JPWO2005027937A1 - Glycolipid-containing composition, use thereof, and production method thereof - Google Patents
Glycolipid-containing composition, use thereof, and production method thereof Download PDFInfo
- Publication number
- JPWO2005027937A1 JPWO2005027937A1 JP2005514037A JP2005514037A JPWO2005027937A1 JP WO2005027937 A1 JPWO2005027937 A1 JP WO2005027937A1 JP 2005514037 A JP2005514037 A JP 2005514037A JP 2005514037 A JP2005514037 A JP 2005514037A JP WO2005027937 A1 JPWO2005027937 A1 JP WO2005027937A1
- Authority
- JP
- Japan
- Prior art keywords
- glycolipid
- containing composition
- fraction
- glycolipids
- present
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 229930186217 Glycolipid Natural products 0.000 title claims abstract description 241
- HVCOBJNICQPDBP-UHFFFAOYSA-N 3-[3-[3,5-dihydroxy-6-methyl-4-(3,4,5-trihydroxy-6-methyloxan-2-yl)oxyoxan-2-yl]oxydecanoyloxy]decanoic acid;hydrate Chemical compound O.OC1C(OC(CC(=O)OC(CCCCCCC)CC(O)=O)CCCCCCC)OC(C)C(O)C1OC1C(O)C(O)C(O)C(C)O1 HVCOBJNICQPDBP-UHFFFAOYSA-N 0.000 title claims abstract description 208
- 239000000203 mixture Substances 0.000 title claims abstract description 158
- 238000004519 manufacturing process Methods 0.000 title claims description 16
- 239000004367 Lipase Substances 0.000 claims abstract description 55
- 108090001060 Lipase Proteins 0.000 claims abstract description 55
- 102000004882 Lipase Human genes 0.000 claims abstract description 55
- 235000019421 lipase Nutrition 0.000 claims abstract description 55
- 238000000034 method Methods 0.000 claims abstract description 50
- 238000011282 treatment Methods 0.000 claims abstract description 33
- 235000009337 Spinacia oleracea Nutrition 0.000 claims abstract description 26
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 70
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 26
- FIJGNIAJTZSERN-DQQGJSMTSA-N monogalactosyl-diacylglycerol Chemical compound CCCCCCCCCCCCCCCC(=O)O[C@H](COC(=O)CCCCCCCCCCCC)CO[C@@H]1O[C@@H](CO)[C@H](O)[C@H](O)[C@@H]1O FIJGNIAJTZSERN-DQQGJSMTSA-N 0.000 claims description 24
- QZXMUPATKGLZAP-DXLAUQRQSA-N [(2S)-1-hexadecanoyloxy-3-[(2R,3R,4S,5R,6R)-3,4,5-trihydroxy-6-[[(2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxymethyl]oxan-2-yl]oxypropan-2-yl] (9Z,12Z)-octadeca-9,12-dienoate Chemical compound O[C@@H]1[C@H](O)[C@@H](O)[C@H](OC[C@@H](COC(=O)CCCCCCCCCCCCCCC)OC(=O)CCCCCCC\C=C/C\C=C/CCCCC)O[C@@H]1CO[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 QZXMUPATKGLZAP-DXLAUQRQSA-N 0.000 claims description 23
- -1 sulfoquinovosyl diacylglycerol Chemical class 0.000 claims description 23
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 22
- 239000000126 substance Substances 0.000 claims description 19
- 241000196324 Embryophyta Species 0.000 claims description 17
- 239000003960 organic solvent Substances 0.000 claims description 14
- 239000002246 antineoplastic agent Substances 0.000 claims description 13
- 239000002994 raw material Substances 0.000 claims description 11
- 241001474374 Blennius Species 0.000 claims description 10
- 239000003112 inhibitor Substances 0.000 claims description 10
- 239000008194 pharmaceutical composition Substances 0.000 claims description 10
- 238000010828 elution Methods 0.000 claims description 9
- 230000002209 hydrophobic effect Effects 0.000 claims description 8
- 150000002759 monoacylglycerols Chemical class 0.000 claims description 8
- 238000004587 chromatography analysis Methods 0.000 claims description 7
- 238000005406 washing Methods 0.000 claims description 7
- 239000004480 active ingredient Substances 0.000 claims description 6
- 239000000419 plant extract Substances 0.000 claims description 4
- 235000013311 vegetables Nutrition 0.000 claims description 4
- 241000219315 Spinacia Species 0.000 claims 1
- 230000002401 inhibitory effect Effects 0.000 abstract description 53
- 206010028980 Neoplasm Diseases 0.000 abstract description 36
- 201000011510 cancer Diseases 0.000 abstract description 27
- 244000300264 Spinacia oleracea Species 0.000 abstract description 25
- 230000010261 cell growth Effects 0.000 abstract description 19
- 235000013376 functional food Nutrition 0.000 abstract description 18
- 230000000694 effects Effects 0.000 abstract description 17
- 230000000259 anti-tumor effect Effects 0.000 abstract description 12
- 230000001766 physiological effect Effects 0.000 abstract description 9
- 230000001093 anti-cancer Effects 0.000 abstract description 5
- 230000002265 prevention Effects 0.000 abstract description 5
- 229940040461 lipase Drugs 0.000 description 52
- 239000000243 solution Substances 0.000 description 21
- 239000000284 extract Substances 0.000 description 19
- 238000002360 preparation method Methods 0.000 description 14
- 238000006243 chemical reaction Methods 0.000 description 12
- 239000002504 physiological saline solution Substances 0.000 description 12
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 10
- 230000005917 in vivo anti-tumor Effects 0.000 description 10
- 239000000469 ethanolic extract Substances 0.000 description 9
- 238000002474 experimental method Methods 0.000 description 9
- 235000013305 food Nutrition 0.000 description 9
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 8
- 235000014113 dietary fatty acids Nutrition 0.000 description 8
- 239000000194 fatty acid Substances 0.000 description 8
- 229930195729 fatty acid Natural products 0.000 description 8
- 238000000746 purification Methods 0.000 description 8
- 208000030507 AIDS Diseases 0.000 description 7
- 241000699666 Mus <mouse, genus> Species 0.000 description 7
- 201000010099 disease Diseases 0.000 description 7
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 7
- 239000003814 drug Substances 0.000 description 7
- 230000005764 inhibitory process Effects 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- 238000004809 thin layer chromatography Methods 0.000 description 7
- 102000003960 Ligases Human genes 0.000 description 6
- 108090000364 Ligases Proteins 0.000 description 6
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 150000004665 fatty acids Chemical class 0.000 description 6
- 239000011347 resin Substances 0.000 description 6
- 229920005989 resin Polymers 0.000 description 6
- 238000003756 stirring Methods 0.000 description 6
- 239000002775 capsule Substances 0.000 description 5
- 238000000605 extraction Methods 0.000 description 5
- 239000008187 granular material Substances 0.000 description 5
- 238000002156 mixing Methods 0.000 description 5
- 241000699800 Cricetinae Species 0.000 description 4
- 229920002472 Starch Polymers 0.000 description 4
- 208000005718 Stomach Neoplasms Diseases 0.000 description 4
- 238000011161 development Methods 0.000 description 4
- 206010017758 gastric cancer Diseases 0.000 description 4
- 230000012010 growth Effects 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 230000002829 reductive effect Effects 0.000 description 4
- 238000010992 reflux Methods 0.000 description 4
- 150000003839 salts Chemical class 0.000 description 4
- 239000008107 starch Substances 0.000 description 4
- 235000019698 starch Nutrition 0.000 description 4
- 201000011549 stomach cancer Diseases 0.000 description 4
- 238000010254 subcutaneous injection Methods 0.000 description 4
- 239000007929 subcutaneous injection Substances 0.000 description 4
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 108010010803 Gelatin Proteins 0.000 description 3
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 3
- 241000699670 Mus sp. Species 0.000 description 3
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 3
- 206010039491 Sarcoma Diseases 0.000 description 3
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 3
- 229930006000 Sucrose Natural products 0.000 description 3
- 208000002495 Uterine Neoplasms Diseases 0.000 description 3
- 230000006907 apoptotic process Effects 0.000 description 3
- 238000007796 conventional method Methods 0.000 description 3
- 238000001514 detection method Methods 0.000 description 3
- 229940088598 enzyme Drugs 0.000 description 3
- 238000001914 filtration Methods 0.000 description 3
- 235000013373 food additive Nutrition 0.000 description 3
- 239000002778 food additive Substances 0.000 description 3
- 239000008273 gelatin Substances 0.000 description 3
- 229920000159 gelatin Polymers 0.000 description 3
- 235000019322 gelatine Nutrition 0.000 description 3
- 235000011852 gelatine desserts Nutrition 0.000 description 3
- 239000003018 immunosuppressive agent Substances 0.000 description 3
- 229940125721 immunosuppressive agent Drugs 0.000 description 3
- 239000007924 injection Substances 0.000 description 3
- 238000002347 injection Methods 0.000 description 3
- 239000003456 ion exchange resin Substances 0.000 description 3
- 229920003303 ion-exchange polymer Polymers 0.000 description 3
- 150000002632 lipids Chemical class 0.000 description 3
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 3
- 238000011580 nude mouse model Methods 0.000 description 3
- 239000000049 pigment Substances 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 239000000741 silica gel Substances 0.000 description 3
- 229910002027 silica gel Inorganic materials 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 239000005720 sucrose Substances 0.000 description 3
- 239000003826 tablet Substances 0.000 description 3
- 150000004670 unsaturated fatty acids Chemical class 0.000 description 3
- 206010046766 uterine cancer Diseases 0.000 description 3
- GVJHHUAWPYXKBD-UHFFFAOYSA-N (±)-α-Tocopherol Chemical compound OC1=C(C)C(C)=C2OC(CCCC(C)CCCC(C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-UHFFFAOYSA-N 0.000 description 2
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 2
- 240000002900 Arthrospira platensis Species 0.000 description 2
- 235000016425 Arthrospira platensis Nutrition 0.000 description 2
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 2
- 230000006820 DNA synthesis Effects 0.000 description 2
- 239000004375 Dextrin Substances 0.000 description 2
- 229920001353 Dextrin Polymers 0.000 description 2
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 2
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 2
- 241000699660 Mus musculus Species 0.000 description 2
- 102000019280 Pancreatic lipases Human genes 0.000 description 2
- 108050006759 Pancreatic lipases Proteins 0.000 description 2
- 241000015177 Saccharina japonica Species 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000003463 adsorbent Substances 0.000 description 2
- MBMBGCFOFBJSGT-KUBAVDMBSA-N all-cis-docosa-4,7,10,13,16,19-hexaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCC(O)=O MBMBGCFOFBJSGT-KUBAVDMBSA-N 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 230000002421 anti-septic effect Effects 0.000 description 2
- 239000003443 antiviral agent Substances 0.000 description 2
- YZXBAPSDXZZRGB-DOFZRALJSA-N arachidonic acid Chemical compound CCCCC\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O YZXBAPSDXZZRGB-DOFZRALJSA-N 0.000 description 2
- 239000011230 binding agent Substances 0.000 description 2
- 125000004432 carbon atom Chemical group C* 0.000 description 2
- 239000001768 carboxy methyl cellulose Substances 0.000 description 2
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 2
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 2
- 229940105329 carboxymethylcellulose Drugs 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 229930002875 chlorophyll Natural products 0.000 description 2
- 235000019804 chlorophyll Nutrition 0.000 description 2
- ATNHDLDRLWWWCB-AENOIHSZSA-M chlorophyll a Chemical compound C1([C@@H](C(=O)OC)C(=O)C2=C3C)=C2N2C3=CC(C(CC)=C3C)=[N+]4C3=CC3=C(C=C)C(C)=C5N3[Mg-2]42[N+]2=C1[C@@H](CCC(=O)OC\C=C(/C)CCC[C@H](C)CCC[C@H](C)CCCC(C)C)[C@H](C)C2=C5 ATNHDLDRLWWWCB-AENOIHSZSA-M 0.000 description 2
- 239000003086 colorant Substances 0.000 description 2
- 238000004440 column chromatography Methods 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- URGJWIFLBWJRMF-JGVFFNPUSA-N ddTTP Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)CC1 URGJWIFLBWJRMF-JGVFFNPUSA-N 0.000 description 2
- 235000019425 dextrin Nutrition 0.000 description 2
- 150000001982 diacylglycerols Chemical class 0.000 description 2
- 239000010432 diamond Substances 0.000 description 2
- 239000007884 disintegrant Substances 0.000 description 2
- 239000002552 dosage form Substances 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- IDGUHHHQCWSQLU-UHFFFAOYSA-N ethanol;hydrate Chemical compound O.CCO IDGUHHHQCWSQLU-UHFFFAOYSA-N 0.000 description 2
- 239000002031 ethanolic fraction Substances 0.000 description 2
- 238000011049 filling Methods 0.000 description 2
- 239000000796 flavoring agent Substances 0.000 description 2
- 235000013402 health food Nutrition 0.000 description 2
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 2
- 239000001341 hydroxy propyl starch Substances 0.000 description 2
- 235000013828 hydroxypropyl starch Nutrition 0.000 description 2
- 208000026278 immune system disease Diseases 0.000 description 2
- 230000001506 immunosuppresive effect Effects 0.000 description 2
- 239000000411 inducer Substances 0.000 description 2
- 239000004615 ingredient Substances 0.000 description 2
- 230000002452 interceptive effect Effects 0.000 description 2
- 239000008101 lactose Substances 0.000 description 2
- 239000000314 lubricant Substances 0.000 description 2
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 2
- 229930014626 natural product Natural products 0.000 description 2
- 229940116369 pancreatic lipase Drugs 0.000 description 2
- 159000000001 potassium salts Chemical class 0.000 description 2
- 239000003755 preservative agent Substances 0.000 description 2
- 230000002335 preservative effect Effects 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 230000008439 repair process Effects 0.000 description 2
- 230000002441 reversible effect Effects 0.000 description 2
- 150000004671 saturated fatty acids Chemical class 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- 229910052708 sodium Inorganic materials 0.000 description 2
- 238000000638 solvent extraction Methods 0.000 description 2
- 239000003549 soybean oil Substances 0.000 description 2
- 235000012424 soybean oil Nutrition 0.000 description 2
- 239000003381 stabilizer Substances 0.000 description 2
- 239000000829 suppository Substances 0.000 description 2
- 239000004094 surface-active agent Substances 0.000 description 2
- 229940124597 therapeutic agent Drugs 0.000 description 2
- 238000002054 transplantation Methods 0.000 description 2
- 235000021122 unsaturated fatty acids Nutrition 0.000 description 2
- YUFFSWGQGVEMMI-JLNKQSITSA-N (7Z,10Z,13Z,16Z,19Z)-docosapentaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCCCCC(O)=O YUFFSWGQGVEMMI-JLNKQSITSA-N 0.000 description 1
- HOBAELRKJCKHQD-UHFFFAOYSA-N (8Z,11Z,14Z)-8,11,14-eicosatrienoic acid Natural products CCCCCC=CCC=CCC=CCCCCCCC(O)=O HOBAELRKJCKHQD-UHFFFAOYSA-N 0.000 description 1
- OYHQOLUKZRVURQ-NTGFUMLPSA-N (9Z,12Z)-9,10,12,13-tetratritiooctadeca-9,12-dienoic acid Chemical compound C(CCCCCCC\C(=C(/C\C(=C(/CCCCC)\[3H])\[3H])\[3H])\[3H])(=O)O OYHQOLUKZRVURQ-NTGFUMLPSA-N 0.000 description 1
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 description 1
- JLPULHDHAOZNQI-ZTIMHPMXSA-N 1-hexadecanoyl-2-(9Z,12Z-octadecadienoyl)-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCC\C=C/C\C=C/CCCCC JLPULHDHAOZNQI-ZTIMHPMXSA-N 0.000 description 1
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 1
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 description 1
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 description 1
- 244000215068 Acacia senegal Species 0.000 description 1
- 241000222518 Agaricus Species 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 241000584629 Aosa Species 0.000 description 1
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 101100263837 Bovine ephemeral fever virus (strain BB7721) beta gene Proteins 0.000 description 1
- 235000011299 Brassica oleracea var botrytis Nutrition 0.000 description 1
- 235000017647 Brassica oleracea var italica Nutrition 0.000 description 1
- 240000003259 Brassica oleracea var. botrytis Species 0.000 description 1
- 235000008534 Capsicum annuum var annuum Nutrition 0.000 description 1
- 240000008384 Capsicum annuum var. annuum Species 0.000 description 1
- 241000195585 Chlamydomonas Species 0.000 description 1
- 241000195649 Chlorella <Chlorellales> Species 0.000 description 1
- 241000195651 Chlorella sp. Species 0.000 description 1
- 241000195628 Chlorophyta Species 0.000 description 1
- 241000251730 Chondrichthyes Species 0.000 description 1
- 229920002261 Corn starch Polymers 0.000 description 1
- 241000192700 Cyanobacteria Species 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- 230000004543 DNA replication Effects 0.000 description 1
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 description 1
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 description 1
- 235000021298 Dihomo-γ-linolenic acid Nutrition 0.000 description 1
- 235000021294 Docosapentaenoic acid Nutrition 0.000 description 1
- 241000257465 Echinoidea Species 0.000 description 1
- 101100316840 Enterobacteria phage P4 Beta gene Proteins 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 239000001856 Ethyl cellulose Substances 0.000 description 1
