CN118001250A - Fat-soluble vitamin liposome precursor, and preparation method and application thereof - Google Patents
Fat-soluble vitamin liposome precursor, and preparation method and application thereof Download PDFInfo
- Publication number
- CN118001250A CN118001250A CN202410168120.6A CN202410168120A CN118001250A CN 118001250 A CN118001250 A CN 118001250A CN 202410168120 A CN202410168120 A CN 202410168120A CN 118001250 A CN118001250 A CN 118001250A
- Authority
- CN
- China
- Prior art keywords
- liposome
- vitamin
- protein
- protease
- precursor
- 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.)
- Pending
Links
- 239000002502 liposome Substances 0.000 title claims abstract description 300
- 239000002243 precursor Substances 0.000 title claims abstract description 130
- 229940088594 vitamin Drugs 0.000 title claims abstract description 77
- 229930003231 vitamin Natural products 0.000 title claims abstract description 77
- 235000013343 vitamin Nutrition 0.000 title claims abstract description 77
- 239000011782 vitamin Substances 0.000 title claims abstract description 77
- 150000003722 vitamin derivatives Chemical class 0.000 title claims abstract description 43
- 238000002360 preparation method Methods 0.000 title claims abstract description 35
- 239000004365 Protease Substances 0.000 claims abstract description 113
- 239000000413 hydrolysate Substances 0.000 claims abstract description 71
- 238000003756 stirring Methods 0.000 claims abstract description 57
- 238000000034 method Methods 0.000 claims abstract description 50
- 108091005804 Peptidases Proteins 0.000 claims abstract description 36
- 238000001694 spray drying Methods 0.000 claims abstract description 34
- 238000005538 encapsulation Methods 0.000 claims abstract description 26
- 238000002156 mixing Methods 0.000 claims abstract description 26
- 108010009736 Protein Hydrolysates Proteins 0.000 claims abstract description 23
- 230000008569 process Effects 0.000 claims abstract description 20
- 239000007788 liquid Substances 0.000 claims abstract description 19
- 239000000945 filler Substances 0.000 claims abstract description 16
- 238000001035 drying Methods 0.000 claims abstract description 11
- 230000001105 regulatory effect Effects 0.000 claims abstract description 11
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 claims abstract 8
- 239000000243 solution Substances 0.000 claims description 82
- 108010046377 Whey Proteins Proteins 0.000 claims description 61
- 102000007544 Whey Proteins Human genes 0.000 claims description 61
- 239000005018 casein Substances 0.000 claims description 57
- BECPQYXYKAMYBN-UHFFFAOYSA-N casein, tech. Chemical compound NCCCCC(C(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(CC(C)C)N=C(O)C(CCC(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(C(C)O)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(COP(O)(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(N)CC1=CC=CC=C1 BECPQYXYKAMYBN-UHFFFAOYSA-N 0.000 claims description 57
- 235000021240 caseins Nutrition 0.000 claims description 57
- 229940111202 pepsin Drugs 0.000 claims description 54
- 229940055729 papain Drugs 0.000 claims description 45
- 235000021119 whey protein Nutrition 0.000 claims description 39
- 235000019419 proteases Nutrition 0.000 claims description 35
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 30
- 230000007062 hydrolysis Effects 0.000 claims description 29
- 238000006460 hydrolysis reaction Methods 0.000 claims description 29
- 102000057297 Pepsin A Human genes 0.000 claims description 28
- 108090000284 Pepsin A Proteins 0.000 claims description 28
- 108090000526 Papain Proteins 0.000 claims description 25
- 235000019834 papain Nutrition 0.000 claims description 25
- 102000004169 proteins and genes Human genes 0.000 claims description 20
- 108090000623 proteins and genes Proteins 0.000 claims description 20
- 235000018102 proteins Nutrition 0.000 claims description 19
- 229920002774 Maltodextrin Polymers 0.000 claims description 16
- 239000005913 Maltodextrin Substances 0.000 claims description 16
- 229940035034 maltodextrin Drugs 0.000 claims description 16
- 239000012460 protein solution Substances 0.000 claims description 16
- 230000001954 sterilising effect Effects 0.000 claims description 15
- 238000004659 sterilization and disinfection Methods 0.000 claims description 14
- 229940088598 enzyme Drugs 0.000 claims description 12
- 102000004190 Enzymes Human genes 0.000 claims description 11
- 108090000790 Enzymes Proteins 0.000 claims description 11
- 230000003301 hydrolyzing effect Effects 0.000 claims description 11
- 238000009928 pasteurization Methods 0.000 claims description 7
- 229910019142 PO4 Inorganic materials 0.000 claims description 5
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims description 5
- DLRVVLDZNNYCBX-UHFFFAOYSA-N Polydextrose Polymers OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(O)O1 DLRVVLDZNNYCBX-UHFFFAOYSA-N 0.000 claims description 4
- 239000012263 liquid product Substances 0.000 claims description 4
- 239000010452 phosphate Substances 0.000 claims description 4
- PVXPPJIGRGXGCY-TZLCEDOOSA-N 6-O-alpha-D-glucopyranosyl-D-fructofuranose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1OC[C@@H]1[C@@H](O)[C@H](O)C(O)(CO)O1 PVXPPJIGRGXGCY-TZLCEDOOSA-N 0.000 claims description 2
- 239000004375 Dextrin Substances 0.000 claims description 2
- 229920001353 Dextrin Polymers 0.000 claims description 2
- 239000004368 Modified starch Substances 0.000 claims description 2
- 229920000881 Modified starch Polymers 0.000 claims description 2
- 229920001100 Polydextrose Polymers 0.000 claims description 2
- 235000019425 dextrin Nutrition 0.000 claims description 2
- 235000021552 granulated sugar Nutrition 0.000 claims description 2
- 235000019426 modified starch Nutrition 0.000 claims description 2
- 239000001259 polydextrose Substances 0.000 claims description 2
- 235000013856 polydextrose Nutrition 0.000 claims description 2
- 229940035035 polydextrose Drugs 0.000 claims description 2
- 238000009835 boiling Methods 0.000 claims 1
- 230000000415 inactivating effect Effects 0.000 claims 1
- 239000002245 particle Substances 0.000 abstract description 54
- 238000000576 coating method Methods 0.000 abstract description 27
- 239000011248 coating agent Substances 0.000 abstract description 25
- 230000008859 change Effects 0.000 abstract description 11
- 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 118
- 229940045997 vitamin a Drugs 0.000 description 80
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 73
- FPIPGXGPPPQFEQ-UHFFFAOYSA-N 13-cis retinol Natural products OCC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-UHFFFAOYSA-N 0.000 description 70
- FPIPGXGPPPQFEQ-BOOMUCAASA-N Vitamin A Natural products OC/C=C(/C)\C=C\C=C(\C)/C=C/C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-BOOMUCAASA-N 0.000 description 70
- FPIPGXGPPPQFEQ-OVSJKPMPSA-N all-trans-retinol Chemical compound OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-OVSJKPMPSA-N 0.000 description 70
- 235000019155 vitamin A Nutrition 0.000 description 70
- 239000011719 vitamin A Substances 0.000 description 70
- 239000003921 oil Substances 0.000 description 60
- 235000019198 oils Nutrition 0.000 description 60
- 108010076119 Caseins Proteins 0.000 description 59
- 102000011632 Caseins Human genes 0.000 description 59
- 229930003427 Vitamin E Natural products 0.000 description 59
- 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 59
- 235000019165 vitamin E Nutrition 0.000 description 59
- 229940046009 vitamin E Drugs 0.000 description 59
- 239000011709 vitamin E Substances 0.000 description 59
- 239000012071 phase Substances 0.000 description 54
- 230000000052 comparative effect Effects 0.000 description 52
- 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 51
- PFRQBZFETXBLTP-RCIYGOBDSA-N 2-[(2e,6e,10e,14e,18e)-3,7,11,15,19,23-hexamethyltetracosa-2,6,10,14,18,22-hexaen-1-yl]-3-methyl-1,4-dihydronaphthalene-1,4-dione Chemical compound C1=CC=C2C(=O)C(C/C=C(C)/CC/C=C(C)/CC/C=C(C)/CC/C=C(C)/CC/C=C(C)/CCC=C(C)C)=C(C)C(=O)C2=C1 PFRQBZFETXBLTP-RCIYGOBDSA-N 0.000 description 45
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 45
- QIVBCDIJIAJPQS-VIFPVBQESA-N L-tryptophane Chemical compound C1=CC=C2C(C[C@H](N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-VIFPVBQESA-N 0.000 description 42
- QIVBCDIJIAJPQS-UHFFFAOYSA-N Tryptophan Natural products C1=CC=C2C(CC(N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-UHFFFAOYSA-N 0.000 description 42
- 239000008213 purified water Substances 0.000 description 31
- 239000000047 product Substances 0.000 description 30
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 29
- 108090000765 processed proteins & peptides Proteins 0.000 description 29
- 102000035195 Peptidases Human genes 0.000 description 28
- 150000003904 phospholipids Chemical group 0.000 description 28
- 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 25
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 description 25
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 24
- 230000000694 effects Effects 0.000 description 23
- 239000000843 powder Substances 0.000 description 23
- 239000005862 Whey Substances 0.000 description 22
- 235000020238 sunflower seed Nutrition 0.000 description 21
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 20
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 19
- 235000019796 monopotassium phosphate Nutrition 0.000 description 19
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 19
- 229930003316 Vitamin D Natural products 0.000 description 18
- 235000019166 vitamin D Nutrition 0.000 description 18
- 239000011710 vitamin D Substances 0.000 description 18
- 150000003710 vitamin D derivatives Chemical class 0.000 description 18
- 229940046008 vitamin d Drugs 0.000 description 18
- 239000011772 phylloquinone Substances 0.000 description 17
- MBWXNTAXLNYFJB-LKUDQCMESA-N phylloquinone Chemical compound C1=CC=C2C(=O)C(C/C=C(C)/CCCC(C)CCCC(C)CCCC(C)C)=C(C)C(=O)C2=C1 MBWXNTAXLNYFJB-LKUDQCMESA-N 0.000 description 17
- 235000013336 milk Nutrition 0.000 description 16
- 239000008267 milk Substances 0.000 description 16
- 210000004080 milk Anatomy 0.000 description 16
- 239000000523 sample Substances 0.000 description 16
- 238000009826 distribution Methods 0.000 description 15
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 13
- 150000001413 amino acids Chemical class 0.000 description 13
- 235000016709 nutrition Nutrition 0.000 description 13
- 229940024606 amino acid Drugs 0.000 description 12
- 235000001014 amino acid Nutrition 0.000 description 12
- 239000007853 buffer solution Substances 0.000 description 12
- 229910000403 monosodium phosphate Inorganic materials 0.000 description 12
- 235000019799 monosodium phosphate Nutrition 0.000 description 12
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 12
- 235000013305 food Nutrition 0.000 description 11
- 238000010438 heat treatment Methods 0.000 description 11
- 230000035764 nutrition Effects 0.000 description 11
- 230000029087 digestion Effects 0.000 description 10
- 239000011780 sodium chloride Substances 0.000 description 10
- 235000002639 sodium chloride Nutrition 0.000 description 10
- 238000002347 injection Methods 0.000 description 9
- 239000007924 injection Substances 0.000 description 9
- 239000000463 material Substances 0.000 description 9
- 235000015097 nutrients Nutrition 0.000 description 9
- 238000010521 absorption reaction Methods 0.000 description 8
- 239000000084 colloidal system Substances 0.000 description 8
- 230000000887 hydrating effect Effects 0.000 description 8
- 239000003531 protein hydrolysate Substances 0.000 description 8
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 description 7
- 230000001133 acceleration Effects 0.000 description 7
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 7
- 230000005540 biological transmission Effects 0.000 description 7
- 239000011162 core material Substances 0.000 description 7
- 238000006911 enzymatic reaction Methods 0.000 description 7
- 235000013922 glutamic acid Nutrition 0.000 description 7
- 239000004220 glutamic acid Substances 0.000 description 7
- 238000010979 pH adjustment Methods 0.000 description 7
- -1 polyoxyethylene lauryl ether Polymers 0.000 description 7
- 235000005282 vitamin D3 Nutrition 0.000 description 7
- 239000011647 vitamin D3 Substances 0.000 description 7
- 229940021056 vitamin d3 Drugs 0.000 description 7
- OZDAOHVKBFBBMZ-UHFFFAOYSA-N 2-aminopentanedioic acid;hydrate Chemical compound O.OC(=O)C(N)CCC(O)=O OZDAOHVKBFBBMZ-UHFFFAOYSA-N 0.000 description 6
- 239000004475 Arginine Substances 0.000 description 6
- 239000004471 Glycine Substances 0.000 description 6
- AGPKZVBTJJNPAG-WHFBIAKZSA-N L-isoleucine Chemical compound CC[C@H](C)[C@H](N)C(O)=O AGPKZVBTJJNPAG-WHFBIAKZSA-N 0.000 description 6
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 6
- 239000004472 Lysine Substances 0.000 description 6
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 229930003448 Vitamin K Natural products 0.000 description 6
- 239000008346 aqueous phase Substances 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
- 239000003623 enhancer Substances 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 238000004108 freeze drying Methods 0.000 description 6
- 229960000310 isoleucine Drugs 0.000 description 6
- AGPKZVBTJJNPAG-UHFFFAOYSA-N isoleucine Natural products CCC(C)C(N)C(O)=O AGPKZVBTJJNPAG-UHFFFAOYSA-N 0.000 description 6
- 230000002503 metabolic effect Effects 0.000 description 6
- 238000003801 milling Methods 0.000 description 6
- SHUZOJHMOBOZST-UHFFFAOYSA-N phylloquinone Natural products CC(C)CCCCC(C)CCC(C)CCCC(=CCC1=C(C)C(=O)c2ccccc2C1=O)C SHUZOJHMOBOZST-UHFFFAOYSA-N 0.000 description 6
- 238000012545 processing Methods 0.000 description 6
- 210000002784 stomach Anatomy 0.000 description 6
- 235000019168 vitamin K Nutrition 0.000 description 6
- 239000011712 vitamin K Substances 0.000 description 6
- 150000003721 vitamin K derivatives Chemical class 0.000 description 6
- 229940046010 vitamin k Drugs 0.000 description 6
- 230000008901 benefit Effects 0.000 description 5
- 238000010586 diagram Methods 0.000 description 5
- 238000004090 dissolution Methods 0.000 description 5
- 238000000338 in vitro Methods 0.000 description 5
- 238000001727 in vivo Methods 0.000 description 5
- 238000004519 manufacturing process Methods 0.000 description 5
- 230000004060 metabolic process Effects 0.000 description 5
- 244000005700 microbiome Species 0.000 description 5
- 229920001184 polypeptide Polymers 0.000 description 5
- 102000004196 processed proteins & peptides Human genes 0.000 description 5
- MTCFGRXMJLQNBG-UHFFFAOYSA-N serine Chemical compound OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 5
- 238000004088 simulation Methods 0.000 description 5
- 239000007787 solid Substances 0.000 description 5
- 235000012424 soybean oil Nutrition 0.000 description 5
- 239000003549 soybean oil Substances 0.000 description 5
- 239000007921 spray Substances 0.000 description 5
- 238000005507 spraying Methods 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- 238000005303 weighing Methods 0.000 description 5
- GUOCOOQWZHQBJI-UHFFFAOYSA-N 4-oct-7-enoxy-4-oxobutanoic acid Chemical compound OC(=O)CCC(=O)OCCCCCCC=C GUOCOOQWZHQBJI-UHFFFAOYSA-N 0.000 description 4
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 4
- 229920001661 Chitosan Polymers 0.000 description 4
- ODKSFYDXXFIFQN-BYPYZUCNSA-P L-argininium(2+) Chemical compound NC(=[NH2+])NCCC[C@H]([NH3+])C(O)=O ODKSFYDXXFIFQN-BYPYZUCNSA-P 0.000 description 4
- HNDVDQJCIGZPNO-YFKPBYRVSA-N L-histidine Chemical compound OC(=O)[C@@H](N)CC1=CN=CN1 HNDVDQJCIGZPNO-YFKPBYRVSA-N 0.000 description 4
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 description 4
- KZSNJWFQEVHDMF-BYPYZUCNSA-N L-valine Chemical compound CC(C)[C@H](N)C(O)=O KZSNJWFQEVHDMF-BYPYZUCNSA-N 0.000 description 4
- 239000002202 Polyethylene glycol Substances 0.000 description 4
- KZSNJWFQEVHDMF-UHFFFAOYSA-N Valine Natural products CC(C)C(N)C(O)=O KZSNJWFQEVHDMF-UHFFFAOYSA-N 0.000 description 4
- 230000002776 aggregation Effects 0.000 description 4
- 235000008452 baby food Nutrition 0.000 description 4
