HPLC 크로마토그램의 피크별로 표기된 바와 같다. 상기 화학식 1 내지 화학식 5의 구조를 가지는 나프토파이론 유도체들(화합물 1 내지 5)을 포함하여 총 29종의 물질은 상기 실시예 1에서 수득한 양의 결명 새싹 추출물로부터 분리 가능한 성분들이다. 또한 HPLC 분석 크로마토그램에 따르면, 상기 29종의 물질은 결명 새싹 추출물의 주요 구성 성분에 해당한다. The HPLC peaks corresponding to all the substances (Compounds 1 to 5, and Compounds C to X) separated in Examples 4 and 5 and Comparative Example 2 are as indicated for each peak of the named bud extract HPLC chromatogram in FIG. 4. A total of 29 substances, including naphthopyrone derivatives having the structures of Formulas 1 to 5 (Compounds 1 to 5), are components that can be separated from the extract of the resulting shoot sprout obtained in Example 1. In addition, according to the HPLC analysis chromatogram, the 29 substances correspond to the main component of the shoot sprout extract.
[[ 실시예Example 6]  6] 결명Fault 새싹 추출물로부터 분리된 화합물 1 내지 5의 구조 동정 Structural identification of compounds 1-5 isolated from bud extract
상기 실시예 4 및 5에서 분리된 화합물 1 내지 5의 구조를 동정하기 위하여 각 화합물들에 대한 질량분석기(Mass Spectrometer, MS) 및 핵자기공명분광기 (NMR)을 이용한 분석을 수행하였다. In order to identify the structures of the compounds 1 to 5 isolated in Examples 4 and 5, an analysis was performed using a mass spectrometer (MS) and a nuclear magnetic resonance spectrometer (NMR) for each compound.
(1) 화합물 1의 동정 (1) Identification of Compound 1
상기 실시예 4에서 분리된 화합물 1의 분자량은 Agilent 1100 고속유체크로마토그래피-질량 분광계(HPLC-ESI-MS)를 이용한 MS 측정을 통하여 664로 결정하였으며, 자외선(PerkinElmer 343 Polarimeter), 적외선(Thermo Scientific Nicolet iS50), 비선광도(PerkinElmer Lambda 35) 분광자료와 핵자기공명기(Bruker 400 MHz, 100 MHz NMR)를 이용한 1H 와 13C NMR 스펙트럼(spectrum) 분석을 통하여 그 구조를 하기 화학식 1의 구조를 가지는 나프토파이론 유도체인 7-하이드록시무시지닐-루브루푸사린-8'-O-글루코사이드로 결정하였다. 상기 화합물 1은 현재까지 보고된 바 없는 신규한 화합물이며, 결명 새싹에서만 주성분 중 하나로 분리되었고, 이는 결명자 또는 완전히 생장한 결명자 식물체에는 함유되어 있지 않거나 매우 미량으로 함유되어 발견되지 않았다.The molecular weight of Compound 1 isolated in Example 4 was determined to be 664 by MS measurement using an Agilent 1100 Fast Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), ultraviolet (PerkinElmer 343 Polarimeter), infrared (Thermo Scientific). Nicolet iS50), 1 H and 13 C NMR spectrum analysis using non-photoluminescence (PerkinElmer Lambda 35) spectroscopy and nuclear magnetic resonance (Bruker 400 MHz, 100 MHz NMR) to obtain the structure of the formula Eggplants were determined with 7-hydroxymusgininyl-rubrufusarin-8'-0-glucoside, a naphthopyrone derivative. Compound 1 is a novel compound that has not been reported to date and has been isolated as one of the main components only in the shoot buds, which was not found or contained in very small amounts in the shooter or fully grown shooter plants.
[화학식 1][Formula 1]
Figure PCTKR2019000399-appb-img-000014
Figure PCTKR2019000399-appb-img-000014
분자식 C 34H 32O 14; ESI-MS: m/z 665 [M+H] +; 적외선 흡수대 ν max 3384, 2936, 1631, 1373, 1204, 1059 cm -1; 자외선 흡수대(MeOH) λ max (log ε) 240 (4.3), 282(4.2); 비선광도 [α] 22 D -20 (c 0.05, MeOH);Molecular formula C 34 H 32 O 14 ; ESI-MS: m / z 665 [M + H] + ; Infrared absorption band ν max 3384, 2936, 1631, 1373, 1204, 1059 cm −1 ; Ultraviolet absorption band (MeOH) λ max (log ε) 240 (4.3), 282 (4.2); Specific light [α] 22 D- 20 (c 0.05, MeOH);
1H NMR (methanol- d 4, 400 MHz): δ 7.18 (1H, s, H-10), 6.97 (1H, d, J = 2.0 Hz, H-5), 6.92 (1H, s, H-9), 6.34 (1H, br s, H-5'), 6.12 (1H, s, H-3), 5.12 (1H, dd, J = 7.5, 3.5 Hz, H-1''), 3.98 (1H, dd, J =12.0, 2.0 Hz, H-6''a), 3.78(1H, dd, J = 12.0, 5.5 Hz, H-6''b), 3.75 (3H, s, OMe-8), 3.58 (1H, t, J = 8.0 Hz, H-2'), 3.55 (1H, dd, J =5.0, 2.0 Hz, H-5''), 3.51 (1H, overlapped, H-3''), 3.48 (1H, t, J = 9.0 Hz, H-4''), 2.63 (3H, s, Me-10'), 2.41 (3H, s, Me-11), 1.98 (3H, s, Me-11'), 13C NMR (CDCl 3, 500 MHz): δ 208.1 (C-9'), 184.1 (C-4), 170.1 (C-2), 161.3 (C-8), 160.9 (C-5), 156.2 (C-6'), 156.1 (C-8'), 154.7 (C-6), 152.4 (C-10a), 150.9 (C-1'), 139.8 (C-9a), 137.1 (C-4'a), 132.4 (C-3'), 122.4 (C-2'), 120.6 (C-4'), 111.9 (C-7), 108.4 (C-8'a), 106.2 (C-5a), 105.8 (C-3), 103.1 (C-1''), 102.9 (C-5'), 102.8 (C-7'), 102.5 (C-4a), 101.4 (C-10), 97.0 (C-9), 77.4 (C-5''), 76.7 (C-3''), 73.5 (C-2''), 69.7 (C-4''), 61.0 (C-6''), 54.8 (OMe-8), 31.4 (C-10'), 19.4 (C-11), 16.1 (C-11') 1 H NMR (methanol- d 4 , 400 MHz): δ 7.18 (1H, s, H-10), 6.97 (1H, d, J = 2.0 Hz, H-5), 6.92 (1H, s, H-9 ), 6.34 (1H, br s, H-5 '), 6.12 (1H, s, H-3), 5.12 (1H, dd, J = 7.5, 3.5 Hz, H-1''), 3.98 (1H, dd, J = 12.0, 2.0 Hz, H-6``a), 3.78 (1H, dd, J = 12.0, 5.5 Hz, H-6``b), 3.75 (3H, s, OMe-8), 3.58 (1H, t, J = 8.0 Hz, H-2 '), 3.55 (1H, dd, J = 5.0, 2.0 Hz, H-5``), 3.51 (1H, overlapped, H-3''), 3.48 (1H, t, J = 9.0 Hz, H-4``), 2.63 (3H, s, Me-10 '), 2.41 (3H, s, Me-11'), 1.98 (3H, s, Me-11 ' ), 13 C NMR (CDCl 3 , 500 MHz): δ 208.1 (C-9 '), 184.1 (C-4), 170.1 (C-2), 161.3 (C-8), 160.9 (C-5), 156.2 (C-6 '), 156.1 (C-8'), 154.7 (C-6), 152.4 (C-10a), 150.9 (C-1 '), 139.8 (C-9a), 137.1 (C-4 'a), 132.4 (C-3'), 122.4 (C-2 '), 120.6 (C-4'), 111.9 (C-7), 108.4 (C-8'a), 106.2 (C-5a) , 105.8 (C-3), 103.1 (C-1``), 102.9 (C-5 '), 102.8 (C-7'), 102.5 (C-4a), 101.4 (C-10), 97.0 (C -9), 77.4 (C-5``), 76.7 (C-3``), 73.5 (C-2 ''), 69.7 (C-4 ''), 61.0 (C-6``), 54.8 (OMe-8), 31.4 (C-10 '), 19.4 (C-11), 16.1 (C-11')
