CN108085303B - 一种活性虫草酵素的制备方法 - Google Patents
一种活性虫草酵素的制备方法 Download PDFInfo
- Publication number
- CN108085303B CN108085303B CN201711096208.8A CN201711096208A CN108085303B CN 108085303 B CN108085303 B CN 108085303B CN 201711096208 A CN201711096208 A CN 201711096208A CN 108085303 B CN108085303 B CN 108085303B
- Authority
- CN
- China
- Prior art keywords
- cordyceps
- cordyceps sinensis
- culture medium
- enzyme
- fermentation
- 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.)
- Expired - Fee Related
Links
- 241001248610 Ophiocordyceps sinensis Species 0.000 title claims abstract description 137
- 102000004190 Enzymes Human genes 0.000 title claims abstract description 117
- 108090000790 Enzymes Proteins 0.000 title claims abstract description 117
- 238000002360 preparation method Methods 0.000 title claims description 9
- 241000190633 Cordyceps Species 0.000 claims abstract description 84
- 238000000855 fermentation Methods 0.000 claims abstract description 46
- 230000004151 fermentation Effects 0.000 claims abstract description 46
- 238000000034 method Methods 0.000 claims abstract description 40
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 claims abstract description 30
- 229920002674 hyaluronan Polymers 0.000 claims abstract description 30
- 229960003160 hyaluronic acid Drugs 0.000 claims abstract description 30
- 239000008367 deionised water Substances 0.000 claims abstract description 27
- 229910021641 deionized water Inorganic materials 0.000 claims abstract description 27
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 27
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims abstract description 22
- 229910000396 dipotassium phosphate Inorganic materials 0.000 claims abstract description 16
- 239000007788 liquid Substances 0.000 claims abstract description 14
- 229910052799 carbon Inorganic materials 0.000 claims abstract description 13
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 claims abstract description 13
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims abstract description 12
- 238000012258 culturing Methods 0.000 claims abstract description 11
- 229910052757 nitrogen Inorganic materials 0.000 claims abstract description 11
- 229910017053 inorganic salt Inorganic materials 0.000 claims abstract description 3
- 239000001963 growth medium Substances 0.000 claims description 86
- 238000009630 liquid culture Methods 0.000 claims description 49
- 239000000306 component Substances 0.000 claims description 38
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 30
- 239000000126 substance Substances 0.000 claims description 27
- 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 claims description 20
- 239000008103 glucose Substances 0.000 claims description 20
- 238000012360 testing method Methods 0.000 claims description 20
- 239000001888 Peptone Substances 0.000 claims description 19
- 108010080698 Peptones Proteins 0.000 claims description 19
- 235000019319 peptone Nutrition 0.000 claims description 19
- OFEZSBMBBKLLBJ-BAJZRUMYSA-N cordycepin Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)C[C@H]1O OFEZSBMBBKLLBJ-BAJZRUMYSA-N 0.000 claims description 17
- OFEZSBMBBKLLBJ-UHFFFAOYSA-N cordycepine Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(CO)CC1O OFEZSBMBBKLLBJ-UHFFFAOYSA-N 0.000 claims description 17
- KQLDDLUWUFBQHP-UHFFFAOYSA-N Cordycepin Natural products C1=NC=2C(N)=NC=NC=2N1C1OCC(CO)C1O KQLDDLUWUFBQHP-UHFFFAOYSA-N 0.000 claims description 16
- 239000002609 medium Substances 0.000 claims description 16
- 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 claims description 15
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 15
- 150000004676 glycans Chemical class 0.000 claims description 13
- 229920001282 polysaccharide Polymers 0.000 claims description 13
- 239000005017 polysaccharide Substances 0.000 claims description 13
- 239000002777 nucleoside Substances 0.000 claims description 12
- 150000003833 nucleoside derivatives Chemical class 0.000 claims description 12
- 239000007787 solid Substances 0.000 claims description 12
- 238000000605 extraction Methods 0.000 claims description 11
- 238000001471 micro-filtration Methods 0.000 claims description 11
- 238000001816 cooling Methods 0.000 claims description 9
- 238000011081 inoculation Methods 0.000 claims description 9
- 230000008569 process Effects 0.000 claims description 9
- 230000001954 sterilising effect Effects 0.000 claims description 9
- -1 VB1 is 0.002-0.005% Chemical compound 0.000 claims description 7
- 238000004321 preservation Methods 0.000 claims description 7
- 238000002054 transplantation Methods 0.000 claims description 7
- 241000255789 Bombyx mori Species 0.000 claims description 6
- 244000000626 Daucus carota Species 0.000 claims description 6
- 235000002767 Daucus carota Nutrition 0.000 claims description 6
- 238000002222 matrix solid-phase dispersion Methods 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 6
- 239000012510 hollow fiber Substances 0.000 claims description 5
- 239000011148 porous material Substances 0.000 claims description 5
- 244000061456 Solanum tuberosum Species 0.000 claims description 4
- 235000002595 Solanum tuberosum Nutrition 0.000 claims description 4
- 239000000706 filtrate Substances 0.000 claims description 4
- 229910052760 oxygen Inorganic materials 0.000 claims description 4
- 238000009423 ventilation Methods 0.000 claims description 4
- 238000011065 in-situ storage Methods 0.000 claims description 3
- OQUKIQWCVTZJAF-UHFFFAOYSA-N phenol;sulfuric acid Chemical compound OS(O)(=O)=O.OC1=CC=CC=C1 OQUKIQWCVTZJAF-UHFFFAOYSA-N 0.000 claims description 3
- 238000001556 precipitation Methods 0.000 claims description 3
- 238000004659 sterilization and disinfection Methods 0.000 claims description 3
- 230000002195 synergetic effect Effects 0.000 claims description 3
- 239000000701 coagulant Substances 0.000 claims description 2
- 125000000600 disaccharide group Chemical group 0.000 claims description 2
- 150000002482 oligosaccharides Polymers 0.000 claims description 2
- 229910052564 epsomite Inorganic materials 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 62
- 239000002537 cosmetic Substances 0.000 abstract description 15
- 235000019797 dipotassium phosphate Nutrition 0.000 abstract description 12
- 230000032683 aging Effects 0.000 abstract description 11
- WRUGWIBCXHJTDG-UHFFFAOYSA-L magnesium sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[O-]S([O-])(=O)=O WRUGWIBCXHJTDG-UHFFFAOYSA-L 0.000 abstract description 11
- 230000003647 oxidation Effects 0.000 abstract description 9
- 238000007254 oxidation reaction Methods 0.000 abstract description 9
- 206010061218 Inflammation Diseases 0.000 abstract description 8
- 230000004054 inflammatory process Effects 0.000 abstract description 8
- 239000000203 mixture Substances 0.000 abstract description 8
- 230000003467 diminishing effect Effects 0.000 abstract description 6
- 238000005562 fading Methods 0.000 abstract description 5
- 239000000463 material Substances 0.000 abstract description 5
- 235000016709 nutrition Nutrition 0.000 abstract description 5
- 230000003321 amplification Effects 0.000 abstract description 4
- 238000003199 nucleic acid amplification method Methods 0.000 abstract description 4
- 230000035764 nutrition Effects 0.000 abstract description 4
- 238000005457 optimization Methods 0.000 abstract description 4
- 230000003698 anagen phase Effects 0.000 abstract description 3
- 238000005273 aeration Methods 0.000 abstract description 2
- 239000002054 inoculum Substances 0.000 abstract 1
- 239000004480 active ingredient Substances 0.000 description 32
- 241001264174 Cordyceps militaris Species 0.000 description 25
- 102000000380 Matrix Metalloproteinase 1 Human genes 0.000 description 21
- 108010016113 Matrix Metalloproteinase 1 Proteins 0.000 description 21
- 239000000047 product Substances 0.000 description 21