- ZZSNKZQZMQGXPY-UHFFFAOYSA-N Ethyl cellulose Chemical compound CCOCC1OC(OC)C(OCC)C(OCC)C1OC1C(O)C(O)C(OC)C(CO)O1 ZZSNKZQZMQGXPY-UHFFFAOYSA-N 0.000 description 1
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 1
- KRHYYFGTRYWZRS-UHFFFAOYSA-N Fluorane Chemical compound F KRHYYFGTRYWZRS-UHFFFAOYSA-N 0.000 description 1
- 241000206671 Gelidium amansii Species 0.000 description 1
- 241001467355 Gigartina Species 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 241000206581 Gracilaria Species 0.000 description 1
- 241000703939 Gracilariopsis longissima Species 0.000 description 1
- 206010053759 Growth retardation Diseases 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 240000005979 Hordeum vulgare Species 0.000 description 1
- 235000007340 Hordeum vulgare Nutrition 0.000 description 1
- 229920002153 Hydroxypropyl cellulose Polymers 0.000 description 1
- 206010020880 Hypertrophy Diseases 0.000 description 1
- 108010093096 Immobilized Enzymes Proteins 0.000 description 1
- 102100034343 Integrase Human genes 0.000 description 1
- 240000008415 Lactuca sativa Species 0.000 description 1
- 238000000134 MTT assay Methods 0.000 description 1
- 231100000002 MTT assay Toxicity 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 229910002651 NO3 Inorganic materials 0.000 description 1
- 235000017879 Nasturtium officinale Nutrition 0.000 description 1
- 240000005407 Nasturtium officinale Species 0.000 description 1
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 description 1
- 239000005642 Oleic acid Substances 0.000 description 1
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 description 1
- 235000021314 Palmitic acid Nutrition 0.000 description 1
- 235000019483 Peanut oil Nutrition 0.000 description 1
- 240000009164 Petroselinum crispum Species 0.000 description 1
- 241000199919 Phaeophyceae Species 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 229920001214 Polysorbate 60 Polymers 0.000 description 1
- 241000206613 Pyropia yezoensis Species 0.000 description 1
- 108010092799 RNA-directed DNA polymerase Proteins 0.000 description 1
- 241000264279 Sargassum fusiforme Species 0.000 description 1
- 241001260874 Sargassum horneri Species 0.000 description 1
- 229920005654 Sephadex Polymers 0.000 description 1
- 239000012507 Sephadex™ Substances 0.000 description 1
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 1
- 235000021355 Stearic acid Nutrition 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 description 1
- 241000192589 Synechococcus elongatus PCC 7942 Species 0.000 description 1
- PPWHTZKZQNXVAE-UHFFFAOYSA-N Tetracaine hydrochloride Chemical compound Cl.CCCCNC1=CC=C(C(=O)OCCN(C)C)C=C1 PPWHTZKZQNXVAE-UHFFFAOYSA-N 0.000 description 1
- 244000269722 Thea sinensis Species 0.000 description 1
- 241000078013 Trichormus variabilis Species 0.000 description 1
- 241000196252 Ulva Species 0.000 description 1
- 241001261506 Undaria pinnatifida Species 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- 229930003427 Vitamin E Natural products 0.000 description 1
- YKTSYUJCYHOUJP-UHFFFAOYSA-N [O--].[Al+3].[Al+3].[O-][Si]([O-])([O-])[O-] Chemical compound [O--].[Al+3].[Al+3].[O-][Si]([O-])([O-])[O-] YKTSYUJCYHOUJP-UHFFFAOYSA-N 0.000 description 1
- JLKDXNDMDDENMB-ARFHVFGLSA-N [[(2r,3s,5r)-5-[2-(4-butylanilino)-6-oxo-3h-purin-9-yl]-3-hydroxyoxolan-2-yl]methoxy-hydroxyphosphoryl] phosphono hydrogen phosphate Chemical compound C1=CC(CCCC)=CC=C1NC(N1)=NC(=O)C2=C1N([C@@H]1O[C@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)[C@@H](O)C1)C=N2 JLKDXNDMDDENMB-ARFHVFGLSA-N 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- DPXJVFZANSGRMM-UHFFFAOYSA-N acetic acid;2,3,4,5,6-pentahydroxyhexanal;sodium Chemical compound [Na].CC(O)=O.OCC(O)C(O)C(O)C(O)C=O DPXJVFZANSGRMM-UHFFFAOYSA-N 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 229910052783 alkali metal Inorganic materials 0.000 description 1
- JAZBEHYOTPTENJ-JLNKQSITSA-N all-cis-5,8,11,14,17-icosapentaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O JAZBEHYOTPTENJ-JLNKQSITSA-N 0.000 description 1
- DTOSIQBPPRVQHS-PDBXOOCHSA-N alpha-linolenic acid Chemical compound CC\C=C/C\C=C/C\C=C/CCCCCCCC(O)=O DTOSIQBPPRVQHS-PDBXOOCHSA-N 0.000 description 1
- 235000020661 alpha-linolenic acid Nutrition 0.000 description 1
- PNEYBMLMFCGWSK-UHFFFAOYSA-N aluminium oxide Inorganic materials [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 description 1
- CEGOLXSVJUTHNZ-UHFFFAOYSA-K aluminium tristearate Chemical compound [Al+3].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O CEGOLXSVJUTHNZ-UHFFFAOYSA-K 0.000 description 1
- 229940024546 aluminum hydroxide gel Drugs 0.000 description 1
- 229940063655 aluminum stearate Drugs 0.000 description 1
- SMYKVLBUSSNXMV-UHFFFAOYSA-K aluminum;trihydroxide;hydrate Chemical compound O.[OH-].[OH-].[OH-].[Al+3] SMYKVLBUSSNXMV-UHFFFAOYSA-K 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 235000021342 arachidonic acid Nutrition 0.000 description 1
- 229940114079 arachidonic acid Drugs 0.000 description 1
- 239000012298 atmosphere Substances 0.000 description 1
- 235000013871 bee wax Nutrition 0.000 description 1
- 239000012166 beeswax Substances 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 235000008429 bread Nutrition 0.000 description 1
- CJZGTCYPCWQAJB-UHFFFAOYSA-L calcium stearate Chemical compound [Ca+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O CJZGTCYPCWQAJB-UHFFFAOYSA-L 0.000 description 1
- 239000008116 calcium stearate Substances 0.000 description 1
- 235000013539 calcium stearate Nutrition 0.000 description 1
- 229920003123 carboxymethyl cellulose sodium Polymers 0.000 description 1
- 229940084030 carboxymethylcellulose calcium Drugs 0.000 description 1
- 229940063834 carboxymethylcellulose sodium Drugs 0.000 description 1
- 210000000845 cartilage Anatomy 0.000 description 1
- 235000005487 catechin Nutrition 0.000 description 1
- 150000001765 catechin Chemical class 0.000 description 1
- ADRVNXBAWSRFAJ-UHFFFAOYSA-N catechin Natural products OC1Cc2cc(O)cc(O)c2OC1c3ccc(O)c(O)c3 ADRVNXBAWSRFAJ-UHFFFAOYSA-N 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 235000010980 cellulose Nutrition 0.000 description 1
- 239000012539 chromatography resin Substances 0.000 description 1
- 229940121657 clinical drug Drugs 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 235000014510 cooky Nutrition 0.000 description 1
- 235000005687 corn oil Nutrition 0.000 description 1
- 239000002285 corn oil Substances 0.000 description 1
- 239000008120 corn starch Substances 0.000 description 1
- 239000000287 crude extract Substances 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 239000000645 desinfectant Substances 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 235000015872 dietary supplement Nutrition 0.000 description 1
- 230000004069 differentiation Effects 0.000 description 1
- HOBAELRKJCKHQD-QNEBEIHSSA-N dihomo-γ-linolenic acid Chemical compound CCCCC\C=C/C\C=C/C\C=C/CCCCCCC(O)=O HOBAELRKJCKHQD-QNEBEIHSSA-N 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 229910001873 dinitrogen Inorganic materials 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 235000020669 docosahexaenoic acid Nutrition 0.000 description 1
- 229940090949 docosahexaenoic acid Drugs 0.000 description 1
- 235000015071 dressings Nutrition 0.000 description 1
- 239000003651 drinking water Substances 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- JAZBEHYOTPTENJ-UHFFFAOYSA-N eicosapentaenoic acid Natural products CCC=CCC=CCC=CCC=CCC=CCCCC(O)=O JAZBEHYOTPTENJ-UHFFFAOYSA-N 0.000 description 1
- 235000020673 eicosapentaenoic acid Nutrition 0.000 description 1
- 229960005135 eicosapentaenoic acid Drugs 0.000 description 1
- 239000000839 emulsion Substances 0.000 description 1
- 235000019325 ethyl cellulose Nutrition 0.000 description 1
- 229920001249 ethyl cellulose Polymers 0.000 description 1
- 229930003935 flavonoid Natural products 0.000 description 1
- 235000017173 flavonoids Nutrition 0.000 description 1
- 235000019634 flavors Nutrition 0.000 description 1
- 235000013355 food flavoring agent Nutrition 0.000 description 1
- 235000013350 formula milk Nutrition 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 239000003205 fragrance Substances 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 1
- WIGCFUFOHFEKBI-UHFFFAOYSA-N gamma-tocopherol Natural products CC(C)CCCC(C)CCCC(C)CCCC1CCC2C(C)C(O)C(C)C(C)C2O1 WIGCFUFOHFEKBI-UHFFFAOYSA-N 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- PEDCQBHIVMGVHV-UHFFFAOYSA-N glycerol group Chemical group OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 1
- 230000009036 growth inhibition Effects 0.000 description 1
- 230000035876 healing Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 150000003840 hydrochlorides Chemical class 0.000 description 1
- 239000008172 hydrogenated vegetable oil Substances 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 239000001863 hydroxypropyl cellulose Substances 0.000 description 1
- 235000010977 hydroxypropyl cellulose Nutrition 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 238000009776 industrial production Methods 0.000 description 1
- 238000010255 intramuscular injection Methods 0.000 description 1
- 239000007927 intramuscular injection Substances 0.000 description 1
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 description 1
- 239000007951 isotonicity adjuster Substances 0.000 description 1
- 235000015110 jellies Nutrition 0.000 description 1
- 239000008274 jelly Substances 0.000 description 1
- KQQKGWQCNNTQJW-UHFFFAOYSA-N linolenic acid Natural products CC=CCCC=CCC=CCCCCCCCC(O)=O KQQKGWQCNNTQJW-UHFFFAOYSA-N 0.000 description 1
- 229960004488 linolenic acid Drugs 0.000 description 1
- 239000012263 liquid product Substances 0.000 description 1
- 229940031703 low substituted hydroxypropyl cellulose Drugs 0.000 description 1
- 229960003511 macrogol Drugs 0.000 description 1
- HCWCAKKEBCNQJP-UHFFFAOYSA-N magnesium orthosilicate Chemical compound [Mg+2].[Mg+2].[O-][Si]([O-])([O-])[O-] HCWCAKKEBCNQJP-UHFFFAOYSA-N 0.000 description 1
- 239000000391 magnesium silicate Substances 0.000 description 1
- 229910052919 magnesium silicate Inorganic materials 0.000 description 1
- 235000019792 magnesium silicate Nutrition 0.000 description 1
- 235000019359 magnesium stearate Nutrition 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 230000000873 masking effect Effects 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 201000001441 melanoma Diseases 0.000 description 1
- 230000000684 melanotic effect Effects 0.000 description 1
- 229920000609 methyl cellulose Polymers 0.000 description 1
- 239000001923 methylcellulose Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 1
- SEEYREPSKCQBBF-UHFFFAOYSA-N n-methylmaleimide Chemical compound CN1C(=O)C=CC1=O SEEYREPSKCQBBF-UHFFFAOYSA-N 0.000 description 1
- 150000002823 nitrates Chemical class 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- QIQXTHQIDYTFRH-UHFFFAOYSA-N octadecanoic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 1
- OQCDKBAXFALNLD-UHFFFAOYSA-N octadecanoic acid Natural products CCCCCCCC(C)CCCCCCCCC(O)=O OQCDKBAXFALNLD-UHFFFAOYSA-N 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 235000019198 oils Nutrition 0.000 description 1
- 239000002674 ointment Substances 0.000 description 1
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 description 1
- 235000021313 oleic acid Nutrition 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000000312 peanut oil Substances 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 235000011197 perejil Nutrition 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- 239000000825 pharmaceutical preparation Substances 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 150000008442 polyphenolic compounds Chemical class 0.000 description 1
- 235000013824 polyphenols Nutrition 0.000 description 1
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 1
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 1
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- OVARTBFNCCXQKS-UHFFFAOYSA-N propan-2-one;hydrate Chemical compound O.CC(C)=O OVARTBFNCCXQKS-UHFFFAOYSA-N 0.000 description 1
- 235000013772 propylene glycol Nutrition 0.000 description 1
- 235000011962 puddings Nutrition 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 238000004366 reverse phase liquid chromatography Methods 0.000 description 1
- 238000012552 review Methods 0.000 description 1
- 239000003419 rna directed dna polymerase inhibitor Substances 0.000 description 1
- 235000012045 salad Nutrition 0.000 description 1
- 235000003441 saturated fatty acids Nutrition 0.000 description 1
- 239000008159 sesame oil Substances 0.000 description 1
- 235000011803 sesame oil Nutrition 0.000 description 1
- 238000010898 silica gel chromatography Methods 0.000 description 1
- RMAQACBXLXPBSY-UHFFFAOYSA-N silicic acid Chemical compound O[Si](O)(O)O RMAQACBXLXPBSY-UHFFFAOYSA-N 0.000 description 1
- 235000012239 silicon dioxide Nutrition 0.000 description 1
- 235000015424 sodium Nutrition 0.000 description 1
- 235000019333 sodium laurylsulphate Nutrition 0.000 description 1
- 159000000000 sodium salts Chemical class 0.000 description 1
- 235000014214 soft drink Nutrition 0.000 description 1
- 235000021055 solid food Nutrition 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 235000014347 soups Nutrition 0.000 description 1
- 235000013322 soy milk Nutrition 0.000 description 1
- 229940083466 soybean lecithin Drugs 0.000 description 1
- 229940082787 spirulina Drugs 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 229960004793 sucrose Drugs 0.000 description 1
- 150000003871 sulfonates Chemical class 0.000 description 1
- 150000003467 sulfuric acid derivatives Chemical class 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000006491 synthase reaction Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 239000006188 syrup Substances 0.000 description 1
- 235000020357 syrup Nutrition 0.000 description 1
- 239000000454 talc Substances 0.000 description 1
- 229910052623 talc Inorganic materials 0.000 description 1
- 235000013616 tea Nutrition 0.000 description 1
- TUNFSRHWOTWDNC-HKGQFRNVSA-N tetradecanoic acid Chemical compound CCCCCCCCCCCCC[14C](O)=O TUNFSRHWOTWDNC-HKGQFRNVSA-N 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 210000001541 thymus gland Anatomy 0.000 description 1
- 235000015112 vegetable and seed oil Nutrition 0.000 description 1
- 239000008158 vegetable oil Substances 0.000 description 1
- 230000009385 viral infection Effects 0.000 description 1
- 230000003612 virological effect Effects 0.000 description 1
- 229940046009 vitamin E Drugs 0.000 description 1
- 235000019165 vitamin E Nutrition 0.000 description 1
- 239000011709 vitamin E Substances 0.000 description 1
- 238000003809 water extraction Methods 0.000 description 1
- 239000001993 wax Substances 0.000 description 1
- 235000013618 yogurt Nutrition 0.000 description 1
- HBOMLICNUCNMMY-XLPZGREQSA-N zidovudine Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](N=[N+]=[N-])C1 HBOMLICNUCNMMY-XLPZGREQSA-N 0.000 description 1
- 229960002555 zidovudine Drugs 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/21—Amaranthaceae (Amaranth family), e.g. pigweed, rockwort or globe amaranth
-
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
- A61K31/7032—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a polyol, i.e. compounds having two or more free or esterified hydroxy groups, including the hydroxy group involved in the glycosidic linkage, e.g. monoglucosyldiacylglycerides, lactobionic acid, gangliosides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
-
- 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
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Public Health (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Medicinal Chemistry (AREA)
- Veterinary Medicine (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Epidemiology (AREA)
- Botany (AREA)
- Mycology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- General Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Alternative & Traditional Medicine (AREA)
- Medical Informatics (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Nutrition Science (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Medicines Containing Plant Substances (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
本発明の糖脂質含有組成物は、例えばホウレン草（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）から簡便な方法で精製することができ、モノガラクトシルジアシルグリセロール、ジガラクトシルジアシルグリセロール、およびスルホキノボシルジアシルグリセロールの３種類の糖脂質を高い割合で含有する。この糖脂質含有組成物は、ＤＮＡ合成酵素阻害活性、癌細胞増殖抑制活性、抗腫瘍活性といった有用な生理活性を有するが、リパーゼ処理によって更に強い生理活性が得られ、例えば抗癌作用または癌予防効果をもつ機能性食品などに有用である。The glycolipid-containing composition of the present invention can be purified by, for example, spinach (Spinacia oleracea L.) by a simple method. Contains a high proportion of glycolipids. This glycolipid-containing composition has useful physiological activities such as DNA synthase inhibitory activity, cancer cell growth inhibitory activity, and antitumor activity, but a stronger physiological activity can be obtained by lipase treatment, such as anticancer activity or cancer prevention. It is useful for functional foods with effects.