(2) 화합물 2의 동정 (2) Identification of Compound 2
상기 실시예 5에서 분리된 화합물 2의 분자량은 Agilent 1100 고속유체크로마토그래피-질량 분광계(HPLC-ESI-MS)를 이용한 MS 측정을 통하여 272로 결정하였으며, 핵자기공명기(Bruker 400 MHz NMR)를 이용한 1H NMR 스펙트럼(spectrum) 분석을 통하여 하기 화학식 2와 같은 구조의 나프토파이론 유도체인 이소토락톤(isotoralactone)으로 추정하였다. 또한 NMR 자료를 기존문헌(Kitanaka, S. et al. Phytochemistry, 1981, 20, 1951-1953)의 자료와 비교하여 그 구조를 동정하였다.The molecular weight of Compound 2 isolated in Example 5 was determined to be 272 by MS measurement using an Agilent 1100 High-Speed Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR). 1 H NMR spectra were used to estimate isotoractone, a naphthopyrone derivative having the structure shown in Formula 2 below. In addition, the structure of the NMR data was compared with that of the existing literature (Kitanaka, S. et al. Phytochemistry, 1981, 20, 1951-1953).
[화학식 2][Formula 2]
Figure PCTKR2019000399-appb-img-000015
Figure PCTKR2019000399-appb-img-000015
분자식 C 15H 12O 5; ESI-MS: m/z 273 [M+H] +; Molecular formula C 15 H 12 O 5 ; ESI-MS: m / z 273 [M + H] + ;
1H NMR (CDCl 3, 400 MHz): δ 13.30 (1H, s, OH-10), 9.41 (1H, s, OH-9), 6.92 (1H, s, H-5), 6.59 (1H, d, J = 2.5 Hz, H-6), 6.56 (1H, d, J = 2.0 Hz, H-8), 4.93 (1H, d, J = 2.0 Hz, H-11), 4.62 (1H, d, J = 2.0 Hz, H-11), 3.90 (3H, s, OMe-7), 3.80 (1H, s, H-4) 1 H NMR (CDCl 3 , 400 MHz): δ 13.30 (1H, s, OH-10), 9.41 (1H, s, OH-9), 6.92 (1H, s, H-5), 6.59 (1H, d , J = 2.5 Hz, H-6), 6.56 (1H, d, J = 2.0 Hz, H-8), 4.93 (1H, d, J = 2.0 Hz, H-11), 4.62 (1H, d, J = 2.0 Hz, H-11), 3.90 (3H, s, OMe-7), 3.80 (1H, s, H-4)
(3) 화합물 3의 동정 (3) Identification of Compound 3
상기 실시예 5에서 분리된 화합물 3의 분자량은 Agilent 1100 고속유체크로마토그래피-질량 분광계(HPLC-ESI-MS)를 이용한 MS 측정을 통하여 272로 결정하였으며, 핵자기공명기(Bruker 400 MHz NMR)를 이용한 1H NMR 스펙트럼 분석을 통하여 그 구조를 하기 화학식 3과 같은 구조의 나프토파이론 유도체인 토라락톤(toralactone)으로 추정하였다. 또한 NMR 자료를 기존문헌(Newman, A. G. et al. J Am Chem Soc 2016, 138, 4219-4228)의 자료와 비교하여 그 구조를 동정하였다.The molecular weight of Compound 3 isolated in Example 5 was determined to be 272 by MS measurement using an Agilent 1100 Fast Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR). The structure was estimated by 1 H NMR spectral analysis as toralactone, a naphthopyrone derivative having the structure shown in Chemical Formula 3 below. In addition, the structure of the NMR data was compared with that of the existing literature (Newman, AG et al. J Am Chem Soc 2016, 138, 4219-4228).
[화학식 3][Formula 3]
Figure PCTKR2019000399-appb-img-000016
Figure PCTKR2019000399-appb-img-000016
분자식 C 15H 12O 5; ESI-MS: m/z 273 [M+H] +;Molecular formula C 15 H 12 O 5 ; ESI-MS: m / z 273 [M + H] + ;
1H NMR (CDCl 3, 400 MHz): δ 13.56 (1H, s, OH-5), 9.44 (1H, s, OH-9), 7.00 (1H, s, H-5), 6.64 (1H, d, J = 2.5 Hz, H-6), 6.55 (1H, d, J = 2.5 Hz, H-8), 6.25 (1H, s, H-4), 3.92 (3H, s, OMe-7), 2.28 (3H, s, Me-11) 1 H NMR (CDCl 3 , 400 MHz): δ 13.56 (1H, s, OH-5), 9.44 (1H, s, OH-9), 7.00 (1H, s, H-5), 6.64 (1H, d , J = 2.5 Hz, H-6), 6.55 (1H, d, J = 2.5 Hz, H-8), 6.25 (1H, s, H-4), 3.92 (3H, s, OMe-7), 2.28 (3H, s, Me-11)
(4) 화합물 4의 동정 (4) Identification of Compound 4
상기 실시예 5에서 분리된 화합물 4의 분자량은 Agilent 1100 고속유체크로마토그래피-질량 분광계(HPLC-ESI-MS)를 이용한 MS 측정을 통하여 288로 결정하였으며, 핵자기공명기(Bruker 400 MHz NMR)를 이용한 1H NMR 스펙트럼 분석을 통하여 그 구조를 하기 화학식 4와 같은 구조의 나프토파이론 유도체인 토로사크리손(torosachrysone)으로 추정하였다. 또한 NMR 자료를 기존문헌(Gill, M. et al. Aust J Chem 2000, 53, 213-220)의 자료와 비교하여 그 구조를 동정하였다.The molecular weight of Compound 4 isolated in Example 5 was determined to be 288 by MS measurement using an Agilent 1100 Fast Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR). The structure was estimated by 1 H NMR spectral analysis as torosachrysone, a naphthopyron derivative having the structure shown in Chemical Formula 4 below. In addition, the structure of the NMR data was compared with that of the existing literature (Gill, M. et al. Aust J Chem 2000, 53, 213-220).