- 230000012010 growth Effects 0.000 description 19
- 238000002474 experimental method Methods 0.000 description 17
- 102000019197 Superoxide Dismutase Human genes 0.000 description 16
- 108010012715 Superoxide dismutase Proteins 0.000 description 16
- 230000005764 inhibitory process Effects 0.000 description 15
- 239000011550 stock solution Substances 0.000 description 15
- 238000001514 detection method Methods 0.000 description 14
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 13
- 235000019341 magnesium sulphate Nutrition 0.000 description 12
- 229940024606 amino acid Drugs 0.000 description 10
- 235000001014 amino acid Nutrition 0.000 description 10
- 150000001413 amino acids Chemical class 0.000 description 10
- 239000012071 phase Substances 0.000 description 10
- 238000011160 research Methods 0.000 description 10
- 239000000243 solution Substances 0.000 description 10
- 230000037394 skin elasticity Effects 0.000 description 9
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 8
- 230000003712 anti-aging effect Effects 0.000 description 8
- 230000002829 reductive effect Effects 0.000 description 8
- 239000011669 selenium Substances 0.000 description 8
- 241000233866 Fungi Species 0.000 description 7
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 description 7
- 239000003814 drug Substances 0.000 description 7
- 230000006870 function Effects 0.000 description 7
- 230000002035 prolonged effect Effects 0.000 description 7
- 239000000523 sample Substances 0.000 description 7
- 229910052711 selenium Inorganic materials 0.000 description 7
- 208000024891 symptom Diseases 0.000 description 7
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 6
- 210000004027 cell Anatomy 0.000 description 6
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 6
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 6
- 239000000284 extract Substances 0.000 description 6
- XEEYBQQBJWHFJM-UHFFFAOYSA-N iron Substances [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 6
- 235000018102 proteins Nutrition 0.000 description 6
- 102000004169 proteins and genes Human genes 0.000 description 6
- 108090000623 proteins and genes Proteins 0.000 description 6
- 238000006243 chemical reaction Methods 0.000 description 5
- 230000003750 conditioning effect Effects 0.000 description 5
- 230000002401 inhibitory effect Effects 0.000 description 5
- 230000004060 metabolic process Effects 0.000 description 5
- 108090000765 processed proteins & peptides Proteins 0.000 description 5
- 239000002994 raw material Substances 0.000 description 5
- 230000037303 wrinkles Effects 0.000 description 5
- TXFPEBPIARQUIG-UHFFFAOYSA-N 4'-hydroxyacetophenone Chemical compound CC(=O)C1=CC=C(O)C=C1 TXFPEBPIARQUIG-UHFFFAOYSA-N 0.000 description 4
- 239000002126 C01EB10 - Adenosine Substances 0.000 description 4
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 4
- 102000010911 Enzyme Precursors Human genes 0.000 description 4
- 108010062466 Enzyme Precursors Proteins 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- 206010040830 Skin discomfort Diseases 0.000 description 4
- 229930182558 Sterol Natural products 0.000 description 4
- 230000004913 activation Effects 0.000 description 4
- 229960005305 adenosine Drugs 0.000 description 4
- 239000003963 antioxidant agent Substances 0.000 description 4
- 230000003078 antioxidant effect Effects 0.000 description 4
- 235000006708 antioxidants Nutrition 0.000 description 4
- 230000008901 benefit Effects 0.000 description 4
- 239000007853 buffer solution Substances 0.000 description 4
- 239000010949 copper Substances 0.000 description 4
- 230000036039 immunity Effects 0.000 description 4
- 239000011159 matrix material Substances 0.000 description 4
- GUAQVFRUPZBRJQ-UHFFFAOYSA-N n-(3-aminopropyl)-2-methylprop-2-enamide Chemical compound CC(=C)C(=O)NCCCN GUAQVFRUPZBRJQ-UHFFFAOYSA-N 0.000 description 4
- 235000015097 nutrients Nutrition 0.000 description 4
- 229920001184 polypeptide Polymers 0.000 description 4
- 102000004196 processed proteins & peptides Human genes 0.000 description 4
- 150000003254 radicals Chemical class 0.000 description 4
- 150000003432 sterols Chemical class 0.000 description 4
- 235000003702 sterols Nutrition 0.000 description 4
- 239000011573 trace mineral Substances 0.000 description 4
- 235000013619 trace mineral Nutrition 0.000 description 4
- 239000011701 zinc Substances 0.000 description 4
- 229940015975 1,2-hexanediol Drugs 0.000 description 3
- 239000003109 Disodium ethylene diamine tetraacetate Substances 0.000 description 3
- 206010020751 Hypersensitivity Diseases 0.000 description 3
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 241000700605 Viruses Species 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 3
- 208000026935 allergic disease Diseases 0.000 description 3
- 230000007815 allergy Effects 0.000 description 3
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 3
- CDQSJQSWAWPGKG-UHFFFAOYSA-N butane-1,1-diol Chemical compound CCCC(O)O CDQSJQSWAWPGKG-UHFFFAOYSA-N 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 235000012000 cholesterol Nutrition 0.000 description 3
- 229910052802 copper Inorganic materials 0.000 description 3
- 239000006071 cream Substances 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000018109 developmental process Effects 0.000 description 3
- 235000019301 disodium ethylene diamine tetraacetate Nutrition 0.000 description 3
- 239000000839 emulsion Substances 0.000 description 3
- 230000001815 facial effect Effects 0.000 description 3
- 235000013305 food Nutrition 0.000 description 3
- FHKSXSQHXQEMOK-UHFFFAOYSA-N hexane-1,2-diol Chemical compound CCCCC(O)CO FHKSXSQHXQEMOK-UHFFFAOYSA-N 0.000 description 3
- 238000000338 in vitro Methods 0.000 description 3
- 238000011534 incubation Methods 0.000 description 3
- 229910052742 iron Inorganic materials 0.000 description 3
- 235000010355 mannitol Nutrition 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 239000001301 oxygen Substances 0.000 description 3
- 150000003839 salts Chemical class 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- 230000006641 stabilisation Effects 0.000 description 3
- 238000011105 stabilization Methods 0.000 description 3
- 239000011782 vitamin Substances 0.000 description 3
- 235000013343 vitamin Nutrition 0.000 description 3
- 229930003231 vitamin Natural products 0.000 description 3
- 229940088594 vitamin Drugs 0.000 description 3
- 229910052725 zinc Inorganic materials 0.000 description 3
- WTVHAMTYZJGJLJ-UHFFFAOYSA-N (+)-(4S,8R)-8-epi-beta-bisabolol Natural products CC(C)=CCCC(C)C1(O)CCC(C)=CC1 WTVHAMTYZJGJLJ-UHFFFAOYSA-N 0.000 description 2
- RGZSQWQPBWRIAQ-CABCVRRESA-N (-)-alpha-Bisabolol Chemical compound CC(C)=CCC[C@](C)(O)[C@H]1CCC(C)=CC1 RGZSQWQPBWRIAQ-CABCVRRESA-N 0.000 description 2
- LXAHHHIGZXPRKQ-UHFFFAOYSA-N 5-fluoro-2-methylpyridine Chemical compound CC1=CC=C(F)C=N1 LXAHHHIGZXPRKQ-UHFFFAOYSA-N 0.000 description 2
- NIXOWILDQLNWCW-UHFFFAOYSA-N Acrylic acid Chemical compound OC(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 description 2
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 244000063299 Bacillus subtilis Species 0.000 description 2
- 235000014469 Bacillus subtilis Nutrition 0.000 description 2
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 2
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 2
- SNPLKNRPJHDVJA-ZETCQYMHSA-N D-panthenol Chemical compound OCC(C)(C)[C@@H](O)C(=O)NCCCO SNPLKNRPJHDVJA-ZETCQYMHSA-N 0.000 description 2
- 108010016626 Dipeptides Proteins 0.000 description 2
- 241000196324 Embryophyta Species 0.000 description 2
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 2
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 2
- 230000005526 G1 to G0 transition Effects 0.000 description 2
