CN109464657A - A kind of method that block copolymer carries insulin control release - Google Patents
A kind of method that block copolymer carries insulin control release Download PDFInfo
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- CN109464657A CN109464657A CN201811418470.4A CN201811418470A CN109464657A CN 109464657 A CN109464657 A CN 109464657A CN 201811418470 A CN201811418470 A CN 201811418470A CN 109464657 A CN109464657 A CN 109464657A
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- insulin
- block copolymer
- glucose
- release
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- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 title claims abstract description 173
- 102000004877 Insulin Human genes 0.000 title claims abstract description 87
- 108090001061 Insulin Proteins 0.000 title claims abstract description 87
- 229940125396 insulin Drugs 0.000 title claims abstract description 87
- 229920001400 block copolymer Polymers 0.000 title claims abstract description 49
- 238000000034 method Methods 0.000 title claims abstract description 25
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims abstract description 35
- 239000008103 glucose Substances 0.000 claims abstract description 33
- 108010015776 Glucose oxidase Proteins 0.000 claims abstract description 15
- 239000004366 Glucose oxidase Substances 0.000 claims abstract description 15
- 229940116332 glucose oxidase Drugs 0.000 claims abstract description 15
- 235000019420 glucose oxidase Nutrition 0.000 claims abstract description 15
- 229920000642 polymer Polymers 0.000 claims abstract description 9
- 238000002360 preparation method Methods 0.000 claims abstract description 6
- 239000007788 liquid Substances 0.000 claims abstract description 5
- 230000035945 sensitivity Effects 0.000 claims abstract description 4
- 238000005253 cladding Methods 0.000 claims abstract description 3
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 claims description 13
- 230000002209 hydrophobic effect Effects 0.000 claims description 13
- 239000002253 acid Substances 0.000 claims description 12
- -1 hydroxyl ethyl Chemical group 0.000 claims description 11
- 239000000243 solution Substances 0.000 claims description 11
- 239000007853 buffer solution Substances 0.000 claims description 9
- 238000012360 testing method Methods 0.000 claims description 9
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 150000001875 compounds Chemical class 0.000 claims description 7
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 6
- 239000002202 Polyethylene glycol Substances 0.000 claims description 6
- 229920001223 polyethylene glycol Polymers 0.000 claims description 6
- 230000008569 process Effects 0.000 claims description 6
- 239000004698 Polyethylene Substances 0.000 claims description 5
- 238000000502 dialysis Methods 0.000 claims description 5
- 238000004108 freeze drying Methods 0.000 claims description 5
- 229910052757 nitrogen Inorganic materials 0.000 claims description 5
- 229920000573 polyethylene Polymers 0.000 claims description 5
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 claims description 4
- 238000006243 chemical reaction Methods 0.000 claims description 4
- 239000003960 organic solvent Substances 0.000 claims description 4
- 229910021591 Copper(I) chloride Inorganic materials 0.000 claims description 3
- 238000010560 atom transfer radical polymerization reaction Methods 0.000 claims description 3
- 229920001577 copolymer Polymers 0.000 claims description 3
- OXBLHERUFWYNTN-UHFFFAOYSA-M copper(I) chloride Chemical compound [Cu]Cl OXBLHERUFWYNTN-UHFFFAOYSA-M 0.000 claims description 3
- 238000006392 deoxygenation reaction Methods 0.000 claims description 3
- 238000007710 freezing Methods 0.000 claims description 3
- 230000008014 freezing Effects 0.000 claims description 3
- 239000005457 ice water Substances 0.000 claims description 3
- 239000012528 membrane Substances 0.000 claims description 3
- WKBOTKDWSSQWDR-UHFFFAOYSA-N Bromine atom Chemical compound [Br] WKBOTKDWSSQWDR-UHFFFAOYSA-N 0.000 claims description 2
- YNAVUWVOSKDBBP-UHFFFAOYSA-N Morpholine Natural products C1COCCN1 YNAVUWVOSKDBBP-UHFFFAOYSA-N 0.000 claims description 2
- WNLRTRBMVRJNCN-UHFFFAOYSA-L adipate(2-) Chemical compound [O-]C(=O)CCCCC([O-])=O WNLRTRBMVRJNCN-UHFFFAOYSA-L 0.000 claims description 2