Description
本発明は、医薬および食品（特に機能性食品）への利用が可能な有用な生理活性を有する糖脂質含有組成物、その用途およびその製造方法に関するものである。 The present invention relates to a glycolipid-containing composition having useful physiological activity that can be used in medicines and foods (especially functional foods), its use, and a production method thereof.
真核生物のＤＮＡ合成酵素（ＤＮＡポリメラーゼ）は、これまでα、β、γ、δ、ε、ζ、η、θ、ι、κ、λ、μ、及びσの１３種類のＤＮＡ合成酵素が知られている。これらのＤＮＡ合成酵素群は、細胞の増殖、***、分化などに関与しているが、α型はＤＮＡ複製、β型は修復と組換え、δ型及びε型は複製と修復の双方を担うといった具合にタイプによって異なる機能を有することが知られている。 As for eukaryotic DNA synthase (DNA polymerase), 13 kinds of DNA synthases of α, β, γ, δ, ε, ζ, η, θ, ι, κ, λ, μ, and σ are known so far. It has been. These DNA synthase groups are involved in cell proliferation, division, differentiation, etc., α-type is responsible for DNA replication, β-type is responsible for repair and recombination, and δ-type and ε-type are responsible for both replication and repair. It is known that it has different functions depending on the type.
このようにＤＮＡ合成酵素は細胞の増殖等に関与することから、その酵素活性を阻害するＤＮＡ合成酵素阻害物質は、例えば、癌に対して癌細胞の増殖抑制作用を示し、エイズに対してＨＩＶ由来逆転写酵素に対する阻害作用を示し、あるいは免疫担当細胞において抗原に対する特異的抗体産生を抑制する免疫抑制作用を示すことが考えられる。また、ＤＮＡ合成酵素の活性を阻害することで細胞周期に影響を与え、細胞死（アポトーシス）を誘導する作用を示すことも考えられる。このため、ＤＮＡ合成酵素阻害物質を、抗癌剤（制癌剤）、エイズ治療剤、抗ウイルス剤、免疫抑制剤、アポトーシス誘導剤などとして利用することが期待されており、同物質を利用し、種々の癌、エイズ等のウイルス疾患、免疫疾患など各種疾患の予防・治療に効果のある医薬品の開発、さらには同様の効果を有する機能性食品の開発が期待されている。 Since DNA synthase is involved in cell growth and the like in this way, a DNA synthase inhibitor that inhibits the enzyme activity exhibits, for example, cancer cell growth inhibitory action against cancer and HIV against AIDS. It is considered that it exhibits an inhibitory action on the derived reverse transcriptase or an immunosuppressive action that suppresses the production of specific antibodies against the antigen in immunocompetent cells. It is also conceivable that the activity of the cell synthase is affected by inhibiting the activity of DNA synthase and induces cell death (apoptosis). For this reason, it is expected that a DNA synthase inhibitor is used as an anticancer agent (anticancer agent), an AIDS therapeutic agent, an antiviral agent, an immunosuppressive agent, an apoptosis inducer, and the like. Development of pharmaceuticals effective for the prevention and treatment of various diseases such as viral diseases such as AIDS and immune diseases, and the development of functional foods having similar effects are expected.
例えば、ＤＮＡ合成酵素阻害活性を有する紅藻類由来の糖脂質が、抗癌剤、ＨＩＶ由来逆転写酵素阻害剤、免疫抑制剤として有用であることが報告されている（下記特許文献１参照）。現在、ＤＮＡ合成酵素阻害剤として、ジデオキシＴＴＰ（ｄｄＴＴＰ）、Ｎ−メチルマレイミド、ブチルフェニル−ｄＧＴＰなどが知られている（下記非特許文献１参照）。また植物由来の糖脂質であるスルホキノボシルアシルグリセロールにもＤＮＡ合成酵素阻害作用が見出されている（下記特許文献２参照）。 For example, red alga-derived glycolipids having DNA synthase inhibitory activity have been reported to be useful as anticancer agents, HIV-derived reverse transcriptase inhibitors, and immunosuppressive agents (see
さらに、ウニより見出された糖脂質スルホキノボシルモノアシルグリセロール（ＳＱＭＧ）の抗腫瘍効果、ホウレン草（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）から精製されたモノガラクトシルジアシルグリセロール（ＭＧＤＧ）、ジガラクトシルジアシルグリセロール（ＤＧＤＧ）、スルホキノボシルジアシルグリセロール（ＳＱＤＧ）など各種糖脂質のＤＮＡ合成酵素阻害活性についても報告されている（下記非特許文献２・３参照）。 Furthermore, antitumor effect of glycolipid sulfoquinovosyl monoacylglycerol (SQMG) found from sea urchin, monogalactosyl diacylglycerol (MGDG) purified from spinach (Spinacia oleracea L.), digalactosyl diacylglycerol (DGDG) In addition, DNA synthase inhibitory activity of various glycolipids such as sulfoquinovosyl diacylglycerol (SQDG) has been reported (see Non-Patent
最近は、生活習慣病など種々の疾病に対する予防・改善効果をもつものとして機能性食品に注目が集まっている。機能性食品は、ＡＨＣＣ（Ａｃｔｉｖｅ Ｈｅｘｏｓｅ−Ｃｏｒｒｅｌａｔｅｄ Ｃｏｍｐｏｕｎｄ）、サメ軟骨、あるいはアガリクスなどといった天然成分を含み、この天然成分の生理活性により人の免疫力や自然治癒力を高め、病気の予防・改善を図るものである。そこで、上述のＤＮＡ合成酵素阻害活性を有する天然物由来の糖脂質を、医薬のみならず、このような機能性食品へ適用することが考えられるが、従来のこれら糖脂質の精製方法は各々の糖脂質を単独に精製するものであるため、精製が比較的煩雑で時間がかかる上、収量も少なく、機能性食品への利用には不向きであった。 Recently, attention has been focused on functional foods as having preventive and ameliorating effects on various diseases such as lifestyle-related diseases. Functional foods contain natural ingredients such as Active Hexose-Correlated Compound (AHCC), shark cartilage, or agaricus, and the physiological activity of these natural ingredients enhances human immunity and natural healing power to prevent and improve disease. It is intended. Therefore, it is conceivable to apply the above-mentioned glycolipid derived from natural products having DNA synthase inhibitory activity not only to pharmaceuticals but also to such functional foods. Since the glycolipid is purified alone, the purification is relatively complicated and time-consuming, and the yield is small, which is not suitable for use in functional foods.
本発明は、上記の事情に鑑み、天然物から簡便な操作で効率よく得られ、しかも有用な生理活性を有する糖脂質を高収量かつ高純度に含有する糖脂質含有組成物を提供すること、およびその用途と製造方法を提供することをその課題とするものである。 In view of the above circumstances, the present invention provides a glycolipid-containing composition that is efficiently obtained from natural products by a simple operation, and that contains glycolipids having useful physiological activity in high yield and high purity, It is another object of the present invention to provide an application and a manufacturing method thereof.
本発明者は、上記の課題に鑑み鋭意研究を進めた結果、（１）製造条件を工夫することにより、後述の３種類の糖脂質を高収量かつ高純度に含有する糖脂質含有組成物を原料（ホウレン草）から簡便な方法で製造し得ること、（２）得られた糖脂質含有組成物は、ＤＮＡ合成酵素阻害活性、癌細胞増殖抑制活性、ｉｎ ｖｉｖｏにおける抗腫瘍活性といった有用な生理活性を有すること、（３）上記糖脂質含有組成物をリパーゼ処理することにより少なくともモノアシルグルセロールタイプの２種類の糖脂質を含有する糖脂質含有組成物が得られ、当該組成物はさらに強力なＤＮＡ合成酵素阻害活性、癌細胞増殖抑制活性、抗腫瘍活性を有すること、等を見出し、本発明を完成するに至った。 As a result of intensive studies in view of the above problems, the present inventor has (1) a glycolipid-containing composition containing the following three types of glycolipids in high yield and high purity by devising production conditions. (2) The obtained glycolipid-containing composition has useful physiological activities such as DNA synthase inhibitory activity, cancer cell growth inhibitory activity, and in vivo antitumor activity. (3) By subjecting the glycolipid-containing composition to lipase treatment, a glycolipid-containing composition containing at least two monoacylglycerol type glycolipids can be obtained, and the composition is more powerful. The present inventors have found that it has a DNA synthase inhibitory activity, a cancer cell growth inhibitory activity, an antitumor activity and the like, and has completed the present invention.
即ち、本発明は、産業上有用な下記Ａ）〜Ｌ）の発明を含むものである。
Ａ）海藻または陸生植物から精製され、少なくとも糖脂質としてモノガラクトシルジアシルグリセロール、ジガラクトシルジアシルグリセロール、およびスルホキノボシルジアシルグリセロールを含有する糖脂質含有組成物（以下、特に「第１の糖脂質含有組成物」という場合がある）。
Ｂ）上記第１の糖脂質含有組成物をリパーゼ処理することにより得られ、少なくとも糖脂質としてモノガラクトシルモノアシルグリセロール、およびスルホキノボシルモノアシルグリセロールを含有する糖脂質含有組成物（以下、特に「第２の糖脂質含有組成物」という場合がある）。
Ｃ）上記第１又は第２の糖脂質含有組成物を有効成分とするＤＮＡ合成酵素阻害剤。
Ｄ）上記第１又は第２の糖脂質含有組成物を有効成分とする抗癌剤。
Ｅ）上記第１又は第２の糖脂質含有組成物を有効成分とする医薬用組成物。
Ｆ）上記第１又は第２の糖脂質含有組成物を含む食用組成物。
Ｇ）海藻または陸生植物を原料として、少なくとも糖脂質としてモノガラクトシルジアシルグリセロール、ジガラクトシルジアシルグリセロール、およびスルホキノボシルジアシルグリセロールを含有する糖脂質含有組成物を製造する方法。
Ｈ）上記Ｇ）記載の方法により製造された糖脂質含有組成物をリパーゼ処理することにより、少なくとも糖脂質としてモノガラクトシルモノアシルグリセロール、およびスルホキノボシルモノアシルグリセロールを含有する糖脂質含有組成物を製造する方法。
Ｉ）上記Ｇ）記載の方法において、疎水クロマトグラフィーを用いて植物抽出物（海藻または陸生植物からの抽出物の意味。以下同じ。）から糖脂質画分を精製する工程を含む方法。
Ｊ）上記Ｉ）記載の方法において、糖脂質の溶出にエタノール等のアルコールまたはアセトン等の有機溶媒を用い、５０％〜７５％の含水有機溶媒にて水溶性物質を溶出後、８５％〜１００％の含水有機溶媒もしくは有機溶媒にて糖脂質を溶出する工程を含む方法。
Ｋ）上記Ｇ）記載の方法において、植物抽出物を得る前に、植物を４０℃〜８０℃の温水で洗浄し、水溶性成分を除去する工程を含む方法。
Ｌ）上記Ｇ）記載の方法において、原料にホウレン草（Ｓｐｉｎａｃｉａ）等の緑黄色野菜を使用する方法。That is, the present invention includes the following industrially useful inventions A) to L).
A) Glycolipid-containing composition purified from seaweed or terrestrial plant and containing at least monogalactosyl diacylglycerol, digalactosyl diacylglycerol, and sulfoquinovosyl diacylglycerol as glycolipids (hereinafter, particularly “first glycolipid-containing” Sometimes referred to as a "composition").
B) A glycolipid-containing composition obtained by lipase treatment of the first glycolipid-containing composition and containing at least monogalactosyl monoacylglycerol and sulfoquinovosyl monoacylglycerol as glycolipids (hereinafter, particularly “ It may be referred to as “second glycolipid-containing composition”).
C) A DNA synthase inhibitor comprising the first or second glycolipid-containing composition as an active ingredient.
D) An anticancer agent containing the first or second glycolipid-containing composition as an active ingredient.
E) A pharmaceutical composition comprising the first or second glycolipid-containing composition as an active ingredient.
F) An edible composition comprising the first or second glycolipid-containing composition.
G) A method for producing a glycolipid-containing composition containing at least monogalactosyl diacylglycerol, digalactosyl diacylglycerol, and sulfoquinovosyl diacylglycerol as glycolipids using seaweed or terrestrial plants as raw materials.
H) A glycolipid-containing composition containing at least monogalactosyl monoacylglycerol and sulfoquinovosyl monoacylglycerol as glycolipids by subjecting the glycolipid-containing composition produced by the method described in G) to lipase treatment How to manufacture.
I) A method comprising the step of purifying a glycolipid fraction from a plant extract (meaning an extract from a seaweed or a terrestrial plant; the same shall apply hereinafter) using hydrophobic chromatography in the method described in G) above.
J) In the method described in I) above, an alcohol such as ethanol or an organic solvent such as acetone is used for elution of glycolipid, and a water-soluble substance is eluted with 50% to 75% hydrous organic solvent, and then 85% to 100% A method comprising a step of eluting glycolipids with a water-containing organic solvent or an organic solvent.
K) In the method described in G) above, a method comprising washing a plant with warm water at 40 ° C. to 80 ° C. to remove water-soluble components before obtaining a plant extract.
L) A method of using a green-yellow vegetable such as spinach as a raw material in the method described in G) above.