- BIVBRWYINDPWKA-VLQRKCJKSA-L Glycyrrhizinate dipotassium Chemical compound [K+].[K+].O([C@@H]1[C@@H](O)[C@H](O)[C@H](O[C@@H]1O[C@H]1CC[C@]2(C)[C@H]3C(=O)C=C4[C@@H]5C[C@](C)(CC[C@@]5(CC[C@@]4(C)[C@]3(C)CC[C@H]2C1(C)C)C)C(O)=O)C([O-])=O)[C@@H]1O[C@H](C([O-])=O)[C@@H](O)[C@H](O)[C@H]1O BIVBRWYINDPWKA-VLQRKCJKSA-L 0.000 description 2
- 239000004354 Hydroxyethyl cellulose Substances 0.000 description 2
- 229920000663 Hydroxyethyl cellulose Polymers 0.000 description 2
- ARIWANIATODDMH-UHFFFAOYSA-N Lauric acid monoglyceride Natural products CCCCCCCCCCCC(=O)OCC(O)CO ARIWANIATODDMH-UHFFFAOYSA-N 0.000 description 2
- 102000002274 Matrix Metalloproteinases Human genes 0.000 description 2
- 108010000684 Matrix Metalloproteinases Proteins 0.000 description 2
- 206010028980 Neoplasm Diseases 0.000 description 2
- PXHVJJICTQNCMI-UHFFFAOYSA-N Nickel Chemical compound [Ni] PXHVJJICTQNCMI-UHFFFAOYSA-N 0.000 description 2
- DFPAKSUCGFBDDF-UHFFFAOYSA-N Nicotinamide Chemical compound NC(=O)C1=CC=CN=C1 DFPAKSUCGFBDDF-UHFFFAOYSA-N 0.000 description 2
- 208000002193 Pain Diseases 0.000 description 2
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 2
- 239000004698 Polyethylene Substances 0.000 description 2
- 229920002125 Sokalan® Polymers 0.000 description 2
- 229930006000 Sucrose Natural products 0.000 description 2
- 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 2
- GSEJCLTVZPLZKY-UHFFFAOYSA-N Triethanolamine Chemical compound OCCN(CCO)CCO GSEJCLTVZPLZKY-UHFFFAOYSA-N 0.000 description 2
- ISAKRJDGNUQOIC-UHFFFAOYSA-N Uracil Chemical compound O=C1C=CNC(=O)N1 ISAKRJDGNUQOIC-UHFFFAOYSA-N 0.000 description 2
- DRTQHJPVMGBUCF-XVFCMESISA-N Uridine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-XVFCMESISA-N 0.000 description 2
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 2
- CKUAXEQHGKSLHN-UHFFFAOYSA-N [C].[N] Chemical compound [C].[N] CKUAXEQHGKSLHN-UHFFFAOYSA-N 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000013543 active substance Substances 0.000 description 2
- RGZSQWQPBWRIAQ-LSDHHAIUSA-N alpha-Bisabolol Natural products CC(C)=CCC[C@@](C)(O)[C@@H]1CCC(C)=CC1 RGZSQWQPBWRIAQ-LSDHHAIUSA-N 0.000 description 2
- 230000001093 anti-cancer Effects 0.000 description 2
- 230000000975 bioactive effect Effects 0.000 description 2
- 230000004071 biological effect Effects 0.000 description 2
- 229940036350 bisabolol Drugs 0.000 description 2
- HHGZABIIYIWLGA-UHFFFAOYSA-N bisabolol Natural products CC1CCC(C(C)(O)CCC=C(C)C)CC1 HHGZABIIYIWLGA-UHFFFAOYSA-N 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 229960001631 carbomer Drugs 0.000 description 2
- 238000004140 cleaning Methods 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 125000004122 cyclic group Chemical group 0.000 description 2
- GVJHHUAWPYXKBD-UHFFFAOYSA-N d-alpha-tocopherol Natural products OC1=C(C)C(C)=C2OC(CCCC(C)CCCC(C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-UHFFFAOYSA-N 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 238000007865 diluting Methods 0.000 description 2
- 229940101029 dipotassium glycyrrhizinate Drugs 0.000 description 2
- POULHZVOKOAJMA-UHFFFAOYSA-N dodecanoic acid Chemical compound CCCCCCCCCCCC(O)=O POULHZVOKOAJMA-UHFFFAOYSA-N 0.000 description 2
- 238000006911 enzymatic reaction Methods 0.000 description 2
- 239000003797 essential amino acid Substances 0.000 description 2
- 235000020776 essential amino acid Nutrition 0.000 description 2
- 230000005284 excitation Effects 0.000 description 2
- 210000002744 extracellular matrix Anatomy 0.000 description 2
- 230000002349 favourable effect Effects 0.000 description 2
- 239000007850 fluorescent dye Substances 0.000 description 2
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 2
- 239000000499 gel Substances 0.000 description 2
- 239000008273 gelatin Substances 0.000 description 2
- 229920000159 gelatin Polymers 0.000 description 2
- 125000003976 glyceryl group Chemical group [H]C([*])([H])C(O[H])([H])C(O[H])([H])[H] 0.000 description 2
- UYTPUPDQBNUYGX-UHFFFAOYSA-N guanine Chemical compound O=C1NC(N)=NC2=C1N=CN2 UYTPUPDQBNUYGX-UHFFFAOYSA-N 0.000 description 2
- IPCSVZSSVZVIGE-UHFFFAOYSA-N hexadecanoic acid Chemical compound CCCCCCCCCCCCCCCC(O)=O IPCSVZSSVZVIGE-UHFFFAOYSA-N 0.000 description 2
- 235000019447 hydroxyethyl cellulose Nutrition 0.000 description 2
- 238000011031 large-scale manufacturing process Methods 0.000 description 2
- 239000006210 lotion Substances 0.000 description 2
- 239000011777 magnesium Substances 0.000 description 2
- 229910052749 magnesium Inorganic materials 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000002207 metabolite Substances 0.000 description 2
- UPSFMJHZUCSEHU-JYGUBCOQSA-N n-[(2s,3r,4r,5s,6r)-2-[(2r,3s,4r,5r,6s)-5-acetamido-4-hydroxy-2-(hydroxymethyl)-6-(4-methyl-2-oxochromen-7-yl)oxyoxan-3-yl]oxy-4,5-dihydroxy-6-(hydroxymethyl)oxan-3-yl]acetamide Chemical compound CC(=O)N[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@H]1[C@H](O)[C@@H](NC(C)=O)[C@H](OC=2C=C3OC(=O)C=C(C)C3=CC=2)O[C@@H]1CO UPSFMJHZUCSEHU-JYGUBCOQSA-N 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 229940101267 panthenol Drugs 0.000 description 2
- 235000020957 pantothenol Nutrition 0.000 description 2
- 239000011619 pantothenol Substances 0.000 description 2
- 230000035699 permeability Effects 0.000 description 2
- 229910052698 phosphorus Inorganic materials 0.000 description 2
- 239000011574 phosphorus Substances 0.000 description 2
- 229920000573 polyethylene Polymers 0.000 description 2
- 229910052700 potassium Inorganic materials 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 230000001105 regulatory effect Effects 0.000 description 2
- YGSDEFSMJLZEOE-UHFFFAOYSA-N salicylic acid Chemical compound OC(=O)C1=CC=CC=C1O YGSDEFSMJLZEOE-UHFFFAOYSA-N 0.000 description 2
- 230000009759 skin aging Effects 0.000 description 2
- 229910052708 sodium Inorganic materials 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- 239000000600 sorbitol Substances 0.000 description 2
- PRAKJMSDJKAYCZ-UHFFFAOYSA-N squalane Chemical compound CC(C)CCCC(C)CCCC(C)CCCCC(C)CCCC(C)CCCC(C)C PRAKJMSDJKAYCZ-UHFFFAOYSA-N 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- 239000005720 sucrose Substances 0.000 description 2
- 230000008961 swelling Effects 0.000 description 2
- 239000003826 tablet Substances 0.000 description 2
- RWQNBRDOKXIBIV-UHFFFAOYSA-N thymine Chemical compound CC1=CNC(=O)NC1=O RWQNBRDOKXIBIV-UHFFFAOYSA-N 0.000 description 2
- 229960001295 tocopherol Drugs 0.000 description 2
- 235000010384 tocopherol Nutrition 0.000 description 2
- 229930003799 tocopherol Natural products 0.000 description 2
- 239000011732 tocopherol Substances 0.000 description 2
- GVJHHUAWPYXKBD-IEOSBIPESA-N α-tocopherol Chemical compound OC1=C(C)C(C)=C2O[C@@](CCC[C@H](C)CCC[C@H](C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-IEOSBIPESA-N 0.000 description 2
- OILXMJHPFNGGTO-UHFFFAOYSA-N (22E)-(24xi)-24-methylcholesta-5,22-dien-3beta-ol Natural products C1C=C2CC(O)CCC2(C)C2C1C1CCC(C(C)C=CC(C)C(C)C)C1(C)CC2 OILXMJHPFNGGTO-UHFFFAOYSA-N 0.000 description 1
- RQOCXCFLRBRBCS-UHFFFAOYSA-N (22E)-cholesta-5,7,22-trien-3beta-ol Natural products C1C(O)CCC2(C)C(CCC3(C(C(C)C=CCC(C)C)CCC33)C)C3=CC=C21 RQOCXCFLRBRBCS-UHFFFAOYSA-N 0.000 description 1
- FDKWRPBBCBCIGA-REOHCLBHSA-N (2r)-2-azaniumyl-3-$l^{1}-selanylpropanoate Chemical compound [Se]C[C@H](N)C(O)=O FDKWRPBBCBCIGA-REOHCLBHSA-N 0.000 description 1
- IEQAICDLOKRSRL-UHFFFAOYSA-N 2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-(2-dodecoxyethoxy)ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethanol Chemical compound CCCCCCCCCCCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCOCCO IEQAICDLOKRSRL-UHFFFAOYSA-N 0.000 description 1
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 1
- MSWZFWKMSRAUBD-IVMDWMLBSA-N 2-amino-2-deoxy-D-glucopyranose Chemical compound N[C@H]1C(O)O[C@H](CO)[C@@H](O)[C@@H]1O MSWZFWKMSRAUBD-IVMDWMLBSA-N 0.000 description 1
- UIVPNOBLHXUKDX-UHFFFAOYSA-N 3,5,5-trimethylhexyl 3,5,5-trimethylhexanoate Chemical compound CC(C)(C)CC(C)CCOC(=O)CC(C)CC(C)(C)C UIVPNOBLHXUKDX-UHFFFAOYSA-N 0.000 description 1