- 239000008280 blood Substances 0.000 description 4
- 210000004369 blood Anatomy 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 4
- 239000013065 commercial product Substances 0.000 description 4
- 239000013078 crystal Substances 0.000 description 4
- 230000001804 emulsifying effect Effects 0.000 description 4
- 235000013373 food additive Nutrition 0.000 description 4
- 239000002778 food additive Substances 0.000 description 4
- 235000020189 fortified milk Nutrition 0.000 description 4
- 230000002496 gastric effect Effects 0.000 description 4
- 238000000265 homogenisation Methods 0.000 description 4
- 150000002632 lipids Chemical class 0.000 description 4
- 238000005259 measurement Methods 0.000 description 4
- 238000001000 micrograph Methods 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 210000000214 mouth Anatomy 0.000 description 4
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 4
- 229920001223 polyethylene glycol Polymers 0.000 description 4
- 230000001681 protective effect Effects 0.000 description 4
- 239000002994 raw material Substances 0.000 description 4
- 229940080237 sodium caseinate Drugs 0.000 description 4
- 229940080313 sodium starch Drugs 0.000 description 4
- 239000004474 valine Substances 0.000 description 4
- XUJNEKJLAYXESH-UHFFFAOYSA-N Cysteine Chemical compound SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- ZDXPYRJPNDTMRX-UHFFFAOYSA-N Glutamine Chemical compound OC(=O)C(N)CCC(N)=O ZDXPYRJPNDTMRX-UHFFFAOYSA-N 0.000 description 3
- 241000699670 Mus sp. Species 0.000 description 3
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Chemical compound OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 3
- 229930006000 Sucrose Natural products 0.000 description 3
- AYFVYJQAPQTCCC-UHFFFAOYSA-N THREONINE Chemical compound CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 description 3
- PFRQBZFETXBLTP-UHFFFAOYSA-N Vitamin K2 Natural products C1=CC=C2C(=O)C(CC=C(C)CCC=C(C)CCC=C(C)CCC=C(C)CCC=C(C)CCC=C(C)C)=C(C)C(=O)C2=C1 PFRQBZFETXBLTP-UHFFFAOYSA-N 0.000 description 3
- 230000009471 action Effects 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 230000037396 body weight Effects 0.000 description 3
- 239000000872 buffer Substances 0.000 description 3
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- 235000012000 cholesterol Nutrition 0.000 description 3
- 238000001514 detection method Methods 0.000 description 3
- 150000002148 esters Chemical class 0.000 description 3
- 210000004211 gastric acid Anatomy 0.000 description 3
- 210000001035 gastrointestinal tract Anatomy 0.000 description 3
- 239000004519 grease Substances 0.000 description 3
- 238000000227 grinding Methods 0.000 description 3
- 230000000968 intestinal effect Effects 0.000 description 3
- 235000021388 linseed oil Nutrition 0.000 description 3
- 239000000944 linseed oil Substances 0.000 description 3
- 238000001819 mass spectrum Methods 0.000 description 3
- 150000004667 medium chain fatty acids Chemical class 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- DKHGMERMDICWDU-GHDNBGIDSA-N menaquinone-4 Chemical compound C1=CC=C2C(=O)C(C/C=C(C)/CC/C=C(C)/CC/C=C(C)/CCC=C(C)C)=C(C)C(=O)C2=C1 DKHGMERMDICWDU-GHDNBGIDSA-N 0.000 description 3
- 239000003960 organic solvent Substances 0.000 description 3
- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Chemical compound OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 description 3
- 239000008055 phosphate buffer solution Substances 0.000 description 3
- 239000000758 substrate Substances 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 238000009210 therapy by ultrasound Methods 0.000 description 3
- 210000001519 tissue Anatomy 0.000 description 3
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 3
- MECHNRXZTMCUDQ-RKHKHRCZSA-N vitamin D2 Chemical compound C1(/[C@@H]2CC[C@@H]([C@]2(CCC1)C)[C@H](C)/C=C/[C@H](C)C(C)C)=C\C=C1\C[C@@H](O)CCC1=C MECHNRXZTMCUDQ-RKHKHRCZSA-N 0.000 description 3
- 235000019143 vitamin K2 Nutrition 0.000 description 3
- 239000011728 vitamin K2 Substances 0.000 description 3
- DCXYFEDJOCDNAF-UHFFFAOYSA-N Asparagine Natural products OC(=O)C(N)CC(N)=O DCXYFEDJOCDNAF-UHFFFAOYSA-N 0.000 description 2
- 108010004103 Chylomicrons Proteins 0.000 description 2
- 241000196324 Embryophyta Species 0.000 description 2
- DCXYFEDJOCDNAF-REOHCLBHSA-N L-asparagine Chemical compound OC(=O)[C@@H](N)CC(N)=O DCXYFEDJOCDNAF-REOHCLBHSA-N 0.000 description 2
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 description 2
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 2
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 2
- ROHFNLRQFUQHCH-UHFFFAOYSA-N Leucine Natural products CC(C)CC(N)C(O)=O ROHFNLRQFUQHCH-UHFFFAOYSA-N 0.000 description 2
- 239000004367 Lipase Substances 0.000 description 2
- 102000004882 Lipase Human genes 0.000 description 2
- 108090001060 Lipase Proteins 0.000 description 2
- 108010084695 Pea Proteins Proteins 0.000 description 2
- 229920002472 Starch Polymers 0.000 description 2
- 239000004473 Threonine Substances 0.000 description 2
- 230000002378 acidificating effect Effects 0.000 description 2
- 238000005054 agglomeration Methods 0.000 description 2
- 238000004220 aggregation Methods 0.000 description 2
- 125000000539 amino acid group Chemical group 0.000 description 2
- 235000009582 asparagine Nutrition 0.000 description 2
- 229960001230 asparagine Drugs 0.000 description 2
- 238000003556 assay Methods 0.000 description 2
- 239000003833 bile salt Substances 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000013329 compounding Methods 0.000 description 2
- 235000005687 corn oil Nutrition 0.000 description 2
- 239000002285 corn oil Substances 0.000 description 2
- 235000018417 cysteine Nutrition 0.000 description 2
- 230000003111 delayed effect Effects 0.000 description 2
- 235000014113 dietary fatty acids Nutrition 0.000 description 2
- 235000015872 dietary supplement Nutrition 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 239000000839 emulsion Substances 0.000 description 2
- 230000007613 environmental effect Effects 0.000 description 2
- 229930195729 fatty acid Natural products 0.000 description 2
- 239000000194 fatty acid Substances 0.000 description 2
- 239000007888 film coating Substances 0.000 description 2
- 238000009501 film coating Methods 0.000 description 2
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 2
- 230000006870 function Effects 0.000 description 2
- 239000008187 granular material Substances 0.000 description 2
- 238000009775 high-speed stirring Methods 0.000 description 2
- 230000006872 improvement Effects 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 235000019421 lipase Nutrition 0.000 description 2
- 210000004185 liver Anatomy 0.000 description 2
- 229910052751 metal Inorganic materials 0.000 description 2
- 239000002184 metal Substances 0.000 description 2
- 229930182817 methionine Natural products 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000007935 neutral effect Effects 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 235000019702 pea protein Nutrition 0.000 description 2
- 229940075999 phytosterol ester Drugs 0.000 description 2
- 235000002378 plant sterols Nutrition 0.000 description 2
- 238000003672 processing method Methods 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 229940032147 starch Drugs 0.000 description 2
- 239000008107 starch Substances 0.000 description 2
- 235000019698 starch Nutrition 0.000 description 2
- 238000003860 storage Methods 0.000 description 2
- 238000005728 strengthening Methods 0.000 description 2
- 239000005720 sucrose Substances 0.000 description 2
- 239000004094 surface-active agent Substances 0.000 description 2
- GETQZCLCWQTVFV-UHFFFAOYSA-N trimethylamine Chemical compound CN(C)C GETQZCLCWQTVFV-UHFFFAOYSA-N 0.000 description 2
- 239000012588 trypsin Substances 0.000 description 2
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 2
- JIAARYAFYJHUJI-UHFFFAOYSA-L zinc dichloride Chemical compound [Cl-].[Cl-].[Zn+2] JIAARYAFYJHUJI-UHFFFAOYSA-L 0.000 description 2
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 description 1
- KILNVBDSWZSGLL-KXQOOQHDSA-N 1,2-dihexadecanoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCCCCCCCCC KILNVBDSWZSGLL-KXQOOQHDSA-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
- GOJUJUVQIVIZAV-UHFFFAOYSA-N 2-amino-4,6-dichloropyrimidine-5-carbaldehyde Chemical group NC1=NC(Cl)=C(C=O)C(Cl)=N1 GOJUJUVQIVIZAV-UHFFFAOYSA-N 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
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 description 1
- 239000004382 Amylase Substances 0.000 description 1
- 102000013142 Amylases Human genes 0.000 description 1
- 108010065511 Amylases Proteins 0.000 description 1
- 239000000120 Artificial Saliva Substances 0.000 description 1
- 201000001320 Atherosclerosis Diseases 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- QIIDATRCGITYRZ-UHFFFAOYSA-N Catalpol Natural products OCC1OC(OC2OC=CC3C(O)C(=C(CO)C23)O)C(O)C(O)C1O QIIDATRCGITYRZ-UHFFFAOYSA-N 0.000 description 1
- 229920002261 Corn starch Polymers 0.000 description 1
- ZAKOWWREFLAJOT-CEFNRUSXSA-N D-alpha-tocopherylacetate Chemical compound CC(=O)OC1=C(C)C(C)=C2O[C@@](CCC[C@H](C)CCC[C@H](C)CCCC(C)C)(C)CCC2=C1C ZAKOWWREFLAJOT-CEFNRUSXSA-N 0.000 description 1
- 239000011626 DL-alpha-tocopherylacetate Substances 0.000 description 1
- 235000001809 DL-alpha-tocopherylacetate Nutrition 0.000 description 1
- 208000004232 Enteritis Diseases 0.000 description 1
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 description 1
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 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
- 240000007472 Leucaena leucocephala Species 0.000 description 1
- 235000010643 Leucaena leucocephala Nutrition 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 102000014171 Milk Proteins Human genes 0.000 description 1
- 108010011756 Milk Proteins Proteins 0.000 description 1
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 108010073771 Soybean Proteins Proteins 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
- 238000003917 TEM image Methods 0.000 description 1
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 description 1
- 208000010011 Vitamin A Deficiency Diseases 0.000 description 1
- 201000000839 Vitamin K Deficiency Bleeding Diseases 0.000 description 1
- 206010047634 Vitamin K deficiency Diseases 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 239000000853 adhesive Substances 0.000 description 1
- 230000001070 adhesive effect Effects 0.000 description 1
- 235000004279 alanine Nutrition 0.000 description 1
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 description 1
- 102000004139 alpha-Amylases Human genes 0.000 description 1
- 108090000637 alpha-Amylases Proteins 0.000 description 1
- 229940024171 alpha-amylase Drugs 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
- 235000011114 ammonium hydroxide Nutrition 0.000 description 1
- 235000019418 amylase Nutrition 0.000 description 1
- 229940040526 anhydrous sodium acetate Drugs 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000003012 bilayer membrane Substances 0.000 description 1
- 229940093761 bile salts Drugs 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 230000023555 blood coagulation Effects 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 210000001185 bone marrow Anatomy 0.000 description 1
- 230000002308 calcification Effects 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 108010079058 casein hydrolysate Proteins 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- LHDWRKICQLTVDL-PZYDOOQISA-N catalpol Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@H]1[C@@H]2[C@@]3(CO)O[C@H]3[C@@H](O)[C@@H]2C=CO1 LHDWRKICQLTVDL-PZYDOOQISA-N 0.000 description 1
- UXSACQOOWZMGSE-UHFFFAOYSA-N catalposide Natural products OC1C(O)C(O)C(CO)OC1OC1C2C3(CO)OC3C(OC(=O)C=3C=CC(O)=CC=3)C2C=CO1 UXSACQOOWZMGSE-UHFFFAOYSA-N 0.000 description 1
- 230000001055 chewing effect Effects 0.000 description 1
- 229940099352 cholate Drugs 0.000 description 1
- BHQCQFFYRZLCQQ-OELDTZBJSA-N cholic acid Chemical compound C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)[C@@H](O)C1 BHQCQFFYRZLCQQ-OELDTZBJSA-N 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 239000008120 corn starch Substances 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 230000002950 deficient Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 230000032798 delamination Effects 0.000 description 1
- 230000001079 digestive effect Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- KPPLIMDJGPOMAL-UHFFFAOYSA-K dipotassium;sodium;phosphate Chemical compound [Na+].[K+].[K+].[O-]P([O-])([O-])=O KPPLIMDJGPOMAL-UHFFFAOYSA-K 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 238000012377 drug delivery Methods 0.000 description 1
- 238000004043 dyeing Methods 0.000 description 1
- 235000006694 eating habits Nutrition 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 description 1
- GCFHZZWXZLABBL-UHFFFAOYSA-N ethanol;hexane Chemical compound CCO.CCCCCC GCFHZZWXZLABBL-UHFFFAOYSA-N 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 238000004880 explosion Methods 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 239000012527 feed solution Substances 0.000 description 1
- 239000011888 foil Substances 0.000 description 1
- 235000015203 fruit juice Nutrition 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 210000004051 gastric juice Anatomy 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 229920000669 heparin Polymers 0.000 description 1
- ZFGMDIBRIDKWMY-PASTXAENSA-N heparin Chemical compound CC(O)=N[C@@H]1[C@@H](O)[C@H](O)[C@@H](COS(O)(=O)=O)O[C@@H]1O[C@@H]1[C@@H](C(O)=O)O[C@@H](O[C@H]2[C@@H]([C@@H](OS(O)(=O)=O)[C@@H](O[C@@H]3[C@@H](OC(O)[C@H](OS(O)(=O)=O)[C@H]3O)C(O)=O)O[C@@H]2O)CS(O)(=O)=O)[C@H](O)[C@H]1O ZFGMDIBRIDKWMY-PASTXAENSA-N 0.000 description 1
- 229960001008 heparin sodium Drugs 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 229960000443 hydrochloric acid Drugs 0.000 description 1
- 230000036512 infertility Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 210000000936 intestine Anatomy 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 238000003973 irrigation Methods 0.000 description 1
- 230000002262 irrigation Effects 0.000 description 1
- 235000015110 jellies Nutrition 0.000 description 1
- 239000008274 jelly Substances 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 235000019626 lipase activity Nutrition 0.000 description 1
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 1
- 235000021056 liquid food Nutrition 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
- 210000004072 lung Anatomy 0.000 description 1
- 125000003588 lysine group Chemical group [H]N([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])(N([H])[H])C(*)=O 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 235000019690 meat sausages Nutrition 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 description 1
- LHDWRKICQLTVDL-UHFFFAOYSA-N methyl iridoid glycoside Natural products OC1C(O)C(O)C(CO)OC1OC1C2C3(CO)OC3C(O)C2C=CO1 LHDWRKICQLTVDL-UHFFFAOYSA-N 0.000 description 1
- 239000003094 microcapsule Substances 0.000 description 1
- 235000021239 milk protein Nutrition 0.000 description 1
- 235000003715 nutritional status Nutrition 0.000 description 1
- ZPIRTVJRHUMMOI-UHFFFAOYSA-N octoxybenzene Chemical compound CCCCCCCCOC1=CC=CC=C1 ZPIRTVJRHUMMOI-UHFFFAOYSA-N 0.000 description 1
- 125000002811 oleoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 230000002572 peristaltic effect Effects 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- WTJKGGKOPKCXLL-RRHRGVEJSA-N phosphatidylcholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCC=CCCCCCCCC WTJKGGKOPKCXLL-RRHRGVEJSA-N 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 229920000259 polyoxyethylene lauryl ether Polymers 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 230000002035 prolonged effect Effects 0.000 description 1
- 239000003223 protective agent Substances 0.000 description 1
- 230000007065 protein hydrolysis Effects 0.000 description 1
- 230000002797 proteolythic effect Effects 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 229960000342 retinol acetate Drugs 0.000 description 1