- GDTBXPJZTBHREO-UHFFFAOYSA-N bromine Substances BrBr GDTBXPJZTBHREO-UHFFFAOYSA-N 0.000 claims description 2
- 229910052794 bromium Inorganic materials 0.000 claims description 2
- 239000002131 composite material Substances 0.000 claims description 2
- 229920000359 diblock copolymer Polymers 0.000 claims description 2
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 claims description 2
- YORCZSDBMQPIEU-UHFFFAOYSA-N lead;pyridine Chemical compound [Pb].C1=CC=NC=C1 YORCZSDBMQPIEU-UHFFFAOYSA-N 0.000 claims description 2
- FQPSGWSUVKBHSU-UHFFFAOYSA-N methacrylamide Chemical compound CC(=C)C(N)=O FQPSGWSUVKBHSU-UHFFFAOYSA-N 0.000 claims description 2
- 238000007789 sealing Methods 0.000 claims description 2
- 229920000428 triblock copolymer Polymers 0.000 claims description 2
- 229920001510 poly[2-(diisopropylamino)ethyl methacrylate] polymer Polymers 0.000 claims 2
- OTMSDBZUPAUEDD-UHFFFAOYSA-N Ethane Chemical compound CC OTMSDBZUPAUEDD-UHFFFAOYSA-N 0.000 claims 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims 1
- 238000001035 drying Methods 0.000 claims 1
- 239000001301 oxygen Substances 0.000 claims 1
- 229910052760 oxygen Inorganic materials 0.000 claims 1
- 210000000496 pancreas Anatomy 0.000 claims 1
- 125000003367 polycyclic group Chemical group 0.000 claims 1
- 206010012601 diabetes mellitus Diseases 0.000 abstract description 4
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 abstract description 3
- 201000010099 disease Diseases 0.000 abstract description 2
- 239000002994 raw material Substances 0.000 abstract description 2
- 239000003814 drug Substances 0.000 description 10
- 229940079593 drug Drugs 0.000 description 8
- 239000002953 phosphate buffered saline Substances 0.000 description 8
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- 239000000463 material Substances 0.000 description 5
- 239000008280 blood Substances 0.000 description 4
- 210000004369 blood Anatomy 0.000 description 4
- 239000000693 micelle Substances 0.000 description 4
- RGHNJXZEOKUKBD-UHFFFAOYSA-N D-gluconic acid Natural products OCC(O)C(O)C(O)C(O)C(O)=O RGHNJXZEOKUKBD-UHFFFAOYSA-N 0.000 description 3
- RGHNJXZEOKUKBD-SQOUGZDYSA-N Gluconic acid Natural products OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C(O)=O RGHNJXZEOKUKBD-SQOUGZDYSA-N 0.000 description 3
- 230000002776 aggregation Effects 0.000 description 3
- 238000004220 aggregation Methods 0.000 description 3
- 230000001186 cumulative effect Effects 0.000 description 3
- 108700022423 fluorescein-isothiocyanated- insulin Proteins 0.000 description 3
- 235000012208 gluconic acid Nutrition 0.000 description 3
- 239000000174 gluconic acid Substances 0.000 description 3
- 201000001421 hyperglycemia Diseases 0.000 description 3
- 230000003914 insulin secretion Effects 0.000 description 3
- 230000005588 protonation Effects 0.000 description 3
- 238000001338 self-assembly Methods 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 229920003171 Poly (ethylene oxide) Polymers 0.000 description 2
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 description 2
- 230000003013 cytotoxicity Effects 0.000 description 2
- 231100000135 cytotoxicity Toxicity 0.000 description 2
- 238000005538 encapsulation Methods 0.000 description 2
- 230000010030 glucose lowering effect Effects 0.000 description 2
- 239000000178 monomer Substances 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- 230000010148 water-pollination Effects 0.000 description 2
- RPAJSBKBKSSMLJ-DFWYDOINSA-N (2s)-2-aminopentanedioic acid;hydrochloride Chemical compound Cl.OC(=O)[C@@H](N)CCC(O)=O RPAJSBKBKSSMLJ-DFWYDOINSA-N 0.000 description 1
- 208000007342 Diabetic Nephropathies Diseases 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 206010018473 Glycosuria Diseases 0.000 description 1
- WOBHKFSMXKNTIM-UHFFFAOYSA-N Hydroxyethyl methacrylate Chemical compound CC(=C)C(=O)OCCO WOBHKFSMXKNTIM-UHFFFAOYSA-N 0.000 description 1
- 208000007976 Ketosis Diseases 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000004204 blood vessel Anatomy 0.000 description 1
- 230000036760 body temperature Effects 0.000 description 1
- 230000031709 bromination Effects 0.000 description 1
- 238000005893 bromination reaction Methods 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 239000008121 dextrose Substances 0.000 description 1