- ZGSCRDSBTNQPMS-UJURSFKZSA-N 3-O-Ethylascorbic acid Chemical compound CCOC1=C(O)C(=O)O[C@@H]1[C@@H](O)CO ZGSCRDSBTNQPMS-UJURSFKZSA-N 0.000 description 1
- VFKZECOCJCGZQK-UHFFFAOYSA-M 3-hydroxypropyl(trimethyl)azanium;chloride Chemical compound [Cl-].C[N+](C)(C)CCCO VFKZECOCJCGZQK-UHFFFAOYSA-M 0.000 description 1
- 229940120145 3-o-ethylascorbic acid Drugs 0.000 description 1
- OQMZNAMGEHIHNN-UHFFFAOYSA-N 7-Dehydrostigmasterol Natural products C1C(O)CCC2(C)C(CCC3(C(C(C)C=CC(CC)C(C)C)CCC33)C)C3=CC=C21 OQMZNAMGEHIHNN-UHFFFAOYSA-N 0.000 description 1
- 229930024421 Adenine Natural products 0.000 description 1
- GFFGJBXGBJISGV-UHFFFAOYSA-N Adenine Chemical compound NC1=NC=NC2=C1N=CN2 GFFGJBXGBJISGV-UHFFFAOYSA-N 0.000 description 1
- 241000045403 Astragalus propinquus Species 0.000 description 1
- 241000193830 Bacillus <bacterium> Species 0.000 description 1
- 241000193738 Bacillus anthracis Species 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 1
- 206010008111 Cerebral haemorrhage Diseases 0.000 description 1
- 206010008132 Cerebral thrombosis Diseases 0.000 description 1
- VYZAMTAEIAYCRO-UHFFFAOYSA-N Chromium Chemical compound [Cr] VYZAMTAEIAYCRO-UHFFFAOYSA-N 0.000 description 1
- 235000005976 Citrus sinensis Nutrition 0.000 description 1
- 240000002319 Citrus sinensis Species 0.000 description 1
- 241000193403 Clostridium Species 0.000 description 1
- 235000019750 Crude protein Nutrition 0.000 description 1
- 244000303965 Cyamopsis psoralioides Species 0.000 description 1
- FDKWRPBBCBCIGA-UWTATZPHSA-N D-Selenocysteine Natural products [Se]C[C@@H](N)C(O)=O FDKWRPBBCBCIGA-UWTATZPHSA-N 0.000 description 1
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 1
- 239000004287 Dehydroacetic acid Substances 0.000 description 1
- 201000004624 Dermatitis Diseases 0.000 description 1
- DNVPQKQSNYMLRS-NXVQYWJNSA-N Ergosterol Natural products CC(C)[C@@H](C)C=C[C@H](C)[C@H]1CC[C@H]2C3=CC=C4C[C@@H](O)CC[C@]4(C)[C@@H]3CC[C@]12C DNVPQKQSNYMLRS-NXVQYWJNSA-N 0.000 description 1
- 206010015150 Erythema Diseases 0.000 description 1
- 102000006587 Glutathione peroxidase Human genes 0.000 description 1
- 108700016172 Glutathione peroxidases Proteins 0.000 description 1
- 239000007995 HEPES buffer Substances 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 108060003951 Immunoglobulin Proteins 0.000 description 1
- 238000012404 In vitro experiment Methods 0.000 description 1
- 206010062717 Increased upper airway secretion Diseases 0.000 description 1
- 229930010555 Inosine Natural products 0.000 description 1
- UGQMRVRMYYASKQ-KQYNXXCUSA-N Inosine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C2=NC=NC(O)=C2N=C1 UGQMRVRMYYASKQ-KQYNXXCUSA-N 0.000 description 1
- 201000001429 Intracranial Thrombosis Diseases 0.000 description 1
- 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 1
- 239000005639 Lauric acid Substances 0.000 description 1
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 description 1
- 239000005913 Maltodextrin Substances 0.000 description 1
- 229920002774 Maltodextrin Polymers 0.000 description 1
- 235000006679 Mentha X verticillata Nutrition 0.000 description 1
- 235000002899 Mentha suaveolens Nutrition 0.000 description 1
- 235000001636 Mentha x rotundifolia Nutrition 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 241000186367 Mycobacterium avium Species 0.000 description 1
- 241000186366 Mycobacterium bovis Species 0.000 description 1
- 208000012902 Nervous system disease Diseases 0.000 description 1
- 235000021314 Palmitic acid Nutrition 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 229920000289 Polyquaternium Polymers 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 241000382353 Pupa Species 0.000 description 1
- 230000006819 RNA synthesis Effects 0.000 description 1
- 230000002292 Radical scavenging effect Effects 0.000 description 1
- 241000282806 Rhinoceros Species 0.000 description 1
- 206010039897 Sedation Diseases 0.000 description 1
- XUIMIQQOPSSXEZ-UHFFFAOYSA-N Silicon Chemical compound [Si] XUIMIQQOPSSXEZ-UHFFFAOYSA-N 0.000 description 1
- UIIMBOGNXHQVGW-DEQYMQKBSA-M Sodium bicarbonate-14C Chemical compound [Na+].O[14C]([O-])=O UIIMBOGNXHQVGW-DEQYMQKBSA-M 0.000 description 1
- 235000021355 Stearic acid Nutrition 0.000 description 1
- 241000194017 Streptococcus Species 0.000 description 1
- 206010042496 Sunburn Diseases 0.000 description 1
- OUUQCZGPVNCOIJ-UHFFFAOYSA-M Superoxide Chemical compound [O-][O] OUUQCZGPVNCOIJ-UHFFFAOYSA-M 0.000 description 1
- 206010042674 Swelling Diseases 0.000 description 1
- RTAQQCXQSZGOHL-UHFFFAOYSA-N Titanium Chemical compound [Ti] RTAQQCXQSZGOHL-UHFFFAOYSA-N 0.000 description 1
- 241000223238 Trichophyton Species 0.000 description 1
- 240000000851 Vaccinium corymbosum Species 0.000 description 1
- 235000003095 Vaccinium corymbosum Nutrition 0.000 description 1
- 235000017537 Vaccinium myrtillus Nutrition 0.000 description 1
- 229930003268 Vitamin C Natural products 0.000 description 1
- QYSXJUFSXHHAJI-XFEUOLMDSA-N Vitamin D3 Natural products C1(/[C@@H]2CC[C@@H]([C@]2(CCC1)C)[C@H](C)CCCC(C)C)=C/C=C1\C[C@@H](O)CCC1=C QYSXJUFSXHHAJI-XFEUOLMDSA-N 0.000 description 1
- 241000212749 Zesius chrysomallus Species 0.000 description 1
- DFPAKSUCGFBDDF-ZQBYOMGUSA-N [14c]-nicotinamide Chemical compound N[14C](=O)C1=CC=CN=C1 DFPAKSUCGFBDDF-ZQBYOMGUSA-N 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 239000012190 activator Substances 0.000 description 1
- 229960000643 adenine Drugs 0.000 description 1
- 239000002671 adjuvant Substances 0.000 description 1
- 229910052782 aluminium Inorganic materials 0.000 description 1
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 1
- 239000012491 analyte Substances 0.000 description 1
- 230000000843 anti-fungal effect Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 230000010100 anticoagulation Effects 0.000 description 1
- 229940121375 antifungal agent Drugs 0.000 description 1
- 239000006286 aqueous extract Substances 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 208000006673 asthma Diseases 0.000 description 1
- 235000006533 astragalus Nutrition 0.000 description 1
- 229940065181 bacillus anthracis Drugs 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 229940076810 beta sitosterol Drugs 0.000 description 1
- DRTQHJPVMGBUCF-PSQAKQOGSA-N beta-L-uridine Natural products O[C@H]1[C@@H](O)[C@H](CO)O[C@@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-PSQAKQOGSA-N 0.000 description 1
- LGJMUZUPVCAVPU-UHFFFAOYSA-N beta-Sitostanol Natural products C1CC2CC(O)CCC2(C)C2C1C1CCC(C(C)CCC(CC)C(C)C)C1(C)CC2 LGJMUZUPVCAVPU-UHFFFAOYSA-N 0.000 description 1
- NJKOMDUNNDKEAI-UHFFFAOYSA-N beta-sitosterol Natural products CCC(CCC(C)C1CCC2(C)C3CC=C4CC(O)CCC4C3CCC12C)C(C)C NJKOMDUNNDKEAI-UHFFFAOYSA-N 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 235000021014 blueberries Nutrition 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 235000011148 calcium chloride Nutrition 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 230000005907 cancer growth Effects 0.000 description 1
- 239000001768 carboxy methyl cellulose Substances 0.000 description 1
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 1
- 239000004359 castor oil Substances 0.000 description 1
- 235000019438 castor oil Nutrition 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 229910052804 chromium Inorganic materials 0.000 description 1
- 239000011651 chromium Substances 0.000 description 1
- 230000015271 coagulation Effects 0.000 description 1
- 238000005345 coagulation Methods 0.000 description 1
- 230000001595 contractor effect Effects 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- SASYSVUEVMOWPL-NXVVXOECSA-N decyl oleate Chemical compound CCCCCCCCCCOC(=O)CCCCCCC\C=C/CCCCCCCC SASYSVUEVMOWPL-NXVVXOECSA-N 0.000 description 1