- QGNJRVVDBSJHIZ-QHLGVNSISA-N retinyl acetate Chemical compound CC(=O)OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C QGNJRVVDBSJHIZ-QHLGVNSISA-N 0.000 description 1
- 235000019173 retinyl acetate Nutrition 0.000 description 1
- 239000011770 retinyl acetate Substances 0.000 description 1
- 210000003296 saliva Anatomy 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000001878 scanning electron micrograph Methods 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- 239000002002 slurry Substances 0.000 description 1
- 210000000813 small intestine Anatomy 0.000 description 1
- 239000004402 sodium ethyl p-hydroxybenzoate Substances 0.000 description 1
- 235000010226 sodium ethyl p-hydroxybenzoate Nutrition 0.000 description 1
- QYNMSPKSYXPZHG-UHFFFAOYSA-M sodium;4-ethoxycarbonylphenolate Chemical compound [Na+].CCOC(=O)C1=CC=C([O-])C=C1 QYNMSPKSYXPZHG-UHFFFAOYSA-M 0.000 description 1
- 239000012265 solid product Substances 0.000 description 1
- 229940001941 soy protein Drugs 0.000 description 1
- 229940071440 soy protein isolate Drugs 0.000 description 1
- 239000008347 soybean phospholipid Substances 0.000 description 1
- 210000000952 spleen Anatomy 0.000 description 1
- 230000001502 supplementing effect Effects 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 230000009747 swallowing Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000004885 tandem mass spectrometry Methods 0.000 description 1
- 239000002562 thickening agent Substances 0.000 description 1
- 230000001550 time effect Effects 0.000 description 1
- 229940042585 tocopherol acetate Drugs 0.000 description 1
- 239000002691 unilamellar liposome Substances 0.000 description 1
- 208000016794 vitamin K deficiency hemorrhagic disease Diseases 0.000 description 1
- 238000000733 zeta-potential measurement Methods 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
- 235000005074 zinc chloride Nutrition 0.000 description 1
- 239000011592 zinc chloride Substances 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/5005—Wall or coating material
- A61K9/5021—Organic macromolecular compounds
- A61K9/5052—Proteins, e.g. albumin
-
- 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
- A61K31/045—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates
- A61K31/07—Retinol compounds, e.g. vitamin A
-
- 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
- A61K31/12—Ketones
- A61K31/122—Ketones having the oxygen directly attached to a ring, e.g. quinones, vitamin K1, anthralin
-
- 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
- A61K31/33—Heterocyclic compounds
- A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
- A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
- A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline
- A61K31/353—3,4-Dihydrobenzopyrans, e.g. chroman, catechin
- A61K31/355—Tocopherols, e.g. vitamin E
-
- 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
- A61K31/59—Compounds containing 9, 10- seco- cyclopenta[a]hydrophenanthrene ring systems
- A61K31/592—9,10-Secoergostane derivatives, e.g. ergocalciferol, i.e. vitamin D2
-
- 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
- A61K31/59—Compounds containing 9, 10- seco- cyclopenta[a]hydrophenanthrene ring systems
- A61K31/593—9,10-Secocholestane derivatives, e.g. cholecalciferol, i.e. vitamin D3
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
- A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/10—Dispersions; Emulsions
- A61K9/127—Liposomes
- A61K9/1277—Processes for preparing; Proliposomes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
- A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
- A61K9/5089—Processes
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/02—Nutrients, e.g. vitamins, minerals
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P60/00—Technologies relating to agriculture, livestock or agroalimentary industries
- Y02P60/80—Food processing, e.g. use of renewable energies or variable speed drives in handling, conveying or stacking
- Y02P60/87—Re-use of by-products of food processing for fodder production
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Epidemiology (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Obesity (AREA)
- Organic Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Hematology (AREA)
- Diabetes (AREA)
- Biochemistry (AREA)
- Molecular Biology (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Nutrition Science (AREA)
- Inorganic Chemistry (AREA)
- Dispersion Chemistry (AREA)
- Medicinal Preparation (AREA)
Abstract
The invention discloses a fat-soluble vitamin liposome precursor, a preparation method and application thereof. The method comprises the steps of coating liposome by adopting protease hydrolysate in the process of preparing liposome precursor from liposome; comprising the following steps: regulating pH of protein-protease hydrolysate, mixing with liposome, adding filler, stirring for dissolving, and homogenizing to obtain feed liquid; and drying the feed liquid by adopting spray drying to obtain liposome precursors. The liposome precursor prepared by the invention has low loss rate in the preparation process, high encapsulation efficiency, improved stability after rehydration of the liposome precursor and small particle size change after rehydration.
Description
Technical Field
The invention relates to the technical field of liposome drug delivery, in particular to a fat-soluble vitamin liposome precursor, a preparation method and application thereof.
Background
The fat-soluble vitamins include vitamin A, vitamin D 2, vitamin D 3, vitamin E, vitamin K 1, and vitamin K 2, and are susceptible to attenuation by light, heat, oxygen, and ions. Its absorption is closely related to lipids in the intestinal tract. Fat-soluble vitamins have interactions in vivo, for example, vitamin E can promote the storage of vitamin A in liver, and intake of high dose vitamin E can interfere with the absorption of vitamin K and antagonize the function of vitamin K. Fat-soluble vitamins are commonly deficient due to nutritional status and eating habits (vegetarian) and the like, and are often supplemented together, such as vitamin a and vitamin K deficiency for children. Therefore, fat-soluble vitamins are often added to various foods in the form of nutritional supplements, and there is little individual fortification of one of the fat-soluble vitamins. Most of the currently marketed fat-soluble nutrition enhancers are nutrition enhancer powder or granules of single fat-soluble vitamins, such as vitamin A granules and vitamin E powder, and the fat-soluble vitamins are required to be added after being weighed independently or after being premixed, so that the steps are complicated.
The liposome is a bilayer vesicle composed of lipid molecules, each layer is a phospholipid bilayer, water phases are arranged between the layers and at the core, and fat-soluble components are embedded in the phospholipid bilayer. The liposome has a cell membrane-like structure, good biocompatibility, high safety and low toxicity, has a certain slow release effect, can change pharmacokinetics and tissue distribution after entering a body, can lead the core material to be passively targeted to tissues such as liver, spleen, lung, bone marrow and the like, can also form a reservoir in circulation and tissues, and can slowly release the core material. The liposome is widely applied to the fields of medicines, cosmetics, skin care products and the like at present. However, liposomes are currently used less in nutritional supplements because: at present, organic solvents are mostly used in the process of preparing liposome. In order to obtain better stability and processing tolerance or smaller particle size, surfactants such as oleoyl trimethylamine cholesterol, polyoxyethylene lauryl ether, polyethylene glycol octyl phenyl ether and the like are often used, artificial modified phospholipids such as dioleoyl propyl dioleoyl phosphatidylethanolamine, dipalmitoyl phosphatidylcholine and the like are often used for preparation, and liposome coatings such as chitosan, polyethylene glycol and the like are often used. The chitosan and the polyethylene glycol belong to food additives, but the application range does not comprise food types which are daily ingested by children and old people, for example, the application range of the chitosan is a film coating agent and a thickening agent of western ham and meat sausage, and the application range of the polyethylene glycol is a film coating agent of candy; the rest substances are not food or food additives, and cannot be used in food. Infants and preschool children have vigorous metabolic demands, while the elderly have reduced absorption capacity due to swallowing and chewing disorders, and are all people with easily lacking fat-soluble nutrients. Milk powder and food with milk protein as main raw material are good nutrition supplementing carrier. Some documents attempt to replace the above substances by other reagents, but the application scope is still limited, or the products are not suitable for infant foods, or the elderly foods, or the products do not comprise milk powder, and are not suitable for enteritis and gastrointestinal tract sensitive people, etc.
Furthermore, liposomes are thermodynamically unstable systems, which, because they are usually present in suspension, are physically and chemically unstable, such as agglomeration, sedimentation, degradation, phospholipid hydrolysis or oxidation, and the like, further limiting the use of liposomes. Liposome precursors are defined as solid, powdered particles that can be used as drug carriers and which can rapidly hydrate in water to form liposomes. Compared with liposome, the liposome precursor has obviously improved physical and chemical stability. At present, liposome is prepared into liposome precursor by adopting spray drying or freeze drying and other processes, and the liposome precursor is influenced by heat and ice crystal formation, so that the problems of high loss of a liposome precursor core material, easiness in demulsification and the like exist.
Disclosure of Invention
According to one embodiment of the invention, the invention aims to provide a fat-soluble vitamin liposome precursor, a preparation method and application thereof, so as to improve encapsulation efficiency, improve stability after rehydration and maintain particle size before spraying. The above object can be achieved by the following embodiments of the present invention:
According to one aspect of the invention, the invention provides a preparation method of a liposome precursor of fat-soluble vitamins, wherein protease hydrolysate is adopted to coat the liposome in the process of preparing the liposome precursor of the liposome; comprising the following steps: regulating the pH of the protein-protease hydrolysate to 7.0-7.5, mixing the protein-protease hydrolysate with liposome, adding a filler, stirring and dissolving, and homogenizing to obtain a feed liquid; and drying the feed liquid by adopting spray drying to obtain liposome precursors.
According to another aspect of the invention, the fat-soluble vitamin liposome precursor is prepared by adopting the preparation method of the fat-soluble vitamin liposome precursor.
According to still another aspect of the present invention, the present invention provides the use of a lipid-soluble vitamin liposome precursor in a liquid product, wherein the lipid-soluble vitamin liposome precursor is prepared by the preparation method.
The beneficial effects are that: according to one embodiment of the invention, the liposome is coated by protease hydrolysate in the process of preparing liposome precursor by liposome, and the liposome is mixed with the protease hydrolysate after the pH of the protease-protease hydrolysate is adjusted to 7.0-7.5, and the liposome is added with filler, stirred, dissolved and homogenized to form a feed liquid; the liposome precursor is obtained by drying the feed liquid through spray drying, and has the advantages of high encapsulation rate, stable particle size and stable core material, the stability of the prepared liposome precursor is further improved, and the particle size change is small after rehydration. The following advantages are also provided in some embodiments of the present invention over the prior art:
1) The pH of the protein-protease hydrolysate is adjusted to 7.0-7.5, so that the hydrolysate and the liposome are mixed at the pH, and the liposome has better protection effect. The liposome itself becomes less stable under acidic conditions, and when the pH is higher than 7.5, the coating effect becomes less, and the obtained product has a larger variation in particle size.
2) The method adopts the step-by-step adjustment of the pH of the protein-protease hydrolysate, solves the problems that the existing solution changes rapidly, the overstock is easy to appear, the buffer capacity for acid-base change is avoided, and the like under the condition that the pH is adjusted to the range by using the reagent alone.
3) Under the pH condition, the liposome is coated by pepsin or papain hydrolysate, so that the optimal coating effect can be obtained, the particle size distribution is narrow after the subsequent spray drying, the encapsulation rate of liposome precursors is high and can reach more than 95%, the liposome precursors are placed for a long time after rehydration without agglomeration phenomenon, and a transmission electron microscope image shows no rupture after rehydration, so that the usable humidity range is increased. Especially when applied to liquid products, the vitamin has better stability compared with the commercial products and liposomes, greatly reduces the loss rate of the vitamin and expands the applicable range.
4) The liposome is prepared from the fat-soluble vitamins, and then the liposome is coated by adopting protein-protease hydrolysate, so that the spray drying preparation of liposome precursors is realized, and the problems of easiness in demulsification and high heat loss rate in the prior art are solved. The vitamin loss rate of the prepared precursor after rehydration is reduced compared with that of uncoated liposome. Compared with freeze drying, the method has the advantages of low energy consumption, short production period, little influence of environmental humidity on the spray drying effect, and wide preparation application humidity range.
5) The temperature and time of tolerance are obviously improved, the liposome can only resist pasteurization before coating, and the UHT sterilization process can be resisted after coating, so that the product can resist the processing heat treatment process of most foods.
6) The raw materials and the auxiliary materials are food-grade, and can be safely, conveniently and conveniently applied to various foods including infant foods. The method does not use an organic solvent, does not have the risk of organic solvent residue, does not need to remove the solvent in the subsequent step, and has low requirements on workshop explosion prevention and the like; the use of no cholesterol or other non-food related regulations allows the use of surfactants or coatings; the food additive with smaller application range such as Tween, chitosan and the like is not used, and the food additive can be safely and limitlessly applied to various foods.
7) Has better slow release effect and higher biological absorption and utilization rate, in particular to casein-gastric protein hydrolysate precursor.
8) The liposome contains two or more fat-soluble nutrients, and the weighing steps are reduced during application, so that the liposome is convenient to use.
Drawings
FIGS. 1-1 to 1-4 are graphs showing the loss rates of four vitamins in each of the comparative vitamin decay tests, respectively; and comprises two states of solution and powder, and two conditions of normal temperature and acceleration.
Figures 2-1 and 2-2 are graphs comparing the attenuation of four vitamins in each case added to a nutritional fortified milk powder product under wet and dry processes, respectively.