- 208000033679 diabetic kidney disease Diseases 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- UAOMVDZJSHZZME-UHFFFAOYSA-N diisopropylamine Chemical compound CC(C)NC(C)C UAOMVDZJSHZZME-UHFFFAOYSA-N 0.000 description 1
- 125000006202 diisopropylaminoethyl group Chemical group [H]C([H])([H])C([H])(N(C([H])([H])C([H])([H])*)C([H])(C([H])([H])[H])C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical group P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 1
- 208000016097 disease of metabolism Diseases 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 230000004064 dysfunction Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000003792 electrolyte Substances 0.000 description 1
- 230000009881 electrostatic interaction Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 229940088598 enzyme Drugs 0.000 description 1
- 125000004494 ethyl ester group Chemical group 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 210000001508 eye Anatomy 0.000 description 1
- 125000000524 functional group Chemical group 0.000 description 1
- 229920001002 functional polymer Polymers 0.000 description 1
- 125000002791 glucosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)[C@H](O1)CO)* 0.000 description 1
- 210000002216 heart Anatomy 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 150000002433 hydrophilic molecules Chemical class 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 230000004140 ketosis Effects 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 208000030159 metabolic disease Diseases 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 229940125395 oral insulin Drugs 0.000 description 1
- 229920002939 poly(N,N-dimethylacrylamides) Polymers 0.000 description 1
- 229920002338 polyhydroxyethylmethacrylate Polymers 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 description 1
- 230000002787 reinforcement Effects 0.000 description 1
- 230000004043 responsiveness Effects 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/43—Enzymes; Proenzymes; Derivatives thereof
- A61K38/44—Oxidoreductases (1)
- A61K38/443—Oxidoreductases (1) acting on CH-OH groups as donors, e.g. glucose oxidase, lactate dehydrogenase (1.1)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/22—Hormones
- A61K38/28—Insulins
-
- 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
- A61K47/34—Macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyesters, polyamino acids, polysiloxanes, polyphosphazines, copolymers of polyalkylene glycol or poloxamers
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/001—Preparation for luminescence or biological staining
- A61K49/0013—Luminescence
- A61K49/0017—Fluorescence in vivo
- A61K49/0019—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules
- A61K49/0021—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules the fluorescent group being a small organic molecule
-
- 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
- A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
- A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y101/00—Oxidoreductases acting on the CH-OH group of donors (1.1)
- C12Y101/03—Oxidoreductases acting on the CH-OH group of donors (1.1) with a oxygen as acceptor (1.1.3)
- C12Y101/03004—Glucose oxidase (1.1.3.4)
Abstract
The invention belongs to technological field of biochemistry, disclose a kind of method that block copolymer carries insulin control release.The described method includes: step 1: preparation pH sensitivity amphipathic nature block polymer makes block copolymer cladding insulin and glucose oxidase step 2: insulin, glucose oxidase and above-mentioned block copolymer are interacted；Step 3: being used for the above-mentioned block copolymer for being loaded with insulin in the liquid environment of the glucose containing various concentration, the release conditions of insulin are different in the solution of different concentration of glucose.The block copolymer preparation for being loaded with insulin in the present invention is simple, and raw material is extensive, easy to spread；The release of concentration control insulin based on glucose, when concentration of glucose increases, the rate of release of insulin is improved；Prospect with the treatment for applying the present invention to the diseases such as diabetes.