[화학식 4][Formula 4]
Figure PCTKR2019000399-appb-img-000017
Figure PCTKR2019000399-appb-img-000017
분자식 C 16H 16O 5; ESI-MS: m/z 289 [M+H] +;Molecular formula C 16 H 16 O 5 ; ESI-MS: m / z 289 [M + H] + ;
1H NMR (methanol-d 4, 400 MHz): δ 9.80 (1H, s, OH-9), 6.90 (1H, s, H-5), 6.57 (1H, d, J = 2.0 Hz, H-6), 6.51 (1H, d, J = 2.0 Hz, H-8), 3.91 (3H, s, OMe-7), 3.08 (2H, br s, H-4), 2.86 (2H, br s, H-2), 1.44 (3H, s, Me-11) 1 H NMR (methanol-d 4 , 400 MHz): δ 9.80 (1H, s, OH-9), 6.90 (1H, s, H-5), 6.57 (1H, d, J = 2.0 Hz, H-6 ), 6.51 (1H, d, J = 2.0 Hz, H-8), 3.91 (3H, s, OMe-7), 3.08 (2H, br s, H-4), 2.86 (2H, br s, H- 2), 1.44 (3H, s, Me-11)
(5) 화합물 5의 동정 (5) Identification of compound 5
상기 실시예 5에서 분리된 화합물 5의 분자량은 Agilent 1100 고속유체크로마토그래피-질량 분광계(HPLC-ESI-MS)를 이용한 MS 측정을 통하여 288로 결정하였으며, 핵자기공명기(Bruker 400 MHz NMR)를 이용한 1H NMR 스펙트럼 분석을 통하여 그 구조를 하기 화학식 5와 같은 구조의 나프토파이론 유도체인 루브로푸사린(rubrofusarin)으로 추정하였다. 또한 NMR 자료를 기존문헌(Alemayehu, G. et al. Phytochemistry, 1993, 32, 1273-1277)의 자료와 비교하여 그 구조를 동정하였다.The molecular weight of Compound 5 isolated in Example 5 was determined to be 288 by MS measurement using an Agilent 1100 High-Speed Fluid Chromatography-Mass Spectrometer (HPLC-ESI-MS), using a nuclear magnetic resonance analyzer (Bruker 400 MHz NMR). Through 1 H NMR spectroscopic analysis, the structure was estimated to be rubrofusarin, a naphthopyron derivative having the structure shown in Chemical Formula 5 below. In addition, the structure of NMR data was compared with that of the existing literature (Alemayehu, G. et al. Phytochemistry, 1993, 32, 1273-1277).
[화학식 5][Formula 5]
Figure PCTKR2019000399-appb-img-000018
Figure PCTKR2019000399-appb-img-000018
분자식 C 15H 12O 5; ESI-MS: m/z 273 [M+H] +;Molecular formula C 15 H 12 O 5 ; ESI-MS: m / z 273 [M + H] + ;
1H NMR (CDCl 3, 400 MHz): δ 9.68(1H, s, OH-9), 7.00 (1H, s, H-10), 6.59 (1H, d, J = 2.5 Hz, H-9), 6.48 (1H, d, J = 2.0 Hz, H-7), 6.04 (1H, s, H-3), 3.91 (3H, s, OMe-7), 2.41 (3H, s, Me-11) 1 H NMR (CDCl 3 , 400 MHz): δ 9.68 (1H, s, OH-9), 7.00 (1H, s, H-10), 6.59 (1H, d, J = 2.5 Hz, H-9), 6.48 (1H, d, J = 2.0 Hz, H-7), 6.04 (1H, s, H-3), 3.91 (3H, s, OMe-7), 2.41 (3H, s, Me-11)
[시험예 1] 결명 새싹 추출물의 항산화 효능 평가[Test Example 1] Evaluation of Antioxidant Efficacy of Extract of Sprout Bud
(1) 결명자 추출물과 결명 새싹 추출물의 DPPH 자유라디컬 소거 효능 비교 (1) Comparison of DPPH free radical scavenging efficacy between the extract of ginseng and the extract of shoots
상기 비교예 1에서 제조한 결명자 추출물(ST)과 상기 실시예 1에서 제조한 결명 새싹 추출물(STS)을 메탄올에 각각 1000 ppm, 500 ppm, 250 ppm, 125 ppm의 농도별로 용해시킨 후, 상기 비교예 1의 결명자 추출 용액 100 μL 및 상기 실시예 1의 결명 새싹 추출물 100 μL 각각을 DPPH 100 μL와 암조건에서 40분 동안 반응시켜 517nm에서 흡광도를 측정하였고 양성 대조군으로는 아스코르브산(L-ascorbic acid)을 사용하였다. 상기 흡광도 측정을 통해 하기 식으로 도출한 DPPH 자유라디컬 소거 효능을 도 1에 나타내었다.After dissolving the Clarifier extract prepared in Comparative Example 1 (ST) and the Clarified sprout extract prepared in Example 1 (STS) in the concentration of 1000 ppm, 500 ppm, 250 ppm, 125 ppm respectively in methanol, the comparison The absorbance was measured at 517 nm by reacting 100 μL of the No. 1 extract of Example 1 and 100 μL of the No. 2 extract of Example 1 with DPPH at 100 μL for 40 minutes under dark conditions. As a positive control, ascorbic acid (L-ascorbic acid) was used. ) Was used. DPPH free radical scavenging efficacy derived by the following formula through the absorbance measurement is shown in FIG. 1.
Figure PCTKR2019000399-appb-img-000019
Figure PCTKR2019000399-appb-img-000019
도 1에 나타난 바와 같이, 상기 실시예 1의 결명 새싹 추출물(STS)의 DPPH 자유라디컬 소거 효과가 상기 비교예 1의 결명자 추출물(ST)에 비해 각 농도에서 1.7배 이상 높은 것을 알 수 있었다.As shown in FIG. 1, it can be seen that the DPPH free radical scavenging effect of the budding extract (STS) of Example 1 was 1.7 times higher in each concentration than the budding extract (ST) of Comparative Example 1.
(2) 결명자 추출물과 결명 새싹 추출믈의 ABTS 온라인 항산화 크로마토그래피 측정 결과 비교 (2) Comparison of the results of ABTS online antioxidant chromatography measurement of the extracts of Cassias japonica and the extracts of Cassia keratus.
상기 비교예 1에서 제조한 결명자 추출물(ST)과 상기 실시예 1에서 제조한 결명 새싹 추출물(STS)을 동일한 농도로 알코올 수용액에 용해시켜, 하기 분석 조건에 따라 ABTS 온라인 항산화 고성능액체크로마토그래피 (HPLC)를 수행하였다. Dissolver extract (ST) prepared in Comparative Example 1 and the crystallized sprout extract (STS) prepared in Example 1 was dissolved in an aqueous solution of alcohol at the same concentration, according to the following analysis conditions ABTS online antioxidant high performance liquid chromatography (HPLC ) Was performed.