本発明の第１および第２の糖脂質含有組成物はいずれも、ＤＮＡ合成酵素阻害活性、癌細胞増殖抑制活性、抗腫瘍活性といった有用な生理活性を有し、ＤＮＡ合成酵素阻害剤、抗癌剤その他の医薬用組成物および食用組成物（食品または食品添加物）に利用できる。しかも、本発明の糖脂質含有組成物は、海藻または陸生植物から簡便な操作で効率よく製造することができ、高収量であるので大量かつ安価に生産することも可能であり、機能性食品などに好適に用いることができ、例えば抗癌作用、癌予防効果をもつ機能性食品として利用できる。 Both the first and second glycolipid-containing compositions of the present invention have useful physiological activities such as DNA synthase inhibitory activity, cancer cell growth inhibitory activity, and antitumor activity, and DNA synthase inhibitors, anticancer agents and others It can be used for pharmaceutical compositions and edible compositions (food or food additives). Moreover, the glycolipid-containing composition of the present invention can be efficiently produced from seaweed or terrestrial plants by a simple operation and can be produced in large quantities and at low cost because of high yield, such as functional foods. For example, it can be used as a functional food having an anticancer effect and a cancer prevention effect.
以下、図面を参照しながら本発明の実施の一形態について説明する。
（１）本発明の糖脂質含有組成物およびその製造方法
前述のとおり、本発明の第１の糖脂質含有組成物は、海藻または陸生植物から精製され、少なくとも糖脂質としてモノガラクトシルジアシルグリセロール、ジガラクトシルジアシルグリセロール、およびスルホキノボシルジアシルグリセロール（以下、これら糖脂質をそれぞれ「ＭＧＤＧ」、「ＤＧＤＧ」、「ＳＱＤＧ」と略称する）を含有するものである。また、本発明の第２の糖脂質含有組成物は、本発明の第１の糖脂質含有組成物をリパーゼ処理することにより得られ、少なくとも糖脂質としてモノガラクトシルモノアシルグリセロール、およびスルホキノボシルモノアシルグリセロール（以下、これら糖脂質をそれぞれ「ＭＧＭＧ」、「ＳＱＭＧ」と略称する）を含有するものである。Hereinafter, an embodiment of the present invention will be described with reference to the drawings.
(1) Glycolipid-containing composition of the present invention and method for producing the same As described above, the first glycolipid-containing composition of the present invention is purified from seaweed or terrestrial plants, and at least as a glycolipid, monogalactosyl diacylglycerol, di It contains galactosyl diacylglycerol and sulfoquinovosyl diacylglycerol (hereinafter, these glycolipids are abbreviated as “MGDG”, “DGDG”, and “SQDG”, respectively). The second glycolipid-containing composition of the present invention is obtained by subjecting the first glycolipid-containing composition of the present invention to a lipase treatment, and at least monogalactosylmonoacylglycerol and sulfoquinovosylmono as glycolipids. Acylglycerol (hereinafter, these glycolipids are abbreviated as “MGMG” and “SQMG”, respectively).
図１の（ａ）〜（ｅ）には、それぞれ、上記ＭＧＤＧ、ＤＧＤＧ、ＳＱＤＧ、ＭＧＭＧ、ＳＱＭＧの化学構造が示される。図中、Ｒ１〜Ｒ８は互いに独立した脂肪酸であり、好ましくは炭素数１４〜２２の飽和脂肪酸または一価もしくは多価の不飽和脂肪酸である。飽和脂肪酸としては、ミリスチン酸、パルミチン酸、ステアリン酸等を例示でき、不飽和脂肪酸としては、パルミトオレイン酸、オレイン酸、リノール酸、リノレン酸（ω３）、ジホモガンマリノレン酸、オクタデカテトラエン酸、アラキドン酸、エイコサペンタエン酸、ドコサペンタエン酸、ドコサヘキサエン酸等を例示できる。なお、各糖脂質におけるグリセロール骨格の１位と２位の構成脂肪酸の種類は特に限定されるものではなく、ジアシルグルセロールタイプの糖脂質においては、両脂肪酸は同じものであっても、異なるものであってもよい。
また、各糖脂質は薬学的に許容され得る塩の状態であってもよい。このような塩としては、フッ化水素酸塩、塩酸塩などのハロゲン化水素酸塩、硫酸塩、硝酸塩などの無機酸塩、ナトリウム塩、カリウム塩などのアルカリ金属塩、スルホン酸塩、有機酸塩、およびアミノ酸塩が挙げられ、好適には塩酸塩、硫酸塩、硝酸塩、ナトリウム塩、カリウム塩を挙げることができる。1A to 1E show chemical structures of the MGDG, DGDG, SQDG, MGMG, and SQMG, respectively. In the figure, R 1 to R 8 are fatty acids independent of each other, preferably a saturated fatty acid having 14 to 22 carbon atoms or a monovalent or polyvalent unsaturated fatty acid. Examples of saturated fatty acids include myristic acid, palmitic acid, stearic acid, and the like. Examples of unsaturated fatty acids include palmitooleic acid, oleic acid, linoleic acid, linolenic acid (ω3), dihomogamma linolenic acid, octadecatetraene. Examples thereof include acid, arachidonic acid, eicosapentaenoic acid, docosapentaenoic acid, and docosahexaenoic acid. The types of constituent fatty acids at the 1st and 2nd positions of the glycerol skeleton in each glycolipid are not particularly limited. In diacylglycerol type glycolipids, both fatty acids are the same, but are different. It may be a thing.
Each glycolipid may be in the form of a pharmaceutically acceptable salt. Examples of such salts include hydrohalides such as hydrofluoride and hydrochloride, inorganic acid salts such as sulfate and nitrate, alkali metal salts such as sodium and potassium salts, sulfonates, and organic acids. Salts, and amino acid salts. Preferred examples include hydrochlorides, sulfates, nitrates, sodium salts, and potassium salts.
本発明の第１の糖脂質含有組成物は、例えば以下の方法によりホウレン草（Ｓｐｉｎａｃｉａ）から抽出・精製することができる。まず、原料には市販のホウレン草（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）を使用し、図２に示すように、その乾燥物１００ｇを細断した後、水溶性成分をできるだけ除去するために、６０℃の温水１Ｌで２回洗浄する。次に、濾紙を使用して濾過後、水分を除去し、得られた固形物（残渣）に１Ｌのエタノールを加え、撹拌しながら６０℃で還流抽出を２回行う。抽出液は濾過後、減圧下で濃縮を行うことで、ホウレン草のオイル状抽出物が得られる。実際、この方法により乾燥ホウレン草１００ｇからオイル状抽出物２０．５ｇが得られた。 The first glycolipid-containing composition of the present invention can be extracted and purified from spinach, for example, by the following method. First, commercially available spinach (Spinacia oleracea L.) is used as a raw material, and as shown in FIG. 2, 1 g of hot water at 60 ° C. is used to remove water-soluble components as much as possible after chopping 100 g of the dried product. Wash twice with. Next, after filtration using filter paper, moisture is removed, 1 L of ethanol is added to the obtained solid (residue), and reflux extraction is performed twice at 60 ° C. with stirring. The extract is filtered and then concentrated under reduced pressure to obtain an oily extract of spinach. In fact, 20.5 g of an oily extract was obtained from 100 g of dried spinach by this method.
次に、得られた抽出物を７０％エタノール溶液（エタノールと水の体積比が７０：３０の溶液。以下同様。）に溶解した後、逆相クロマトグラフィーにより糖脂質画分を精製し、本発明の第１の糖脂質含有組成物を得る。本発明者が行った方法では、上記抽出物を７０％エタノール溶液に溶解した後、これを疎水クロマトグラフィー用樹脂５００ｇ（ダイヤイオンＨＰ−２０、三菱化学社製）に注入し、その後、７０％エタノールで未吸着物質を洗浄・溶出した画分Ｉ（水溶性画分、１３．０ｇ）、９５％エタノールで溶出される画分ＩＩ（糖脂質画分すなわち本発明の糖脂質含有組成物、６．５ｇ）、クロロホルムで溶出される画分ＩＩＩ（クロロフィル（葉緑素）を含む色素画分、１．０ｇ）の３つに分画した。 Next, the obtained extract was dissolved in a 70% ethanol solution (a solution having a volume ratio of ethanol and water of 70:30, the same applies hereinafter), and then the glycolipid fraction was purified by reverse phase chromatography. A first glycolipid-containing composition of the invention is obtained. In the method performed by the present inventor, the above extract was dissolved in a 70% ethanol solution and then injected into 500 g of a resin for hydrophobic chromatography (Diaion HP-20, manufactured by Mitsubishi Chemical Corporation), and then 70% Fraction I (water-soluble fraction, 13.0 g) obtained by washing and eluting unadsorbed substances with ethanol, Fraction II (glycolipid fraction, ie, glycolipid-containing composition of the present invention, 6) eluted with 95% ethanol 0.5 g), and fraction III (a pigment fraction containing chlorophyll (chlorophyll), 1.0 g) eluted with chloroform.
上記画分ＩＩ、すなわち本発明の糖脂質含有組成物に含まれる成分を薄層クロマトグラフィーにより解析した結果、ＭＧＤＧ、ＤＧＤＧおよびＳＱＤＧの３つの糖脂質が含まれていることが確認された（図３参照）。また、分析の結果、上記糖脂質画分（画分ＩＩ）２５０ｍｇには、ＭＧＤＧが８４．７ｍｇ（３３．９％）、ＤＧＤＧが３５．０ｍｇ（１４．０％）、ＳＱＤＧが９２．３ｍｇ（３６．９％）含まれていた。このように、本発明の第１の糖脂質含有組成物において、ＭＧＤＧ、ＤＧＤＧおよびＳＱＤＧの３つの糖脂質の含有量は好ましくは重量比７０％以上、より好ましくは７５％以上、さらに好ましくは８０％以上である。 As a result of analyzing the fraction II, that is, the components contained in the glycolipid-containing composition of the present invention by thin layer chromatography, it was confirmed that three glycolipids of MGDG, DGDG and SQDG were contained (FIG. 3). As a result of the analysis, 250 mg of the glycolipid fraction (Fraction II) contained 84.7 mg (33.9%) of MGDG, 35.0 mg (14.0%) of DGDG, and 92.3 mg of SQDG ( 36.9%). Thus, in the first glycolipid-containing composition of the present invention, the content of the three glycolipids MGDG, DGDG, and SQDG is preferably 70% or more by weight, more preferably 75% or more, and still more preferably 80%. % Or more.
さらに、上記糖脂質画分（画分ＩＩ）のＤＮＡ合成酵素阻害活性およびヒト癌細胞増殖抑制活性を検討したところ、上記糖脂質画分はＤＮＡ合成酵素αを阻害し、ヒト胃癌細胞株ＮＵＧＣ−３細胞の増殖を抑制し、ｉｎ ｖｉｖｏにおける抗腫瘍活性も認められた（図４、図５および図９〜図１２参照）。なお、これら実験の詳細については、後述の実施例で説明する。 Further, when the DNA synthase inhibitory activity and the human cancer cell growth inhibitory activity of the glycolipid fraction (Fraction II) were examined, the glycolipid fraction inhibited the DNA synthetase α, and the human gastric cancer cell line NUGC- Inhibition of the growth of 3 cells and anti-tumor activity in vivo were also observed (see FIGS. 4, 5 and 9 to 12). The details of these experiments will be described in the examples described later.
このように、本発明の糖脂質含有組成物は上記方法によりホウレン草から抽出・精製することができるが、上記製造方法に限定されるものではなく、種々の変更が可能である。例えば、上記画分ＩＩの精製方法では原料にホウレン草（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）を使用したが、これに限らず他の植物または海藻を原料に使用してもよい。好ましくは、他の緑黄色野菜（小松菜、クレソン、パセリ、ブロッコリー、ピーマン等）や大麦の葉等の陸生植物、藍藻類（Ａｎａｂａｅｎａ ｖａｒｉａｂｉｌｉｓ、Ａｎａｃｙｓｔｉｓ ｎｉｄｕｌａｎｓ、Ｓｐｉｒｕｌｉｎａ ｐｌａｔｅｎｓｉｓ：スピルリナ等）、紅藻類（Ｐｏｒｐｈｙｒａ ｙｅｚｏｅｎｓｉｓ：スサビノリ、Ｇｅｌｉｄｉｕｍ ａｍａｎｓｉｉ：マクサ、Ｇｉｇａｒｔｉｎａ ｔｅｎｅｌｌａ：スギノリ、Ｇｒａｃｉｌａｒｉａ ｖｅｒｒｕｃｏｓａ：オゴノリ等）、褐藻類（Ｕｎｄａｒｉａ ｐｉｎｎａｔｉｆｉｄａ：ワカメ、Ｈｉｚｉｋｉａ ｆｕｓｉｆｏｒｍｉｓ：ヒジキ、Ｌａｍｉｎａｒｉａ ｊａｐｏｎｉｃａ：マコンブ、Ｓａｒｇａｓｓｕｍ ｈｏｒｎｅｒｉ：アカモク等）または緑藻類（Ｃｈｌａｍｉｄｏｍｏｎａｓ ｓｐ．：クラミドモナスの一種、Ｅｎｔｅｒｏｍｏｒｐｈａ ｓｐ．：アオサの一種、Ｃｈｌｏｒｅｌｌａ ｓｐ．：クロレラ等）に属する藻の藻体を挙げることができる。 As described above, the glycolipid-containing composition of the present invention can be extracted and purified from spinach by the above method, but is not limited to the above production method, and various modifications can be made. For example, in the purification method of fraction II, spinach (Spinia oleracea L.) is used as a raw material, but not limited thereto, other plants or seaweed may be used as a raw material. Preferably, other green-yellow vegetables (Komatsuna, watercress, parsley, broccoli, green pepper, etc.), terrestrial plants such as barley leaves, cyanobacteria (Anabaena variabilis, Anacystis nidulans, Spirulina platensis: Spirulina, etc.) Porphyra yezoensis, Gelidium amansii: MAXA, Gigartina tenella: Suginori, Gracilaria verrucosa: Gracilaria etc.), brown algae (Undaria pinnatifida: seaweed, Hizikia fusiformis: hijiki, Laminaria japonica: Laminaria japonica, Sargassum horneri: akamoku etc.) or green algae (Chlamid omonas sp .: a kind of Chlamydomonas, Enteromorpha sp .: a kind of Aosa, Chlorella sp .: Chlorella, etc.).
これらいずれか一つ又は複数の原料を必要に応じて細断ないしは粉砕し、ヘキサン、クロロホルム、アセトン、メタノール、エタノール等の脂質成分抽出用有機溶媒を用いて抽出処理し、該抽出液から溶媒を除去して全脂質を得る。ついで、シリカゲル、アルミナ、セファデックス、逆相吸着剤（オクタデシルシリル化シリカゲル等）、イオン交換樹脂、合成吸着剤等を用いたカラムクロマトグラフィーで分画処理して濃縮画分を採取し、さらに必要に応じて濃縮精製し、目的物である本発明の糖脂質含有組成物を調製することができる。さらに、得られた糖脂質含有組成物を後述のようにリパーゼ処理することによって、モノアシルグリセロールタイプの糖脂質を含有する本発明の第２の糖脂質含有組成物を調製することができる。 Any one or more of these raw materials are shredded or pulverized as necessary, extracted with an organic solvent for lipid component extraction such as hexane, chloroform, acetone, methanol, ethanol, etc., and the solvent is removed from the extract. Remove to obtain total lipids. Next, fractionation is performed by column chromatography using silica gel, alumina, Sephadex, reverse phase adsorbent (octadecylsilylated silica gel, etc.), ion exchange resin, synthetic adsorbent, etc. The glycolipid-containing composition of the present invention which is the target product can be prepared by concentration and purification according to the above. Furthermore, the second glycolipid-containing composition of the present invention containing a monoacylglycerol type glycolipid can be prepared by subjecting the obtained glycolipid-containing composition to lipase treatment as described below.