- 230000007123 defense Effects 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 235000019258 dehydroacetic acid Nutrition 0.000 description 1
- 229940061632 dehydroacetic acid Drugs 0.000 description 1
- JEQRBTDTEKWZBW-UHFFFAOYSA-N dehydroacetic acid Chemical compound CC(=O)C1=C(O)OC(C)=CC1=O JEQRBTDTEKWZBW-UHFFFAOYSA-N 0.000 description 1
- PGRHXDWITVMQBC-UHFFFAOYSA-N dehydroacetic acid Natural products CC(=O)C1C(=O)OC(C)=CC1=O PGRHXDWITVMQBC-UHFFFAOYSA-N 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 201000001981 dermatomyositis Diseases 0.000 description 1
- 208000010643 digestive system disease Diseases 0.000 description 1
- 239000004205 dimethyl polysiloxane Substances 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 230000000857 drug effect Effects 0.000 description 1
- 229960001484 edetic acid Drugs 0.000 description 1
- 230000009982 effect on human Effects 0.000 description 1
- 239000007938 effervescent tablet Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- DNVPQKQSNYMLRS-SOWFXMKYSA-N ergosterol Chemical compound C1[C@@H](O)CC[C@]2(C)[C@H](CC[C@]3([C@H]([C@H](C)/C=C/[C@@H](C)C(C)C)CC[C@H]33)C)C3=CC=C21 DNVPQKQSNYMLRS-SOWFXMKYSA-N 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000004088 foaming agent Substances 0.000 description 1
- 235000003599 food sweetener Nutrition 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 125000005456 glyceride group Chemical group 0.000 description 1
- ZEMPKEQAKRGZGQ-XOQCFJPHSA-N glycerol triricinoleate Natural products CCCCCC[C@@H](O)CC=CCCCCCCCC(=O)OC[C@@H](COC(=O)CCCCCCCC=CC[C@@H](O)CCCCCC)OC(=O)CCCCCCCC=CC[C@H](O)CCCCCC ZEMPKEQAKRGZGQ-XOQCFJPHSA-N 0.000 description 1
- 235000019674 grape juice Nutrition 0.000 description 1
- 235000013402 health food Nutrition 0.000 description 1
- 230000036074 healthy skin Effects 0.000 description 1
- 230000023597 hemostasis Effects 0.000 description 1
- 241000411851 herbal medicine Species 0.000 description 1
- 235000008216 herbs Nutrition 0.000 description 1
- 230000000147 hypnotic effect Effects 0.000 description 1
- 210000002865 immune cell Anatomy 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 102000018358 immunoglobulin Human genes 0.000 description 1
- 230000007365 immunoregulation Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 229960003786 inosine Drugs 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 238000007917 intracranial administration Methods 0.000 description 1
- 230000009545 invasion Effects 0.000 description 1
- 229940100554 isononyl isononanoate Drugs 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- PYIDGJJWBIBVIA-UYTYNIKBSA-N lauryl glucoside Chemical compound CCCCCCCCCCCCO[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O PYIDGJJWBIBVIA-UYTYNIKBSA-N 0.000 description 1
- 229940048848 lauryl glucoside Drugs 0.000 description 1
- AGBQKNBQESQNJD-UHFFFAOYSA-M lipoate Chemical compound [O-]C(=O)CCCCC1CCSS1 AGBQKNBQESQNJD-UHFFFAOYSA-M 0.000 description 1
- 235000019136 lipoic acid Nutrition 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 210000002540 macrophage Anatomy 0.000 description 1
- 229940035034 maltodextrin Drugs 0.000 description 1
- 229910052748 manganese Inorganic materials 0.000 description 1
- 239000011572 manganese Substances 0.000 description 1
- WPBNNNQJVZRUHP-UHFFFAOYSA-L manganese(2+);methyl n-[[2-(methoxycarbonylcarbamothioylamino)phenyl]carbamothioyl]carbamate;n-[2-(sulfidocarbothioylamino)ethyl]carbamodithioate Chemical compound [Mn+2].[S-]C(=S)NCCNC([S-])=S.COC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OC WPBNNNQJVZRUHP-UHFFFAOYSA-L 0.000 description 1
- 239000012092 media component Substances 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 229940044591 methyl glucose dioleate Drugs 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- LXCFILQKKLGQFO-UHFFFAOYSA-N methylparaben Chemical compound COC(=O)C1=CC=C(O)C=C1 LXCFILQKKLGQFO-UHFFFAOYSA-N 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 210000000865 mononuclear phagocyte system Anatomy 0.000 description 1
- WQEPLUUGTLDZJY-UHFFFAOYSA-N n-Pentadecanoic acid Natural products CCCCCCCCCCCCCCC(O)=O WQEPLUUGTLDZJY-UHFFFAOYSA-N 0.000 description 1
- JXTPJDDICSTXJX-UHFFFAOYSA-N n-Triacontane Natural products CCCCCCCCCCCCCCCCCCCCCCCCCCCCCC JXTPJDDICSTXJX-UHFFFAOYSA-N 0.000 description 1
- 210000000653 nervous system Anatomy 0.000 description 1
- 229910052759 nickel Inorganic materials 0.000 description 1
- 229960003966 nicotinamide Drugs 0.000 description 1
- 235000005152 nicotinamide Nutrition 0.000 description 1
- 239000011570 nicotinamide Substances 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
- WWZKQHOCKIZLMA-UHFFFAOYSA-N octanoic acid Chemical compound CCCCCCCC(O)=O WWZKQHOCKIZLMA-UHFFFAOYSA-N 0.000 description 1
- 125000002801 octanoyl group Chemical group C(CCCCCCC)(=O)* 0.000 description 1
- 229940049964 oleate Drugs 0.000 description 1
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 description 1
- 239000004006 olive oil Substances 0.000 description 1
- 235000008390 olive oil Nutrition 0.000 description 1
- 230000014207 opsonization Effects 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- 230000004792 oxidative damage Effects 0.000 description 1
- FJKROLUGYXJWQN-UHFFFAOYSA-N papa-hydroxy-benzoic acid Natural products OC(=O)C1=CC=C(O)C=C1 FJKROLUGYXJWQN-UHFFFAOYSA-N 0.000 description 1
- UWJJYHHHVWZFEP-UHFFFAOYSA-N pentane-1,1-diol Chemical compound CCCCC(O)O UWJJYHHHVWZFEP-UHFFFAOYSA-N 0.000 description 1
- 238000011056 performance test Methods 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- WVDDGKGOMKODPV-ZQBYOMGUSA-N phenyl(114C)methanol Chemical compound O[14CH2]C1=CC=CC=C1 WVDDGKGOMKODPV-ZQBYOMGUSA-N 0.000 description 1
- 208000026435 phlegm Diseases 0.000 description 1
- 229920000435 poly(dimethylsiloxane) Polymers 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 235000015277 pork Nutrition 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 235000012015 potatoes Nutrition 0.000 description 1
- 239000013630 prepared media Substances 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 239000012744 reinforcing agent Substances 0.000 description 1
- 230000000284 resting effect Effects 0.000 description 1
- 229960004889 salicylic acid Drugs 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 108700004121 sarkosyl Proteins 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000036280 sedation Effects 0.000 description 1
- 229940055619 selenocysteine Drugs 0.000 description 1
- ZKZBPNGNEQAJSX-UHFFFAOYSA-N selenocysteine Natural products [SeH]CC(N)C(O)=O ZKZBPNGNEQAJSX-UHFFFAOYSA-N 0.000 description 1
- 235000016491 selenocysteine Nutrition 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 229910052710 silicon Inorganic materials 0.000 description 1
- 239000010703 silicon Substances 0.000 description 1
- KZJWDPNRJALLNS-VJSFXXLFSA-N sitosterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CC[C@@H](CC)C(C)C)[C@@]1(C)CC2 KZJWDPNRJALLNS-VJSFXXLFSA-N 0.000 description 1
- 229950005143 sitosterol Drugs 0.000 description 1
- 230000005808 skin problem Effects 0.000 description 1
- WXMKPNITSTVMEF-UHFFFAOYSA-M sodium benzoate Chemical compound [Na+].[O-]C(=O)C1=CC=CC=C1 WXMKPNITSTVMEF-UHFFFAOYSA-M 0.000 description 1
- 235000010234 sodium benzoate Nutrition 0.000 description 1
- 239000004299 sodium benzoate Substances 0.000 description 1
- 239000001509 sodium citrate Substances 0.000 description 1
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 1
- 229940079776 sodium cocoyl isethionate Drugs 0.000 description 1
- KSAVQLQVUXSOCR-UHFFFAOYSA-M sodium lauroyl sarcosinate Chemical compound [Na+].CCCCCCCCCCCC(=O)N(C)CC([O-])=O KSAVQLQVUXSOCR-UHFFFAOYSA-M 0.000 description 1
- 229940045885 sodium lauroyl sarcosinate Drugs 0.000 description 1
- 229940048109 sodium methyl cocoyl taurate Drugs 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 229940032094 squalane Drugs 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 239000008117 stearic acid Substances 0.000 description 1