ABTS 온라인 항산화 HPLC 분석 조건ABTS online antioxidant HPLC analysis conditions
사용장비: Agilent 1200 systemEquipment: Agilent 1200 system
컬럼: RP C-18 HPLC 컬럼 (4.6x150 mm, 5μm)Column: RP C-18 HPLC Columns (4.6x150 mm, 5μm)
용매: (a) 0.02% TFA (Trifluoroacetic acid)를 함유한 아세토니트릴Solvent: (a) Acetonitrile containing 0.02% TFA (Trifluoroacetic acid)
(b) 0.02% TFA를 함유한 물      (b) water containing 0.02% TFA
이동상 조성: 0분 (a)용매:(b)용매 부피비 10:90으로 용출 시작Mobile phase composition: 0 min (a) Solvent: (b) Elution starts with solvent volume ratio 10:90
0분에서 30분 (a)용매 비율을 10%에서 100%로 증가             0 to 30 minutes (a) Increased solvent ratio from 10% to 100%
30분에서 37분 (a)용매 100%로 용출             30 to 37 minutes (a) Elution with 100% solvent
이동상 유속: 0.7 mL/minMobile phase flow rate: 0.7 mL / min
ABTS 조성: 0.08mM ABTS와 0.12 mM 과황화칼륨 (potassium persulfate)를 함유한 물ABTS composition: water containing 0.08 mM ABTS and 0.12 mM potassium persulfate
ABTS 유속: 0.35 mL/minABTS flow rate: 0.35 mL / min
검출기 검출 파장: 자외선 254 nm; 734 nmDetector detection wavelength: ultraviolet 254 nm; 734 nm
그 결과, 결명자 추출물(ST)과 결명 새싹 추출물(STS)은 매우 상이한 항산화 성분 양상을 나타내었다. 결명자 추출물(ST)과 결명 새싹 추출물(STS) 모두 머무름 시간 15분 이전에 황산화 활성을 가진 플라보노이드, 나프탈렌 유도체들로 구성된 성분들이 관찰되었으나, 결명 새싹 추출물(STS)에서는 결명자 추출물(ST)에 비해 상기의 성분들의 함량이 증가하는 동시에 15분 이후에 항산화 활성을 가진 나프탈렌, 안트라퀴논 유도체들이 추가적으로 관찰되었다. 도 2의 (A)와 (B)는 결명자와 결명 새싹 추출물의 ABTS 온라인항산화 HPLC를 수행한 결과이며, 자외선(UV) 검출기 254 nm (위 방향으로 표기된 파란색 선의 크로마토그램), 734 nm (아래 방향으로 표기된 붉은색 선의 크로마토그램) 파장을 각각 검출하여 얻어진 크로마토그램이다. As a result, Cassiae extract (ST) and Cassiae sprout extract (STS) showed very different antioxidant components. The components of the Cassia vulgaris extract (ST) and Cassia vulgaris extract (STS) were found to be composed of flavonoids and naphthalene derivatives having sulfated activity 15 minutes before the retention time. At the same time as the contents of the above components were increased, naphthalene and anthraquinone derivatives having antioxidant activities were additionally observed after 15 minutes. Figure 2 (A) and (B) is a result of performing the ABTS online antioxidant HPLC of the deficiency and the germinated bud extract, ultraviolet (UV) detector 254 nm (chromatic gram of the blue line indicated in the upward direction), 734 nm (downward) Chromatogram obtained by detecting wavelengths of red lines indicated by?).
[시험예 2] 결명자 추출물(ST)과 다양한 빛 조건 하에서 재배된 결명 새싹 추출물의 ABTS 온라인 항산화 크로마토그래피 측정 결과 비교[Test Example 2] Comparison of measurement results of ABTS on-line antioxidant chromatography of Cassia tora extract (ST) and Cassia tora sprout extract grown under various light conditions
상기 실시예 1에서 제조한 결명 새싹 추출물(STS)과 상기 실시예 2에서 제조한 7종의 다양한 빛 조건 하에서 재배된 결명 새싹 추출물(STS-C, STS-385, STS-465, STS-645 및 STS-780)을 알코올 수용액에 용해시켜, 상기 시험예 1과 동일한 분석 조건에 따라 ABTS 온라인 항산화 고성능액체크로마토그래피(HPLC)를 수행하였다. Clarified sprout extract (STS) prepared in Example 1 and Clarified sprout extract grown under various light conditions prepared in Example 2 (STS-C, STS-385, STS-465, STS-645 and STS-780) was dissolved in an aqueous alcohol solution, and subjected to ABTS online antioxidant high performance liquid chromatography (HPLC) according to the same analysis conditions as in Test Example 1.
그 결과, 상기 실시예 2의 5종의 다양한 빛 조건 하에서 재배된 결명 새싹 추출물에서 항산화 활성을 지닌 구성 성분들이 공통적으로 관찰되었으나 빛 조건에 따라 성분들의 함량 변화가 나타났다. 도 3의 A, B, C, D, E, F는 각각 상기 6종의 결명 새싹 추출물인 STS, STS-C, STS-385, STS-465, STS-645, STS-780 추출물의 ABTS 온라인 항산화 HPLC를 수행한 결과이며, 자외선(UV) 검출기 254 nm (UV 254nm를 흡수하는 성분들의 피크를 나타냄) 및 734 nm 파장 (ABTS에 대한 항산화 작용을 나타내는 성분의 피크들과 피크들의 면적으로 항산화도를 나타냄)을 각각 검출하여 얻어진 크로마토그램이다. As a result, although the components with antioxidant activity were commonly observed in the Cultivated Sprout Extract cultivated under the various light conditions of Example 2, the contents of the components were changed according to the light conditions. A, B, C, D, E, F of Figure 3 are ABTS online antioxidants of the six species of Sprout bud extracts STS, STS-C, STS-385, STS-465, STS-645 and STS-780, respectively. As a result of HPLC, the degree of antioxidant activity was determined by the area of the peaks and peaks of the UV detector 254 nm (showing peaks of the components absorbing UV 254 nm) and 734 nm wavelengths (antioxidative action against ABTS). It is a chromatogram obtained by detecting each).
도 3의 크로마토그램들을 바탕으로 차광 조건과 형광등 및 각 특정 파장의 빛 조건 하에서 재배한 결명 새싹 추출물의 항산화 성분 프로파일을 살펴본 결과, 재배 과정 중에 조사된 빛 파장별로 항산화 성분들의 함량과 조성은 차이가 있었으나 특정 조건에서 사라지거나 새롭게 나타나는 성분은 발견되지 않았다. 도 3의 (A) 내지 (F)의 모든 크로마토그램에서 본 발명의 상기 화학식 1의 구조를 가지는 신규 나프토파이론 성분(실시예 4에서 분리된 화합물 1)이 가장 주목할 만한 항산화 피크 또는 주요 항산화 피크로 검출되었으며, 상기 화학식 2 내지 화학식 5의 구조를 가지는 4종의 나프토파이론 성분들(실시예 5에서 분리된 화합물 2 내지 5) 또한 도 3의 (A) 내지 (F)의 크로마토그램에서 항산화 성분으로 검출되었다. 일반 빛 조건에서 재배한 결명 새싹 추출물(STS-C)의 크로마토그램 (B)와 465 nm LED 조명 하에서 재배한 결명 새싹 추출물(STS-465)의 크로마토그램 (D)를 제외하고, (A), (C), (E) 및 (F)의 크로마토그램에서 상기 실시예 4 및 5에서 분리된 화합물 1 내지 5의 유효성분들이 상대적으로 높은 함량으로 검출되었다. Based on the chromatograms of FIG. 3, the antioxidant component profiles of Cultivated Sprout extracts grown under light-shielding conditions, fluorescent lamps, and light conditions of specific wavelengths were examined. However, no disappearing or new components were found under certain conditions. In all the chromatograms of Figures (A) to (F), the novel naphthopyron component (Compound 1 isolated in Example 4) having the structure of Formula 1 of the present invention is the most notable antioxidant peak or the main antioxidant peak. 4 naphthopyron components (compounds 2 to 5 isolated in Example 5) having the structure of Formulas 2 to 5 were also detected as antioxidants in the chromatograms of (A) to (F) of FIG. It was detected as a component. (A), except for chromatogram (B) of inflorescence bud extract (STS-C) grown under normal light conditions and chromatogram (D) of inflorescence bud extract (STS-465) grown under 465 nm LED illumination, In the chromatograms of (C), (E) and (F), the active ingredients of the compounds 1 to 5 isolated in Examples 4 and 5 were detected in a relatively high content.