より具体的に、植物から本発明の糖脂質含有組成物を製造する方法として２つの好ましい方法について説明すると、第１の方法では、まず植物の乾燥物を細断して、温水、好ましくは４０℃〜８０℃（より好ましくは５０℃〜７０℃）で洗浄する。その後、濾過により水分を除去し、得られた固形物（残渣）に５０％以上のエタノール溶液もしくはエタノールを加え、撹拌しながら４０℃〜８０℃（より好ましくは５０℃〜７０℃）で還流抽出を行う。抽出液を濾過後、減圧下で濃縮を行い、得られた抽出物を５０％〜７５％のエタノール溶液に溶解し、疎水性クロマトグラフィー樹脂に注入する。そして水−エタノールの混合比において段階的に溶出を行う。まず５０％〜７５％のエタノール溶液では樹脂に吸着しない水溶性物質が溶出され、それに続く８５％以上のエタノール溶液（またはアセトンなどの有機溶媒）によって、糖脂質など脂溶性物質が溶出される。さらに脂溶性色素や脂肪酸などを除去して糖脂質画分を精製してもよいが、８５％〜１００％エタノール溶液で溶出することによって糖脂質だけを効率よく分画することが可能である。本発明者が行った前述の画分ＩＩの精製方法も基本的にこの第１の方法であり、簡易迅速に高純度かつ高収量で本発明の糖脂質含有組成物を調製する方法として好ましい方法である。 More specifically, two preferred methods for producing the glycolipid-containing composition of the present invention from a plant will be described. In the first method, first, a dried product of a plant is first shredded and warm water, preferably 40 Washing is performed at a temperature of 80 ° C. to 80 ° C. (more preferably 50 ° C. to 70 ° C.). Thereafter, water is removed by filtration, and 50% or more ethanol solution or ethanol is added to the obtained solid (residue), followed by reflux extraction at 40 ° C. to 80 ° C. (more preferably 50 ° C. to 70 ° C.) with stirring. I do. The extract is filtered and then concentrated under reduced pressure. The obtained extract is dissolved in a 50% to 75% ethanol solution and injected into a hydrophobic chromatography resin. Then, elution is performed stepwise at a water-ethanol mixing ratio. First, a 50% to 75% ethanol solution elutes a water-soluble substance that does not adsorb to the resin, and a subsequent 85% or more ethanol solution (or an organic solvent such as acetone) elutes a fat-soluble substance such as a glycolipid. Further, the lipid-soluble pigment and fatty acid may be removed to purify the glycolipid fraction, but it is possible to efficiently fractionate only the glycolipid by elution with an 85% to 100% ethanol solution. The above-mentioned method for purifying fraction II performed by the present inventor is also basically this first method, which is a preferred method for easily and rapidly preparing the glycolipid-containing composition of the present invention with high purity and high yield. It is.
第２の方法は、まず植物の乾燥物を細断して、含水アセトンもしくはアセトンを加え、撹拌しながら３０℃〜５０℃で還流抽出を行う。抽出液は濾過後、減圧下で濃縮を行い、得られた抽出物を含水アセトン溶液に溶解し、イオン交換樹脂（ＳＫ１Ｂイオン交換樹脂（三菱化学社製）など）に注入する。水−アセトンの混合比において段階的に溶出を行うことにより、糖脂質含有画分は例えば９５％以上のアセトン溶液による溶出によって得ることができる。 In the second method, a dried plant product is first shredded, hydrous acetone or acetone is added, and reflux extraction is performed at 30 ° C. to 50 ° C. with stirring. The extract is filtered and concentrated under reduced pressure, and the resulting extract is dissolved in a water-containing acetone solution and injected into an ion exchange resin (SK1B ion exchange resin (manufactured by Mitsubishi Chemical Corporation)). By elution stepwise at a water-acetone mixing ratio, the glycolipid-containing fraction can be obtained, for example, by elution with an acetone solution of 95% or more.
本発明者が行った前述の画分ＩＩの精製方法は、（１）粗抽出物（オイル状抽出物）を得る前に、温水抽出すなわち温水で洗浄することにより水溶性成分をできるだけ事前除去して、糖脂質含有組成物の精製を容易にしたこと、（２）疎水クロマトグラフィーを用いたこと、（３）エタノール溶液で糖脂質画分を溶出させるときの水−エタノールの比率を厳密に検討したこと、を特徴としている。上記（１）については、植物（ホウレン草）を６０℃の温水１Ｌで２回洗浄し、水溶性成分を除去したが、４０℃〜８０℃の温水で洗浄してもよい。また、温水の量、洗浄回数などは原料の量などに応じて適宜設定すればよい。上記（２）については、三菱化学社製のダイヤイオンＨＰ−２０を使用したが、他の樹脂等を使用して疎水（逆相）クロマトグラフィーを行ってもよい。上記（３）については、７０％エタノール溶液で水溶性画分（画分Ｉ）を溶出後、９５％エタノール溶液で糖脂質画分（画分ＩＩ）を溶出したが、この方法以外に、エタノール等のアルコールまたはアセトン等の有機溶媒を用い、５０％〜７５％（より好ましくは６５％〜７５％）の含水有機溶媒にて水溶性物質を溶出後、８５％〜１００％（より好ましくは９２％〜９８％）の含水有機溶媒もしくは有機溶媒にて糖脂質を溶出してもよい。 The purification method of the above-mentioned fraction II performed by the present inventor is as follows. (1) Before obtaining a crude extract (oil-like extract), water-soluble components are removed in advance by hot water extraction, that is, washing with warm water. (2) Use of hydrophobic chromatography, (3) Strict examination of water-ethanol ratio when elution of glycolipid fraction with ethanol solution It is characterized by that. About said (1), although the plant (spinach) was washed twice with 1 L of warm water at 60 ° C. to remove water-soluble components, it may be washed with warm water at 40 ° C. to 80 ° C. Further, the amount of warm water, the number of times of washing, etc. may be set as appropriate according to the amount of raw material. As for (2) above, Diaion HP-20 manufactured by Mitsubishi Chemical Corporation was used, but hydrophobic (reverse phase) chromatography may be performed using another resin or the like. Regarding (3) above, the water-soluble fraction (fraction I) was eluted with a 70% ethanol solution, and then the glycolipid fraction (fraction II) was eluted with a 95% ethanol solution. Using an organic solvent such as alcohol or acetone, the water-soluble substance is eluted with 50% to 75% (more preferably 65% to 75%) water-containing organic solvent, and then 85% to 100% (more preferably 92). % -98%) hydrous organic solvent or organic solvent may be used for elution.
後述のように、上記方法により得られた画分ＩＩには、ＤＮＡ合成酵素阻害活性および癌細胞増殖抑制活性などの活性が認められる一方、単なるエタノール抽出液（前記オイル状抽出物）にはこのような活性は認められず、水溶性画分（画分Ｉ）および色素画分（画分ＩＩＩ）にもこのような活性は認められなかった。これらの実験結果から、水溶性画分または色素画分に活性妨害物質（マスキング物質）の存在する可能性が想定され、もしそうであれば、上記の方法により糖脂質画分を精製し、活性妨害物質を取り除くことは非常に有効な方法といえる。また、上記の方法により本発明の糖脂質含有組成物を製造することは、（１）各々の糖脂質を精製するよりも簡便であり、（２）各々の糖脂質を精製するよりも収量が多く、（３）精製した糖脂質よりも機能性食品として利用しやすい、などの利点がある。 As will be described later, the fraction II obtained by the above method has activities such as a DNA synthase inhibitory activity and a cancer cell growth inhibitory activity, while a simple ethanol extract (the oily extract) does not have this activity. Such activity was not observed, and such activity was not observed in the water-soluble fraction (fraction I) and the dye fraction (fraction III). From these experimental results, it is assumed that there is a possibility that an activity interfering substance (masking substance) exists in the water-soluble fraction or the dye fraction. If so, the glycolipid fraction is purified by the above-described method, Removing the interfering substances is a very effective method. In addition, the production of the glycolipid-containing composition of the present invention by the above method is simpler than (1) purifying each glycolipid, and (2) yield is higher than purifying each glycolipid. There are many advantages such as (3) being easier to use as a functional food than purified glycolipids.
本発明の第１の糖脂質含有組成物において、ＭＧＤＧ、ＤＧＤＧおよびＳＱＤＧの各々が含まれる割合は特に限定されるものではない。これら３つの糖脂質の含有比は原料となる植物・海藻によっても異なり、例えばホウレン草の場合、通常これら糖脂質の含有比は、ＭＧＤＧ：ＤＧＤＧ：ＳＱＤＧ＝およそ１：２．１５：１．７５（およそ２０：４３：３５）である。他の野菜などの場合はこれら糖脂質の含有比はかなり異なるものがある。本発明者が行った実験結果によると、ＳＱＤＧの比率が高い糖脂質画分はＤＮＡ合成酵素阻害活性が強いので、ＳＱＤＧ含量が多い方が好ましい。 The ratio in which each of MGDG, DGDG, and SQDG is contained in the 1st glycolipid containing composition of this invention is not specifically limited. The content ratio of these three glycolipids varies depending on the plant and seaweed used as a raw material. For example, in the case of spinach, the content ratio of these glycolipids is usually MGDG: DGDG: SQDG = approximately 1: 2.15: 1.75 ( Approximately 20:43:35). In the case of other vegetables, the content ratio of these glycolipids is quite different. According to the results of experiments conducted by the present inventor, a glycolipid fraction having a high SQDG ratio has a strong DNA synthase inhibitory activity, and therefore preferably has a high SQDG content.
本発明の糖脂質含有組成物に含まれる各糖脂質の脂肪酸鎖の種類についても特に限定されるものではないが、脂肪酸鎖は比較的長い（例えば炭素数１６以上）ほうが好ましく、不飽和脂肪酸鎖における二重結合の数は１つまたは２つであるときにＤＮＡ合成酵素阻害活性が高いので、両者いずれかであることが好ましい。 The type of fatty acid chain of each glycolipid contained in the glycolipid-containing composition of the present invention is not particularly limited, but the fatty acid chain is preferably relatively long (for example, having 16 or more carbon atoms), and the unsaturated fatty acid chain. Since the DNA synthase inhibitory activity is high when the number of double bonds in is one or two, it is preferable to be either of them.
前述のとおり、本発明の第１の糖脂質含有組成物において、ＭＧＤＧ、ＤＧＤＧおよびＳＱＤＧの３つの糖脂質の含有量は好ましくは重量比７０％以上、より好ましくは７５％以上、さらに好ましくは８０％以上であり、糖脂質以外の成分はできる限り含まれていないことが好ましい。もっとも、抗酸化活性のあるフラボノイド化合物（カテキンやポリフェノールなどの脂溶性成分）は脂肪酸の二重結合の酸化を防ぐ性質を有するので、本発明の糖脂質含有組成物は同化合物などの物質を含むものであってもよい。 As described above, in the first glycolipid-containing composition of the present invention, the content of the three glycolipids MGDG, DGDG, and SQDG is preferably 70% or more by weight, more preferably 75% or more, and still more preferably 80%. It is preferable that components other than glycolipids are contained as much as possible. However, since flavonoid compounds having antioxidative activity (lipid-soluble components such as catechins and polyphenols) have the property of preventing the oxidation of fatty acid double bonds, the glycolipid-containing composition of the present invention contains substances such as the same compounds. It may be a thing.
さらに、以上説明した製法により得られた本発明の第１の糖脂質含有組成物をリパーゼ処理することにより、モノアシルグリセロールタイプの糖脂質を含有する本発明の第２の糖脂質含有組成物を製造することができる。 Furthermore, the second glycolipid-containing composition of the present invention containing a monoacylglycerol type glycolipid is obtained by subjecting the first glycolipid-containing composition of the present invention obtained by the production method described above to lipase treatment. Can be manufactured.
本発明者が行った方法は、図１３に示すように、上記糖脂質画分（画分ＩＩ）１０ｍｇ／ｍｌおよびブタ膵臓リパーゼ１０ｍｇ／ｍｌを反応液（０．２Ｍ Ｔｒｉｓ−ＨＣｌ（ｐＨ７．６）、０．２５Ｍ ＣａＣｌ２）において３７℃、２０分間振とう反応させ、糖脂質をリパーゼにより加水分解し、その後６ＮのＨＣｌで反応を止める。次にｎ−ブタノール（ｎ−ＢｕＯＨ）と水で分配することにより、糖脂質はｎ−ブタノール層へ、リパーゼは水層へ移動するので、リパーゼ分解された糖脂質を含有する本発明の第２の糖脂質含有組成物を調製することができる。As shown in FIG. 13, the method performed by the present inventor was carried out using 10 mg / ml of the glycolipid fraction (fraction II) and 10 mg / ml of porcine pancreatic lipase as a reaction solution (0.2 M Tris-HCl (pH 7.6). ), Shaking reaction at 37 ° C. for 20 minutes in 0.25 M CaCl 2 ), hydrolyzing the glycolipid with lipase, and then stopping the reaction with 6N HCl. Next, by partitioning with n-butanol (n-BuOH) and water, the glycolipid moves to the n-butanol layer and the lipase moves to the water layer. Therefore, the second of the present invention containing the lipase-decomposed glycolipid is used. A glycolipid-containing composition can be prepared.
薄層クロマトグラフィーにより解析した結果、リパーゼ反応後の上記糖脂質含有組成物には、ＭＧＭＧおよびＳＱＭＧの２つの糖脂質が含まれ、リパーゼ反応前の画分ＩＩ中の糖脂質ＭＧＤＧ・ＳＱＤＧは、リパーゼ加水分解反応により、それぞれＭＧＭＧ・ＳＱＭＧに分解された。一方、ＤＧＤＧは、リパーゼにより加水分解されず、リパーゼ反応後の上記糖脂質含有組成物中にも検出された（図６参照）。 As a result of analysis by thin layer chromatography, the glycolipid-containing composition after the lipase reaction contains two glycolipids, MGMG and SQMG, and the glycolipid MGDG / SQDG in fraction II before the lipase reaction is The lipase was hydrolyzed into MGMG and SQMG, respectively. On the other hand, DGDG was not hydrolyzed by lipase and was also detected in the glycolipid-containing composition after the lipase reaction (see FIG. 6).
このように、本発明の第１の糖脂質含有組成物をリパーゼ処理することにより得られた本発明の第２の糖脂質含有組成物についてＤＮＡ合成酵素阻害活性などを検討したところ、ＤＮＡ合成酵素阻害活性、癌細胞増殖抑制活性、抗腫瘍活性のいずれにおいても第１の糖脂質含有組成物に比べて高い活性が認められた（図７〜図９参照）。したがって、本発明の第２の糖脂質含有組成物は、ＤＮＡ合成酵素阻害剤、抗癌剤（制癌剤）、機能性食品などの各用途に、より有効なものといえる。 Thus, when the second glycolipid-containing composition of the present invention obtained by subjecting the first glycolipid-containing composition of the present invention to lipase treatment was examined, the DNA synthase inhibitory activity and the like were examined. Higher activity was observed in all of the inhibitory activity, cancer cell growth inhibitory activity, and antitumor activity as compared with the first glycolipid-containing composition (see FIGS. 7 to 9). Therefore, it can be said that the second glycolipid-containing composition of the present invention is more effective for each use such as a DNA synthase inhibitor, an anticancer agent (anticancer agent), and a functional food.
なお、本発明の第１の糖脂質含有組成物から第２の糖脂質含有組成物を得るためのリパーゼ処理は、図１３に示される方法に限定されるものではなく、他の公知のリパーゼ処理によって第２の糖脂質含有組成物を製造してもよい。 Note that the lipase treatment for obtaining the second glycolipid-containing composition from the first glycolipid-containing composition of the present invention is not limited to the method shown in FIG. 13, but other known lipase treatments. The second glycolipid-containing composition may be produced by
本発明の第１および第２の糖脂質含有組成物を工業生産する場合は、以上説明した方法を工業生産により適したものとするため各工程の具体的方法に適宜変更を加えてもよい。例えば、疎水クロマトグラフィーによる糖脂質画分の精製工程においては、カラム法（クロマト樹脂を筒状のカラムに詰めて上から組成物溶液を流す方法）のほかに、バッチ法（組成物溶液の中にクロマト樹脂を入れてかき混ぜる方法）が工業化に有利であると考えられる。また、リパーゼ処理については、酵素（リパーゼ）溶液を組成物溶液に加えて攪拌しながら反応させる方法のほかに、リパーゼを固定化したカラムに組成物を流して反応させる、いわゆる「固定化酵素」を用いる精製方法が大量生産には有利であると考えられる。 When the first and second glycolipid-containing compositions of the present invention are industrially produced, the specific methods in each step may be appropriately changed in order to make the above-described method more suitable for industrial production. For example, in the purification process of glycolipid fractions by hydrophobic chromatography, in addition to the column method (a method in which a chromatographic resin is packed in a cylindrical column and the composition solution is flowed from above), a batch method (in the composition solution) is used. It is considered that the method of adding a chromatographic resin and stirring the mixture is advantageous for industrialization. As for the lipase treatment, in addition to a method in which an enzyme (lipase) solution is added to a composition solution and reacted while stirring, a so-called “immobilized enzyme” in which the composition is allowed to flow through a column on which lipase is immobilized and reacted. It is considered that a purification method using the is advantageous for mass production.