Figure 3 is a graph of the attenuation of four vitamins in each instance of a nutritional fortified milk powder product, respectively.
Figures 4-1 to 4-4 are graphs comparing the vitamin release rates of four vitamins in each of the in vitro metabolism comparison tests, respectively.
FIG. 5-1 is a liposome transmission electron microscope image of example 6; FIGS. 5-2 and 5-3 are transmission electron micrographs of example 6 after rehydration of the liposome precursors; FIGS. 5-4 are scanning electron micrographs of the liposome precursors of example 6.
FIG. 6 is a schematic diagram of the principle of the coating of liposomes with a proteolytic liquid.
FIGS. 7-1 to 7-7 are particle size distribution diagrams of typical liposomes, examples 6, 3,4, 5, and comparative examples 21, 28, respectively, after rehydration of the liposome precursors;
FIG. 8 is a plot of zeta potential versus pH for the liposome precursors of examples 3-6.
Detailed Description
The technical solutions of the present invention will be clearly and completely described in conjunction with the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, but not all embodiments. The following description of at least one exemplary embodiment is merely exemplary in nature and is in no way intended to limit the invention, its application, or uses. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
As previously described, the present inventors recognized that: compared with liposome, liposome precursor has advantages in stability and other aspects, but liposome precursor obtained by the prior liposome precursor preparation process can not ensure that the original particle size is still maintained or the loss rate of core material is high after rehydration. The preparation method of the fat-soluble vitamin liposome precursor provided by the application comprises the steps of coating the liposome by adopting protease hydrolysate in the process of preparing the liposome precursor from the liposome, regulating the pH of protein-protease hydrolysate, mixing the protein-protease hydrolysate with the liposome, adding a filler, stirring, dissolving and homogenizing to obtain a feed liquid; and then the feed liquid is dried by spray drying, so that the obtained liposome precursor has high encapsulation efficiency, little heat loss, small particle size change after rehydration, high stability and reduced vitamin loss rate; overcomes the limitation that the prior liposome precursor can not be prepared by spray drying.
In the invention, the fat-soluble vitamin is firstly prepared into liposome, and then the liposome is coated by adopting protein-protease hydrolysate. The invention uses phospholipids of natural origin to prepare liposomes which are negatively charged at said pH conditions and which can be coated with protease hydrolysates to form a protective effect at said pH conditions.
In an alternative embodiment of the invention, the oil phase is injected into the water phase by adopting an injection method under a high-speed stirring state, and the liposome is prepared by stirring and hydrating and then high-pressure micro-jet, in particular to the liposome containing two or more fat-soluble nutrients and compound vitamins. Specifically, the method may include the steps of: 1) Weighing fat-soluble vitamins, phospholipids and grease (or plant catalpol), grinding for 8-12 min, for example 10min, to obtain oil phase. 2) Dissolving dipotassium hydrogen phosphate and potassium dihydrogen phosphate or disodium hydrogen phosphate and sodium dihydrogen phosphate and a certain amount of sodium chloride in purified water to prepare a buffer solution with pH of 7.0-7.5, and preparing a water phase. 3) The oil phase is put into a colloid mill for cyclic grinding, and then is rapidly injected into a water phase, namely phosphate buffer solution by using a syringe or compressed air under the high-speed stirring state, such as the stirring slurry rotating speed of 400r/min to 800r/min, for example 600r/min, and is stirred and hydrated at the temperature of 60 ℃ to 70 ℃, for example 65 ℃ for 15min to 30min, for example 30min, so as to form the primary emulsion. 4) The primary emulsion is subjected to high-pressure micro-jet, the micro-jet pressure is 70 to 110mPa, for example 90mPa, and the micro-jet times are 3 to 5 times, so that the vitamin complex liposome is prepared.
Wherein the fat-soluble vitamins can be vitamin A, vitamin D (vitamin D 2, vitamin D 3), vitamin E, and vitamin K (vitamin K 1, vitamin K 2). In addition, at least two of the vitamin complex liposome can be selected, so that the vitamin complex liposome can be obtained, corresponding precursors can be obtained, the weighing steps in application are reduced, and the use is more convenient. In addition, when at least two vitamins are selected for compounding, the mass ratio may be in accordance with vitamin a: vitamin D: vitamin E: vitamin K 1: vitamin K 2 is 1 (0.015-0.030), 4-10, 0.006-0.05 and 0.05-0.1, and the vitamin A and vitamin K 2 are compounded according to 1: (0.05-0.1) compounding. Vitamin D 2 and vitamin D 3 have the same physiological functions and are regarded as the same vitamin; vitamin K 1 is mainly used for regulating the synthesis of blood coagulation proteins, and vitamin K 2 is mainly used for regulating calcification of bone tissues and has different functions, so that two vitamins are considered, vitamin K 1 is mainly reinforced in infant foods, and vitamin K 2 is mainly reinforced in teenagers, children and elderly foods. The water phase can be a prepared phosphate buffer solution, wherein the total ionic strength of phosphate in the phosphate buffer solution is 0.05-0.15 mmol/L, and the ionic strength of sodium chloride is 0.025-0.1 mol/L. In addition, the ratio of the oil phase to the phosphate buffer may be 4.5 to 9:1. The phospholipid is phospholipid with phosphatidylcholine content more than or equal to 80%. In this embodiment, soybean phospholipid is used. The mass ratio of the phospholipid to the fat-soluble nutrient (calculated by taking vitamin A as a reference) is as follows: 40-70:1. The grease comprises: one or more of medium chain fatty acid triglyceride, sunflower seed oil, soybean oil, corn oil and linseed oil. The mass ratio of the grease to the fat-soluble nutrient (calculated by taking the vitamin A as the reference) is as follows: 60-120:1. In addition, medium chain fatty acid triglycerides can only be used without vitamin D, and when vitamin D is added, one or more of sunflower seed oil, soybean oil, and linseed oil are used as oils. When the crowd is middle-aged and elderly people, the plant Zialcohol ester can be added during grinding to prepare oil phase. The plant sterol ester has a structure similar to cholesterol, is favorable for maintaining the stability of a liposome double-layer membrane structure to a certain extent, has a certain auxiliary effect on reducing blood fat and preventing atherosclerosis, and can not be used for infant foods, so that the plant sterol ester is only selected to be reinforced in products of which the target population is middle-aged and elderly people. In the case of selective fortification, the phytosterol esters: fat-soluble nutrient (calculated by taking vitamin A as a reference) 2-5:1.
In the present invention, the protein-protease hydrolysate is obtained by adding a protease to a protein solution and hydrolyzing at the protease hydrolysis temperature. In an alternative embodiment, the preparation of the protein-protease hydrolysate comprises: 1) Dissolving protein in hydrochloric acid or phosphate solution with proper pH for enzymolysis to obtain protein solution, specifically, firstly adopting hydrochloric acid or phosphate to regulate pH of purified water to proper pH for enzymolysis, weighing protein and dissolving in water to prepare 2% -3% w/w solution, for example, protein concentration is 2.5%. 2) Adding a protease to the protein solution and hydrolyzing at the protease hydrolysis temperature. Furthermore, before this step, it further comprises: the concentration of protease and the hydrolysis time were determined according to the kind of protein. In the embodiment, the preparation and hydrolysis conditions, such as enzymolysis temperature, pH, substrate concentration (i.e. protein concentration) and the like, are further controlled according to the protease or the protein, the obtained hydrolysate has better chain length and configuration, the liposome can be effectively protected, the obtained liposome precursor has high encapsulation rate, small particle size change after rehydration and improved stability.
In a preferred embodiment, the protease may be one or more of pepsin and papain. Wherein, the pH of the purified water can be adjusted to 1.0-1.8 by hydrochloric acid, and the pepsin enzymolysis is suitable for pH 1.0-1.8. The enzymolysis of papain is suitable for pH 6-7, and the pH of the purified water can be adjusted to 6-7 by using dipotassium hydrogen phosphate and potassium dihydrogen phosphate or disodium hydrogen phosphate and sodium dihydrogen phosphate. In addition, the hydrolysis temperature of pepsin is 35-40 ℃; the hydrolysis temperature of papain is 50-60 ℃. The pepsin activity unit is 3000 NF.U/g, and the papain activity unit is 10 ten thousand U/g. Further, the protein may be one or more of casein, concentrated whey protein. Wherein, the protease concentration of the casein is 0.2-0.3%, the preferable enzyme concentration is 0.3%, the hydrolysis time is 20-60 min, and the preferable hydrolysis time is 30min; the protease concentration of the concentrated whey protein is 0.1% -0.3%, the preferable enzyme concentration is 0.3%, the hydrolysis time is 10-40 min, and the preferable hydrolysis time is 30min. In the preferred embodiment, the particle size distribution of the liposome precursor after rehydration is narrow, the encapsulation efficiency is higher and can reach more than 95%, the liposome precursor is free from aggregation phenomenon after long-term placement, the transmission electron microscope image shows no rupture after rehydration, and the liposome precursor has better stability compared with the commercial products and the liposome when applied to liquid products, the vitamin loss rate after rehydration is greatly reduced, and the applicable range is enlarged.
Before the protein-protease hydrolysate is mixed with liposome, the pH of the protein-protease hydrolysate is adjusted after the hydrolysis is completed. Further, the pH of the protein-protease hydrolysate is adjusted to be near neutral, specifically to be 7.0 to 7.5, for example, to be 7.1, 7.2, 7.3, 7.4, 7.5, etc., under the condition that basic amino acids such as lysine, arginine, histidine, etc. in the protease hydrolysate peptide chain are positively charged, the protease hydrolysate peptide chain forms a positively charged region, and the protease hydrolysate peptide chain is combined with negatively charged liposome to form a coating, thereby protecting the liposome. In addition, after the pH adjustment, the solution needs to be boiled and deactivated to prevent continuous hydrolysis.
In a preferred embodiment, the step-wise adjustment of the pH of the protein-protease hydrolysate to near neutral comprises: the pH of the protein-protease hydrolysate is adjusted to 6.6-6.8, and then adjusted to 7.0-7.5. The pH regulator may be sodium hydroxide, dipotassium hydrogen phosphate, disodium hydrogen phosphate, etc. In particular when the protease is pepsin, the inventors noted that: the pH value is regulated by singly using sodium hydroxide, the solution changes rapidly, the over-head is easy to appear, and the buffer capacity for acid-base change is avoided; thus, the pH was adjusted in two steps using sodium hydroxide and dipotassium (sodium) phosphate.
According to the invention, the protein-protease hydrolysate with the pH adjusted is mixed with the liposome, the filler is added, the mixture is stirred and homogenized to obtain the feed liquid, and then the feed liquid is subjected to spray drying by using a spray drying tower to prepare the liposome precursor, so that the liposome precursor is coated, and meanwhile, the liposome precursor can be prepared by adopting a spray drying process based on the protection effect, so that the limitation in the prior art is overcome.
In an alternative embodiment, to enhance the coating effect of the protease hydrolysate on the liposome, the protease hydrolysate and the liposome are mixed in a mass ratio of 4-6:1 (the mass ratio of the polypeptide to the phospholipid is 3.2-5.625:1), for example, preferably 5:1. Further, the two materials are mixed and stirred for 5 to 10 minutes, the filling agent is added and stirred for 5 to 10 minutes for dissolution, a homogenizer is adopted for homogenization after dissolution, and when the homogenizer is adopted for homogenization, the homogenization pressure is 30 to 50mPa, and the homogenization times are 2 to 4 times. Furthermore, for the spray drying tower, the air inlet temperature can be controlled to be 90-110 ℃ according to different environmental humidity, and the air inlet frequency and the air flow rate can be controlled to ensure that the air outlet temperature is 50-70 ℃. In addition, the filler can be one or two of maltodextrin, white granulated sugar, isomaltulose, modified starch, polydextrose and resistant dextrin. The filler is mainly used for adjusting the solid content of the spray feed liquid, the solid content of the feed liquid is controlled to be 15% -20%, and the proportion of the filler to the liposome is 0.45-0.90:1 correspondingly. In the embodiment, the coating effect of the protein hydrolysate on the liposome can be improved, the stability can be improved, the particle size change after rehydration is small, the applicable humidity range is wide, and spray drying can be adopted by controlling the proportion of the protein-enzyme hydrolysate and the liposome and optimizing the homogenizing and spray drying conditions.
In an alternative embodiment, the feed solution may optionally be pasteurized or UHT sterilized prior to spray drying. Wherein the pasteurization conditions: the temperature is 65-85 ℃ and the time is 10-30 min. UHT sterilization conditions: the temperature is 125-135 ℃ and the time is 5-15 s. In this embodiment, the liposome precursors are significantly more resistant to temperature and time, and still have better stability after sterilization.
The invention is further illustrated, but should not be construed as being limited, by the following examples.
The implementation materials are as follows: vitamin a (retinol acetate, crystals), vitamin D 3 (cholecalciferol, crystals), vitamin E (dl-alpha-tocopherol acetate, oil), vitamin K 1 (phytomenaquinone, oil), vitamin K 2 (menatemenaquinone, 5% oil), phospholipids (PC > 85%), medium chain fatty acid triglycerides, sunflower seed oil, soybean oil, corn oil, linseed oil, phytosterol esters, dipotassium hydrogen phosphate, potassium dihydrogen phosphate, sodium chloride, casein, concentrated whey protein, pepsin, papain, hydrochloric acid, water.
The implementation effect evaluation method comprises the following steps: determination of vitamins in liposomes. Wherein, the determination of vitamin A and vitamin E: the detection wavelength is determined by GB 5009.82, a C 30 chromatographic column is used, a mobile phase is a water and methanol gradient system, the column temperature is 20 ℃, the flow rate is 0.8 mL/min: vitamin A is 325nm, vitamin E is 294nm, and the sample injection amount is 10 mu L. Determination of vitamin D: using a C 18 chromatographic column, the mobile phase was methanol-water solution (ratio 95:5) using GB 5009.82 assay, column temperature 35 ℃, flow rate 1mL/min, detection wavelength: 264nm, and the sample injection amount is 100 mu L. Determination of vitamin K 1: using a C 18 chromatographic column and a zinc reduction column according to GB 5009.158, mixing the above materials uniformly, adding 1.5g of zinc chloride, 0.5g of anhydrous sodium acetate, preparing the mixture at a flow rate of 1mL/min, and detecting the wavelength: excitation wavelength 243nm, emission wavelength 430nm, and sample injection amount 10 μl. Determination of vitamin K 2: the wavelength was measured by the method described in bulletin 8 of Weijie, 2016, using a C 18 column with a flow of methanol at a flow rate of 1 mL/min: 254nm, and the sample injection amount is 10 mu L. Determination of free vitamins: taking 0.1g of liposome or liposome precursor, adding 1.2mL of n-hexane, mixing for 10s by vortex vibration, centrifuging for 10min by 12000r/min, sucking 200 mu L of n-hexane on the upper layer, drying by nitrogen, adding absolute ethyl alcohol to a volume of 1mL, and carrying out sample injection measurement.
Determination of the total content: wherein, liposome: taking 0.2mL of liposome, adding an ethanol-n-hexane solvent (v: v=3:1), carrying out ultrasonic treatment for 15min, drying by nitrogen, adding absolute ethanol to a volume of 1mL, and carrying out sample injection measurement. Liposome precursors: taking 0.5mL of liposome precursor, adding 4mL of 10% ammonia water solution, performing ultrasonic treatment at 50 ℃ for 15min, adding absolute ethyl alcohol to fix the volume to 20mL, performing ultrasonic treatment at 50 ℃ for 15min, and performing sample injection measurement.