Description
Technical field
The invention belongs to the sides that technological field of biochemistry more particularly to a kind of block copolymer carry insulin control release
Method.
Background technique
Insulin is a kind of hormone for reducing blood glucose, secretes to promote sugar to utilize when blood glucose rise and reduces blood glucose.Glycosuria
Disease is to cash the metabolic disease being characterized for hyperglycemia since the opposite or absolute shortage of insulin causes.Hyperglycemia is then
Since defect of insulin secretion or its biological effect are impaired, or both have concurrently and cause.Long-standing hyperglycemia, leads when diabetes
Cause various tissues, especially eye, kidney, heart, blood vessel, the chronic lesion of nerve, dysfunction.Under normal circumstances, Cong Yuhou and
For price, preferred oral medicine glucose-lowering treatment, but there is complication, such as ketosis, diabetic nephropathy or pregnant woman, big hand
When art, need using insulin glucose-lowering treatment.Insulin is protein, can be subcutaneously injected or intravenous drip, directly
Connect oral insulin can be digested enzyme metabolism fall, do not have blood sugar reducing function.
Stimuli responsive polymers belong to one of functional polymer, mostly amphiphilic structure, produce shape in water
The different aggregation of state, such aggregation is under extraneous environmental stimulus, such as pH, temperature, light, molecule, electrolyte, voltage or other
The variation of factor can generate the response variation of specificity.Stimuli responsive nano-micelle medicament carrier system has obtained extensive pass
Note and research reduce the obvious excellent of cytotoxicity, and reinforcement therapeutic effect etc. especially in the targeting conveying for realizing drug
Gesture.
Glucose responding material has potential application in insulin control release field, the Portugal of load insulin
Grape sugar responsiveness material can responsively discharge drug according to concentration of glucose variation in environment, in terms for the treatment of diabetes
It is the research hotspot in macromolecule and biomedical materials field with certain application potential.
The high molecular material of load insulin, the amphipathic copolymer self assembly being made of hydrophilic molecules and hydrophobic molecule,
Each block of amphipathic nature block polymer is usually thermodynamically incompatible, it can be self-assembled into specific in selective solvent
One micella of supramolecular ordered aggregation.The forming process of micella is since hydrophobic effect, electrostatic interaction and hydrogen bond etc. are intermolecular
Active force dredges solvent area and attracts each other association together, forms fine and close kernel, which is equivalent to nano container, hydrophobicity
Drug can greatly improve difficulty and being chemically combined with interior internuclear physics synergistic effect or with hydrophobic section into micelle inner core
The solubility of soluble drug, and the then orderly arrangement of solvophilic area, freely protrude into a large amount of solvent phase, form coating,
To which micella can exist within the scope of a certain concentration with long-time stable.Amphipathic nature block polymer, since it possesses hydrophilic chain
Section and hydrophobic segment, can be self-assembly of the nano-micelle with hydrophobic core hydrophilic outer shell, this special construction in aqueous solution
Make it a kind of important drug conveying carrier.
Summary of the invention
In view of the above technical problems existing in the prior art, the present invention, which provides a kind of block copolymer and carries insulin control, releases
The method put, the method can be realized the control release of insulin.The concentration of glucose plays " switch " function, when glucose is dense
When spending higher, insulin releasing speed is fast, and when concentration of glucose is lower, insulin releasing speed is slow.
The technical solution adopted by the present invention is as described below.
The present invention provides a kind of method that block copolymer carries insulin control release, which comprises
Step 1: preparing PH sensitivity amphipathic nature block polymer using atom transfer radical polymerization；
Step 2: insulin, glucose oxidase and above-mentioned block copolymer are interacted, coat block copolymer
Insulin and glucose oxidase；
Step 3: the above-mentioned block copolymer for being loaded with insulin to be used for the liquid environment of the glucose containing various concentration
In, the release conditions of insulin are different in the solution of different concentration of glucose.