- 150000003431 steroids Chemical class 0.000 description 1
- 229910052712 strontium Inorganic materials 0.000 description 1
- CIOAGBVUUVVLOB-UHFFFAOYSA-N strontium atom Chemical compound [Sr] CIOAGBVUUVVLOB-UHFFFAOYSA-N 0.000 description 1
- 150000005846 sugar alcohols Polymers 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000012353 t test Methods 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
- 229960002663 thioctic acid Drugs 0.000 description 1
- 229940113082 thymine Drugs 0.000 description 1
- 229910052719 titanium Inorganic materials 0.000 description 1
- 239000010936 titanium Substances 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000003053 toxin Substances 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 229940035893 uracil Drugs 0.000 description 1
- DRTQHJPVMGBUCF-UHFFFAOYSA-N uracil arabinoside Natural products OC1C(O)C(CO)OC1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-UHFFFAOYSA-N 0.000 description 1
- 229940045145 uridine Drugs 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 229960004295 valine Drugs 0.000 description 1
- 235000019154 vitamin C Nutrition 0.000 description 1
- 239000011718 vitamin C Substances 0.000 description 1
- QYSXJUFSXHHAJI-YRZJJWOYSA-N vitamin D3 Chemical compound C1(/[C@@H]2CC[C@@H]([C@]2(CCC1)C)[C@H](C)CCCC(C)C)=C\C=C1\C[C@@H](O)CCC1=C QYSXJUFSXHHAJI-YRZJJWOYSA-N 0.000 description 1
- 235000005282 vitamin D3 Nutrition 0.000 description 1
- 239000011647 vitamin D3 Substances 0.000 description 1
- 229940021056 vitamin d3 Drugs 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 210000004885 white matter Anatomy 0.000 description 1
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 1
- 229910000368 zinc sulfate Inorganic materials 0.000 description 1
- 239000011686 zinc sulphate Substances 0.000 description 1
- 235000009529 zinc sulphate Nutrition 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
- A61K8/66—Enzymes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/02—Preparations for care of the skin for chemically bleaching or whitening the skin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Zoology (AREA)
- Animal Behavior & Ethology (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Organic Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- Dermatology (AREA)
- General Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Microbiology (AREA)
- Medicinal Chemistry (AREA)
- Biochemistry (AREA)
- Birds (AREA)
- Gerontology & Geriatric Medicine (AREA)
- Molecular Biology (AREA)
- Epidemiology (AREA)
- Botany (AREA)
- Mycology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Virology (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Cosmetics (AREA)
Abstract
本发明提供活性虫草酵素制备的优化方法,包括:1)培养基的选择:液体培养基成分的组成如下:a、碳源100‑200重量份,b、氮源100‑200重量份,c、无机盐包括K2HPO4,MgSO4.7H2O,K2HPO4为0.4‑1.0重量份,MgSO4.7H2O为0.1‑0.7重量份,d、其他包括VB1,透明质酸和去离子水,VB1为0.02‑0.05重量份,透明质酸2.0‑5.0重量份,去离子水补到1000重量份;2)虫草菌种的移植和保藏,移植时间为3个月;3)虫草菌种的接种,接种量为3cm2;4)虫草菌种的摇床上培养,选取培养72h的处于对数生长期的虫草菌种进行扩大培养;5)虫草菌种的扩大发酵,培养10h后通气,通气量为1.8(v/v·min)。本方法制备的虫草酵素可以以重量比0.01‑10.0%添加到任意化妆品配方中使用,其中的主要营养及有效成份具有淡斑抗氧化、抗衰老、抑菌消炎等功效。
Description
技术领域
本发明本发明涉及一种生物酵素,尤其是一种虫草酵素制备的优化方法。
背景技术
酵素是一种由氨基酸组成的具有特殊生物活性的物质,几乎参与所有的生命活动,生物若无酵素将无法生存。目前为止人体内已发现酵素多达1000种以上,酵素又称为具有生物活性的酶,它是健康的源泉,也是生命的源泉,酵素具有排毒解毒,提高免疫的功能,它是生物体内各种反应的媒介,酵素广泛地存在于动植物食物中但加热到48℃以上时这些酵素就会被破坏,所以生产制备酵素的过程中要尤为注意。科学研究表明酵素进入人体30分钟后,就开始发生作用,分解老化细胞,制造新生细胞;同时酵素还可以分解血液中的废物,使血液保持弱碱性,促进血液循环;人体中的氨基酸会产生一些有毒有害的物质,而体内的酵素能将这些有害物质转变成低毒性的尿液排出体外;酵素还有防感染的防御能力,可以有效地抵抗异物侵袭,达到防皱美容和永葆青春魅力的作用。
北虫草是北冬虫夏草的简称,也叫蛹虫草或蛹草,俗名不老草。是虫、菌结合的药用真菌,现代珍稀中草药,北冬虫夏草与产自青藏高原的中华冬虫夏草同属于麦角菌科真菌类植物,主要生长在我国的北方地区,其生长需要高寒环境,要求温差大,这样高寒环境生长的北虫草营养价值最高。其主要的化学成分药理、药效与冬虫夏草极为相似,经检测表明北虫草中的一些主要营养及药物成份如虫草素、虫草多糖、虫草酸、核苷类物质、虫草SOD、蛋白质等还要高于冬虫夏草。现代医学研究表明,北虫草不仅含有丰富的蛋白质(含量为39.37%,是猪肉白质含量的1.8倍)和氨基酸,而且含有30多种人体所需的微量元素,其中P(磷)的含量是冬虫夏草的3.5倍,Zn(锌)、Cu(铜)、Fe(铁三种元素的含量是28种补益药平均值的1.8、2.1和8.8倍,Se(硒)的含量与黄芪的含量相当。所以,北虫草具有催眠镇静、益肝肾、补虚损、防癌、抗癌、止血、化痰、平喘等多种功效,与人参、鹿茸并称为中药宝库中的三大补药。我国现有的北虫草一般通过固态发酵法或液态发酵法得到,一般的液态发酵虫草制备得到的虫草酵素中的活性成分含量偏低,而且得到的虫草酵素主要以食用或药用为主,在化妆品领域却很少有涉及。
随着生物技术的融入和发酵技术的迅猛发展,以及市场的需求,我国的酵素行业也逐渐更新换代。目前市场上的酵素产品多种多样已经涉及到食品、保健食品、化妆品等多个领域。本发明采用优化液体深层发酵的方法制备虫草酵素,得到的虫草酵素除了活性成分含量普遍提高外,也具有一般酵素的基本作用,在融合了北虫草的主要营养及药物成份后,还具有淡斑抗氧化、抗衰老、抑菌消炎等活性功效,在化妆品领域具有光明的发展前景。
发明内容
为解决现有虫草酵素制备和应用的技术问题,为得到一种活性成分更多、效果更好、应用更广泛的虫草酵素。
本发明提供的第一种技术方案是这样实现的:
一种虫草酵素制备的优化方法,其特征在于包括如下步骤:
1)培养基的选择目前,北虫草是虫草属中唯一接种成功和可以人工栽培进行大规模生产的菌种。其培养基的配方多种多样,选择适于虫草菌种发酵产生虫草酵素活性成分的液体培养基的组成和配比至关重要。本发明使用的液体培养基有五种配方,具体如下,
1号培养基(1000g):葡萄糖10.0-20.0g,蛋白胨5.0-15.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;
2号培养基(1000g):葡萄糖10.0-20.0g,胡萝卜100.0-200.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;
3号培养基(1000g):葡萄糖10.0-20.0g,鲜蚕蛹30.0-50.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;
4号培养基(1000g):马铃薯100.0-200.0g,蛋白胨5.0-15.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;5号培养基(1000g):葡萄糖10.0-20.0g,蛋白胨5.0-15.0g,透明质酸2.0-5.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g。
经试验结果表明5号液体培养基为最佳培养基。
2)虫草菌种的移植和保藏
移植培养的操作过程,将加了凝固剂的液体培养基pH值调到6.0左右,灭菌后倾斜放入无菌操作台得到斜面固体培养基,冷却后接种虫草菌种,接种后的试管放入恒温培养箱中,在25℃下培养7d;保藏,菌种放在4℃-6℃冰箱内避光保存,移植期为2-3个月;移植培养,将虫草菌种转接到新配制的斜面固体培养基中,重复上述操作;
3)虫草菌种的接种
按照步骤1)中的液体培养基成分配制1000g液体培养基,装于带侧臂管的三角烧瓶内,灭菌冷却到室温后从斜面固体培养基中取出面积为1-3cm2的块状虫草菌苔,接入液体培养基中;
4)虫草菌种的摇床培养
将已接入液体培养基中的虫草菌种,在24-26℃的恒温震荡培养箱中以转速为120-150r/min避光培养68-72h,期间pH控制在5.0-7.0;
5)虫草菌种的扩大发酵
将配制好的液体培养基倒入发酵罐,在位灭菌25-35min,降温后,迅速倒入摇床培养得到虫草菌种培养液开始扩大发酵;设置发酵温度24℃~26℃,转速120-150r/min,培养8-12h后通气,通气量1.5-1.8(v/v·min),避光培养1~2天,期间pH控制在5.0-7.0;
所述优选摇床培养72h左右的处于对数生长期的虫草菌种进行扩大培养,所述虫草菌种扩大深层液体培养10h后通气,通气量为1.8(v/v·min)
6)虫草酵素活性成分检测
将上述发酵后发酵液置于离心机中,以3000-4500r/min的转速离心15-30min,收集发酵上清液,对发酵上清液用孔径为0.22μm中空纤维微滤系统进行微滤,收集发酵滤过液得到的便是活性虫草酵素,备用;取上述发酵滤过液,分别用基质固相分散提取法测定虫草素、分级沉淀法测定虫草SOD含量、苯酚-硫酸法测定虫草多糖含量、乙醇-微波协同提取法测定虫草酸含量及基质固相分散提取法测定核苷类物质含量。所述选取孔径为0.22μm中空纤维微滤系统进行微滤。
本发明的制备方法优化的具体体现:
①菌种保藏中移植培养的优化,3个月之后菌种在斜面固体培养基上会长满而堆积,严重影响菌种活性,此时直接接种会影响菌种的培养;
②液体培养基的优化选择,选用本方法提供的最优培养基培养得到的虫草酵素,其活性成分中虫草素、虫草SOD、虫草多糖和核苷类物质的含量增加非常显著;
③接种量的控制,接种量的多少会直接影响后期菌种的培养和发酵,接种太多会大量消耗培养基营养成分,可能造成菌种生长还未达到理想时期就被迫停止,而接种太少会影响产量;
④虫草菌种的二次扩大深层液体培养及通气量的控制,取摇床培养到对数生长期后期的虫草菌种进行扩大发酵,可以延长此时菌种的对数生长期,增加产物产量,同时在深层发酵过程中适时适当的通气也可以提高产物的产量;
⑤微孔过滤和活性检测方法的选择,对发酵后发酵液进行过滤,去除发酵菌丝体和剩余培养基等成分,可以提高虫草酵素的纯度,方便后续活性成分检测,同时也选择最佳的活性成分检测方法进行检测。
本发明还提供依据上述制备方法得到的虫草酵素在化妆品中的应用,按照重量比,包括:
虫草酵素0.01-10.0%
基质辅料90.0-99.9%。
本发明制备的虫草酵素可以以重量比0.01-10.0%添加到任意化妆品配方中使用,其中的主要营养及有效成份具有淡斑抗氧化、抗衰老、抑菌消炎等功效,可以对人体皮肤进行深层调理及保护,通过添加相关基质辅料还可以做成不同剂型的日化用品,在该领域具有很广阔的应用前景,包括但不限于水剂、洗剂、粉剂、片剂、面膜、凝胶、膏霜等。
说明书附图
图1为虫草菌种生长曲线图。
具体实施方式
为详细说明本发明的技术内容、构造特征、所实现目的及效果,以下结合实施方式予以说明。
1.方案内容
1.1虫草酵素制备的优化方法
包括如下步骤:
1)培养基的选择
目前,北虫草是虫草属中唯一接种成功和可以人工栽培进行大规模生产的菌种。其培养基的配方多种多样,选择适于虫草菌种发酵产生虫草酵素活性成分的液体培养基的组成和配比至关重要。
液体培养基成分按照重量比,组成如下:a、碳源10-20%,b、氮源10-20%,c、无机盐包括K2HPO4、MgSO4.7H2O,K2HPO4为0.04-0.1%,MgSO4.7H2O为0.01-0.07%,d、其他包括VB1、透明质酸和去离子水,VB1为0.002-0.005%,透明质酸0.2-0.5%,去离子水补到100%。
本发明使用的液体培养基,具体如下有五种配方,
1号培养基(1000g):葡萄糖10.0-20.0g,蛋白胨5.0-15.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;
2号培养基(1000g):葡萄糖10.0-20.0g,胡萝卜100.0-200.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;
3号培养基(1000g):葡萄糖10.0-20.0g,鲜蚕蛹30.0-50.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;
4号培养基(1000g):马铃薯100.0-200.0g,蛋白胨5.0-15.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g;
5号培养基(1000g):葡萄糖10.0-20.0g,蛋白胨5.0-15.0g,透明质酸2.0-5.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g。
经试验结果表明5号液体培养基为最佳培养基,并且最佳配比为:葡萄糖为20g,所述蛋白胨为15g,所述K2HPO4为1g,所述MgSO4.7H2O为0.5g,所述VB1为0.05g,所述透明质酸为5g,余下去离子水补到1000g。
2)虫草菌种的移植和保藏
移植培养的操作过程,将加了凝固剂的步骤1)中的液体培养基pH值调到6.0左右,灭菌后倾斜放入无菌操作台得到斜面固体培养基,冷却后接种虫草菌种,接种后的试管放入恒温培养箱中,在25℃下培养7d;保藏,菌种放在4℃-6℃冰箱内避光保存,移植期为2-3个月;移植培养,将虫草菌种转接到新配制的斜面固体培养基中,重复上述操作;移植期优选为3个月。
3)虫草菌种的接种
按照步骤1)中的液体培养基成分配制1000g液体培养基,装于带侧臂管的三角烧瓶内,灭菌冷却到室温后从斜面固体培养基中取出面积为1-3cm2的块状虫草菌苔,接入液体培养基中;所述接种量优选3cm2的块状虫草菌苔;
4)虫草菌种的摇床培养
将已接入液体培养基中的虫草菌种,在24-26℃的恒温震荡培养箱中以转速为120-150r/min避光培养68-72h,期间pH控制在5.0-7.0;
5)虫草菌种的扩大发酵