Calculation of encapsulation efficiency: the encapsulation efficiency of each fat-soluble vitamin was calculated separately as follows:
Calculation of loss rate: the decay rate of each fat-soluble vitamin was calculated as follows:
In vitro digestion experiments:
1. Preparing simulated digestive juice: simulating saliva: the pH6.8 solution is prepared from sodium dihydrogen phosphate, disodium hydrogen phosphate or potassium dihydrogen phosphate, disodium hydrogen phosphate and purified water. Simulating gastric juice: 2g of NaCl and 7mLHCl (concentration: 12 mol/L) were mixed, dissolved in a suitable amount of water and brought to a volume of 1L, and the pH was adjusted to 1.2. Simulation of intestinal juice: weighing 6.8gKH 2PO4, adding a proper amount of water for dissolution, then adding a proper amount of 0.5mol/L NaOH solution to enable the pH value of the buffer solution to be approximately equal to 7.0, and adding water to fix the volume to 1L.
2. In vitro digestion simulation experiment: oral digestion simulation: 10mL of the test substance and 10mL of artificial saliva are mixed, 0.3g of alpha-amylase (4000 IU/g) is added, the mixture is poured into a beaker, the mixture is incubated for 10min at 37 ℃, 0.2mL of sample is taken, and the release amount of vitamins is detected according to the method for measuring free vitamins in the encapsulation efficiency. Gastric digestion simulation: after digestion and incubation for 10min in the oral cavity, 10mL of simulated gastric buffer is added, the pH is adjusted to 2.0, 3mg of pepsin (20000 IU/g) is added, incubation is carried out at 37 ℃, samples are taken at 15min, 30min, 45min, 60min, 75min, 90min, 105min and 120min respectively, and the release amount of vitamins is detected according to the method for measuring the free vitamins in the encapsulation rate. Intestinal digestion simulation: continuously adding 10mL of simulated intestinal buffer solution, 4mL of cholate buffer solution (46 mg/mL) and 1mL of CaCl 2 buffer solution (110 mg/mL), adjusting the pH to 7.0 (0.5 mol/L NaOH solution), adding 2.5mL of lipase buffer solution (2 mg/100mL, 4000IU/mg of lipase activity), incubating at 37 ℃, sampling at 15min, 30min, 45min, 60min, 75min, 90min, 105min and 120min respectively, and detecting the release amount of vitamins.
3. In vivo digestion experiments: 250 adult mice (ICR) were randomly divided into 3 groups (80 mice/group, 10 mice reserved for blank blood collection and spare) and housed in separate cages. The experiment is fasted for 12 hours, water is freely drunk, and the vitamin A & E liposome, the vitamin A & E soybean oil and the basf vitamin A & E solution are respectively administrated by single-dose stomach irrigation, wherein the dose is 450 mug/kg body weight of the vitamin A and 2.1mg/kg body weight of the vitamin E. The samples were lavage after 10-fold dilution, calculated as 0.1mL/10g body weight. 0.25h (15 min), 0.5h (30 min), 1h,2h,4h,6h,8h,12h,24h and 48h after dosing were collected from the eyes and placed in a 1% heparin sodium treated centrifuge tube. After processing the blood sample, the vitamin content of the blood was measured using high performance liquid chromatography tandem mass spectrometry. The experimental data were processed with DAS2.0 and the metabolic kinetic parameters were calculated.
4. The field emission scanning electron microscope processing method comprises the following steps: and taking a small amount of liposome precursor samples, uniformly scattering the liposome precursor samples on the conductive adhesive, performing metal spraying treatment on the samples by using a HITACHI MC-1000 metal spraying instrument, and shooting under the acceleration voltage of 3-5 kV.
5. The low-voltage transmission electron microscope processing method comprises the following steps: the liposome is diluted to a proper concentration or the precursor is prepared to a proper concentration, and the liposome is observed after the negative dyeing treatment and photographed at an accelerating voltage of 100 kV.
6. Particle size measurement: the particle size distribution of the liposome or liposome precursor after rehydration is determined by using an European and American gram Topsizer laser particle sizer, wherein the refractive index of the sample material is 1.65, the absorptivity of the sample material is 0.001, and the sample is injected through a microcirculatory injector after rehydration.
Zeta potential measurement: the Zeta potential of the liposomes or liposome precursors after rehydration is determined using Malvern Zetasizer Nano ZS90,90 laser particle sizer, after adding the sample to a potentiometric sample cell.
8. Polypeptide sequence identification: and obtaining a polypeptide mass spectrum original file by using a Thermo Fisher LC-MS secondary mass spectrum. The mass spectrum original file was used Byonic to retrieve the target protein database and identify the polypeptide sequence.
Example 1
According to vitamin A: vitamin D 3: vitamin E: the mass ratio of the vitamin K 1 is as follows: 1:0.020:5:0.02, phospholipid is weighed according to the mass ratio of the phospholipid to the vitamin A of 50:1, and sunflower seed oil is weighed according to the mass ratio of the sunflower seed oil to the vitamin A of 80:1. Heating vitamin D 3 and sunflower seed oil to 90deg.C under vacuum, mixing well, mixing with other oil phase, and colloid milling. The water phase is prepared into buffer solution with pH value of 7.3 by dipotassium hydrogen phosphate 0.01mol/L, potassium dihydrogen phosphate 0.002mol/L and total molar concentration of 0.012mol/L and purified water, and sodium chloride is added according to 0.05 mol/L. The ratio of the aqueous phase to the oil phase is 9:1. Preheating the water phase to 60 ℃, injecting the oil phase into the water phase under the stirring of 500r/min, hydrating for 20min, performing ultrahigh pressure micro-jet, repeating for 4 times under the pressure of 90mPa, and obtaining the vitamin A, vitamin D 3, vitamin E, and vitamin K 1 liposome.
The pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and casein was added with stirring to prepare a 2.5% (w/w) casein solution. Dissolving casein, adding pepsin, and performing enzymolysis at 37deg.C for 30 min; after the pH of the solution is adjusted to 6.6-6.8 by using sodium hydroxide, the pH of the solution is adjusted to 7.3 by using dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring is continued for 5min at 95 ℃ to completely terminate the enzyme reaction, so as to obtain casein-pepsin hydrolysate after the pH adjustment.
Mixing the vitamin A, vitamin D 3, vitamin E, vitamin K 1 liposome and casein-pepsin hydrolysate in a ratio of 1:5, and stirring for 5min. Maltodextrin was added at a ratio of 0.8:1 to the liposome, dissolved, and passed through a homogenizer under a homogenizing pressure of 40mPa, and repeated 2 times. Spray drying at 100deg.C and 60deg.C to obtain vitamin A, vitamin D 3, vitamin E, and vitamin K 1 liposome precursor.
Example 2
According to vitamin A: vitamin D 3: vitamin E: the mass ratio of the vitamin K 1 is as follows: 1:0.020:5:0.02, phospholipid is weighed according to the mass ratio of the phospholipid to the vitamin A of 50:1, and sunflower seed oil is weighed according to the mass ratio of the sunflower seed oil to the vitamin A of 80:1. Heating vitamin D 3 and sunflower seed oil to 90deg.C under vacuum, mixing well, mixing with other oil phase, and colloid milling. The water phase is prepared into a buffer solution with pH of 7.3 by dipotassium hydrogen phosphate 0.01mol/L, potassium dihydrogen phosphate 0.002mol/L and total molar concentration of 0.012mol/L and purified water, and sodium chloride is added according to 0.05 mol/L; the ratio of the aqueous phase to the oil phase is 9:1. Preheating the water phase to 60 ℃, injecting the oil phase into the water phase under the stirring of 500r/min, hydrating for 20min, performing ultrahigh pressure micro-jet, repeating for 4 times under the pressure of 90mPa, and obtaining the vitamin A, vitamin D 3, vitamin E, and vitamin K 1 liposome.
The pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and casein was added under stirring to prepare a 2.5% (w/w) casein solution. After casein is dissolved, papain is added, after 30min of hydrolysis, the pH of the hydrolysate is adjusted to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring is continued for 5min at 95 ℃ to completely terminate the enzyme reaction. Obtaining casein-papain hydrolysate after pH adjustment.
Mixing vitamin A, vitamin D 3, vitamin E, vitamin K 1 liposome and casein-papain hydrolysate in a ratio of 1:5, and stirring for 5min. Maltodextrin was added at a ratio of 0.8:1 to the liposome, dissolved, and passed through a homogenizer under a homogenizing pressure of 40mPa, and repeated 2 times. Spray drying at 100deg.C and 60deg.C to obtain liposome precursor of vitamin A, vitamin D3, vitamin E, and vitamin K 1.
Example 3
According to vitamin A: vitamin D 3: vitamin E: the mass ratio of the vitamin K 2 is as follows: 1:0.020:5:0.08 (vitamin K 2 is weighed after conversion according to 5% oil solution), phospholipid is weighed according to the mass ratio of the phospholipid to vitamin A of 50:1, and sunflower seed oil is weighed according to the mass ratio of the sunflower seed oil to the vitamin A of 80:1. Heating vitamin D 3 and sunflower seed oil to 90deg.C under vacuum, mixing well, mixing with other oil phase, and colloid milling. The water phase is prepared into buffer solution with pH value of 7.3 by dipotassium hydrogen phosphate 0.01mol/L, potassium dihydrogen phosphate 0.002mol/L and total molar concentration of 0.012mol/L and purified water, and sodium chloride is added according to 0.05 mol/L. The ratio of the aqueous phase to the oil phase is 9:1. Preheating the water phase to 60 ℃, injecting the oil phase into the water phase under the stirring of 500r/min, hydrating for 20min, performing ultrahigh pressure micro-jet, repeating for 4 times under the pressure of 90mPa, and obtaining the vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome.
The pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and casein was added under stirring to prepare a 2.5% (w/w) casein solution. After casein is dissolved, papain is added, after 30min of hydrolysis, the pH of the hydrolysate is adjusted to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring is continued for 5min at 95 ℃ to completely terminate the enzyme reaction. Obtaining casein-papain hydrolysate after pH adjustment.
Mixing vitamin A, vitamin D 3, vitamin E, vitamin K 2 liposome and casein-papain hydrolysate in a ratio of 1:5, and stirring for 5min. Maltodextrin was added at a ratio of 0.8:1 to the liposome, dissolved, and homogenized under 40mPa for 2 times. Spray drying at 100deg.C and 60deg.C to obtain liposome precursor of vitamin A, vitamin D3, vitamin E, and vitamin K 2. The particle size distribution is shown in FIGS. 7-3.
Example 4
According to vitamin A: vitamin D 3: vitamin E: the mass ratio of the vitamin K 2 is as follows: 1:0.020:5:0.08 (vitamin K 2 is weighed after conversion according to 5% oil solution), phospholipid is weighed according to the mass ratio of the phospholipid to vitamin A of 50:1, and sunflower seed oil is weighed according to the mass ratio of the sunflower seed oil to the vitamin A of 80:1. Heating vitamin D 3 and sunflower seed oil to 90deg.C under vacuum, mixing well, mixing with other oil phase, and colloid milling. The water phase is prepared into a buffer solution with pH of 7.3 by dipotassium hydrogen phosphate 0.01mol/L, potassium dihydrogen phosphate 0.002mol/L and total molar concentration of 0.012mol/L and purified water, and sodium chloride is added according to 0.05 mol/L; the ratio of the aqueous phase to the oil phase is 9:1. Preheating the water phase to 60 ℃, injecting the oil phase into the water phase under the stirring of 500r/min, hydrating for 20min, performing ultrahigh pressure micro-jet, repeating for 4 times under the pressure of 90mPa, and obtaining the vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome.
The pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and concentrated whey protein was added under stirring to prepare a 2.5% (w/w) concentrated whey protein solution. After the concentrated whey protein is dissolved, pepsin is added for enzymolysis for 30min at 37 ℃, sodium hydroxide is used for adjusting the pH of the solution to 6.6-6.8, dipotassium hydrogen phosphate or disodium hydrogen phosphate is continuously used for adjusting the pH of the solution to 7.3, and stirring is continuously carried out for 5min at 95 ℃ to completely terminate the enzyme reaction. The concentrated whey protein-pepsin hydrolysate after pH adjustment is obtained.
Mixing the vitamin A, vitamin D 3, vitamin E, vitamin K 2 liposome and the concentrated whey protein-pepsin hydrolysate in a ratio of 1:5, and stirring for 5min. Maltodextrin was added at a ratio of 0.8:1 to the liposome, dissolved, and homogenized under 40mPa for 2 times. Spray drying at 100deg.C and 60deg.C to obtain liposome precursor of vitamin A, vitamin D3, vitamin E, and vitamin K 2. The particle size distribution is shown in FIGS. 7-4.
Example 5
According to vitamin A: vitamin D 3: vitamin E: the mass ratio of the vitamin K 2 is as follows: 1:0.020:5:0.08 (vitamin K 2 is weighed after conversion according to 5% oil solution), phospholipid is weighed according to the mass ratio of the phospholipid to vitamin A of 50:1, and sunflower seed oil is weighed according to the mass ratio of the sunflower seed oil to the vitamin A of 80:1. Heating vitamin D 3 and sunflower seed oil to 90 ℃ under vacuum, mixing uniformly by rotation, mixing with other oil phases, and passing through a colloid mill. The water phase is prepared into buffer solution with pH value of 7.3 by dipotassium hydrogen phosphate 0.01mol/L, potassium dihydrogen phosphate 0.002mol/L and total molar concentration of 0.012mol/L and purified water, and sodium chloride is added according to 0.05 mol/L. The ratio of the aqueous phase to the oil phase is 9:1. Preheating the water phase to 60 ℃, injecting the oil phase into the water phase under the stirring of 500r/min, hydrating for 20min, performing ultrahigh pressure micro-jet, repeating for 4 times under the pressure of 90mPa, and obtaining the vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome.
The pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and concentrated whey protein was added under stirring to prepare a 2.5% (w/w) concentrated whey protein solution. Dissolving concentrated whey protein, adding papain, hydrolyzing for 30min, adjusting pH of the hydrolyzed solution to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring at 95deg.C for 5min to completely terminate enzyme reaction. The concentrated whey protein-papain hydrolysate after pH adjustment is obtained.
Mixing vitamin A, vitamin D 3, vitamin E, vitamin K 2 liposome and concentrated whey protein-papain hydrolysate at a ratio of 1:5, and stirring for 5min. Maltodextrin was added at a ratio of 0.8:1 to the liposome, dissolved, and homogenized under 40mPa for 2 times. Spray drying at 100deg.C and 60deg.C to obtain vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome precursor. The particle size distribution is shown in FIGS. 7-5.