[[ 시험예Test Example 3]  3] 실시예Example 1 및 2의 7종의  7 kinds of 1 and 2 결명Fault 새싹 추출물의  Of sprout extract 망막전구세포Retinal progenitor cells (R28) 보호 효능(R28) Protective effect
상기 비교예 1 및 실시예 1에서 각각 수득된 결명자 추출물(ST)과 결명 새싹 추출물(STS), 및 상기 실시예 2에서 수득된 다양한 빛 조건 하에서 재배된 결명 새싹 추출물(STS-C, STS-385, STS-465, STS-645, STS-780) 의 망막전구세포(R28) 보호 효능을 확인하기 위해 하기와 같은 실험을 수행하였다. Clarifier extract (ST) and Claw sprout extract (STS) obtained in Comparative Example 1 and Example 1, respectively, and Cultivated sprout extract (STS-C, STS-385) grown under various light conditions obtained in Example 2 , STS-465, STS-645, STS-780) was carried out the following experiment to confirm the protective effect of the retinal progenitor cells (R28).
구체적으로, R28 세포는 10% FBS(fetal bovine serum), 100 U/mL 페니실린(penicillin), 100 μg/mL 스트렙토마이신(streptomycin)이 포함된 DMEM(Dulbecco’s modified eagle’s medium)/low glucose 배지에서 37 ℃ 인큐베이터에서 5% CO 2 조건하에서 배양하고, 2일마다 0.05% 트립신(Trypsin)을 사용하여 계대배양한 후, 96 플레이트에 1X10 4의 밀도로 접종하고 24시간 배양하였다. 상기 배양된 R28 세포에 상기 7종의 추출물들을 도 5의 (A) 내지 (C)의 농도로 처리하고 2시간 후, 10 mM 글루타메이트(glutamate)와 0.5 mM BSO(buthionine sulphoximine)를 첨가하여 22시간 배양하였다. 그런 다음, 세포 생존률을 측정하기 위해 10 μL EZ-cytox를 처리하여 2시간 유지하였으며, UV 흡광도 450 nm에서 측정하였다.Specifically, R28 cells at 37 ° C. in Dulbecco's modified eagle's medium (DMEM) / low glucose medium containing 10% fetal bovine serum (FBS), 100 U / mL penicillin, and 100 μg / mL streptomycin. After incubation under 5% CO 2 conditions in an incubator, passaged with 0.05% trypsin every 2 days, inoculated at a density of 1 × 10 4 in 96 plates and cultured for 24 hours. The 7 extracts were treated to the cultured R28 cells at the concentrations of (A) to (C) of FIG. 5, and after 2 hours, 10 mM glutamate and 0.5 mM BSO (buthionine sulphoximine) were added for 22 hours. Incubated. Then, 10 μL EZ-cytox was treated and maintained for 2 hours to measure cell viability, and UV absorbance was measured at 450 nm.
도 5에서 (A)는 글루타메이트에 의해서 유발되는 산화적 스트레스로부터 3가지 농도에서 상기 비교예 1의 결명자 추출물(ST), 상기 실시예 1의 결명 새싹 추출물(STS), 상기 실시예 2의 빛 조건을 다르게 하여 재배된 5종의 결명 새싹 추출물(STS-C, STS-385, STS-465, STS-645, STS-780)의 R28 세포 보호 효능을 세포 생존률로 나타낸 결과이다. 도 5에서 (B) 및 (C)는 각각 5가지 농도에서 글루타메이트에 의해서 유발되는 산화적 스트레스로부터 상기 비교예 1의 결명자 추출물(ST) 및 상기 실시예 1의 결명 새싹 추출물(STS)의 R28 세포 보호 효능을 세포 생존률로 나타낸 결과이다.In Figure 5 (A) is a light extract of Example 2, the extract of the crystallized shooter (ST) of the Comparative Example 1, ST extract of Example 1, at three concentrations from the oxidative stress caused by glutamate This is the result of the cell survival rate of the R28 cell protection effect of the five species of sprout sprouts cultivated differently (STS-C, STS-385, STS-465, STS-645, STS-780). In Figure 5 (B) and (C) are R28 cells of the clarifier extract (ST) of the Comparative Example 1 and the sprout bud extract (STS) of Example 1 from the oxidative stress caused by glutamate at each of five concentrations, respectively Protective efficacy is shown as cell viability.
도 5의 (A)에 나타난 바와 같이, 모든 추출물 처리군에서 글루타메이트에 의해서 유발되는 산화적 스트레스로부터 HT-22 세포를 보호하는 효과를 보였으나, 결명자 추출물에 비해 결명 새싹 추출물(STS, STS-C, STS-385, STS-465, STS-645 및 STS-780)이 거의 대부분의 농도에서 글루타메이트에 의해서 유발되는 산화적 스트레스로부터 R28 세포를 보호하는 효과가 보다 우수함을 보였으며, 특히 도 5의 (B) 및 (C)에 나타난 바와 같이, 상기 실시예 1의 결명 새싹 추출물(STS)은 상기 비교예 1의 결명자 추출물(ST)에 비해 R28 세포 보호 효능이 확연히 뛰어난 것으로 확인되었다.As shown in (A) of FIG. 5, all the extract treatment groups showed an effect of protecting HT-22 cells from oxidative stress caused by glutamate, but the shoot sprout extracts (STS, STS-C) compared to the extracts , STS-385, STS-465, STS-645, and STS-780) showed a better effect of protecting R28 cells from oxidative stress induced by glutamate at almost all concentrations. As shown in B) and (C), it was confirmed that the nodule sprout extract (STS) of Example 1 was significantly superior to the R28 cell protection effect compared to the nodule extract (ST) of Comparative Example 1.
[[ 시험예Test Example 4]  4] 실시예Example 5 및 6의  5 and 6 결명Fault 새싹 추출물로부터 분리된 화합물 1 내지 5의  Of compounds 1 to 5 isolated from sprout extract 망막전구세포Retinal progenitor cells (R28) 보호 효능 확인(R28) Confirmation of protective efficacy
상기 시험예 3의 결명 새싹 추출물(STS)의 망막전구세포(R28) 보호 효능이 결명자 추출물(ST)의 효능보다 뛰어난 이유를 확인하기 위하여, R28 세포에 대한 실시예 5 및 6의 화합물 1 내지 5의 보호 효능을 비교하였다. In order to confirm why the retinal progenitor cells (R28) protective effect of the Sprout bud extract (STS) of Test Example 3 is superior to that of the deficiency extract (ST), the compounds 1 to 5 of Examples 5 and 6 for R28 cells The protective efficacy of was compared.