（２）本発明の糖脂質含有組成物の用途
上述した本発明の第１および第２の糖脂質含有組成物はいずれも、ＤＮＡ合成酵素阻害活性、癌細胞増殖抑制活性、抗腫瘍活性といった有用な生理活性を有することから、ＤＮＡ合成酵素阻害剤、抗癌剤（制癌剤）として利用可能であり、特に機能性食品などに好適に用いることができ、例えば抗癌作用または癌予防効果をもつ機能性食品として利用できる。(2) Use of the glycolipid-containing composition of the present invention The above-described first and second glycolipid-containing compositions of the present invention are all useful such as DNA synthase inhibitory activity, cancer cell growth inhibitory activity, and antitumor activity. Because of its excellent physiological activity, it can be used as a DNA synthase inhibitor and an anticancer agent (anticancer agent), and can be suitably used particularly for functional foods. For example, a functional food having an anticancer action or a cancer prevention effect Available as
また、本発明の第１および第２の糖脂質含有組成物は、ＤＮＡ合成酵素阻害活性を有することから、抗癌剤以外の医薬（医薬用組成物）への利用、機能性食品以外の食用組成物（食品または食品添加物）としての利用も可能であり、癌のほかエイズの発症、進行を予防する作用あるいは治療効果、また臓器移植時等の免疫抑制作用をねらいとして利用することができ、とりわけ前述したエイズ治療剤、抗ウイルス剤、免疫抑制剤、アポトーシス誘導剤などとして有用である。このように、本発明の糖脂質含有組成物は、癌、エイズその他のウイルス感染症、免疫疾患などに対する予防あるいは治療のための手段として利用し得るものである。 In addition, since the first and second glycolipid-containing compositions of the present invention have DNA synthase inhibitory activity, they are used for pharmaceuticals (pharmaceutical compositions) other than anticancer agents, and edible compositions other than functional foods. It can also be used as a (food or food additive) and can be used for the purpose of preventing or developing AIDS as well as cancer and preventing or promoting the disease, as well as immunosuppressive effects such as organ transplantation. It is useful as the aforementioned AIDS therapeutic agent, antiviral agent, immunosuppressive agent, apoptosis inducer and the like. Thus, the glycolipid-containing composition of the present invention can be used as a means for preventing or treating cancer, AIDS and other viral infections, immune diseases and the like.
一例として、ＤＮＡ合成酵素阻害剤であるアジドチミジンのような既に市販の臨床薬によっても、エイズに対して治療効果をもたらし得ることは実証されている（日本医薬情報センター編、株式会社薬事時報社発行、「１９９６年度版 医療薬 日本医薬品集」、第５９９頁）。 As an example, it has been demonstrated that a commercially available clinical drug such as azidothymidine, a DNA synthase inhibitor, can also have a therapeutic effect on AIDS. , "1996 edition of Japanese Medicine", page 599).
次に、本発明の糖脂質含有組成物を配合してなる医薬用組成物および食用組成物について説明する。本発明の糖脂質含有組成物は必要に応じてさらに精製した後、これをそのまま、あるいは慣用の医薬製剤担体とともに医薬用組成物となし、動物およびヒトに投与することができる。医薬用組成物の剤形としては特に制限されるものではなく必要に応じて適宜選択すればよいが、例えば、錠剤、カプセル剤、顆粒剤、細粒剤、散剤等の経口剤、注射剤、坐剤等の非経口剤が挙げられる。投与量は、通常、成人で糖脂質の重量として１日あたり２０ｍｇ以上〜６００ｍｇ以下を数回に分けて服用するのが適当である。 Next, a pharmaceutical composition and an edible composition comprising the glycolipid-containing composition of the present invention will be described. The glycolipid-containing composition of the present invention can be further purified as necessary, and can be administered to animals and humans as it is or as a pharmaceutical composition together with a conventional pharmaceutical preparation carrier. The dosage form of the pharmaceutical composition is not particularly limited and may be appropriately selected as necessary.For example, oral preparations such as tablets, capsules, granules, fine granules, powders, injections, Parenteral preparations such as suppositories are mentioned. In general, for adults, it is appropriate to take 20 mg to 600 mg or less per day as glycolipid weight in several divided doses.
本発明において錠剤、カプセル剤、顆粒剤、細粒剤、散剤としての経口剤は、例えば、デンプン、乳糖、白糖、マンニット、カルボキシメチルセルロース、コーンスターチ、無機塩類等を用いて常法に従って製造される。これらの製剤中の糖脂質の配合量は特に限定されるものではなく適宜設計できる。この種の製剤には本発明の糖脂質含有組成物の他に、結合剤、崩壊剤、界面活性剤、滑沢剤、流動性促進剤、矯味剤、着色剤、香料等を適宜に使用することができる。 In the present invention, oral preparations such as tablets, capsules, granules, fine granules, and powders are produced according to a conventional method using, for example, starch, lactose, sucrose, mannitol, carboxymethylcellulose, corn starch, inorganic salts, and the like. . The blending amount of the glycolipid in these preparations is not particularly limited and can be appropriately designed. In addition to the glycolipid-containing composition of the present invention, a binder, a disintegrant, a surfactant, a lubricant, a fluidity promoter, a corrigent, a coloring agent, a fragrance and the like are appropriately used for this type of preparation. be able to.
ここに、結合剤としてデンプン、デキストリン、アラビアゴム末、ゼラチン、ヒドロキシプロピルスターチ、メチルセルロースナトリウム、ヒドロキシプロピルセルロース、結晶セルロース、エチルセルロース、ポリビニルピロリドン、マクロゴール等を例示できる。崩壊剤としてはデンプン、ヒドロキシプロピルスターチ、カルボキシメチルセルロースナトリウム、カルボキシメチルセルロースカルシウム、カルボキシメチルセルロース、低置換ヒドロキシプロピルセルロース等を例として挙げることができる。界面活性剤の例としてラウリル硫酸ナトリウム、大豆レシチン、蔗糖脂肪酸エステル、ポリオキシエチレンソルビタン脂肪酸エステル等を挙げることができる。滑沢剤では、タルク、ロウ類、水素添加植物油、蔗糖脂肪酸エステル、ステアリン酸マグネシウム、ステアリン酸カルシウム、ステアリン酸アルミニウム、ポリエチレングリコール等を例示できる。流動性促進剤では、軽質無水ケイ酸、乾燥水酸化アルミニウムゲル、合成ケイ酸アルミニウム、ケイ酸マグネシウム等を例として挙げることができる。また、糖脂質含有組成物は懸濁液、エマルション剤、シロップ剤、エリキシル剤としても投与することができ、これらの各種剤形には、矯味矯臭剤、着色剤を含有させてもよい。 Examples of the binder include starch, dextrin, gum arabic powder, gelatin, hydroxypropyl starch, sodium methylcellulose, hydroxypropylcellulose, crystalline cellulose, ethylcellulose, polyvinylpyrrolidone, macrogol and the like. Examples of the disintegrant include starch, hydroxypropyl starch, carboxymethylcellulose sodium, carboxymethylcellulose calcium, carboxymethylcellulose, and low-substituted hydroxypropylcellulose. Examples of the surfactant include sodium lauryl sulfate, soybean lecithin, sucrose fatty acid ester, polyoxyethylene sorbitan fatty acid ester and the like. Examples of lubricants include talc, waxes, hydrogenated vegetable oils, sucrose fatty acid esters, magnesium stearate, calcium stearate, aluminum stearate, polyethylene glycol and the like. Examples of the fluidity promoter include light anhydrous silicic acid, dry aluminum hydroxide gel, synthetic aluminum silicate, magnesium silicate and the like. The glycolipid-containing composition can also be administered as a suspension, emulsion, syrup, or elixir, and these various dosage forms may contain a flavoring agent and a coloring agent.
非経口剤として本発明の所望の効果を発現せしめるには、患者の年齢、体重、疾患の程度により異なるが、通常、成人で糖脂質の重量として１日あたり１〜６０ｍｇの静注、点滴静注、皮下注射、筋肉注射が適当である。この非経口投与剤は常法に従って製造され、希釈剤として一般に注射用蒸留水、生理食塩水、ブドウ糖水溶液、注射用植物油、ゴマ油、ラッカセイ油、大豆油、トウモロコシ油、プロピレングリコール等を用いることができる。さらに必要に応じて、殺菌剤、防腐剤、安定剤を加えてもよい。また、この非経口剤は安定性の点から、バイアル等に充填後冷凍し、通常の凍結乾燥処理により水分を除き、使用直前に凍結乾燥物から液剤を再調製することもできる。さらに必要に応じて、等張化剤、安定剤、防腐剤、無痛化剤を加えてもよい。これら製剤中の糖脂質含有組成物の配合量は特に限定されるものではなく任意に設定できる。その他の非経口剤の例として、外用液剤、軟膏等の塗布剤、直腸内投与のための坐剤等が挙げられ、これらも常法に従って製造される。 In order to achieve the desired effect of the present invention as a parenteral preparation, it varies depending on the age, body weight, and degree of disease of the patient, but it is usually 1-60 mg / day intravenously or intravenously as the weight of glycolipid in adults. Note, subcutaneous injection and intramuscular injection are appropriate. This parenteral preparation is produced according to a conventional method, and generally used as diluent is distilled water for injection, physiological saline, aqueous glucose solution, vegetable oil for injection, sesame oil, peanut oil, soybean oil, corn oil, propylene glycol, etc. it can. Furthermore, you may add a disinfectant, antiseptic | preservative, and a stabilizer as needed. In addition, from the viewpoint of stability, this parenteral preparation can be frozen after filling into a vial or the like, the water can be removed by ordinary freeze-drying treatment, and the liquid preparation can be re-prepared from the freeze-dried product immediately before use. Furthermore, you may add an isotonic agent, a stabilizer, an antiseptic | preservative, and a soothing agent as needed. The amount of the glycolipid-containing composition in these preparations is not particularly limited and can be arbitrarily set. Examples of other parenteral agents include liquid preparations for external use, coating agents such as ointments, suppositories for rectal administration, etc., and these are also produced according to conventional methods.
本発明の糖脂質含有組成物の他の好適な用途は食用組成物である。即ち、本発明の糖脂質含有組成物は必要に応じてさらに精製した後、これをそのまま液状、ゲル状あるいは固形状の食品、例えばジュース、清涼飲料、茶、スープ、豆乳、サラダ油、ドレッシング、ヨーグルト、ゼリー、プリン、ふりかけ、育児用粉乳、ケーキミックス、粉末状または液状の乳製品、パン、クッキー等に添加したり、必要に応じてデキストリン、乳糖、澱粉等の賦形剤や香料、色素等とともにペレット、錠剤、顆粒等に加工したり、またゼラチン等で被覆してカプセルに成形加工して健康食品や栄養補助食品、機能性食品等として利用できる。 Another suitable use of the glycolipid-containing composition of the present invention is an edible composition. That is, the glycolipid-containing composition of the present invention is further purified as necessary, and is then used as it is in a liquid, gel or solid food such as juice, soft drink, tea, soup, soy milk, salad oil, dressing, yogurt Jelly, pudding, sprinkles, infant formula, cake mix, powdered or liquid dairy products, bread, cookies, etc., and if necessary, excipients such as dextrin, lactose, starch, flavorings, pigments, etc. At the same time, it can be processed into pellets, tablets, granules, etc., or coated with gelatin and formed into capsules for use as health foods, dietary supplements, functional foods, and the like.
これらの食品類あるいは食用組成物における本発明の糖脂質含有組成物の配合量は、当該食品や組成物の種類や状態等により一律に規定しがたいが、約０．０１〜５０重量％、より好ましくは０．１〜３０重量％である。配合量が０．０１重量％未満では経口摂取による所望の効果が小さく、５０重量％を超えると食品の種類によっては風味を損ない、または当該食品を調製できなくなる場合がある。 The blending amount of the glycolipid-containing composition of the present invention in these foods or edible compositions is difficult to define uniformly depending on the type or state of the food or composition, but is about 0.01 to 50% by weight, More preferably, it is 0.1 to 30% by weight. If the blending amount is less than 0.01% by weight, the desired effect by oral intake is small, and if it exceeds 50% by weight, the flavor may be impaired or the food may not be prepared depending on the type of food.
以下、図面を参照しながら本発明の実施例について説明するが、本発明はこれら実施例によって何ら限定されるものではない。 Examples of the present invention will be described below with reference to the drawings, but the present invention is not limited to these examples.
〔実施例１：本発明の第１の糖脂質含有組成物（糖脂質画分）の精製〕
本実施例では、図２に示される方法により、原料のホウレン草（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）から本発明の第１の糖脂質含有組成物を精製した。まず、市販ホウレン草の乾燥物１００ｇを細断した後、水溶性成分をできるだけ除去するために、６０℃の温水１Ｌで２回洗浄した。次に、濾紙を使用して濾過後、水分を除去し、得られた固形物（残渣）に１Ｌのエタノールを加え、撹拌しながら６０℃で還流抽出を２回行った。抽出液は濾過後、減圧下で濃縮を行い、ホウレン草のオイル状抽出物２０．５ｇを得た。[Example 1: Purification of the first glycolipid-containing composition (glycolipid fraction) of the present invention]
In this example, the first glycolipid-containing composition of the present invention was purified from the raw material spinach (Spinia oleracea L.) by the method shown in FIG. First, after chopping 100 g of a dried product of commercially available spinach, it was washed twice with 1 L of warm water at 60 ° C. in order to remove water-soluble components as much as possible. Next, after filtration using filter paper, water was removed, 1 L of ethanol was added to the obtained solid (residue), and reflux extraction was performed twice at 60 ° C. with stirring. The extract was filtered and concentrated under reduced pressure to obtain 20.5 g of spinach oily extract.
次に、得られた上記抽出物を７０％含水エタノール溶液に溶解し、疎水クロマトグラフィー用樹脂５００ｇ（ダイヤイオンＨＰ−２０、三菱化学社製）に注入した。その後、７０％エタノールで未吸着物質を洗浄・溶出した画分Ｉ（水溶性画分、１３．０ｇ）、９５％エタノールで溶出される画分ＩＩ（糖脂質画分すなわち本発明の糖脂質含有組成物、６．５ｇ）、クロロホルムで溶出される画分ＩＩＩ（色素画分、１．０ｇ）の３つに分画した。 Next, the obtained extract was dissolved in a 70% aqueous ethanol solution and poured into 500 g of a resin for hydrophobic chromatography (Diaion HP-20, manufactured by Mitsubishi Chemical Corporation). Thereafter, fraction I (water-soluble fraction, 13.0 g) obtained by washing and eluting unadsorbed substances with 70% ethanol, fraction II (glycolipid fraction, ie containing the glycolipid of the present invention) eluted with 95% ethanol The composition was 6.5 g) and fraction III (dye fraction, 1.0 g) eluted with chloroform.