In order to facilitate the release conditions of observation insulin, before above-mentioned second step, by insulin and isosulfocyanic acid fluorescence
Element is compound.
Wherein, by insulin and the compound process of fluorescein isothiocynate are as follows: by insulin, fluorescein isothiocynate
(FITC) co-dissolve in organic solvent, and this solution is slowly added into PBS buffer solution, and 4 DEG C are protected from light 15
Hour, 3 days are dialysed in pure water to remove not compound fluorescein isothiocynate, adjust the pH value of composite solution in 4.5-5.0, so
After be centrifuged, be lyophilized, obtain the insulin compound with fluorescein isothiocynate.Meanwhile it being needed after reaction in second step
Analysis removal fluorescein isothiocynate.
In the above-mentioned first step, the PH sensitivity amphipathic nature block polymer, hydrophilic-structure can be polyethylene glycol
(PEG), one of polyethylene oxide (PEO) or a variety of, hydrophobic structure be polymethylacrylic acid morpholine ethyl ester (PMEMA),
Polydiethylene glycol adipate (PDEA), polymethylacrylic acid diisopropylaminoethyl ethyl ester (PDPA), poly- N, N-2 Methacrylamide
(PDMA), one of poly (glycidylmethacrylate--co-ethylene dimethacrylate) (PGMA), polymethylacrylic acid -2- hydroxyl ethyl ester (PHEMA) or a variety of.
In the above-mentioned first step, the block copolymer is diblock copolymer or triblock copolymer.
In the above-mentioned first step, the block copolymer is block copolymer polyethylene glycol-b- polymethylacrylic acid diisopropyl
Amino ethyl ester (PEG-b-PDPA).Wherein, the block copolymer polyethylene glycol-b- polymethylacrylic acid diisopropylaminoethyl ethyl ester
Preparation process be that bromine replaced into polyethylene glycol (PEG-Br) polymerized monomer and pyridine, under ice-water bath, logical nitrogen deoxygenation, 30 points
Clock is rapidly added CuCl, reacts 5 hours at 50 DEG C, dialyses in pure water later, and freeze-drying obtains block copolymer PEG-b-
PDPA。
In above-mentioned second step, the process of cladding are as follows: be dissolved in above-mentioned block copolymer, insulin, glucose oxidase
Organic solvent in stir 3 hours, react 12 hours, freezing, which is protected from light, dry carries insulin micella.
In above-mentioned third step, it is added by fluophotometer scanning after being loaded with the block copolymer of insulin under different time
The release conditions of insulin, specifically: the block copolymer for being loaded with insulin is placed in test tube, with dialysis membrane sealing end back-off in
In the PBS buffer solution of pH7.4, and keeping temperature is 37 DEG C, and the rate of release and release of insulin are calculated using fluophotometer
Amount.
The present invention is loaded into insulin (IND), glucose oxidase (GOD), fluorescent material FITC by block copolymer
(fluorescein isothiocynate) simulates 37 DEG C of body temperature, and environment is phosphate buffered saline solution (PBS), buffer solution ph 7.4.Pass through
The response of glucose controls the release of insulin.When the concentration of the glucose in liquid environment increases, glucose is able to enter
The inside of micella, because of the catalytic action of glucose oxidase, so that it becomes gluconic acid, reduces the pH value of microenvironment.Block
The hydrophobic side of copolymer is sensitive to pH value, can protonate rapidly as pH value must reduce, and the block copolymer of protonation is dredged
Water end (W.E.) becomes hydrophily from hydrophobicity, micellar structure from closely becoming loose, thus enable to be coated on the insulin at center by
Gradually discharge.In this way, the nano-micelle of self assembly can be according to the partial size of micella, and form, carrying drug ratio, the sensibility of glucose,
The controls such as cytotoxicity discharge insulin.