상기 시험예 3과 동일한 방법으로, R28 세포 생존율을 측정하되, 비교예 1, 실시예 1 및 2의 결명자 추출물(ST), 결명 새싹 추출물(STS, STS-C, STS-385, STS-465, STS-645 및 STS-780) 대신 실시예 5 및 6의 화합물 1 내지 5를 50 μM, 16.6 μM, 5.55 μM로 처리하였으며, 그 결과를 도 6에 나타내었다.In the same manner as in Test Example 3, the cell viability of R28 was measured, but the extract extract (ST), the extract extract of shoot (STS, STS-C, STS-385, STS-465) of Comparative Example 1, Examples 1 and 2, STS-645 and STS-780) were treated with 50 μM, 16.6 μM, and 5.55 μM of compounds 1 to 5 of Examples 5 and 6, and the results are shown in FIG. 6.
도 6에 나타난 바와 같이, 글루타메이트에 의해서 유발되는 신경독성으로부터 화합물 1 내지 5는 R28 세포를 보호하는 효과를 보였으며, 상기 화합물들은 결명 새싹 추출물(STS)에 상대적 함량이 매우 높거나 또는 결명 새싹 추출물(STS)에 주로 존재하는 화합물들이므로 결명 새싹 추출물(STS) 의 R28 세포 보호 효능이 결명자 추출물(ST) 의 효능보다 뛰어난 이유를 확인할 수 있었다.As shown in FIG. 6, Compounds 1 to 5 exhibited an effect of protecting R28 cells from neurotoxicity induced by glutamate, and the compounds have a relatively high content in the Sprout Sprout Extract (STS) or the Sprout Sprout Extract. Since the compounds are mainly present in (STS), it was confirmed that the R28 cell protective effect of the shoot sprout extract (STS) is superior to that of the shooter extract (ST).
[[ 시험예Test Example 5]  5] 실시예Example 1 및 2의 7종의  7 kinds of 1 and 2 결명Fault 새싹 추출물의 해마신경세포(HT-22) 보호 효능 Protective Effect of Sprout Extract on Hippocampal Neuronal Cells (HT-22)
상기 비교예 1 및 실시예 1에서 수득된 결명자 추출물(ST)과 결명 새싹 추출물(STS), 및 상기 실시예 2에서 수득된 다양한 빛 조건 하에서 재배된 결명 새싹 추출물(STS-C, STS-385, STS-465, STS-645, STS-780) 의 해마신경세포(HT-22) 보호 효능을 확인하기 위해 하기와 같은 실험을 수행하였다. Clarifier extract (ST) and Clarified sprout extract (STS) obtained in Comparative Examples 1 and 1, and Cultivated sprout extract (STS-C, STS-385, cultivated under various light conditions obtained in Example 2) STS-465, STS-645, STS-780) was carried out the following experiment to confirm the protective effect of hippocampal neurons (HT-22).
구체적으로, HT-22 세포는 10% FBS(fetal bovine serum), 100 U/mL 페니실린, 100 μg/mL 스트렙토마이신이 포함된 DMEM(Dulbecco’s modified eagle’s medium)/low glucose 배지에서 37.5 ℃ 인큐베이터에서 5% CO 2 조건하에서 배양하고, 2일마다 0.05% 트립신(Trypsin)을 사용하여 계대배양한 후, 96 플레이트에 3X10 3의 밀도로 접종하고 24시간 배양하였다. 상기 배양된 HT-22 세포에 상기 7종의 추출물들을 도 7의 (A) 내지 (C)의 농도로 처리하고 2시간 후, 5 mM 글루타메이트(glutamate)를 첨가하여 22시간 배양하였다. 그런 다음, 세포 생존률을 측정하기 위해 10 μL EZ-cytox를 처리하여 2시간 유지하였으며, UV 흡광도 450 nm에서 측정하였다.Specifically, HT-22 cells are 10% fetal bovine serum (FBS), 100 U / mL penicillin, and 100 μg / mL streptomycin containing DMEM (Dulbecco's modified eagle's medium), 5% in a 37.5 ° C incubator in low glucose medium. Incubated under CO 2 conditions, subcultured with 0.05% trypsin every 2 days, inoculated at a density of 3 × 10 3 in 96 plates and cultured for 24 hours. The 7 extracts were treated to the cultured HT-22 cells at the concentrations of (A) to (C) of FIG. 7, and 2 hours later, 5 mM glutamate was added and cultured for 22 hours. Then, 10 μL EZ-cytox was treated and maintained for 2 hours to measure cell viability, and UV absorbance was measured at 450 nm.
도 7에서 (A)는 글루타메이트에 의해서 유발되는 산화적 스트레스로부터 3가지 농도에서 상기 비교예 1의 결명자 추출물(ST), 상기 실시예 1의 결명 새싹 추출물(STS), 상기 실시예 2의 빛 조건을 다르게 하여 재배된 5종의 결명 새싹 추출물(STS-C, STS-385, STS-465, STS-645, STS-780)의 HT-22 세포 보호 효능을 세포 생존률로 나타낸 결과이다. 도 7에서 (B) 및 (C)는 5가지 농도에서 글루타메이트에 의해서 유발되는 산화적 스트레스로부터 상기 비교예 1의 결명자 추출물(ST) 및 상기 실시예 1의 결명 새싹 추출물(STS)의 HT-22 세포 보호 효능을 세포 생존률로 나타낸 결과이다.In Figure 7 (A) is the light extract of Example 2, the extract of the glenja extract of Comparative Example 1 (ST), the shoot sprout of Example 1 (STS) at three concentrations from the oxidative stress caused by glutamate HT-22 cell protection effect of the five species of sprout sprouts (STS-C, STS-385, STS-465, STS-645, STS-780) cultivated differently to the cell survival rate. In Figure 7 (B) and (C) is HT-22 of the deficiency extract (ST) of the Comparative Example 1 and the deficiency sprout extract (STS) of Example 1 from the oxidative stress caused by glutamate at five concentrations It is the result of cell protection effect by cell viability.
도 7의 (A)에 나타난 바와 같이, 모든 추출물 처리군에서 글루타메이트에 의해서 유발되는 산화적 스트레스로부터 HT-22 세포를 보호하는 효과를 보였으나, 특히 도 7의 (B), (C)에 나타난 바와 같이, 상기 실시예 1의 결명 새싹 추출물(STS)은 상기 비교예 1의 결명자 추출물(ST)에 비해 우수한 HT-22 세포 보호 효능을 나타내는 것을 확인하였다.As shown in (A) of FIG. 7, all extract treatment groups showed an effect of protecting HT-22 cells from oxidative stress caused by glutamate, but particularly shown in (B) and (C) of FIG. 7. As described above, it was confirmed that the seedling sprout extract (STS) of Example 1 exhibited superior HT-22 cell protection efficacy compared to the extractor extract (ST) of Comparative Example 1.