将配制好的液体培养基倒入发酵罐,在位灭菌25-35min,降温后,迅速倒入摇床培养得到虫草菌种培养液开始扩大发酵;设置发酵温度24℃~26℃,转速120-150r/min,培养8-12h后通气,通气量1.5-1.8(v/v·min),避光培养1~2天,期间pH控制在5.0-7.0,制得活性虫草酵素发酵液;所述优选摇床培养72h左右的处于对数生长期的虫草菌种进行扩大培养,所述虫草菌种扩大深层液体培养10h后通气,通气量为1.8(v/v·min);
6)虫草酵素活性成分检测
将上述发酵后活性虫草酵素发酵液置于离心机中,以3000-4500r/min的转速离心15-30min,收集发酵上清液,对发酵上清液用孔径为0.22μm中空纤维微滤系统进行微滤,收集发酵滤过液,再分别用基质固相分散提取法测定虫草素、分级沉淀法测定虫草SOD含量、苯酚-硫酸法测定虫草多糖含量、乙醇-微波协同提取法测定虫草酸含量及基质固相分散提取法测定核苷类物质含量。所述选取孔径为0.22μm中空纤维微滤系统进行微滤。
1.2虫草酵素的应用
根据上述制备方法得到的虫草酵素在化妆品中的应用,按照重量比,包括:虫草酵素0.01-10.0%,基质辅料90.0-99.9%。
本方法制备的虫草酵素可以以重量比0.01-10.0%添加到任意化妆品配方中使用,其中的主要营养及有效成份具有淡斑抗氧化、抗衰老、抑菌消炎等功效,可以对人体皮肤进行深层调理及保护,通过添加相关基质辅料还可以做成不同剂型的日化用品,包括但不限于水剂、洗剂、粉剂、片剂、面膜、凝胶、膏霜等。
1.2.1虫草酵素原液(重量比例)
虫草酵素0.01-10.0%,加去离子水至100。
1.2.2虫草酵素精华(重量比例)
虫草酵素10.0%、甘油4.0%、丁二醇3.0%、烟酰胺3.0%、蔗糖2.0%、泛醇1.5%、水杨酸1.0%、透明质酸1.0%、辛酰羟肟酸0.7%、1,2-己二醇0.3%、3-o-乙基抗坏血酸0.3%、生育酚0.5%、红没药醇0.1%、羟乙基纤维素0.2%、纤维素胶0.4%、甘草酸二钾0.05%、EDTA二钠0.05%、Solubilisant LRI 5.0%,加去离子水至100。
1.2.3虫草酵素面膜(重量比例)
虫草酵素5.0%、甘油5.0%、丁二醇3.0%、戊二醇1.5%、透明质酸1.0%、聚乙二醇-4001.0%、甘油聚醚-261.0%、山梨糖醇0.2%、卡波姆0.15%、羟乙基纤维素0.08%、硫辛酸0.01%、甘草酸二钾0.1%、氢化橄榄油0.1%、对羟基苯乙酮0.3%、1,2-己二醇0.3%、PEG-40氢化蓖麻油0.1%、三乙醇胺0.14%、EDTA-二钠0.05%、香精0.01%,加去离子水至100。
1.2.4虫草酵素乳液(重量比例)
虫草酵素2.0%、甘油5.0%、丁二醇3.0%、异壬酸异壬酯4.0%、角鲨烷2.0%、甘油聚醚-26 1.0%、聚二甲基硅氧烷1.0%、透明质酸1.0%、MONTANOV L 0.6%、SIMULGELEG 0.5%、烟酰胺0.5%、生育酚0.1%、红没药醇0.2%、卡波0.18%、三乙醇胺0.14%、乙酰壳糖胺0.1%、对羟基苯乙酮0.3%、1,2-己二醇0.3%、EDTA-二钠0.05%、香精0.02%,加去离子水至100。
1.2.5虫草酵素洗面乳(重量比例)
虫草酵素0.1%、甘油25.0%、肉豆蔻酸17.0%、棕榈酸7.5%、硬脂酸7.3%、氢氧化钾7.2%、LC 3.0%、月桂酸2.0%、透明质酸1.0%、羟苯甲酯0.2%、香精0.5%、聚乙二醇-14M 0.1%、EDTA二钠0.1%、瓜儿胶羟丙基三甲基氯化铵0.07%,加去离子水至100。
1.2.6虫草酵素沐浴露(重量比例)
虫草酵素3.5%、甲基椰油酰基牛磺酸钠6.0%、月桂酰两性基乙酸钠6.0%、月桂酰肌氨酸钠5.0%、椰油酰胺甲基MEA3.0%、癸基单油酸甘油酯2.0%、聚甘油-10油酸酯2.0%、辛酸/癸酸甘油酯类聚甘油-10酯类2.0%、PEG-120甲基葡糖二油酸酯1.5%、椰油酰羟乙磺酸酯钠1.0%、月桂基葡糖苷1.0%、MS621 1.0%、透明质酸1.0%、聚季铵盐-100.4%、柠檬酸钠0.1%、EDTA二钠0.1%、甘油月桂酸酯0.3%、柠檬酸0.3%、泛醇0.5%、脱氢乙酸0.5%、苯甲醇0.3%、香精0.25%,加去离子水至100。
1.2.7虫草酵素沐浴泡腾片(重量比例)
虫草酵素0.5%、柠檬酸20.0%、碳酸氢钠20.0%、氯化钠8.5.%、山梨醇8.0%、薄荷5.0%、聚乙二醇4000 2.0%、透明质酸1.0%、氨基酸起泡剂5.0%、苯甲酸钠0.5%、乙醇0.5%、香精0.5%。
1.2.8虫草酵素口服液(重量比例)
虫草酵素2.5%、甜橙汁5.0%、蓝莓汁2.0%、葡萄汁2.0%、维生素C 1.0%、透明质酸1.0%、乳糖0.2%、麦芽糊精0.2%、氨基酸0.1%、氯化钠0.1%,加去离子水至100。
2.虫草酵素活性成分及检测结果
2.1活性成分
虫草酵素:由北虫草经液态深层发酵制得,主要包括虫草素、虫草多糖、虫草酸、虫草SOD、核苷类物质、虫草甾醇、维生素、蛋白质、多肽、氨基酸和微量元素等活性成分,对人体皮肤有多种功效作用,在化妆品领域具有光明的发展前景。
虫草素:虫草素又称虫草菌素、蛹虫草菌素、3’-脱氧腺苷,是腺苷(Adenosine)的类似物,它是第一个从真菌中分离出来的核苷类抗菌素。虫草素可以抑制病毒的RNA合成,对鸟结核杆菌、枯草杆菌、牛型结核分枝杆菌、鼻疽杆菌、炭疽杆菌、梭菌、链球菌及石膏样小芽饱癣菌等均有一定抑制作用,此外还具有一定的抗真菌和抗HIVI型病毒活性,其中对枯草杆菌的抑制作用最强;由于机体代谢过程中产生SAFR、HFR等过量的活性氧,导致体内产生大量的自由基,导致人体衰老,研究表明,虫草素具有特异性清除活性氧SAFR的作用,虫草中含有的D-甘露醇对HFR的清除作用较强,达到延缓衰老、清除体内自由基的作用。
虫草多糖:虫草多糖是虫草酵素的主要活性物质之一,它是高分子复合物,作为药理活性物质虫草多糖含量最高。虫草多糖(EPS)的抗癌活性和免疫活性引起人们广泛关注,研究表明,它对巨噬细胞及网状皮系统有显著的激活作用,是非特异性免疫调节及增强剂,具有调节免疫细胞间信息的传递与感受和促进淋巴细胞的转化作用,具有抗氧化、抗凝血、抗衰老、抗病毒等作用。
虫草酸:虫草酸又名D-甘露密醇、D-甘露醇、D-甘露糖醇。虫草酸是一种自然的多元醇与六个碳糖和蔗糖相比其相对甜度为40-50%,属于非代谢的甜味剂。虫草酸具有一些独特的优势,可以作为一种抗氧化剂,对氧自由基的氧化损伤有保护作用。虫草酸还可以可以显著地降低颅压,促进机体新陈代谢,因而使脑溢血和脑血栓病症得到缓解。
虫草SOD:虫草SOD又称虫草超氧化物歧化酶,是一种生物活性蛋白质,可由人体自身合成的重要抗氧化剂,对人体细胞起一定保护作用。它是目前为止发现的唯一的以自由基为底物的酶,它可以清除人体代谢产生的自由基,避免细胞受到氧化、老化和破坏,对于人体活性氧的代谢具有维护平衡作用。SOD具有抗氧化、抗老化,清除因辐射和药物等诱导因素产生的超氧自由基,可以增强免疫,抑制病菌,辅助医疗神经系统、消化系统疾病等功能,SOD可用于皮肌炎、抗类风湿,用于化妆品及添加在药品、食品中作为抗氧化剂,对于保持青春容颜、提高人体免疫功能等方面均有显著作用。研究发现,人工培养的虫草酵素中含有丰富的SOD,SOD的活性与培养基的种类有很大的关系,适宜的培养基可以使SOD的活性达到147.60U/mL。
核苷类物质:虫草酵素中核苷类物质除虫草素外还有次黄嘌呤、次黄嘌呤核苷、尿嘧啶、尿苷、胸腺嘧啶、鸟嘌呤、腺嘌呤、腺苷等。研究表明,人工培养的北虫草比天然虫草中腺苷含量高许多,核苷类物质对人体具有极其重要的作用,如预防辐射造成的伤害,对心脏具有负收缩效应,可以抑制血小板凝结等作用。
虫草甾醇:虫草酵素中含有麦角甾醇、角甾醇、β-谷甾醇、黄豆醇等甾醇衍生物,虫草甾醇对人体具有较强的抗炎作用,具有能够抑制人体对胆固醇的吸收、促进胆固醇的降解代谢、抑制胆固醇的生化合成等作用,虫草甾醇还是重要的甾体药物和维生素D3的生产原料,虫草甾醇对皮肤具有很高的渗透性,可以保持皮肤表面水份,促进皮肤新陈代谢、抑制皮肤炎症,可防日晒红斑、皮肤老化,还有生发、养发的功效。
维生素:虫草酵素中含VA、VB类、VC、VE等多种维生素,在皮肤抗氧化和调理方面起到功效作用。
蛋白质、多肽、氨基酸:虫草酵素中含有丰富的氨基酸和蛋白质,其中氨基酸含量在22%左右,粗蛋白总量达到30%,虫草酵素中含有18种氨基酸,总量可以达到11145mg/100g,其中人体8种必需氨基酸比例适当且含量较高,含量最高的为谷氨酸和天门冬氨酸,虫草酵素中还分离得到L-丙-L-缬环二肽等六个环二肽类化合物和虫草环A等多肽,可快速被人体皮肤吸收。
微量元素:虫草酵素中已检出含有37种微量元素,以P含量为最高,其次为Na,K,Mg,Mn,Fe,Cu,Zn,尤其富含硒,它是人体必需物质,是谷胱甘肽过氧化酶的活性中心,以硒半胱氨酸的形式连接在酶蛋白的肽链上,保护细胞膜的稳定性和正常的通透性,并刺激免疫球蛋白和抗体的产生,增强机体免疫和抗氧化能力,同时硒可明显地抑制癌细胞的生长,具有良好的抗癌作用,其中冬虫夏草的硒的含量为0.34μg/g,北虫草中的硒的含量比前者高58.82%,达到0.54μg/g。同时北虫草中还有多种无机元素,其中包括含量较高的镁、磷、钙、铁,以及钛、锌、铝、锰、硅、铜、铬和含量丰富的锶、镍、钾。
2.2活性成分检测结果
培养基的成分组成及配比是影响虫草发酵的主要因素,在不同培养基培养条件下,虫草发酵后得到的虫草酵素活性成分的检测结果如下。
表1不同培养基中虫草酵素活性成分的检测结果:
虫草素 | 虫草SOD | 虫草多糖 | 虫草酸 | 核苷类物质 | |
1号培养基 | 14.68mg/mL | 143.80U/mL | 9.56% | 1.11% | 120.24mg/mL |
2号培养基 | 14.25mg/mL | 142.32U/mL | 10.28% | 1.10% | 118.82mg/mL |
3号培养基 | 13.17mg/mL | 141.95U/mL | 10.95% | 1.05% | 119.76mg/mL |
4号培养基 | 12.89mg/mL | 142.05U/mL | 8.44% | 1.01% | 117.69mg/mL |
5号培养基 | 15.10mg/mL | 147.60U/mL | 11.37% | 1.12% | 124.88mg/mL |
结果:由表1对比1号、2号和3号培养基可知,蛋白胨为最佳氮源;对比1号和4号培养基可知,葡萄糖为最佳碳源;选用1号和5号培养基培养制备的虫草酵素的活性成分含量明显高于其他各组;在1号培养基中加入适量的透明质酸的5号培养基可以明显提高虫草酵素的活性成分含量,为本方法的最优培养基,说明透明质酸有利于虫草的发酵;选用本方法提供的最优培养基培养得到的虫草酵素,其活性成分中虫草素、虫草SOD、虫草多糖和核苷类物质的含量增加非常显著。
这是因为虫草菌种的发酵过程要依次经历延迟期、对数期、稳定期、衰亡期四个阶段,以虫草菌种生长速率或数目与培养基配方和培养时间为基础研究虫草发酵后得到的虫草酵素活性成分的含量,可以帮助人们寻找虫草菌种的最佳培养基。
由图1可知,将菌种接入到新配制的培养基中,在最佳条件下培养,每隔5h测定一次吸光度。0-25h菌种处于迟缓期,迟缓期为菌种休眠期,25h后开始进入菌种生长对数期,对数期为菌种快速生长和增加数目的时期,大约72h吸光度值达到最高峰,菌种生长从对数期转为稳定期,稳定期为菌种代谢产物(发酵产物-虫草酵素)最旺盛时期,接近120h吸光度开始下降菌种进入衰亡期,衰亡期的菌种逐渐失去活性。因此在培养过程中可以选择虫草菌种培养72h左右作为扩大培养的转接点,并延长虫草菌种在扩大培养时对数期的时间及提升对数期生长的速率,都可以增加虫草菌种的数目,还可以延长虫草菌种在扩大培养时稳定期的时间,推迟衰亡期的到来,增加发酵产物虫草酵素的含量,从而使得虫草酵素中的各活性成分有所提高。
(1)培养基成分的选择如下:
a、液体培养基碳源的选择
碳源可以为虫草提供碳元素的营养源,大部分可以作为虫草生长所需的能源,碳源谱分布广泛。葡萄糖与马铃薯等其他碳源相比具有稳定性好、菌种好利用、延长稳定期时间的优点,从而可以提高发酵产物活性成分的含量,因此本研究选择葡萄糖作为培养基碳源。
b、液体培养基氮源的选择
氮源可以将氮元素提供给虫草,一般不能作为生长所需要的能源。蛋白胨与胡萝卜、蚕蛹等氮源相比具有稳定性好、菌种好利用、延长稳定期时间的优点,从而可以提高发酵产物活性成分的含量,因此本研究选择蛋白胨作为培养基的氮源。
c、液体培养基无机盐的选择
无机盐可以为虫草的生长提供化学成分,同时在调节细胞渗透压和酸碱度起着重要的作用。研究表明K2HPO4和MgSO4·7H2O对北虫草的对数期生长速率具有明显的提升效果,所以本研究选择K2HPO4和MgSO4·7H2O两种无机盐进行发酵培养。
d、液体培养基其他成分的选择
其他成分包括VB1、透明质酸和去离子水,其中VB1可以增加菌种保藏时间,延缓菌种衰亡期的到来。透明质酸选取3-10个双糖单位、分子量在1000-4000D的小分子透明质酸寡糖片段,这样的透明质酸利于吸收和利用,还可以促进培养基中其他营养成分的利用,添加到培养基中可以为菌种生长提供能源;菌种移植过程中可以提高活化延长移植时间;摇床上培养过程中可以提高菌种对数期生长速率,提高菌种数目;扩大培养时可以提高菌种对数期生长速率、提高菌种数目,并延长菌种生长稳定期的时间,增加发酵产物虫草酵素的含量,从而使得虫草酵素中的各活性成分有所提高。
(2)培养基成分配比的选择如下:
a、液体培养基葡萄糖配比的选择
液体培养基葡萄糖的配比在10-20g范围内,虫草酵素活性成分含量随葡萄糖含量增加而增加,大于20g时葡萄糖含量增加虫草酵素活性成分含量变化不大,这是因为当碳氮比变大,有机物分解速度变慢,抑制虫草菌种的生长速度,故选定葡萄糖20g为最佳添加量。
b、液体培养基蛋白胨配比的选择
液体培养基蛋白胨的配比在5-15g范围内,虫草酵素活性成分含量随蛋白胨含量增加而增加,15-25g时蛋白胨含量增加虫草酵素活性成分含量反而减小,这是因为蛋白胨含量大,碳氮比变小影响代谢产物的积累,抑制虫草菌种的生长,故选定蛋白胨15g为最佳添加量。
c、液体培养基K2HPO4配比的选择
液体培养基K2HPO4的配比在0.4-1.0g范围内,虫草酵素活性成分含量随K2HPO4含量增加而增加,大于1.0g时K2HPO4含量增加虫草酵素活性成分含量反而减小,这是因为K2HPO4过多导致培养基中PH值发生改变,从而抑制虫草菌种的生长,故K2HPO4选取1.0g为最佳添加量。
d、液体培养基MgSO4.7H2O配比的选择
液体培养基MgSO4.7H2O的配比在0.1-0.5g范围内,虫草酵素活性成分含量随MgSO4.7H2O含量增加而增加,0.5-0.7g时虫草酵素活性成分含量基本不变,大于0.7g时MgSO4.7H2O含量增加虫草酵素活性成分含量反而减小,这是因为当MgSO4.7H2O含量过高时会变成了抑制虫草菌种生长的因子,故MgSO4.7H2O选定0.5g为最佳的添加量。
e、液体培养基其他成分配比的选择
液体培养基VB1和透明质酸的配比分别在0.02-0.05g和2.0-5.0g范围内,虫草酵素活性成分含量随液体培养基VB1和透明质酸含量增加而增加,当两者含量到一定程度时虫草酵素活性成分含量基本不变,故液体培养基VB1和透明质酸分别选定0.05g和5.0g为最佳的添加量。
3.功效实验
a、虫草酵素的抗衰老实验
根据基质金属蛋白酶衰老学说得知,一些外界因素可以引起人体内基质金属蛋白酶-1(MMP-1)的活性增强,MMP-1会过度降解细胞外基质,破坏皮肤结构,从而导致皱纹的产生。如果某种成分能够有效抑制基质金属蛋白酶-1的活性,就能减少细胞外基质的破坏和流失,阻止皱纹的产生,达到抵抗皮肤衰老的目的。因此,通过体外抑制MMP-1活性的实验来建立体外功效检测方法,可以用于筛选具有消除细纹、紧致肌肤、抗皱抗衰功效的化妆品原料或产品。
一、实验方法:在不破坏酶反应体系的前提下,加入待测物样品,通过酶反应体系检测产物荧光强度的变化,间接反映酶活力的变化,从而反映功效成分对MMP-1活性的影响作用。
二、具体实验步骤:(1)溶剂配制:①配制缓冲液(50mmol/L HEPES,0.12mol/LNaCl,10mmol/L CaCl2,20μmol/L,ZnSO4,0.05%Brij-35,pH=7.5);②将MMP-1酶原用缓冲液梯度稀释至0.22μg/100μL备用;③将荧光底物DQ-gelatin用缓冲液梯度稀释至0.20μg/100μL备用;④在1L的0.1mol/L NaOH溶液中加入30mmol的酶原激活剂APMA,形成APMA储备液(30mmol/L)备用;⑤将各待测物样品配制成1mg/ml浓度的溶液备用:分别按实施例7-9中五味中药提取物配比比例,分别标识为试验1-3组;实施例1制备的浸膏,标识为试验4组;实施例3制备的浸膏,标识为试验5组;实施例6制备的提取物,按实施例7的中药提取物比例配比,标识为试验6组。各单味原料药的水提物制备方法与实施例1提取方法相同;各单味原料药的醇提物制备方法与实施例3提取方法相同;(2)MMP-1酶原的活化:将MMP-1酶原溶液(0.22μg/100μL)和APMA溶液(30mmol/L)按10:1的体积比(MMP:APMA)混合,在37℃孵育45min活化;(3)加样:取活化后的MMP-1溶液(浓度已降为0.20μg/100μL)40μL加入96孔酶标板中,再加入待测物样品溶液8μL,荧光底物DQ-gelatin溶液(0.20μg/100μL)52μL,使最终反应体系为100μL;(4)恒温孵育:将完成加样的96孔酶标板放入37℃培养箱中恒温孵育600s。
表2体外抑制MMP-1实验加样表:
样品 | 缓冲溶液 | 荧光底物 | MMP-1 | 待测物溶液 | 总体积 |
样品组A2 | 92μL | - | - | 8μL | 100μL |
实验组A1 | - | 52μL | 40μL | 8μL | 100μL |
对照组A0 | 8μL | 52μL | 40μL | - | 100μL |
三、实验结果检测
(1)反应前在激发波长为460nm,发射波长为520nm的检测条件下检测待测物功效成分的荧光强度;(2)反应体系停止孵育后立即用荧光酶标仪检测整个反应体系的荧光强度,激发波长为460nm,发射波长为520nm。
四、实验数据处理:MMP-1抑制百分数(%)=[1-(A1-A2)/A0]*100%,A1—代表添加待测物功效成分抑制组的荧光强度,A2—代表待测物功效成分自身的荧光强度,A0—代表空白对照组的荧光强度。