Example 6
According to vitamin A: vitamin D 3: vitamin E: the mass ratio of the vitamin K 2 is as follows: 1:0.020:5:0.08 (vitamin K 2 is weighed after conversion according to 5% oil solution), phospholipid is weighed according to the mass ratio of 50:1 with vitamin A, and medium chain fatty acid triglyceride is weighed according to the mass ratio of 80:1 with vitamin A. Heating vitamin D 3 and medium chain fatty acid triglyceride to 90deg.C under vacuum, mixing with other oil phase after rotating and mixing, and colloid milling. The water phase is prepared into buffer solution with pH value of 7.3 by dipotassium hydrogen phosphate 0.01mol/L, potassium dihydrogen phosphate 0.002mol/L and total molar concentration of 0.012mol/L and purified water, and sodium chloride is added according to 0.05 mol/L. The ratio of the aqueous phase to the oil phase is 9:1. Preheating the water phase to 60 ℃, injecting the oil phase into the water phase under the stirring of 500r/min, hydrating for 20min, performing ultrahigh pressure micro-jet, repeating for 4 times under the pressure of 90mPa, and obtaining the vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome. The transmission electron microscope of the liposome is shown in FIG. 5-1.
The pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and casein was added under stirring to prepare a 2.5% (w/w) casein solution. After casein is dissolved, pepsin is added for enzymolysis for 30min at 37 ℃, sodium hydroxide is used for adjusting the pH of the solution to 6.6-6.8, dipotassium hydrogen phosphate or disodium hydrogen phosphate is continuously used for adjusting the pH of the solution to 7.3, and stirring is continuously carried out for 5min at 95 ℃ to completely terminate the enzyme reaction. Obtaining casein-pepsin hydrolysate after pH adjustment.
Mixing the vitamin A, vitamin D 3, vitamin E, vitamin K 2 liposome and casein-pepsin hydrolysate in a ratio of 1:5, and stirring for 5min. Maltodextrin was added at a ratio of 0.8:1 to the liposome, dissolved, and passed through a homogenizer under a homogenizing pressure of 40mPa, and repeated 2 times. Spray drying at 100deg.C and 60deg.C to obtain vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome precursor. The particle size distribution of the liposome precursor is shown in FIG. 7-2.
In addition, as shown in fig. 5-2 and 5-3, the transmission electron microscope after rehydration of the liposome precursor of example 6 shows that the polypeptide forms a coating effect on the liposome, and the phenomenon that one peptide chain coats one liposome and one peptide chain coats a plurality of liposomes exists simultaneously. As shown in figures 5-4, the surface of most liposome precursor particles is provided with pits and folds, and no cracks, which are consistent with the characteristics of preparing microcapsules by spray drying.
The use of vitamin K 1 or vitamin K 2 for the preparation of liposomes does not have a significant effect by comparing the indexes such as particle size, encapsulation efficiency, etc. of the liposomes prepared in examples 1 and 6, and examples 2 and 3 and their precursors. Thus, the following comparative examples were each prepared as vitamin a & vitamin D 3 & vitamin E & vitamin K 2 liposomes and then spray dried for comparison.
Comparative example 1
1) Vitamin a & vitamin D 3 & vitamin E & vitamin K 2 liposomes were prepared as in example 6. 2) Adding vitamin A, vitamin D 3, vitamin E, vitamin K 2 liposome and maltodextrin at a ratio of 1.1:1, dissolving, homogenizing under 40mPa, and repeating for 2 times. 3) And (3) putting the materials into an ultralow temperature refrigerator to freeze for 4 hours, transferring into a freeze dryer, and starting freeze drying at the temperature of-60 ℃ and the vacuum degree of 5Pa for 12 hours.
Comparative example 2:
1) Vitamin a & vitamin D 3 & vitamin E & vitamin K 2 liposomes were prepared as in comparative example 1. 2) Adding vitamin A, vitamin D 3, vitamin E, vitamin K 2 liposome and maltodextrin at a ratio of 1.1:1, dissolving, homogenizing under 40mPa, and repeating for 2 times. 3) Using a spray freeze dryer, cold trap temperature-100 ℃, air inlet temperature-20 ℃, vacuum degree 10Pa, spray pressure 4Bar, and preparing the vitamin A, vitamin D 3, vitamin E, vitamin K 2 liposome precursor.
Comparative examples 3 to 27:
1) Vitamin a & vitamin D 3 & vitamin E & vitamin K 2 liposomes were prepared as in comparative example 1. 2) The liposome was mixed with purified water, casein, concentrated whey protein, soy protein isolate, pea protein isolate, low-moisture sodium caseinate, sodium caseinate+sucrose fatty acid ester, casein+sucrose fatty acid ester, casein-neutral protease hydrolysate, casein-alkaline protease hydrolysate, casein-bromelain hydrolysate, casein-trypsin hydrolysate, concentrated whey protein-neutral protease hydrolysate, concentrated whey protein-alkaline protease hydrolysate, concentrated whey protein-bromelain hydrolysate, and concentrated whey protein-trypsin hydrolysate at a mass ratio of 1:5 (see table 1 below), and maltodextrin was used as a filler to be stirred, dissolved and homogenized. 3) Spray drying at 100deg.C and 60deg.C to obtain vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome precursor.
Comparative example 28:
1) According to vitamin A: vitamin D 3: vitamin E: the mass ratio of the vitamin K 1 is as follows: 1:0.020:5:0.02, phospholipid is weighed according to the mass ratio of the phospholipid to the vitamin A of 50:1, and sunflower seed oil is weighed according to the mass ratio of the sunflower seed oil to the vitamin A of 80:1. Heating vitamin D 3 and sunflower seed oil to 90deg.C under vacuum, mixing well, mixing with other oil phase, and colloid milling. 2) The pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and casein was added under stirring to prepare a 2.5% (w/w) casein solution. Dissolving casein, adding pepsin, and performing enzymolysis at 37deg.C for 30 min. 3) And then adjusting the pH of the solution to 6.6-6.8 by using sodium hydroxide, continuously adjusting the pH of the solution to 7.3 by using dipotassium hydrogen phosphate or disodium hydrogen phosphate, and continuously stirring for 5min at 95 ℃ to completely terminate the enzyme reaction, so as to obtain casein-pepsin hydrolysate with the pH adjusted, and directly using the casein-pepsin hydrolysate as a water phase for preparing the liposome. 4) Cooling the water phase to 60 ℃, injecting the oil phase into the water phase under the stirring of 500r/min, hydrating for 20min, performing ultrahigh pressure micro-jet, repeating for 4 times under the pressure of 90mPa, and obtaining the vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome. 5) Maltodextrin was added at a ratio of 0.8:1 to the liposome, dissolved, and homogenized under 40mPa for 2 times. 6) Spray drying with spray dryer at 100deg.C and 60deg.C to obtain vitamin A, vitamin D 3, vitamin E, and vitamin K 2 liposome precursor. The particle size distribution diagram after rehydration is shown in figures 7-7.
Fig. 7-1 shows particle size distribution diagrams of typical liposomes, and fig. 7-2 to 7-7 show particle size distribution diagrams of liposome precursors of example 6, example 3, example 4, example 5, comparative example 21, and comparative example 28, respectively, after rehydration. As can be seen from fig. 7-1 to fig. 7-7: the particle sizes of examples 6, 3, 4 and 5 after rehydration remained largely within the size range of the liposomes, and comparative examples 21 and 28 had severe tailing, probably due to fusion and rupture during preparation.
The particle size, encapsulation efficiency, loss rate index of the liposome precursors corresponding to examples 1 to 6 and comparative examples 1 to 28 were compared, specifically as shown in table 1 below.
Table 1 comparison of particle size, encapsulation efficiency, loss rate of examples and comparative products
The liposome can be divided into unilamellar liposome with particle size of 10-1000 nm and multilamellar liposome with particle size of 1000-5000 nm according to bilayer membrane number; the particle size of the multivesicular liposome is more than 1000nm. Examples are compared with comparative examples by combining the indexes of particle diameter, encapsulation efficiency, loss rate and the like. The vitamin A, vitamin D3, vitamin E and vitamin K2 liposome prepared by the invention has larger drug-loading rate which can reach 4 percent at most. Based on this factor, maltodextrin and the like are probably used as fillers and freeze-drying protective agents, and the precursor prepared by a common freeze-drying method is used, so that the structure of the liposome is damaged by ice crystals in the freeze-drying process, the content leaks, and the liposome structure cannot be recovered after rehydration (such as comparative examples 1 and 2). The precursor prepared by spray drying with maltodextrin as filler has liposome structure destroyed at high temperature to leak content, and vitamin A, vitamin K, etc. have high loss of light-sensitive and heat-sensitive vitamins (as in comparative examples 3 to 3). Since saccharides such as maltodextrin, white sugar and trehalose have no emulsifying property or surface charge, they can only be used as fillers.
Sodium caseinate has a certain emulsifying property, and can prepare liposome precursors (such as comparative example 8) with most of liposome structures and particle sizes maintained after rehydration under a low humidity environment (relative humidity < 30% RH). However, in a high humidity environment, the ability of hot air to carry moisture is reduced, and liposome precursors (e.g., comparative example 9) cannot be prepared. This feature limits the production of liposome precursors in summer in the south and north. The isolated soy protein and pea protein per se have a larger particle size, poor water solubility and poor emulsifying properties, and thus have no protective effect on liposomes during spray drying (as in comparative examples 6 and 7). The particle size of casein itself is large, the water solubility is good under alkaline condition, the performance is similar to that of sodium caseinate, and liposome precursor (such as comparative example 4) can not be prepared under high humidity environment. The concentrated whey protein itself has small particle size and good water solubility, but is characterized by poor heat resistance, the protein is denatured during spraying, and the protection effect on liposome is reduced (as in comparative example 5).
Comparative example 28 has a particle size distribution close to that of liposomes prepared with PBS solution. However, after spray drying to prepare liposome precursors, the particle size distribution is as described in the accompanying table, with severe tailing as shown in figures 7-7. It is inferred that the formation of liposome is that peptide chains are wrapped in an inner core of a water phase, and the longer peptide chains break through a liposome double-layer membrane after drying and water loss in the precursor preparation process, so that the particle size is increased, the encapsulation rate is reduced, and the loss rate is increased.
The comparison shows that: after enzyme treatment, casein and concentrated whey protein change their original characteristics and can strengthen their protective action on liposome. And the enzymes with the best treatment effect are pepsin and papain. By combining a transmission electron microscope image, the possible protection mechanism is deduced that casein and concentrated whey protein are hydrolyzed into peptide by pepsin or papain, and then positively charged amino acid residues are exposed at pH of 7.0-7.5, and the surface of the liposome is negatively charged, so that the coating of peptide chains on the liposome is realized, and the protection of the liposome is realized. Zeta potential is an important factor affecting liposome stability and is also a prerequisite for liposome to be coated. The zeta potentials of examples 1 to 6 and some of comparative examples 1, 5 to 6 and 20 to 27 were compared, and the comparison results are shown in Table 2 below.
TABLE 2 zeta potential comparison of examples and part of comparative examples
Taking casein as an example, the isoelectric point (pI) of casein is 4.6, the surface of casein is positively charged below the isoelectric point, above the isoelectric point, and the surface of casein is negatively charged. Liposomes are often negative in zeta potential due to the presence of phosphate ions in the phospholipid membrane. If the casein is to be coated on the liposome, the pH should be adjusted to 4.6 or less. However, under this pH condition, phospholipids tend to oxidize and become unstable. The liposome has a suitable pH of 7.0-8.0, and at pH7.3, casein (comparative example 4) cannot form a coating on the liposome, and the zeta potential of the liposome does not change significantly. Likewise, the remaining protein-protease hydrolysates also did not have a significant effect on the zeta potential of the liposomes. However, as shown in examples 1 to 6, the absolute value of zeta potential of the system can be significantly improved by performing enzyme treatment on casein/concentrated whey protein by pepsin/papain, and thus liposome coating can be realized.
As shown in FIG. 8, in examples 3 to 6, the absolute value of zeta potential was significantly increased at pH7.0 to 7.5. At pH <7, the absolute value of zeta potential is lower than that of uncoated liposome, and it is presumed that at this pH, the hydrolysate forms a coating on the liposome, and the zeta potential tends to be positive potential because the amino acid residue is charged more positively than pH7.0 or more, but the liposome at pH <7 is unstable. Therefore, pH7.0 to 7.5 is a preferable pH for binding, and can ensure the stability of the liposome while forming a coating on the liposome.
Different enzymes have different sites of action, and their hydrolysis is directed to different amino acid sequences, so that the selectivity of different enzymes for proteins is different, and the peptide chains hydrolyzed by them are also different. The amino acid is an amphoteric substance, at least one amino (NH 3+) group capable of releasing protons and one carboxyl (COO -) group capable of accepting protons are arranged on the same molecule, the acidic amino acid contains 1 amino acid and 2 carboxyl groups, the isoelectric point is about 2.8-3.2, the basic amino acid contains 2 amino groups and 1 carboxyl group, and the isoelectric point is about 9.7-10.7. Amino acids form peptide chains through peptide bonds, peptide chains further form proteins with larger molecular weights through the modes of helix, folding and the like, and the proteins or peptide chains can be positively charged or negatively charged at a certain pH according to the difference of the exposed residues. As described above, in the present invention, pepsin and papain hydrolysates have a better protective effect on liposomes, and the peptide chains produced by the hydrolysis of proteins by these two enzymes form a positively charged region at a suitable pH, as shown in FIG. 6, which can be bound to liposomes with negatively charged surfaces, thus forming a coating.
In addition, the invention further identifies the peptide chain in the protease hydrolysate. Tables 3 and 4 below show the relatively large number of 5 peptide chains identified in the hydrolysates after hydrolysis of casein with pepsin and papain, respectively. Corresponding amino acid abbreviations: g: glycine; a: alanine; v: valine; l: leucine; i: isoleucine; p: proline; f: phenylalanine; y: tyrosine; w: tryptophan; s: serine; t: threonine; c: cysteine; m: methionine; n: asparagine; q: glutamine; e: glutamic acid; k: lysine; r: arginine; h: histidine.
Table 3 casein-pepsin hydrolysate peptide chain:
| Peptide chain | Duty cycle (%) |
| LLRLKKYKVPQL | 10.28 |
| VRSPAQILQWQVL | 5.41 |
| INNQFLPYPYYAKPAAVRSPAQILQ | 4.87 |
| TKVIPYVRYL | 4.22 |
| YVLSRYPSYGLN | 3.59 |
Table 4 casein-papain hydrolysate peptide chain:
| Peptide chain | Duty cycle (%) |
| FLKKISQRYQKFALPQYLKTVY | 7.78 |
| INNQFLPYPYYAKPAAVR | 5.86 |
| GYLEQLLRLK | 4.09 |
| FPPQSVLSLSQSKVLPVPQKAVPYPQRDMPIQAFLLYQEPVLGPVR | 2.69 |
| FALPQYLKTVY | 2.30 |
As shown in tables 3 and 4 above, the casein-pepsin hydrolysate peptide chain and the casein-papain hydrolysate peptide chain have basic amino acids therein. Basic amino acids such as lysine, arginine, histidine, etc. have pI values of 9.74, 10.76, 7.59, respectively, and the pI values of the rest amino acids are below 7. Under the condition that the pH is 7.0-7.5, lysine, arginine, histidine and the like have positive charges, and the rest amino acids have negative charges. It is assumed that the positively charged region is composed of lysine, arginine, histidine in the peptide chain by the action of helix, folding, etc., and thus the liposome is coated. The negatively charged amino acids account for a relatively large proportion at pH7.3, so that the zeta potential of the system as a whole is negative.