[[ 시험예Test Example 6]  6] 실시예Example 5 및 6의  5 and 6 결명Fault 새싹 추출물로부터 분리된 화합물 1 내지 5의 해마신경세포(HT-22) 보호 효능 확인 Confirmation of Protective Effect of Hippocampal Neuronal Cells (HT-22) of Compounds 1 to 5 Isolated from Sprout Extract
상기 시험예 5의 결명 새싹 추출물(STS)의 해마신경세포(HT-22) 보호 효능이 결명자 추출물(ST)보다 뛰어난 이유를 확인하기 위하여, R28 세포에 대한 실시예 5 및 6의 화합물 1 내지 5 및 비교예 2의 화합물 A 내지 X의 보호 효능을 비교하였다. In order to confirm why hippocampal nerve cell (HT-22) protective effect of the Sprout bud extract (STS) of Test Example 5 is superior to the deficiency extract (ST), the compounds 1 to 5 of R28 cells 1 to 5 And protective efficacy of Compounds A to X of Comparative Example 2.
상기 시험예 5와 동일한 방법으로 HT-22 세포 생존율을 측정하되, 비교예 1, 실시예 1 및 2의 결명자 추출물(ST), 결명 새싹 추출물(STS, STS-C, STS-385, STS-465, STS-645 및 STS-780) 대신 실시예 5 및 6의 화합물 1 내지 5를 50 μM, 16.6 μM, 5.55 μM로 처리하였으며, 그 결과를 도 8에 나타내었다.HT-22 cell viability was measured in the same manner as in Test Example 5, Comparative Example 1, Example 1, Example 2 extract gyeongjab (ST), the shoot sprout extract (STS, STS-C, STS-385, STS-465 , STS-645 and STS-780) instead of compounds 1 to 5 of Examples 5 and 6 were treated with 50 μM, 16.6 μM, 5.55 μM, and the results are shown in FIG. 8.
도 8에 나타난 바와 같이, 글루타메이트에 의해서 유발되는 신경독성으로부터 실시예 5 및 6의 화합물 1 내지 5와 비교예 2의 피크 K와 피크 T에 각각 해당하는 화합물 K 및 화합물 T이 HT-22 세포를 보호하는 효과를 나타냈다. 즉, 상기 화합물 1 내지 5는 결명 새싹 추출물(STS)에 상대적 함량이 매우 높거나 또는 결명 새싹 추출물(STS)에 주로 존재하는 화합물들이므로 결명 새싹 추출물(STS)의 HT-22 세포 보호 효능이 결명자 추출물(ST)의 효능보다 뛰어난 이유를 확인할 수 있었다.As shown in FIG. 8, compounds K and T corresponding to peak K and peak T of compounds 1 to 5 and comparative example 2 of Examples 5 and 6 and neurotoxicity induced by glutamate, respectively, were used for HT-22 cells. It showed a protective effect. That is, the compounds 1 to 5 are compounds having a relatively high content in the Sprout Sprout Extract (STS) or are mainly present in the Sprout Sprout Extract (STS), and thus the HT-22 cell protection effect of the Sprout Sprout Extract (STS) is deficiency. Reasons superior to the efficacy of the extract (ST) could be confirmed.
상기 시험예 1, 시험예 4 및 시험예 6에서 평가한 성분별 생리활성은 하기 표 1과 같다. 하기 표 1은 실시예 1의 결명 새싹 추출물(STS)로부터 분리한 성분들의 생리활성에 관한 것이다.The physiological activity of each component evaluated in Test Example 1, Test Example 4 and Test Example 6 is shown in Table 1 below. Table 1 below relates to the physiological activity of the components isolated from the bud extract (STS) of Example 1.
Figure PCTKR2019000399-appb-img-000020
Figure PCTKR2019000399-appb-img-000020

Claims (17)

  1. 하기 화학식 1의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물.A naphthopyrone derivative of Formula 1, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof.
    [화학식 1][Formula 1]
    Figure PCTKR2019000399-appb-img-000021
    Figure PCTKR2019000399-appb-img-000021
  2. 하기 화학식 1 내지 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물의 제조방법으로서, 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹(sprout)으로부터 상기 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상을 분리하는 과정을 포함하는, 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물의 제조방법.To a method for manufacturing a formula 1-5 of naphthyl topa theory derivatives naphthyl topa theory derivative at least one selected from the group consisting of, their stereoisomers, their pharmaceutically acceptable salt, hydrate, or solvate thereof, gyeolmyeong (Cassia obtusifolia L. or Cassia tora L.) A process for separating at least one selected from the group consisting of the naphthopyron derivatives, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof from sprouts A method for producing a naphthopyron derivative, a stereoisomer thereof, a pharmaceutically acceptable salt thereof, a hydrate thereof, or a solvate thereof, comprising a.
    [화학식 1][Formula 1]
    Figure PCTKR2019000399-appb-img-000022
    Figure PCTKR2019000399-appb-img-000022
    [화학식 2][Formula 2]
    Figure PCTKR2019000399-appb-img-000023
    Figure PCTKR2019000399-appb-img-000023
    [화학식 3][Formula 3]
    Figure PCTKR2019000399-appb-img-000024
    Figure PCTKR2019000399-appb-img-000024
    [화학식 4][Formula 4]
    Figure PCTKR2019000399-appb-img-000025
    Figure PCTKR2019000399-appb-img-000025
    [화학식 5][Formula 5]
    Figure PCTKR2019000399-appb-img-000026
    Figure PCTKR2019000399-appb-img-000026
  3. 제2항에 있어서, 상기 제조방법은 결명 새싹에서 물, 탄소수 1 내지 6의 알코올, 및 이들의 혼합 용매로 이루어진 군으로부터 선택된 용매로 추출하는 과정을 더 포함하는 것인, 제조방법.The method according to claim 2, wherein the manufacturing method further comprises extracting a solvent selected from the group consisting of water, alcohol having 1 to 6 carbon atoms, and a mixed solvent thereof in the shoot sprout.
  4. 제2항에 있어서, 상기 제조방법은 물, 에틸아세테이트, 헥산, 염화메틸렌, 클로로포름, 메탄올, 에탄올, 아세톤, 및 이들의 혼합 용매로 이루어진 군으로부터 선택된 하나 이상으로 분획하는 과정을 더 포함하는 것인, 제조방법.According to claim 2, wherein the method further comprises the step of fractionating with one or more selected from the group consisting of water, ethyl acetate, hexane, methylene chloride, chloroform, methanol, ethanol, acetone, and a mixed solvent thereof. , Manufacturing method.
  5. 하기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 산화적 스트레스로부터의 신경세포 보호용 또는 신경세포 사멸 억제용 조성물.One or more naphthopyrone derivatives selected from the group consisting of naphthopyrone derivatives of the following Chemical Formulas 1 to 5, stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof. , Cassia obtusifolia L. (or Cassia tora L.) bud extract comprising the same, or a bud fraction containing the sprout as an active ingredient, for the protection of neurons from oxidative stress or for the suppression of neuronal cell death.
    [화학식 1][Formula 1]
    Figure PCTKR2019000399-appb-img-000027
    Figure PCTKR2019000399-appb-img-000027
    [화학식 2][Formula 2]
    Figure PCTKR2019000399-appb-img-000028
    Figure PCTKR2019000399-appb-img-000028
    [화학식 3][Formula 3]
    Figure PCTKR2019000399-appb-img-000029
    Figure PCTKR2019000399-appb-img-000029
    [화학식 4][Formula 4]
    Figure PCTKR2019000399-appb-img-000030
    Figure PCTKR2019000399-appb-img-000030
    [화학식 5][Formula 5]
    Figure PCTKR2019000399-appb-img-000031
    Figure PCTKR2019000399-appb-img-000031
  6. 제5항에 있어서, 상기 산화적 스트레스는 글루타메이트에 의해 유발된 것인, 조성물.The composition of claim 5, wherein the oxidative stress is caused by glutamate.