Glucose oxidase is in glucose induction
Gluconic acid, to reduce the pH of microenvironment.Its reaction process are as follows:
The hydrophobic side of block high polymer is the functional group sensitive to pH, can be hydrionic in the case where pH is reduced
The lower protonation of effect.Hydrophobic side after protonation is changed into hydrophily, but the hydrophobic structure on strand still has hydrophobicity.
Therefore, the hydrophilic end of hydrophobic end conversion can extend, but not micellar structure is made to scatter completely, so that control is slowly released
Insulin is released, rather than disposably all release.
For the block copolymer for being loaded with insulin of the invention, the switching function of its insulin releasing is detected
Concrete mode are as follows:
Entire test tube is placed in constant temperature oscillator, 37 DEG C of water-bath, will be inverted in equipped with the inside test tube for carrying insulin micella
In the external test tube of concentration (choosing 0mg/ml, 10mg/ml) PBS buffer solution equipped with different glucose, and internal test tube
Opening is blocked with bag filter (MWCO8000-14000), surveys insulin Cumulative release amount and release speed using FITC fluorescence calibration
Rate.
The invention has the following beneficial effects:
1, preparation is simple, and raw material is extensive, easy to spread；
2, the release of the concentration control insulin based on glucose, when concentration of glucose increases, the release speed of insulin
Rate improves；
3, with the prospect for applying the present invention to diabetes energy disease.
Detailed description of the invention
Fig. 1 is that the control of insulin in the embodiment of the present invention discharges schematic diagram；
Fig. 2 is the Cumulative release amount curve of insulin in the embodiment of the present invention；
Fig. 3 is the schematic device of detection insulin control release in the embodiment of the present invention；
Fig. 4 is that " ON-OFF " tests switching function figure in the embodiment of the present invention.
Specific embodiment
In order to illustrate more clearly of technical solution of the present invention, it is illustrated below in conjunction with specific embodiments and the drawings,
It should be evident that the embodiment in being described below is only some embodiments of the present invention, those of ordinary skill in the art are come
It says, without creative efforts, other examples can also be obtained according to these embodiments.
Embodiment 1
The block copolymer selected in the present embodiment is block copolymer polyethylene glycol-b- polymethylacrylic acid diisopropyl ammonia
Base ethyl ester (PEG-b-PDPA).
1, atom transfer radical polymerization, bromination well prepared in advance the preparation of block copolymer PEG-b-PDPA: are used
Polyethylene glycol, as the releaser of high polymer, specific synthesis process is as follows: PEG-Br (0.18mmol), polymerized monomer
(2.7mmol) and pyridine lead to nitrogen deoxygenation, 30 minutes, are rapidly added CuCl (0.18mmol) under ice-water bath, anti-at 50 DEG C
It answers 5 hours, dialyses in pure water later, freeze-drying obtains block copolymer PEG-b-PDPA.
2, insulin (148mg), FITC (fluorescein isothiocynate) (20mg) co-dissolve are being dissolved in DMSO (4mL)
In, this total solution is slowly added into PBS buffer solution (PBS, pH 7.4), and 4 DEG C are protected from light 15 hours, dialyse in pure water
3 days removal FITC, adjust the pH value of FITC-insulin in 4.5-5.0, are then centrifuged for, freeze-drying.
3, by above-mentioned block copolymer (50mg), FITC-insulin (15mg) and glucose oxidase (25mg) dissolution
It stirs 3 hours, reacts 12 hours in the DMSO of 2.5mL, dialysis removes FITC, and freezing is protected from light so dry that carry insulin micella.
4, test nanometer carries insulin micella and obtains carrying drug ratio and encapsulation rate
Excitation wavelength and absorbing wavelength are respectively 485nm, 519nm, carrying drug ratio (LE) and encapsulation rate (EE) by calculating
It arrives.
5, the accumulative release of insulin micella FITC-insulin is carried
The accumulative release of insulin is by dialysis membrane (molecular cut-off 8,000-14,000) in pH7.4
It is carried out in PBS buffer solution, concentration of the main glucose in buffer solution is 10mg/ml and 0mg/ml, adds up release
Insulin concentration is come out by fluophotometer (Shimazu RF-5301PC) measuring and calculation, test device such as Fig. 3 institute
Show.The control release principle of insulin is as shown in Figure 1, wherein IND is insulin, and GOD is glucose oxidase, and glucose is
Glucose, glucose acid are gluconic acid.