The following describes the tolerance to heat treatment before and after liposome coating, the vitamin attenuation, release characteristics, half-life and bioavailability of each type of product in each state, with specific examples and comparative examples:
resistance to heat treatment before and after liposome encapsulation:
the change of liposome stability before and after coating is compared under the common sterilization temperature and treatment time.
TABLE 3 comparison of the resistance to Heat treatment before and after coating of liposomes with protein hydrolysates
Note that: "-" is a distinct layered demulsification, with no particle size detected. The treatment at 65-95 ℃ is pasteurization treatment, so that most bacteria can be killed, and the temperature and time which can be tolerated by the coated liposome are obviously increased under the pasteurization condition through comparison. The treatment at 121 ℃ is high-temperature high-pressure sterilization, so that all microorganisms can be killed, and the requirement of commercial sterility is met. However, under these conditions, the liposome samples all exhibited delamination and demulsification, and were not able to withstand the sterilization conditions. The method is used for carrying out ultra-high temperature instantaneous sterilization at 135 ℃, has high treatment temperature and short treatment time, and can kill most microorganisms. Uncoated liposomes failed to withstand UHT treatment and after coating with protein hydrolysates, the liposomes were significantly more tolerant to UHT. At present, most liquid foods adopt UHT except the fruit jelly added with preservative and the fresh milk and fruit juice products with short shelf life transported and preserved by a cold chain. After the liposome precursor is prepared by coating the protein-protease hydrolysate, the application range of the liposome is further enlarged.
Microbial changes before and after sterilization:
And (3) detecting microorganisms on the sterilized sample with the particle size not changed obviously and evaluating the sterilization effect.
TABLE 4 microbial changes before and after sterilization
As shown in the table above, pasteurization at 72℃killed mold, yeast and most of the microorganisms in the liposomes, which were left at ambient temperature for 6 months, the uncoated liposomes were occasionally gel-like masses, and no such phenomenon occurred after coating. Pasteurization at 85deg.C or below and UHT treatment can kill all microorganisms in liposome, and can be placed at room temperature for 6 months without gel cake. The liposome stability is further improved by sterilization treatment after being coated by protein hydrolysate.
Vitamin decay (both solution and solid state):
Commercial products of vitamin A, vitamin D and vitamin K 2 are solid products, and commercial product 1 is a product produced by a double embedding process, and main wall materials of the commercial product are corn starch, acacia, sodium starch octenyl succinate and the like; the commercial 2 is used for representing vitamin A, vitamin D, vitamin K 2 products produced by a single embedding process, and the main wall materials are lactose, sodium starch octenyl succinate, maltodextrin and the like; commercial vitamin E is a single-embedding process product, and the main wall material is sodium starch octenyl succinate. Dissolving commercial product, rehydrating liposome precursor, pasteurizing liposome, adding sodium ethyl parahydroxybenzoate, sealing with glass bottle, respectively placing at room temperature and 37deg.C under 75% RH acceleration condition, and comparing and inspecting stability difference in solution state; commercial products and liposome precursors were sealed with aluminum foil bags and placed under room temperature conditions and under 37 ℃ and 75% RH acceleration conditions, respectively, to compare and examine differences in stability in the solid state. Namely, the loss rate results of the four vitamins under the conditions of normal temperature of the solution, acceleration of the solution, powder and acceleration of the powder are shown in figures 1-1 to 1-4 respectively. As can be seen from the accompanying drawings: the stability of vitamin A is the worst, in a solution system, the loss rate of a single embedded product is the fastest, the loss rate of liposome is close to that of the commercial 2, the loss rate of liposome precursor is the lowest, and the several precursors have no obvious difference; in the powder state, the loss rate of the liposome precursor is higher than that of the liposome precursor sold in the market 1 and lower than that of the liposome precursor sold in the market 2, and the loss rate is probably caused by instability of the liposome due to the fact that the temperature is close to the temperature point of the phospholipid at 45 ℃. Vitamin K 2 has more double bonds and is unstable in storage environment, and the comparison result is similar to that of vitamin A, and the loss rate is higher than that of the commercial 1 and lower than that of the commercial 2. The loss rate of the vitamin D and the vitamin E in the liposome is close to the loss rate of the vitamin D and the vitamin E in the liposome which is smaller than the loss rate of the vitamin D and the vitamin E in the liposome precursor and is smaller than the loss rate of the vitamin D and the vitamin E in the liposome precursor which is smaller than the loss rate of the vitamin D and the vitamin E in the liposome precursor. Overall, it is seen that: liposome precursors have better stability in solution and little difference between the examples, which are more suitable for liquid systems. Since the stability of each protein hydrolysate coating is not very different, one of the examples was chosen for stability comparison with the commercial product in the application product.
Attenuation of vitamins in nutrient fortified milk powder products:
At present, a milk powder production enterprise generally adopts two modes of wet method and dry method to produce the nutrition-enhanced milk powder. Wet production is to use a spray drying tower to produce milk powder, other raw materials and a nutrition enhancer after rehydration and dissolution, and double-embedding type namely commercial 1 nutrition enhancer is commonly used for strengthening because of high-temperature treatment involved in spray drying. The dry production is to mix the milk powder, other raw materials and the nutrition enhancer by using a three-dimensional mixer, a horizontal mixer and other equipment, and the commercial type 1 nutrition enhancer with a single embedded type 2 particle size close to that of the milk powder is used for enhancement because the powder cannot be mixed uniformly due to larger particle size and the deviation of detection results is easy to cause. In the decay experiments, there is no obvious difference between the examples, so that the stability in milk powder is compared with the stability in milk powder by adding and strengthening in milk powder according to two methods respectively only in the example 6. Wherein, the condition of wet spray drying is that the inlet air temperature is 175 ℃, the outlet air is stable at 100 ℃, the inlet air frequency is 30Hz, and the rotating speed of a peristaltic pump is 40r/min. The prepared sample is placed in a constant temperature and humidity box for acceleration treatment under the condition of 37 ℃ and 75 percent RH. The wet process and dry process treatment results of the four vitamins are shown in figures 2-1 and 2-2, respectively. As can be seen from fig. 2-1: the processing loss of vitamin A in the liposome and the liposome precursor of the nutrition-enriched milk powder prepared by the wet process is smaller than the loss rate of the vitamin A in the market at the observation period of 1. The loss rates of vitamin D, vitamin E and vitamin K are also lower than those of the commercial 1, but the loss rates are not far different. As can be seen from fig. 2-2: the nutrient-enriched milk powder prepared by the dry process has a vitamin loss rate which is lower than that of the commercial 2 in the liposome precursor, but has smaller difference. And (3) injection: wet-produced milk powder, data for "0" month is processing loss.
Attenuation of vitamins in nutrient fortified milk:
The attenuation in the nutrition-enriched milk adopts ultrahigh temperature instantaneous sterilization commonly used for dairy products, namely, the stability comparison of vitamins is carried out after UHT treatment of the nutrition-enriched milk, the UHT sterilization temperature is 135 ℃, and the time is 5s. The samples were subjected to accelerated observation at 37℃and 75% RH. The results are shown in FIG. 3. As can be seen from fig. 3: the most loss of vitamin processing and the worst stability of commercial 2, liposome stability is near or slightly lower than commercial 1, and liposome precursors are the best.
In vitro metabolism assay (release profile):
The digestion environment of the oral cavity, stomach and intestinal tract is simulated in vitro, and the release characteristics of vitamin A, vitamin D, vitamin E, vitamin K 2 commercial products, liposome and liposome precursor are compared. Wherein 0 min-10 min is simulated oral environment, 10 min-115 min is simulated gastric environment, and 115 min-220 min is simulated small intestine environment. The comparison results are shown in FIGS. 4-1 through 4-4. From fig. 4-1 to fig. 4-4, it can be seen that: both commercial products 1 and 2 used starch or sodium starch octenyl succinate as emulsifying and embedding agents, and when they simulate oral digestion, starch may be enzymatically hydrolyzed by salivary amylase, thus being released in large amounts in simulated environments of the oral cavity and stomach. The liposome has strong tolerance to oral cavity and stomach environment, but can be affected by gastric acid in stomach environment, and a small amount of phospholipid releases vitamins after hydrolysis. In the intestine, there is a large release under the influence of lipases and bile salts. The "concentrated whey protein hydrolysate coated liposomes" (i.e. examples 4 and 5) also released slightly in the stomach environment, but at a lower release rate than uncoated liposomes. The "casein hydrolysate coated liposomes" (i.e. examples 3 and 6) may retain the property of partially casein to aggregate with acid and release less in simulated gastric environment.
In vivo metabolism experiments (half-life and bioavailability):
The in vivo metabolism experiment of animals is carried out by selecting commercial products of vitamin A, vitamin D, vitamin E and vitamin K 2 double embedding process, liposome-casein-pepsin hydrolysate precursor, liposome-concentrated whey protein-pepsin hydrolysate precursor, liposome and pure product. Double embedding process products, precursor rehydration, liposome dilution with deionized water, pure soybean oil dissolution. The results of the vitamin metabolism kinetic parameters are shown in the following tables 5-1 to 5-4. As can be seen from the table: compared with the commercial aqueous solution and the pure oil solution, the absorption half-life (T1/2 alpha) of the liposome and the precursor thereof after rehydration is obviously improved, the peak reaching concentration (Cmax) is obviously improved, the peak reaching time (Tmax) is respectively delayed by 4h and 2h, the surface liposome and the precursor thereof have longer in-vivo circulation time and lower clearance, the release is slower, and the effective action is prolonged. Compared with bioavailability, the liposome and the precursor thereof are obviously improved relative to commercial products and oil solutions, and particularly for vitamin D, the bioavailability improvement effect is more obvious, and the bioavailability improvement effect is probably caused by the fact that the vitamin D is difficult to emulsify relative to other vitamins and the solubility in oil is lower. The low bioavailability of commercial products may be due to the lack of lipid material as a carrier and the inability to form sufficient chylomicrons. The liposome imitates chylomicron structurally, is not limited by lipid and bile salt in the absorption process, and has better tolerance to gastric acid environment. (example 6) the absorption half-life of the liposome-casein-pepsin hydrolysate precursor and (example 4) the liposome-concentrated whey protein-pepsin hydrolysate precursor were slightly pushed relative to the liposomes, and the release of vitamins should be delayed for digestion of the protein coated on the outer layer of the liposomes; whereas example 6 has a somewhat higher relative availability, probably due to the retention of the properties of part of the casein, due to aggregation by gastric acid, after the half-life of absorption compared to example 4.
TABLE 5-1 vitamin A Metabolic kinetic parameters
TABLE 5-2 vitamin D Metabolic kinetic parameters
| Parameters (parameters) | Unit (B) | Commercially available | Oil solution | Liposome | Example 6 | Example 4 |
| t1/2α | h | 2.13 | 2.84 | 6.84 | 6.90 | 6.71 |
| AUC(0-t) | ug/L*h | 676 | 875 | 1232 | 1471 | 1385 |
| Cmax | ng/L | 33.60 | 38.19 | 38.37 | 50.36 | 46.73 |
| Tmax | h | 4 | 6 | 8 | 8 | 8 |
| Relative availability | ％ | 100 | 129 | 182 | 217 | 205 |
TABLE 5-3 vitamin E Metabolic kinetic parameters
| Parameters (parameters) | Unit (B) | Commercially available | Oil solution | Liposome | Example 6 | Example 4 |
| t1/2α | h | 1.74 | 4.42 | 5.38 | 6.56 | 6.67 |
| AUC(0-t) | mg/L*h | 1115 | 1695 | 1979 | 2041 | 1809 |
| Cmax | ug/L | 56.29 | 67.58 | 70.03 | 71.83 | 62.38 |
| Tmax | h | 4 | 6 | 8 | 8 | 8 |
| Relative availability | ％ | 100 | 152 | 177 | 183 | 162 |
TABLE 5-4 vitamin K Metabolic kinetic parameters
| Parameters (parameters) | Unit (B) | Commercially available | Oil solution | Liposome | Example 6 | Example 4 |
| t1/2α | h | 1.80 | 3.41 | 5.12 | 6.11 | 5.78 |
| AUC(0-t) | ug/L*h | 2517 | 3297 | 4070 | 4734 | 4742 |
| Cmax | ng/L | 103.7 | 127.7 | 132.1 | 175.16 | 165.60 |
| Tmax | h | 4 | 6 | 8 | 8 | 8 |
| Relative availability | ％ | 100 | 131 | 162 | 188 | 188 |
The optimal hydrolysis conditions according to the present application are further described by the following specific examples and comparative examples:
Hydrolysis time effect:
Comparative example 29 comparative example 46 vitamin a & vitamin D 3 & vitamin E & vitamin K 2 liposomes were prepared according to the procedure of example 6; the protein-protease hydrolysates of the respective comparative examples were prepared according to the following procedures; finally, liposome precursors were prepared by the procedure of example 6 using liposomes and hydrolysates.
Preparation of comparative examples 27-31: the pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and casein was added under stirring to prepare a 2.5% (w/w) casein solution. After casein is dissolved, 0.3% pepsin (according to the weight of the solution) is added for enzymolysis at 37 ℃ for 10min, 20min, 40min, 60min and 120min respectively, the pH of the solution is adjusted to 6.6-6.8 by using sodium hydroxide, the pH of the solution is adjusted to 7.3 by continuously using dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring is continued for 5min at 95 ℃ to obtain casein-pepsin hydrolysate.
Preparation of comparative examples 32-36: the pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and casein was added under stirring to prepare a 2.5% (w/w) casein solution. Dissolving casein, adding 0.3% papain (by weight of the solution), hydrolyzing for 10min, 20min, 40min, 60min, and 120min, adjusting pH to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring at 95deg.C for 5min to obtain casein-papain hydrolysate.
Preparation of comparative examples 37-41: the pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and concentrated whey protein was added under stirring to prepare a 2.5% (w/w) concentrated whey protein solution. Dissolving the concentrated whey protein, adding 0.3% pepsin (according to the weight of the solution), respectively carrying out enzymolysis for 10min,20min,40 min, 60min and 120min at 37 ℃, regulating the pH of the solution to 6.6-6.8 by using sodium hydroxide, regulating the pH of the solution to 7.3 by using dipotassium hydrogen phosphate or disodium hydrogen phosphate, and continuously stirring for 5min at 95 ℃ to obtain the concentrated whey protein-pepsin hydrolysate.
Preparation of comparative examples 42-46: the pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and concentrated whey protein was added under stirring to prepare a 2.5% (w/w) concentrated whey protein solution. Dissolving the concentrated whey protein, adding 0.3% papain (based on the weight of the solution), hydrolyzing for 10min, 20min, 40min, 60min, and 120min, adjusting pH to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring at 95deg.C for 5min to obtain concentrated whey protein-papain hydrolysate.
The particle size, encapsulation efficiency, loss rate of the above four comparative liposome precursors are shown in tables 6-1 to 6-4.