図３は、上記画分Ｉ〜ＩＩＩに含まれる成分を薄層クロマトグラフィーにより解析した結果を示す図である。薄層クロマトグラフィーは、シリカゲルプレートの開始点に試料２μｇを添加して、クロロホルム：メタノール＝３：１で展開後、５０％硫酸スプレー・１１０℃加熱で検出した。図中、レーン１〜４は、それぞれ、エタノール抽出液（上記オイル状抽出物のこと。以下同じ。）、画分ＩＩＩ、画分Ｉ、画分ＩＩの検出結果である。同図に示すように、レーン４の画分ＩＩには、ＭＧＤＧ、ＤＧＤＧおよびＳＱＤＧの３つの糖脂質が含まれていることが分かった。一方、画分Ｉおよび画分ＩＩＩではこれら糖脂質はいずれも検出されなかった。 FIG. 3 is a diagram showing the results of analyzing the components contained in the fractions I to III by thin layer chromatography. In thin layer chromatography, 2 μg of sample was added to the starting point of the silica gel plate, developed with chloroform: methanol = 3: 1, and then detected with 50% sulfuric acid spray and heating at 110 ° C. In the figure,
次にシリカゲルカラムクロマトグラフィーを用いて、糖脂質画分である画分ＩＩからＭＧＤＧ、ＤＧＤＧおよびＳＱＤＧを単離・精製した。その結果、糖脂質画分２５０ｍｇには、ＭＧＤＧが８４．７ｍｇ（３３．９％）、ＤＧＤＧが３５．０ｍｇ（１４．０％）、ＳＱＤＧが９２．３ｍｇ（３６．９％）含まれていることが分かった。さらに、精製したＭＧＤＧ、ＤＧＤＧ、ＳＱＤＧについて１Ｈ−ＮＭＲおよび１３Ｃ−ＮＭＲを使用して構造の確認と純度の検定を行った結果、ＭＧＤＧ、ＤＧＤＧ、ＳＱＤＧともに化学構造を確認でき、純度はいずれも９５％以上であった。Next, MGDG, DGDG, and SQDG were isolated and purified from fraction II, which is a glycolipid fraction, using silica gel column chromatography. As a result, the glycolipid fraction 250 mg contained MGDG 84.7 mg (33.9%), DGDG 35.0 mg (14.0%), and SQDG 92.3 mg (36.9%). I understood that. Furthermore, as a result of confirming the structure and purity of purified MGDG, DGDG, and SQDG using 1 H-NMR and 13 C-NMR, the chemical structure of MGDG, DGDG, and SQDG can be confirmed. Was 95% or more.
〔実施例２：上記糖脂質画分のＤＮＡ合成酵素阻害活性〕
上記方法により得られた糖脂質画分（画分ＩＩ）すなわち本発明の第１の糖脂質含有組成物のＤＮＡ合成酵素阻害活性を調べた。ＤＮＡ合成酵素には哺乳動物由来のＤＮＡ合成酵素α、βを使用した。より詳細には、ＤＮＡ合成酵素αは、牛胸腺の抽出液を抗体カラムクロマトグラフィーで精製した標品を、ＤＮＡ合成酵素βは、ラットのＤＮＡ合成酵素β遺伝子を導入した組換え大腸菌を培養後、破砕して精製した標品を用いた。[Example 2: DNA synthase inhibitory activity of the glycolipid fraction]
The DNA synthase inhibitory activity of the glycolipid fraction (Fraction II) obtained by the above method, that is, the first glycolipid-containing composition of the present invention was examined. Mammal-derived DNA synthases α and β were used as DNA synthases. More specifically, DNA synthase α is a sample obtained by purifying bovine thymus extract by antibody column chromatography, and DNA synthase β is obtained after culturing recombinant Escherichia coli into which a rat DNA synthase β gene is introduced. The sample was crushed and purified.
上記ＤＮＡ合成酵素α、βに対する阻害作用の測定には、一般的なＤＮＡ合成酵素反応系（日本生化学会編、新生化学実験講座２，核酸ＩＶ、東京化学同人、第６３頁〜６６頁）を用いた。すなわち、放射性同位元素で標識した［３Ｈ］−ＴＴＰを含む系においてＤＮＡ合成反応を行い、放射比活性を生成物（合成ＤＮＡ鎖）量の指標とするものである。阻害率は、（ａ）コントロールでの合成ＤＮＡ量、（ｂ）調査対象のホウレン草画分（またはエタノール抽出液）存在下での合成ＤＮＡ量について、
（ａ−ｂ）／ａ×１００＝阻害率（％）
として評価した。得られた結果を図４に示す。同図は、ＤＮＡ合成酵素αに対する阻害結果を示すグラフであり、図中、三角印はエタノール抽出液存在下での阻害結果、菱形印、丸印、四角印はそれぞれ画分Ｉ〜ＩＩＩ存在下での阻害結果である。同図に示すように、画分ＩＩには強いＤＮＡ合成酵素α阻害活性が認められ、画分ＩＩの５０％阻害濃度（ＩＣ５０）はおよそ４０μｇ／ｍｌであった。一方、エタノール抽出液、画分ＩおよびＩＩＩは、ＤＮＡ合成酵素αを殆ど阻害しなかった。For the measurement of the inhibitory action on the above-mentioned DNA synthases α and β, a general DNA synthase reaction system (edited by the Japanese Biochemical Society, New
(Ab) / a × 100 = inhibition rate (%)
As evaluated. The obtained results are shown in FIG. This figure is a graph showing the inhibition results for DNA synthase α. In the figure, triangles indicate the inhibition results in the presence of the ethanol extract, diamonds, circles, and squares indicate the presence of fractions I to III, respectively. It is the inhibition result in. As shown in the figure, a strong DNA synthetase α inhibitory activity was observed in fraction II, and the 50% inhibitory concentration (IC 50 ) of fraction II was approximately 40 μg / ml. On the other hand, the ethanol extract and fractions I and III hardly inhibited DNA synthase α.
〔実施例３：上記糖脂質画分のヒト癌細胞増殖抑制活性〕
次に、上記ホウレン草各画分およびエタノール抽出液が癌細胞増殖抑制作用を有するかどうかを検討した。実験には、ヒト胃癌細胞株であるＮＵＧＣ−３細胞を使用し、種々の濃度のホウレン草画分（またはエタノール抽出液）を添加してインキュベーションし、４８時間後、各場合における癌細胞の生存率を通常のＭＴＴアッセイにより決定した。[Example 3: Human cancer cell growth inhibitory activity of the glycolipid fraction]
Next, it was examined whether each of the above spinach fractions and the ethanol extract had cancer cell growth inhibitory action. In the experiment, human gastric cancer cell line NUGC-3 cells were used, incubated with various concentrations of spinach herb fraction (or ethanol extract), and after 48 hours, the survival rate of cancer cells in each case Was determined by routine MTT assay.
その結果を図５に示す。図中、三角印はエタノール抽出液存在下での増殖抑制結果、菱形印、丸印、四角印はそれぞれ画分Ｉ〜ＩＩＩ存在下での増殖抑制結果である。同図に示すように、画分ＩＩにはヒト胃癌細胞に対する強い増殖抑制活性が認められたが、エタノール抽出液、画分ＩおよびＩＩＩには増殖抑制活性は認められなかった。この結果はＤＮＡ合成酵素α阻害活性の実験結果とよく符合するものであった。 The result is shown in FIG. In the figure, triangles indicate the results of growth inhibition in the presence of the ethanol extract, and diamonds, circles, and squares indicate the results of growth suppression in the presence of fractions I to III, respectively. As shown in the figure, fraction II showed strong growth inhibitory activity against human gastric cancer cells, but ethanol extract and fractions I and III showed no growth inhibitory activity. This result was in good agreement with the experimental result of the DNA synthase α inhibitory activity.
〔実施例４：上記糖脂質画分のリパーゼ処理による第２の糖脂質含有組成物の精製〕
以下の方法により、上記糖脂質画分（画分ＩＩ）をリパーゼ処理し、第２の糖脂質含有組成物を精製した。
図１３に示すように、糖脂質画分（画分ＩＩ）１０ｍｇ／ｍｌおよびブタ膵臓リパーゼ１０ｍｇ／ｍｌを反応液（０．２Ｍ Ｔｒｉｓ−ＨＣｌ（ｐＨ７．６）、０．２５Ｍ ＣａＣｌ２）において３７℃、２０分間振とう反応させ、糖脂質をリパーゼにより加水分解し、ジアシルグリセロールタイプのものからモノアシルグリセロールタイプのものを生成し、その後６ＮのＨＣｌで反応を止めた。[Example 4: Purification of second glycolipid-containing composition by lipase treatment of glycolipid fraction]
The glycolipid fraction (fraction II) was treated with lipase by the following method to purify the second glycolipid-containing composition.
As shown in FIG. 13, 10 mg / ml of glycolipid fraction (fraction II) and 10 mg / ml of porcine pancreatic lipase were reacted in a reaction solution (0.2 M Tris-HCl (pH 7.6), 0.25 M CaCl 2 ). The mixture was shaken at 20 ° C. for 20 minutes, and the glycolipid was hydrolyzed with lipase to produce a monoacylglycerol type from a diacylglycerol type, and then the reaction was stopped with 6N HCl.
次にｎ−ブタノール（ｎ−ＢｕＯＨ）と水で分配することにより、糖脂質はｎ−ブタノール層へ、リパーゼは水層へ移動するので、リパーゼ分解された糖脂質（本発明の第２の糖脂質含有組成物）を精製することができる。 Next, by partitioning with n-butanol (n-BuOH) and water, the glycolipid moves to the n-butanol layer, and the lipase moves to the water layer. Therefore, the lipase-degraded glycolipid (the second sugar of the present invention) The lipid-containing composition) can be purified.
図６は、糖脂質画分（画分ＩＩ）のリパーゼ反応前（第１の糖脂質含有組成物）およびリパーゼ反応後（第２の糖脂質含有組成物）にそれぞれ含まれる成分を、図３と同様に薄層クロマトグラフィーにより解析した結果を示す図である。図中、レーン１はリパーゼ反応前、レーン２はリパーゼ反応後の検出結果である。同図に示すように、レーン２の第２の糖脂質含有組成物には、ＭＧＭＧおよびＳＱＭＧの２つの糖脂質が含まれ、第１の糖脂質含有組成物中の糖脂質ＭＧＤＧ・ＳＱＤＧは、リパーゼ加水分解反応により、それぞれＭＧＭＧ・ＳＱＭＧに分解された。一方、ＤＧＤＧは、リパーゼにより加水分解されず、第２の糖脂質含有組成物中にも検出された。 FIG. 6 shows components contained in the glycolipid fraction (fraction II) before the lipase reaction (first glycolipid-containing composition) and after the lipase reaction (second glycolipid-containing composition), respectively. It is a figure which shows the result analyzed by thin layer chromatography similarly to FIG. In the figure,
〔実施例５：第２の糖脂質含有組成物のＤＮＡ合成酵素阻害活性およびヒト癌細胞増殖抑制活性〕
図４の実験と同様の方法により、リパーゼ処理前の糖脂質画分（第１の糖脂質含有組成物）およびリパーゼ処理後の糖脂質画分（第２の糖脂質含有組成物）のＤＮＡ合成酵素阻害活性を調べ、両者を比較した。その結果を図７に示す。図中、黒丸印は、ＤＮＡ合成酵素αに対するリパーゼ処理前の糖脂質画分、黒四角印は、ＤＮＡ合成酵素αに対するリパーゼ処理後の糖脂質画分、白丸印は、ＤＮＡ合成酵素βに対するリパーゼ処理前の糖脂質画分、白四角印は、ＤＮＡ合成酵素βに対するリパーゼ処理後の糖脂質画分、の各阻害結果を示すものである。[Example 5: DNA synthase inhibitory activity and human cancer cell growth inhibitory activity of the second glycolipid-containing composition]
DNA synthesis of the glycolipid fraction before the lipase treatment (first glycolipid-containing composition) and the glycolipid fraction after the lipase treatment (second glycolipid-containing composition) by the same method as in the experiment of FIG. Enzyme inhibitory activity was examined and compared. The result is shown in FIG. In the figure, black circles indicate the glycolipid fraction before lipase treatment for DNA synthase α, black squares indicate the glycolipid fraction after lipase treatment for DNA synthase α, and white circles indicate lipase for DNA synthase β. The glycolipid fraction before treatment and the white squares indicate the results of inhibition of the glycolipid fraction after lipase treatment for DNA synthase β.
同図に示すように、リパーゼ処理前およびリパーゼ処理後のいずれの糖脂質画分も、ＤＮＡ合成酵素βよりＤＮＡ合成酵素αに対してより強いＤＮＡ合成酵素阻害活性を有していた。また、リパーゼ処理後の糖脂質画分（即ち、第２の糖脂質含有組成物）のほうが、リパーゼ処理前の糖脂質画分（即ち、第１の糖脂質含有組成物）よりもＤＮＡ合成酵素αおよびβを強く阻害した。 As shown in the figure, both glycolipid fractions before and after lipase treatment had stronger DNA synthetase inhibitory activity against DNA synthase α than with DNA synthase β. In addition, the glycolipid fraction after lipase treatment (that is, the second glycolipid-containing composition) is more DNA synthase than the glycolipid fraction before lipase treatment (that is, the first glycolipid-containing composition). α and β were strongly inhibited.
次に、図５の実験と同様の方法により、リパーゼ処理前の糖脂質画分（第１の糖脂質含有組成物）およびリパーゼ処理後の糖脂質画分（第２の糖脂質含有組成物）のヒト癌細胞増殖抑制作用を調べ、両者を比較した。その結果を図８に示す。図中、黒丸印はリパーゼ処理前の糖脂質画分、黒四角印はリパーゼ処理後の糖脂質画分の結果である。同図に示すように、リパーゼ処理後の糖脂質画分（即ち、第２の糖脂質含有組成物）のほうが、リパーゼ処理前の糖脂質画分（即ち、第１の糖脂質含有組成物）よりもヒト胃癌細胞（ＮＵＧＣ−３）の増殖を強く抑制・阻害した。 Next, by the same method as the experiment of FIG. 5, the glycolipid fraction before the lipase treatment (first glycolipid-containing composition) and the glycolipid fraction after the lipase treatment (second glycolipid-containing composition) The human cancer cell growth inhibitory action of was investigated, and both were compared. The result is shown in FIG. In the figure, black circles indicate the glycolipid fraction before lipase treatment, and black squares indicate the glycolipid fraction after lipase treatment. As shown in the figure, the glycolipid fraction after the lipase treatment (ie, the second glycolipid-containing composition) is the glycolipid fraction before the lipase treatment (ie, the first glycolipid-containing composition). The growth of human gastric cancer cells (NUGC-3) was strongly suppressed / inhibited.
以上のように、リパーゼ処理した第２の糖脂質含有組成物のほうが、リパーゼ処理していない第１の糖脂質含有組成物に比べて、ＤＮＡ合成酵素阻害活性およびヒト癌細胞増殖抑制活性いずれも強かった。 As described above, the second glycolipid-containing composition treated with lipase has both a DNA synthase inhibitory activity and a human cancer cell growth inhibitory activity compared to the first glycolipid-containing composition not treated with lipase. It was strong.
〔実施例６：本発明の第１および第２の糖脂質含有組成物の抗腫瘍活性〕
さらに、上記第１および第２の糖脂質含有組成物のｉｎ ｖｉｖｏ抗腫瘍活性を調査した。実験動物にはシリアン（ゴールデン）ハムスターを用い、同ハムスターへ黒色腫（Ｍｅｌａｎｏｔｉｃ Ｎｏ．１７９，Ｄ１−１７９）片を皮下移植して、一週間定着させてから、糖脂質含有組成物を生理食塩水に溶かして、一日おきに１０ｍｇ／ｋｇを皮下注射により投与して、腫瘍体積を測定した。その結果を図９および図１０に示す。[Example 6: Antitumor activity of first and second glycolipid-containing compositions of the present invention]
Furthermore, the in vivo antitumor activity of the first and second glycolipid-containing compositions was investigated. A Syrian (Golden) hamster was used as an experimental animal, and a melanoma (Melanotic No. 179, D1-179) fragment was subcutaneously implanted into the hamster and allowed to settle for a week, and then the glycolipid-containing composition was added to physiological saline. The tumor volume was measured by dissolving 10 mg / kg every other day by subcutaneous injection. The results are shown in FIG. 9 and FIG.