  7. 제5항에 있어서, 상기 신경세포는 망막신경세포 또는 해마신경세포인, 조성물.The composition of claim 5, wherein the nerve cells are retinal nerve cells or hippocampal nerve cells.
  8. 하기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 망막신경세포 또는 해마신경세포의 손상 또는 사멸에 의해 유발되는 신경 손상성 질환의 예방 또는 치료용 조성물.At least one naphthopyrone derivative selected from the group consisting of naphthopyrone derivatives of Formula 1 to Formula 5, at least one selected from the group consisting of stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof Nerve damage caused by injury or death of retinal nerve cells or hippocampal nerve cells, comprising as an active ingredient, Cassia obtusifolia L. or Cassia tora L. sprout extract, or associative bud fraction comprising the same Composition for the prevention or treatment of sexual diseases.
    [화학식 1][Formula 1]
    Figure PCTKR2019000399-appb-img-000032
    Figure PCTKR2019000399-appb-img-000032
    [화학식 2][Formula 2]
    Figure PCTKR2019000399-appb-img-000033
    Figure PCTKR2019000399-appb-img-000033
    [화학식 3][Formula 3]
    Figure PCTKR2019000399-appb-img-000034
    Figure PCTKR2019000399-appb-img-000034
    [화학식 4][Formula 4]
    Figure PCTKR2019000399-appb-img-000035
    Figure PCTKR2019000399-appb-img-000035
    [화학식 5][Formula 5]
    Figure PCTKR2019000399-appb-img-000036
    Figure PCTKR2019000399-appb-img-000036
  9. 제8항에 있어서, 상기 신경 손상성 질환은 기억력 감퇴, 학습능력 감퇴, 우울장애, 근위축성 측삭 경화증(루게릭병), 파킨슨병, 뇌허혈성 신경손상, 당뇨병성 신경병증, 황반변성, 망막세포 손상에 의한 시력장애, 망막색소변성, 망막박리, 망막혈관폐쇄 및 녹내장으로 이루어진 군으로부터 선택된 하나 이상인, 조성물.The method of claim 8, wherein the neurological damage disorder is memory loss, learning ability decline, depressive disorder, amyotrophic lateral sclerosis (Lou Gehrig's disease), Parkinson's disease, cerebral ischemic nerve damage, diabetic neuropathy, macular degeneration, retinal cell damage At least one selected from the group consisting of visual acuity disorders, retinal pigmentation, retinal detachment, retinal vascular occlusion and glaucoma.
  10. 제8항에 있어서, 상기 망막신경세포 또는 해마신경세포의 손상 또는 사멸은 글루타메이트 신경독성에 의한 것인, 조성물.The composition of claim 8, wherein the damage or killing of the retinal neurons or hippocampal neurons is by glutamate neurotoxicity.
  11. 제8항에 있어서, 상기 나프토파이론 유도체는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹으로부터 분리 또는 정제된 것인, 조성물.The composition of claim 8, wherein the naphthopyrone derivative is isolated or purified from the shoot ( Cassia obtusifolia L. or Cassia tora L.) sprout.
  12. 제11항에 있어서, 상기 나프토파이론 유도체는 결명 새싹 에탄올 추출물로부터 분리 또는 정제된 것인, 조성물.12. The composition of claim 11, wherein the naphthopyrone derivative is isolated or purified from the shoot sprout ethanol extract.
  13. 제8항에 있어서, 상기 나프토파이론 유도체는 결명 새싹 에탄올 추출물을 에틸아세테이트로 분획한 뒤, 상기 에틸아세테이트 분획물을 크로마토그래피로 분리하거나 또는 상기 에틸아세테이트 분획물을 다시 헥산, 염화메틸렌 및 메탄올의 혼합 용매로 분획하고 이를 크로마토그래피로 분리한 것인, 조성물.According to claim 8, wherein the naphthopyron derivatives are fractionated sprout ethanol extract fractions with ethyl acetate, the ethyl acetate fractions are separated by chromatography or the ethyl acetate fractions again mixed solvent of hexane, methylene chloride and methanol Fractions and separated by chromatography.
  14. 제8항에 있어서, 상기 추출물은 물, 탄소수 1 내지 6의 알코올, 및 이들의 혼합 용매로 구성된 군에서 선택된 용매로 추출한 것인, 조성물.The composition of claim 8, wherein the extract is extracted with a solvent selected from the group consisting of water, alcohols having 1 to 6 carbon atoms, and mixed solvents thereof.
  15. 제8항에 있어서, 상기 분획물은 물, 에틸아세테이트, 헥산, 염화메틸렌, 클로로포름, 메탄올, 에탄올, 아세톤, 및 이들의 혼합 용매로 이루어진 군으로부터 선택된 하나 이상으로 분획한 것인, 조성물.The composition of claim 8, wherein the fraction is fractionated with one or more selected from the group consisting of water, ethyl acetate, hexane, methylene chloride, chloroform, methanol, ethanol, acetone, and mixed solvents thereof.
  16. 제8항에 있어서, 상기 조성물은 약학 또는 식품 조성물인, 조성물.The composition of claim 8, wherein the composition is a pharmaceutical or food composition.
  17. 하기 화학식 1 내지 화학식 5의 나프토파이론 유도체로 이루어진 군으로부터 선택된 1종 이상의 나프토파이론 유도체, 이의 입체이성질체, 이의 약학적으로 허용가능한 염, 이의 수화물, 또는 이의 용매화물로 이루어진 군으로부터 선택된 하나 이상, 그를 포함하는 결명( Cassia obtusifolia L. 또는 Cassia tora L.) 새싹 추출물, 또는 그를 포함하는 결명 새싹 분획물을 유효성분으로 포함하는, 항산화용 조성물.At least one naphthopyrone derivative selected from the group consisting of naphthopyrone derivatives of Formula 1 to Formula 5, at least one selected from the group consisting of stereoisomers thereof, pharmaceutically acceptable salts thereof, hydrates thereof, or solvates thereof , Cassia obtusifolia L. or Cassia tora L. Sprout extract containing it, or Claw sprout fraction comprising the same as an active ingredient, antioxidant composition.
    [화학식 1][Formula 1]
    Figure PCTKR2019000399-appb-img-000037
    Figure PCTKR2019000399-appb-img-000037
    [화학식 2][Formula 2]
    Figure PCTKR2019000399-appb-img-000038
    Figure PCTKR2019000399-appb-img-000038
    [화학식 3][Formula 3]
    Figure PCTKR2019000399-appb-img-000039
    Figure PCTKR2019000399-appb-img-000039
    [화학식 4][Formula 4]
    Figure PCTKR2019000399-appb-img-000040
    Figure PCTKR2019000399-appb-img-000040
    [화학식 5][Formula 5]
    Figure PCTKR2019000399-appb-img-000041
    Figure PCTKR2019000399-appb-img-000041
PCT/KR2019/000399 2018-09-03 2019-01-10 Neuron-protective composition comprising cassia obtusifolia shoot-derived naphthopyrone derivative WO2020050462A1 (en)

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