TABLE 6-1 Casein-pepsin
TABLE 6-2 Casein-papain
TABLE 6-3 concentration of whey protein-pepsin
Tables 6-4 concentrated whey protein-papain
As can be seen from the table above: the hydrolysis time is a key factor affecting the hydrolysate, and the length and configuration of the peptide chain of the hydrolysate can be key factors affecting the particle size, encapsulation efficiency and loss rate of the precursor after rehydration. The proper hydrolysis time of casein is 20 min-60 min, and the proper hydrolysis time of concentrated whey protein is 10 min-40 min. Possibly related to the molecular weight of casein, the concentrated whey protein itself. According to the data of the particle sizes, the encapsulation efficiency and the loss rate of the examples and the comparative examples, the smaller the particle size is, the higher the encapsulation efficiency is, the lower the loss rate is, and in order to rapidly screen the most suitable proportion and process, the particle size is only used as a comparative element, and the encapsulation efficiency and the loss rate are not detected and calculated any more.
Protein concentration effects:
Comparative example 47-comparative example 62 vitamin a & vitamin D3& vitamin E & vitamin K2 liposomes were prepared according to the procedure of example 6; the protein-protease hydrolysates of the respective comparative examples were prepared according to the following procedures; finally, liposome precursors were prepared by the procedure of example 6 using liposomes and hydrolysates.
Preparation of comparative examples 47-50: the pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and casein was added under stirring to prepare 1%, 2%, 3% and 4% (w/w) casein solutions, respectively. After casein is dissolved, 0.3% pepsin (according to the weight of the solution) is added for enzymolysis for 30min at 37 ℃, sodium hydroxide is used for adjusting the pH of the solution to 6.6-6.8, dipotassium hydrogen phosphate or disodium hydrogen phosphate is continuously used for adjusting the pH of the solution to 7.3, and stirring is continuously carried out for 5min at 95 ℃ to obtain casein-pepsin hydrolysate.
Preparation of comparative examples 51-54: the pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and casein was added under stirring to prepare 1%, 2%, 3% and 4% (w/w) casein solutions. Dissolving casein, adding 0.3% papain (based on the weight of the solution), hydrolyzing at 55deg.C for 30min, adjusting pH of the hydrolyzed solution to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring at 95deg.C for 5min to obtain casein-papain hydrolyzed solution.
Preparation of comparative examples 55-58: the pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and concentrated whey protein was added under stirring to prepare 1%, 2%, 3%, 4% (w/w) concentrated whey protein solutions. Dissolving the concentrated whey protein, adding 0.3% pepsin (according to the weight of the solution), carrying out enzymolysis for 30min at 37 ℃, adjusting the pH of the solution to 6.6-6.8 by using sodium hydroxide, continuously adjusting the pH of the solution to 7.3 by using dipotassium hydrogen phosphate or disodium hydrogen phosphate, and continuously stirring for 5min at 95 ℃ to obtain the concentrated whey protein-pepsin hydrolysate.
Preparation of comparative examples 59-62: the pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and concentrated whey protein was added under stirring to prepare 1%, 2%, 3% and 4% (w/w) concentrated whey protein solutions. Dissolving the concentrated whey protein, adding 0.3% papain (based on the weight of the solution), hydrolyzing at 55deg.C for 30min, adjusting pH of the hydrolyzed solution to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring at 95deg.C for 5min to obtain concentrated whey protein-papain hydrolyzed solution.
The particle size, encapsulation efficiency, loss rate of the above four comparative liposome precursors are shown in tables 7-1 to 7-4.
TABLE 7-1 Casein-pepsin
TABLE 7-2 Casein-papain
TABLE 7-3 concentration of whey protein-pepsin
TABLE 7-4 concentrated whey protein-papain
As can be seen from the table above: the proper concentration of casein and concentrated whey protein is 2% -3%. At 1% concentration, protection of the liposomes may not be formed due to too little protein hydrolysate produced; at a concentration of 4%, the unhydrolyzed protein has a large influence on the particle size.
Protease addition amount:
Comparative examples 63-74 vitamin a & vitamin D3& vitamin E & vitamin K2 liposomes were prepared as in example 6; the protein-protease hydrolysates of the respective comparative examples were prepared according to the following procedures; finally, liposome precursors were prepared as in example 6 using liposomes and hydrolysates.
Preparation of comparative examples 63-65: the pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and casein was added under stirring to prepare 2.5% (w/w) casein solutions, respectively. After casein is dissolved, 0.1%, 0.2% and 0.4% pepsin (based on the weight of the solution) are added respectively, enzymolysis is carried out for 30min at 37 ℃, after the pH of the solution is adjusted to 6.6-6.8 by using sodium hydroxide, the pH of the solution is adjusted to 7.3 by continuously using dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring is continued for 5min at 95 ℃ to obtain casein-pepsin hydrolysate.
Preparation of comparative examples 66-68: the pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and casein was added under stirring to prepare a 2.5% (w/w) casein solution. Dissolving casein, adding 0.1%, 0.2% and 0.4% papain (based on the weight of the solution), hydrolyzing at 55deg.C for 30min, adjusting pH of the hydrolyzed solution to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring at 95deg.C for 5min to obtain casein-papain hydrolyzed solution.
Preparation of comparative examples 69-71: the pH of the purified water was adjusted to 1.5 using hydrochloric acid, and preheated to 37℃and concentrated whey protein was added under stirring to prepare a 2.5% (w/w) concentrated whey protein solution. Dissolving the concentrated whey protein, adding 0.1%, 0.2% and 0.4% pepsin (by weight of the solution), carrying out enzymolysis for 30min at 37 ℃, regulating the pH of the solution to 6.6-6.8 by using sodium hydroxide, regulating the pH of the solution to 7.3 by using dipotassium hydrogen phosphate or disodium hydrogen phosphate, and continuously stirring for 5min at 95 ℃ to obtain the concentrated whey protein-pepsin hydrolysate.
Preparation of comparative examples 72-74: the pH of the purified water was adjusted to 6.5 using potassium dihydrogen phosphate or sodium dihydrogen phosphate, preheated to 55℃and concentrated whey protein was added under stirring to prepare a 2.5% (w/w) concentrated whey protein solution. Dissolving the concentrated whey protein, adding 0.1%, 0.2% and 0.4% papain (based on the weight of the solution), hydrolyzing at 55deg.C for 30min, adjusting pH of the hydrolyzed solution to 7.3 with dipotassium hydrogen phosphate or disodium hydrogen phosphate, and stirring at 95deg.C for 5min to obtain concentrated whey protein-papain hydrolyzed solution.
The particle size, encapsulation efficiency, loss rate of the above four comparative liposome precursors are shown in tables 8-1 to 8-4.
TABLE 8-1 Casein-pepsin
TABLE 8-2 Casein-papain
TABLE 8-3 concentration of whey protein-pepsin
TABLE 8-4 concentrated whey protein-papain
As can be seen from the table above: at a casein concentration of 2.5%, the amount of pepsin and papain added was suitably 0.3%. The proper addition amount of pepsin and papain is 0.1% -0.3% under the condition that the concentration of the concentrated whey protein is 2.5%. Of course, the enzyme is a high concentration of active substance, and fluctuations within the error range are acceptable.
According to the embodiment, the hydrolysate is prepared under the preferable conditions, the liposome is coated by the hydrolysate, and the liposome precursor obtained after coating has the advantages of smaller particle size, narrow distribution, high encapsulation efficiency, low loss rate and improved stability after rehydration. The main factors influencing the enzymatic hydrolysis reaction are temperature, pH and substrate concentration, i.e. the concentration of the protein solution. In the application, the protein concentration is a key factor influencing the preparation of liposome precursors, so that the operation is convenient, and the liposome precursors are not concentrated or diluted and then mixed with the liposome for spraying. Of course, other applications of hydrolyzing the substrate and the protease in a similar proportion and concentrating or diluting the hydrolyzed product or selecting the protease with different enzyme activity units to prepare the hydrolyzed product are also within the protection scope of the scheme.
The foregoing description of the invention has been presented for purposes of illustration and description, and is not intended to be exhaustive or limited to the invention in the form disclosed. Many modifications/variations will be apparent to those of ordinary skill in the art. The embodiments were chosen and described in order to best explain the principles of the invention and the practical application, to thereby enable others skilled in the art to understand the invention for various embodiments with various modifications as are suited to the particular use contemplated.
Claims (10)
1. A preparation method of a fat-soluble vitamin liposome precursor is characterized in that a protease hydrolysate is adopted to coat a liposome in the process of preparing the liposome precursor; comprising the following steps:
Regulating the pH of the protein-protease hydrolysate to 7.0-7.5, mixing the protein-protease hydrolysate with liposome, adding a filler, stirring and dissolving, and homogenizing to obtain a feed liquid;
And drying the feed liquid by adopting spray drying to obtain liposome precursors.
2. The method for preparing a lipid-soluble vitamin liposome precursor according to claim 1, wherein the protease is one or more of pepsin and papain.
3. The method for preparing a fat-soluble vitamin liposome precursor according to claim 2, wherein the protein is one or more of casein and concentrated whey protein.
4. The method for preparing a lipid-soluble vitamin liposome precursor according to claim 1, wherein the protein-protease hydrolysate and the liposome are mixed according to a mass ratio of 4-6:1.
5. The method of preparing a fat-soluble vitamin liposome precursor according to claim 1, wherein the adjusting of the pH of the protein-protease hydrolysate in two steps comprises: the pH is adjusted to 6.6-6.8, and then the pH is adjusted to 7.0-7.5.
6. The method of preparing a lipid-soluble vitamin liposome precursor according to any one of claims 1 to 5, wherein the preparation of the protein-protease hydrolysate comprises: dissolving protein in hydrochloric acid or phosphate solution with proper pH for enzymolysis to obtain protein solution; determining the concentration of protease and the hydrolysis time according to the protein type, adding protease to the protein solution and hydrolyzing at the protease hydrolysis temperature;
wherein the concentration of the protein solution is 2-3%w/w of the protein solution; the proper pH value of the enzymolysis of the pepsin is 1.0-1.8, and the proper pH value of the enzymolysis of the papain is 6-7;
The hydrolysis temperature of pepsin is 35-40 ℃; the hydrolysis temperature of papain is 50-60 ℃; the concentration of the protease of the casein is 0.2-0.3 percent, and the hydrolysis time is 20-60 min; the protease concentration of the concentrated whey protein is 0.1-0.3%, and the hydrolysis time is 10-40 min.
7. The method for preparing a precursor of fat-soluble vitamin liposome according to claim 6, wherein when the feed liquid is dried by spray drying, the air inlet temperature is 90-110 ℃ and the air outlet temperature is 55-75 ℃;
Homogenizing by using a homogenizer, wherein the homogenizing pressure is 30-50 mPa, and the homogenizing times are 2-4 times;
The ratio of the filler to the liposome is 0.45-0.90:1; the filler is one or two of maltodextrin, white granulated sugar, isomaltulose, modified starch, polydextrose and resistant dextrin.
8. The method for preparing a lipid-soluble vitamin liposome precursor according to any one of claim 1 to 5,
After the step of adjusting the pH of the protein-protease hydrolysate, the method further comprises: boiling and inactivating enzyme of the protein-protease hydrolysate;
and/or, before the step of drying the feed liquid by spray drying, further comprising: pasteurizing or UHT sterilizing the feed liquid; wherein the pasteurization conditions: the temperature is 65-85 ℃ and the time is 10-30 min; UHT sterilization conditions: the temperature is 125-135 ℃ and the time is 5-15 s.
9. A liposoluble vitamin liposome precursor prepared by the method for preparing a liposoluble vitamin liposome precursor according to any one of claims 1-8, wherein the encapsulation efficiency of the liposome precursor is not less than 92%.
10. Use of a fat-soluble vitamin liposome precursor in a liquid product, characterized in that the fat-soluble vitamin liposome precursor is prepared by the preparation method of the fat-soluble vitamin liposome precursor according to any one of claims 1-8.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410168120.6A CN118001250A (en) | 2024-02-06 | 2024-02-06 | Fat-soluble vitamin liposome precursor, and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410168120.6A CN118001250A (en) | 2024-02-06 | 2024-02-06 | Fat-soluble vitamin liposome precursor, and preparation method and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN118001250A true CN118001250A (en) | 2024-05-10 |
Family
ID=90942528
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410168120.6A Pending CN118001250A (en) | 2024-02-06 | 2024-02-06 | Fat-soluble vitamin liposome precursor, and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN118001250A (en) |
-
2024
- 2024-02-06 CN CN202410168120.6A patent/CN118001250A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Wang et al. | Spray drying strategy for encapsulation of bioactive peptide powders for food applications | |
| Jash et al. | Liposomes for oral delivery of protein and peptide-based therapeutics: Challenges, formulation strategies, and advances | |
| Aguilar-Toalá et al. | Encapsulation of bioactive peptides: A strategy to improve the stability, protect the nutraceutical bioactivity and support their food applications | |
| Xiong et al. | Construction of food-grade pH-sensitive nanoparticles for delivering functional food ingredients | |
| Chai et al. | The intelligent delivery systems for bioactive compounds in foods: Physicochemical and physiological conditions, absorption mechanisms, obstacles and responsive strategies | |
| Tang | Assembly of food proteins for nano-encapsulation and delivery of nutraceuticals (a mini-review) | |
| Sarabandi et al. | Protection of casein hydrolysates within nanoliposomes: Antioxidant and stability characterization | |
| FI58579C (en) | ANALYSIS OF HOUSE-AMYLOSSTAERKELSE VID FRAMSTAELLNING AV EN DIETFOEDOAEMNESKOMPOSITION | |
| US20220040258A1 (en) | Gastro-resistant microencapsulates, and uses thereof to stimulate in-vivo ileal glp-1 release in a mammal | |
| Reyhani Poul et al. | Physicochemical and antioxidant properties of chitosan-coated nanoliposome loaded with bioactive peptides produced from shrimp wastes hydrolysis | |
| Wang et al. | The stability and bioavailability of curcumin loaded α-lactalbumin nanocarriers formulated in functional dairy drink | |
| Sarabandi et al. | Fractionation of flaxseed-derived bioactive peptides and their influence on nanoliposomal carriers | |
| Mosquera et al. | Incorporation of liposomes containing squid tunic ACE‐inhibitory peptides into fish gelatin | |
| CN108850790A (en) | A kind of monascorubin microcapsules and preparation method thereof | |
| Xu et al. | Encapsulation of oyster protein hydrolysates in nanoliposomes: Vesicle characteristics, storage stability, in vitro release, and gastrointestinal digestion | |
| CN108578357A (en) | A kind of protein-polysaccharide self-assembled nanometer gel and the preparation method and application thereof with nucleocapsid structure | |
| CN105617358A (en) | Medical foods with antitumor function and preparation method thereof | |
| CN110679953A (en) | Preparation method of nano liposome embedded with egg white source active peptide | |
| CN106978462A (en) | A kind of bionic enzymatic prepares the production method of corn Gly-His-Lys | |
| Jiao et al. | Improvement in entrapment efficiency and in vitro digestion stability of lutein by zein nanocarriers with pepsin hydrolysis | |
| Yang et al. | Stability and bioavailability of protein matrix‐encapsulated astaxanthin ester microcapsules | |
| CN113397175B (en) | Apostichopus japonicus oligopeptide cross-linked microcapsule and preparation method and application thereof | |
| Cao et al. | Chitosan‐coated nanoliposomes for the enhanced stability of walnut angiotensin‐converting enzyme (ACE) inhibitory peptide | |
| Sudareva et al. | Alginate-containing systems for oral delivery of superoxide dismutase. Comparison of various configurations and their properties | |
| JP6410909B2 (en) | Cartilage formation promoter |
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 |