表3待测物功效成分对MMP-1活性的抑制率
结论:本实验中对MMP-1活性的抑制率越大说明待测物功效成分的抗衰老效果越好。由表3可以看出,浓度在0.01-10%的虫草酵素原液对MMP-1活性均有抑制作用,而且随着虫草酵素浓度的增加虫草酵素原液对MMP-1活性的抑制作用也越来越强,但当虫草酵素浓度达到5.0%左右时这种抑制作用的增幅变缓;不同浓度的虫草酵素配制成的日化护肤产品同样对MMP-1活性有抑制作用;相同浓度的虫草酵素配制成的日化护肤产品对MMP-1活性的抑制作用比虫草酵素原液强烈,如虫草酵素精华比10.0%的虫草酵素原液对MMP-1活性的抑制作用强,这可能是某种或某些成分与虫草酵素在对MMP-1活性的抑制作用上产生了协同增强的作用。
b、皮肤弹性测试实验
人体皮肤弹性会随皮肤的衰老而降低,因此皮肤弹性是判断皮肤衰老的重要标志之一,是抗衰老化妆品原料或产品功效检测必不可少的一项。
一、实验方法:选择健康肌肤正常、无化妆品过敏史受试人群,年龄20-35岁,其中男性10位,女性20位。以周为1个疗程,总共4个疗程,每次涂抹前将面部清洁干净,且每次洁面的时间都相同,待皮肤晾干后涂抹对应产品,各组每天涂抹相对应产品2次,每次2g。
二、测量:选择面颊部作为测试区域,待使用产品后10min进行测试。使用皮肤弹性测试仪测定皮肤弹性曲线,根据皮肤的拉伸量和回弹性数据,可对化妆品的抗衰老功效进行评价。
三、实验结果:皮肤弹性性能测试是基于吸力和拉伸原理,在被测皮肤的表面产生一个负压,将皮肤吸进一个特定测试探头内,皮肤被吸进的深度是通过一个非接触式的光学测试系统测得,测量结果通常用一个指数来表示,单位为1,数值越大,说明皮肤弹性越强,抗衰老效果越好。抗衰老产品对皮肤弹性的影响如表4所示。
表4皮肤弹性测试值
结论:由表4可以看出,使用浓度在0.01-10%的虫草酵素原液4周后人体皮肤的弹性值都有所提升,说明不同浓度的虫草酵素原液抗衰老效果很好;不同浓度的虫草酵素配制成的日化护肤产品同样可以提升皮肤的弹性值;相同浓度的虫草酵素配制成的日化护肤产品对皮肤弹性值的提升作用比虫草酵素原液强烈,由虫草酵素精华与10.0%的虫草酵素原液对比可知,这可能是某种或某些成分与虫草酵素在对皮肤弹性值的提升作用上产生了协同增强的作用;虫草酵素使用4周内的第2、3周对人体皮肤弹性值的提升更有效。
c、虫草酵素的皮肤调理性实验
现采用人体实验的方法检测虫草酵素在人体的生物活性,即淡斑抗氧化、抗衰老、抑菌消炎的活性,研究该虫草酵素以预防和调理皮肤不适,具体指皮肤感觉不适、过敏、炎症、红肿、疼痛、细纹、斑点、异味、污渍等不适现象。
本实验所有试验者均为不同程度的皮肤不适者,实验组1是0.01%的虫草酵素原液每晚睡前涂抹1次;实验组2是1.0%的虫草酵素原液每晚睡前涂抹1次;实验组3是5.0%的虫草酵素原液每晚睡前涂抹1次;实验组4是10.0%的虫草酵素原液每晚睡前涂抹1次;实验组5是虫草酵素精华每晚睡前涂抹1次;实验组6是虫草酵素面膜精华每晚睡前涂抹1次;实验组7是虫草酵素乳液每晚睡前涂抹1次;阴性对照采用不含虫草酵素的普通乳液每晚睡前涂抹1次;比较五组调理7天后症状改善情况。
皮肤感觉不适、过敏、炎症、红肿、疼痛、细纹、斑点、异味、污渍等不适现象的症状评价:
显效:症状显著改善或消失,试验者满意;有效:症状有改善或减轻,试验者仍有不适,要求继续调理;无效:症状无明显减轻,试验者不满意。
统计学方法:采用SPSS12.0统计学软件进行统计学处理,计量和计数资料组间比较采用t检验和x2检验,以p<0.05为有统计学意义差异。
结果如下:
如表5所示,第8天症状改善情况:实验组1显效4例,有效2例,无效4例,总有效率60%,显效率与实验组3比较P<0.01;实验组2显效6例,有效4例,总有效率100%;实验组3显效10例,总有效率100%;实验组4显效10例,总有效率100%;实验组5显效10例,总有效率100%;实验组6显效10例,总有效率100%;实验组7显效10例,总有效率100%。
表5第8天各组试验者症状改善情况比较如下:
结论:各实验组检测结果均比阴性对照有效。本发明的虫草酵素用人体实验表明浓度在0.01-10.0%的虫草酵素原液及产品对预防和调理皮肤不适效果显著;该虫草酵素在短期内对淡斑抗氧化、抗衰老、抑菌消炎等皮肤问题有一定功效,长期使用效果会更佳,可以广泛的用在化妆品领域。
以上所述仅为本发明的实施例,并非因此限制本发明的专利保护范围,凡是利用本发明说明书及附图内容所作的等效结构或等效流程变换,或直接或间接运用在其他相关的技术领域,均同理包括在本发明的专利保护范围内。
Claims (7)
1.一种活性虫草酵素的制备方法,其特征在于包括如下步骤:
1)液体培养基的选择
液体培养基成分按照重量比,组成如下:
a、碳源10-20%,
b、氮源10-20%,
c、无机盐为K2HPO4和MgSO4·7H2O,K2HPO4为0.04-0.1%,MgSO4·7H2O为0.01-0.07%,
d、其他为VB1,透明质酸和去离子水,VB1为0.002-0.005%,透明质酸0.2-0.5%,去离子水补到100%;
2)虫草菌种的移植和保藏
移植培养的操作过程,将加了凝固剂的液体培养基pH值调到6.0,灭菌后倾斜放入无菌操作台得到斜面固体培养基,冷却后接种虫草菌种,接种后的试管放入恒温培养箱中,在25℃下培养7d;保藏,菌种放在4℃-6℃冰箱内避光保存,移植前保藏期为2-3个月;移植培养,将虫草菌种转接到新配制的斜面固体培养基中,重复上述操作;
3)虫草菌种的接种
按照步骤1)中的液体培养基成分配制1000g液体培养基,装于带侧臂管的三角烧瓶内,灭菌冷却到室温后从斜面固体培养基中取出面积为1-3cm2的块状虫草菌苔,接入液体培养基中;
4)虫草菌种的摇床上培养
将步骤3中接入液体培养基中的虫草菌种,在24-26℃的恒温震荡培养箱中以转速为120-150r/min避光培养68-72h,期间pH控制在5.0-7.0;
5)虫草菌种的扩大发酵
将步骤1配制好的液体培养基倒入发酵罐,原位灭菌25-35min,降温后,迅速将步骤4中摇床上培养得到的虫草菌种培养液倒入发酵罐中,开始扩大发酵;设置发酵温度24℃~26℃,转速120-150r/min,培养8-12h后通气,通气量1.5-1.8v/v·min,避光培养1~2天,期间pH控制在5.0-7.0,制得活性虫草酵素发酵液。
2.如权利要求1所述的活性虫草酵素的制备方法,其特征在于,还包括步骤6)虫草酵素活性成分检测,
将上述步骤5)发酵后制得的活性虫草酵素发酵液置于离心机中,以3000-4500r/min的转速离心15-30min,收集发酵上清液,对发酵上清液用孔径为0.22μm中空纤维微滤系统进行微滤,收集发酵滤过液,再分别用基质固相分散提取法测定虫草素、分级沉淀法测定虫草SOD含量、苯酚-硫酸法测定虫草多糖含量、乙醇-微波协同提取法测定虫草酸含量及基质固相分散提取法测定核苷类物质含量。
3.如权利要求1所述的活性虫草酵素的制备方法,其特征在于,所述液体培养基组成成分中碳源为葡萄糖、马铃薯其中之一;氮源为蛋白胨、胡萝卜、蚕蛹其中之一。
4.如权利要求1所述的活性虫草酵素的制备方法,其特征在于,所述透明质酸为3-10个双糖单位、分子量在1000-4000D的小分子透明质酸寡糖片段。
5.如权利要求1或3所述的活性虫草酵素的制备方法,其特征在于,所述液体培养基为:
葡萄糖10.0-20.0g,蛋白胨5.0-15.0g,透明质酸2.0-5.0g,K2HPO4 0.4-1.0g,MgSO4·7H2O 0.1-0.7g,VB1 0.02-0.05g,余下去离子水补到1000g。
6.如权利要求5所述的活性虫草酵素的制备方法,其特征在于,所述液体培养基组成成分中葡萄糖为20g,所述蛋白胨为15g,所述K2HPO4为1g,所述MgSO4·7H2O为0.5g,所述VB1为0.05g,所述透明质酸为5g,余下去离子水补到1000g。
7.如权利要求1或2所述的活性虫草酵素的制备方法,其特征在于,所述菌种保藏中移植时间3个月,所述接种量取3cm2的块状虫草菌苔,所述步骤4中接入液体培养基中的虫草菌种摇床培养时间为72h,步骤5扩大发酵过程中所述虫草菌种扩大深层液体培养10h后通气,通气量为1.8v/v·min。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201711096208.8A CN108085303B (zh) | 2017-11-09 | 2017-11-09 | 一种活性虫草酵素的制备方法 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201711096208.8A CN108085303B (zh) | 2017-11-09 | 2017-11-09 | 一种活性虫草酵素的制备方法 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN108085303A CN108085303A (zh) | 2018-05-29 |
CN108085303B true CN108085303B (zh) | 2021-03-19 |
Family
ID=62172041
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN201711096208.8A Expired - Fee Related CN108085303B (zh) | 2017-11-09 | 2017-11-09 | 一种活性虫草酵素的制备方法 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN108085303B (zh) |
Families Citing this family (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN109355211A (zh) * | 2018-12-13 | 2019-02-19 | 蒋涛 | 一种辣木蛹虫草及其培养方法及抑菌喷剂 |
CN111635864B (zh) * | 2020-05-20 | 2022-02-01 | 中国科学院动物研究所 | 一种用于培养冬虫夏草菌的液体培养基及其制法 |
CN113425663A (zh) * | 2021-08-18 | 2021-09-24 | 皖西学院 | 一种石斛花大豆美肤面膜的制备方法 |
CN114469824A (zh) * | 2021-12-27 | 2022-05-13 | 北京电子科技职业学院 | 虫草精华多效臻养乳液及其制备方法 |
CN115141760A (zh) * | 2022-08-23 | 2022-10-04 | 北京焉支山科技有限公司 | 一种桦褐孔菌发酵提取液及其制备方法和在制备化妆品中的应用 |
CN116747176A (zh) * | 2023-08-10 | 2023-09-15 | 珠海市华喜生物科技有限公司 | 莲花发酵物、制备方法及在抗皱紧致化妆品中的应用 |
Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN103330258A (zh) * | 2013-07-17 | 2013-10-02 | 黄雄 | 一种液体深层发酵制备的蛹虫草保健饮料及其制备方法 |
KR20140062456A (ko) * | 2014-05-09 | 2014-05-23 | 주식회사 엘지생활건강 | 주름개선용 화장료 조성물 |
CN104173389A (zh) * | 2014-08-20 | 2014-12-03 | 厦门元尊生物工程有限公司 | 一种蛹虫草酵素粉及其制备方法 |
CN104846018A (zh) * | 2015-05-23 | 2015-08-19 | 吉林省方平科技有限公司 | 虫草酵素深层液体发酵及其扩培方法 |
CN107058119A (zh) * | 2016-12-29 | 2017-08-18 | 徐州鸿宇农业科技有限公司 | 一种提高蛹虫草液态发酵生产虫草素和耐热蛋白酶产量的方法 |
-
2017
- 2017-11-09 CN CN201711096208.8A patent/CN108085303B/zh not_active Expired - Fee Related
Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN103330258A (zh) * | 2013-07-17 | 2013-10-02 | 黄雄 | 一种液体深层发酵制备的蛹虫草保健饮料及其制备方法 |
KR20140062456A (ko) * | 2014-05-09 | 2014-05-23 | 주식회사 엘지생활건강 | 주름개선용 화장료 조성물 |
CN104173389A (zh) * | 2014-08-20 | 2014-12-03 | 厦门元尊生物工程有限公司 | 一种蛹虫草酵素粉及其制备方法 |
CN104846018A (zh) * | 2015-05-23 | 2015-08-19 | 吉林省方平科技有限公司 | 虫草酵素深层液体发酵及其扩培方法 |
CN107058119A (zh) * | 2016-12-29 | 2017-08-18 | 徐州鸿宇农业科技有限公司 | 一种提高蛹虫草液态发酵生产虫草素和耐热蛋白酶产量的方法 |
Also Published As
Publication number | Publication date |
---|---|
CN108085303A (zh) | 2018-05-29 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN108085303B (zh) | 一种活性虫草酵素的制备方法 | |
EP2444480B1 (en) | Method for fermentation and cultivation, fermented plant extract, fermented plant extract powder, and composition containing the extract of fermented plant | |
KR101353054B1 (ko) | 고로쇠 나무와 황칠 나무를 이용한 발효물 및 이의 제조방법 그리고 이를 이용한 식품 조성물 및 화장료 조성물 | |
KR101852047B1 (ko) | 식물세포 복합발효추출물을 포함하는 주름 및 미백 개선 기능성 조성물 및 이를 포함하는 기능성 화장품 | |
CN107898743A (zh) | 全天然多植物酶解提取的美白嫩肤除皱全能美容液 | |
KR101705482B1 (ko) | 세리포리아 락세라타가 포함된 아토피 개선용 로션 또는 크림 | |
WO2021158179A1 (en) | SCHIZOPHYLLUM COMMUNE STRAIN PLNP13 AND β-GLUCAN OBTAINED FROM CO-CULTURING THE STRAIN WITH GANODERMA LUCIDUM | |
CN108042436B (zh) | 蓝铜胜肽抗衰保湿精华霜 | |
ES2526873T3 (es) | Procedimiento para la preparación de un producto natural fermentado | |
KR101214872B1 (ko) | 버섯균사체 배양액을 유효성분으로 함유하는 화장료 조성물 | |
KR100438009B1 (ko) | 잎새버섯 균사체의 추출물을 함유하는 화장료 조성물 | |
KR101127271B1 (ko) | 동충하초균의 배양방법 및 이에 따른 배양물의 용도 | |
TWI747488B (zh) | 佛手柑發酵汁液用於製備改善皮膚老化組合物的用途 | |
KR101778853B1 (ko) | 버섯을 이용한 삼투압 효소 발효물의 제조방법, 이로부터 제조된 발효물 및 이를 포함하는 화장료, 식품 또는 약학 조성물 | |
KR102507651B1 (ko) | 전통주 효모 함유 천연발효효소복합액을 이용한 숙취해소, 알콜성간질환 또는 간기능 개선용 멀꿀발효물과 그 제조방법 | |
TWM590969U (zh) | 具有仿生物間質系統的芸香科植物發酵液微粒結構 | |
JP2006075083A (ja) | ゴーヤから得られる発酵物 | |
KR101750619B1 (ko) | 여주와 어성초를 이용한 삼투압 효소 발효물의 제조방법, 이로부터 제조된 발효물 및 이를 포함하는 화장료, 식품 또는 약학 조성물 | |
KR20160121632A (ko) | 효모 발효 허브 복합체을 이용하여 항산화 효과가 우수하고 저자극인 추출물의 제조 방법 | |
JP2006089408A (ja) | レモンから得られる発酵物 | |
KR20220039942A (ko) | 블루베리로부터 분리된 티로시나아제 저해 활성 및 항산화 활성이 있는 사카로마이세스 세레비지애 ft4-4 균주 및 이의 용도 | |
KR20170114574A (ko) | 세리포리아 락세라타가 함유된 아토피 피부염 개선 비누 | |
CN111658575A (zh) | 美白酵素面膜 | |
CN113616584A (zh) | 含双菌发酵产物与活性葡萄籽提取物抵御城市污染的抗炎护肤品组合物 | |
CN110638738A (zh) | 一种低温浴处理玫瑰制备生态营养液的方法及应用 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
GR01 | Patent grant | ||
GR01 | Patent grant | ||
CF01 | Termination of patent right due to non-payment of annual fee | ||
CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20210319 |