図９のグラフ中、白四角印はコントロールであり、生理食塩水を投与した場合の結果、黒三角印、黒丸印、黒菱形印は、それぞれ、エタノール抽出液、リパーゼ処理前の糖脂質画分（第１の糖脂質含有組成物）、リパーゼ処理後の糖脂質画分（第２の糖脂質含有組成物）、を投与した場合の結果である。ＤＮＡ合成酵素阻害活性およびヒト癌細胞増殖抑制活性と同様に、抗腫瘍活性についても第２の糖脂質含有組成物のほうが第１の糖脂質含有組成物よりも強かった。また、図１０の（ａ）は生理食塩水を投与した場合の結果、（ｂ）は第２の糖脂質含有組成物を生理食塩水に溶かして投与した場合の結果であり、リパーゼ処理した第２の糖脂質含有組成物を投与したハムスターでは腫瘍の肥大化は強く抑制された。 In the graph of FIG. 9, the white square mark is a control, and as a result of the administration of physiological saline, the black triangle mark, the black circle mark, and the black rhombus mark are the ethanol lipid extract and the glycolipid fraction before lipase treatment, respectively. It is a result at the time of administering (1st glycolipid containing composition) and the glycolipid fraction (2nd glycolipid containing composition) after a lipase process. Similar to the DNA synthase inhibitory activity and human cancer cell growth inhibitory activity, the anti-tumor activity of the second glycolipid-containing composition was stronger than that of the first glycolipid-containing composition. FIG. 10 (a) shows the results when physiological saline was administered, and FIG. 10 (b) shows the results when the second glycolipid-containing composition was dissolved in physiological saline. In the hamster administered with the glycolipid-containing composition of No. 2, tumor enlargement was strongly suppressed.
また、ヒト由来の腫瘍をマウスに移植した実験により、本発明の糖脂質含有組成物のｉｎ ｖｉｖｏ抗腫瘍活性を調査した。図１１は、ヒト子宮癌細胞（ＨｅＬａ）を移植したヌードマウス（ｂａｌｂ／ｃ，−／−）において、本発明の糖脂質含有組成物のｉｎ ｖｉｖｏ抗腫瘍活性を調査した結果である。実験は、ヌードマウス（ｂａｌｂ／ｃ，−／−）へヒト由来の子宮癌細胞（ＨｅＬａ）を皮下注射により移植後、１７日目に腫瘍体積が２５ｍｍ３になったところで、３日に１回、ホウレン草の画分ＩＩ（第１の糖脂質含有組成物）またはリパーゼ処理後の画分ＩＩ（第２の糖脂質含有組成物）を生理食塩水に溶かして皮下注射することにより行った。生理食塩水には５ｍｇ／ｍｌの割合で本発明の糖脂質含有組成物を溶かし、１回当たり、本発明の糖脂質含有組成物を１０ｍｇ／ｋｇマウスに投与した。その結果、図１１に示すように、生理食塩水だけを注射したコントロールと比べて、ホウレン草の画分ＩＩおよびリパーゼ処理後の画分ＩＩを注射したマウスでは腫瘍肥大化の抑制が見られた。また、ホウレン草の画分ＩＩよりもリパーゼ処理後の画分ＩＩの方がより強い抗腫瘍活性が認められた。
以上の結果より、本発明の糖脂質含有組成物は、ヒト由来の腫瘍に対してもハムスター由来の腫瘍と同様に抑制活性が認められることから、健康食品の開発などに有用である。Further, the in vivo antitumor activity of the glycolipid-containing composition of the present invention was investigated by an experiment in which a human-derived tumor was transplanted into a mouse. FIG. 11 shows the results of investigating the in vivo antitumor activity of the glycolipid-containing composition of the present invention in nude mice (balb / c, − / −) transplanted with human uterine cancer cells (HeLa). The experiment was conducted once every 3 days when the tumor volume became 25 mm 3 on the 17th day after transplantation of human-derived uterine cancer cells (HeLa) into nude mice (balb / c, − / −) by subcutaneous injection. Spinach fraction II (first glycolipid-containing composition) or lipase-treated fraction II (second glycolipid-containing composition) was dissolved in physiological saline and injected subcutaneously. The glycolipid-containing composition of the present invention was dissolved in physiological saline at a rate of 5 mg / ml, and the glycolipid-containing composition of the present invention was administered to a 10 mg / kg mouse at one time. As a result, as shown in FIG. 11, tumor hypertrophy was suppressed in mice injected with spinach fraction II and lipase-treated fraction II, compared to controls injected with physiological saline alone. Further, stronger antitumor activity was observed in the fraction II after lipase treatment than in the spinach fraction II.
From the above results, the glycolipid-containing composition of the present invention is useful for the development of health foods and the like because it has inhibitory activity on human-derived tumors as well as hamster-derived tumors.
次に、本発明の糖脂質含有組成物を経口投与した場合にもｉｎ ｖｉｖｏ抗腫瘍活性が認められるか調査した。図１２は、マウス由来の肉腫細胞（Ｓ−１８０）を移植したＩＣＲマウスにおいて、本発明の糖脂質含有組成物を経口投与した場合のｉｎ ｖｉｖｏ抗腫瘍活性を調査した結果である。実験は、マウス由来の肉腫細胞（Ｓ−１８０）を皮下注射により移植後、４日目に腫瘍体積が１００ｍｍ３になったところで、毎日１回、ホウレン草の画分ＩＩ（第１の糖脂質含有組成物）を生理食塩水に溶かして経口投与することにより行った。生理食塩水には５ｍｇ／ｍｌの割合で本発明の糖脂質含有組成物を溶かし、１回当たり、本発明の糖脂質含有組成物を７０ｍｇ／ｋｇマウスに投与した。その結果、図１２に示すように、生理食塩水だけを飲ませたコントロールと比べて、ホウレン草の画分ＩＩを経口投与したマウスでは６日目から有意な抗腫瘍活性が観察され、３０日でほぼ腫瘍が消失した。
以上の結果より、本発明の糖脂質含有組成物は、飲料水などとして経口投与する抗癌機能性食品の開発などに有用である。Next, it was investigated whether or not in vivo antitumor activity was observed when the glycolipid-containing composition of the present invention was orally administered. FIG. 12 shows the results of investigating the in vivo antitumor activity when the glycolipid-containing composition of the present invention was orally administered to ICR mice transplanted with mouse-derived sarcoma cells (S-180). In the experiment, a mouse-derived sarcoma cell (S-180) was transplanted by subcutaneous injection, and when the tumor volume reached 100 mm 3 on the fourth day, spinach fraction II (containing the first glycolipid) was once a day. The composition was dissolved in physiological saline and orally administered. The glycolipid-containing composition of the present invention was dissolved in physiological saline at a rate of 5 mg / ml, and the glycolipid-containing composition of the present invention was administered to a 70 mg / kg mouse at one time. As a result, as shown in FIG. 12, a significant antitumor activity was observed from the 6th day in mice orally administered spinach Fraction II as compared to the control in which only physiological saline was drunk. The tumor almost disappeared.
From the above results, the glycolipid-containing composition of the present invention is useful for the development of anticancer functional foods that are orally administered as drinking water or the like.
〔実施例７：医薬用組成物、あるいは、食用組成物の試作〕
上記の方法により調製した本発明の糖脂質含有組成物１５０ｍｇ、精製大豆油１２５ｍｇ、ミツロウ１５ｍｇおよびビタミンＥ１０ｍｇを窒素ガス雰囲気下で約４０℃に加温し、十分に混合して均質な液状物とした。これをカプセル充填機に供給して１粒内容量３００ｍｇのゼラチン被覆カプセル製剤を試作した。この製剤は医薬用組成物または食用組成物として利用できるものである。[Example 7: Preparation of pharmaceutical composition or edible composition]
150 mg of the glycolipid-containing composition of the present invention prepared by the above method, 125 mg of purified soybean oil, 15 mg of beeswax and 10 mg of vitamin E are heated to about 40 ° C. in a nitrogen gas atmosphere and mixed thoroughly to obtain a homogeneous liquid product. did. This was supplied to a capsule filling machine, and a gelatin-coated capsule preparation with an inner volume of 300 mg was made as a prototype. This preparation can be used as a pharmaceutical composition or an edible composition.
以上のように、本発明の糖脂質含有組成物は、ＤＮＡ合成酵素阻害活性、癌細胞増殖抑制活性、抗腫瘍活性といった有用な生理活性を有することから、ＤＮＡ合成酵素阻害剤、抗癌剤（制癌剤）として利用可能であり、特に機能性食品などに好適に用いることができ、例えば抗癌作用または癌予防効果をもつ機能性食品として利用できるほか、前述したとおり、他の医薬用組成物および食用組成物（食品または食品添加物）としても利用できるものである。 As described above, the glycolipid-containing composition of the present invention has useful physiological activities such as a DNA synthase inhibitory activity, a cancer cell growth inhibitory activity, and an antitumor activity. Therefore, a DNA synthase inhibitor and an anticancer agent (anticancer agent) In particular, it can be suitably used for functional foods and the like. For example, it can be used as a functional food having an anticancer effect or cancer prevention effect, and as described above, other pharmaceutical compositions and edible compositions. It can also be used as a food (food or food additive).
Claims (12)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2005514037A JP4744297B2 (en) | 2003-09-22 | 2004-09-15 | Glycolipid-containing composition, use thereof, and production method thereof |
Applications Claiming Priority (4)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2003330682 | 2003-09-22 | ||
| JP2003330682 | 2003-09-22 | ||
| PCT/JP2004/013432 WO2005027937A1 (en) | 2003-09-22 | 2004-09-15 | Glycolipid-containing composition, use thereof and process for producing the same |
| JP2005514037A JP4744297B2 (en) | 2003-09-22 | 2004-09-15 | Glycolipid-containing composition, use thereof, and production method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPWO2005027937A1 true JPWO2005027937A1 (en) | 2007-11-15 |
| JP4744297B2 JP4744297B2 (en) | 2011-08-10 |
Family
ID=34373023
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP2005514037A Expired - Fee Related JP4744297B2 (en) | 2003-09-22 | 2004-09-15 | Glycolipid-containing composition, use thereof, and production method thereof |
Country Status (2)
| Country | Link |
|---|---|
| JP (1) | JP4744297B2 (en) |
| WO (1) | WO2005027937A1 (en) |
Families Citing this family (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP4839436B2 (en) * | 2005-11-02 | 2011-12-21 | 国立大学法人徳島大学 | Gastrointestinal mucosa protective agent, caveolin gene expression promoter and anti-stress agent |
| ATE524483T1 (en) * | 2006-07-03 | 2011-09-15 | Hyben Vital Licens Aps | METHOD FOR PRODUCING A GLYCOSIDE OF A MONO- OR DIACYL-GLYCEROL PRODUCT FROM A PLANT MATERIAL |
| WO2011087403A1 (en) * | 2010-01-18 | 2011-07-21 | Lipidor Ab | Alcoholysis of glycolipid and corresponding lysis product |
| EP2389816A1 (en) | 2010-05-25 | 2011-11-30 | Nestec S.A. | Synergistic antioxidant composition |
| JP5676702B2 (en) * | 2012-08-17 | 2015-02-25 | 株式会社サラヴィオ化粧品 | Compositions containing extracts extracted from algal bodies, cosmetic compositions, therapeutic / preventive agents for inflammatory diseases, and novel microorganisms |
| US10123986B2 (en) | 2012-12-24 | 2018-11-13 | Qualitas Health, Ltd. | Eicosapentaenoic acid (EPA) formulations |
| US9629820B2 (en) | 2012-12-24 | 2017-04-25 | Qualitas Health, Ltd. | Eicosapentaenoic acid (EPA) formulations |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2002338474A (en) * | 2001-05-23 | 2002-11-27 | Bizen Chemical Co Ltd | Caspase inhibitor |
-
2004
- 2004-09-15 WO PCT/JP2004/013432 patent/WO2005027937A1/en active Application Filing
- 2004-09-15 JP JP2005514037A patent/JP4744297B2/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2002338474A (en) * | 2001-05-23 | 2002-11-27 | Bizen Chemical Co Ltd | Caspase inhibitor |
Non-Patent Citations (1)
| Title |
|---|
| JPN6011001495, Ｂｉｏｃｈｅｍｉｃａｌ Ｐｈａｒｍａｃｏｌｏｇｙ, 2003, Ｖｏｌ．６５, ｐ．２５９−２６７ * |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2005027937A1 (en) | 2005-03-31 |
| JP4744297B2 (en) | 2011-08-10 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP2014028858A (en) | Maillard reaction inhibitor | |
| EP2682122A1 (en) | Growth hormone secretion promoter | |
| JP4744297B2 (en) | Glycolipid-containing composition, use thereof, and production method thereof | |
| ES2732247T3 (en) | Olive extract containing desramnosilacteósido | |
| JP4629834B2 (en) | Nerve cell activator and use thereof | |
| KR101722297B1 (en) | ISOLATION OF SINGLE COMPOUND FROM Albatrellus fletti ITS APPLICATION IN TREATING AND PREVENTING OBESITY | |
| JP4913289B2 (en) | Caspase inhibitor | |
| KR20210146590A (en) | Pharmaceutical composition for preventing or treating cancer comprising belamcandae rhizoma extract as an active ingredient | |
| JP2006016340A (en) | Blood uric acid level reduction agent having extract of punica granatum l. as active ingredient | |
| JP2006022033A (en) | Neovascularization inhibitor | |
| KR102114510B1 (en) | Composition for modulating immunity comprising coenzyme Q10 and omega 3 | |
| KR101691256B1 (en) | COMPOSITION COMPRISING ALCOHOL EXTRACTS FROM Oligoporus stipticus, FOR TREATING OR PREVENTING HYPERLIPIDEMIA | |
| KR20190142672A (en) | Pharmaceutical composition for preventing or treating liver damage comprising Curcuma longa extract | |
| KR102607716B1 (en) | A composition for preventing or improving circadian rhythm disorders comprising Artemisia annua plant extracts or artemisinin | |
| KR101625280B1 (en) | COMPOSITION COMPRSING ALCOHOL EXRACTS FROM Albatrellus fletti FOR TREATING OR PREVENTING OBESITY | |
| KR101773221B1 (en) | Composition comprising alcohol extracts from russula japonica for treating or preventing hyperlipidemia | |
| KR101651106B1 (en) | ISOLATED SINGLE COMPOUND FROM Mori Cortex Radicis ITS APPLICATION IN TREATING AND PREVENTING OBESITY | |
| JP2024064707A (en) | Fat decomposition promoter | |
| KR101576916B1 (en) | Compositon containing sulfuretin for treatment or improvement of dental diseases | |
| KR20240078554A (en) | Composition for improving inflammation or pruritis comprising an extract of Sikbangpung as an active ingredient | |
| KR101569254B1 (en) | composition comprising water extracts from oudemansiella platyphylla for treating or preventing hyperlipidemia | |
| KR20200020354A (en) | Pharmaceutical composition for preventing or treating inflammatory disease comprising extract of Pilea martinii as effective component | |
| KR20190142674A (en) | Pharmaceutical composition for preventing or treating liver damage comprising Curcuma longa extract | |
| KR20100045581A (en) | Composition comprising the fraction of starfishes having anti-inflammatory or anti-allergic activity | |
| JP2005162634A (en) | Dna synthetase inhibitor |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A621 | Written request for application examination |
Free format text: JAPANESE INTERMEDIATE CODE: A621 Effective date: 20070904 |
|
| RD03 | Notification of appointment of power of attorney |
Free format text: JAPANESE INTERMEDIATE CODE: A7423 Effective date: 20080307 |
|
| RD04 | Notification of resignation of power of attorney |
Free format text: JAPANESE INTERMEDIATE CODE: A7424 Effective date: 20080321 |
|
| A521 | Written amendment |
Free format text: JAPANESE INTERMEDIATE CODE: A821 Effective date: 20080307 |
|
| A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20110118 |
|
| A521 | Written amendment |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20110316 |
|
| TRDD | Decision of grant or rejection written | ||
| A01 | Written decision to grant a patent or to grant a registration (utility model) |
Free format text: JAPANESE INTERMEDIATE CODE: A01 Effective date: 20110419 |
|
| A01 | Written decision to grant a patent or to grant a registration (utility model) |
Free format text: JAPANESE INTERMEDIATE CODE: A01 |
|
| A61 | First payment of annual fees (during grant procedure) |
Free format text: JAPANESE INTERMEDIATE CODE: A61 Effective date: 20110510 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20140520 Year of fee payment: 3 |
|
| R150 | Certificate of patent or registration of utility model |
Ref document number: 4744297 Country of ref document: JP Free format text: JAPANESE INTERMEDIATE CODE: R150 Free format text: JAPANESE INTERMEDIATE CODE: R150 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| LAPS | Cancellation because of no payment of annual fees |