Fig. 2 is the Cumulative release amount curve of insulin, as seen from the figure, when concentration of glucose is 10mg/ml, insulin
Rate of release is very fast, and when concentration of glucose is 0mg/ml, the rate of release of insulin is slower.Pass through 10mg/ml and 0mg/ml
It can be concluded that the conclusion of " ON-OFF " experiment, as shown in Figure 4 (" ON " is the insulin release that concentration of glucose is 10mg/ml,
" OFF " is the insulin release that concentration of glucose is 0mg/ml), i.e., the insulin releasing in the glucose solution of higher concentration
Comparatively fast, in the dextrose buffer solution of low concentration, insulin releasing is slow.
The foregoing description of the disclosed embodiments enables those skilled in the art to implement or use the present invention.
Various modifications to these embodiments will be readily apparent to those skilled in the art, as defined herein
General Principle can be realized in other embodiments without departing from the spirit or scope of the present invention.Therefore, of the invention
It is not intended to be limited to the embodiments shown herein, and is to fit to and the principles and novel features disclosed herein phase one
The widest scope of cause.
Claims (9)
1. a kind of method that block copolymer carries insulin control release, which is characterized in that the described method includes:
Step 1: preparing pH sensitivity amphipathic nature block polymer using atom transfer radical polymerization；
Step 2: insulin, glucose oxidase and above-mentioned block copolymer are interacted, block copolymer is made to coat pancreas islet
Element and glucose oxidase；
Step 3: the above-mentioned block copolymer for being loaded with insulin is used in the liquid environment of the glucose containing various concentration,
The release conditions of insulin are different in the solution of different concentration of glucose.
2. the method that block copolymer according to claim 1 carries insulin control release, which is characterized in that above-mentioned the
It is before two steps, insulin is compound with fluorescein isothiocynate.
3. the method that block copolymer according to claim 2 carries insulin control release, which is characterized in that by insulin
The compound process with fluorescein isothiocynate are as follows: in organic solvent by insulin, fluorescein isothiocynate co-dissolve, and will
This solution is slowly added into PBS buffer solution, and 4 DEG C are protected from light 15 hours, dialyse in pure water 3 days with remove it is not compound
Fluorescein isothiocynate adjusts the pH value of composite solution in 4.5-5.0, is then centrifuged for, and freeze-drying obtains and fluorescein isothiocynate
Compound insulin；Meanwhile it needing to dialyse after reaction in second step to remove fluorescein isothiocynate.
4. the method that block copolymer according to claim 1 carries insulin control release, which is characterized in that above-mentioned first
In step, the block copolymer being made of hydrophilic-structure and hydrophobic structure, hydrophilic-structure can be polyethylene glycol, polycyclic oxygen
One of ethane is a variety of, and hydrophobic structure is polymethylacrylic acid morpholine ethyl ester, polydiethylene glycol adipate, poly- methyl
Acrylic acid diisopropylaminoethyl ethyl ester, poly- N, N-2 Methacrylamide, poly (glycidylmethacrylate--co-ethylene dimethacrylate), polymethylacrylic acid -2-
One of hydroxyl ethyl ester is a variety of.
5. the method that block copolymer according to claim 1 carries insulin control release, which is characterized in that above-mentioned first
In step, the block copolymer is diblock copolymer or triblock copolymer.
6. the method that block copolymer according to claim 4 carries insulin control release, which is characterized in that above-mentioned first
In step, the block copolymer is block copolymer polyethylene glycol-b- polymethylacrylic acid diisopropylaminoethyl ethyl ester (PEG-b-
PDPA)。
7. the method that block copolymer according to claim 6 carries insulin control release, which is characterized in that the block
The preparation process of copolymer polyethylene glycol-b- polymethylacrylic acid diisopropylaminoethyl ethyl ester is that bromine is replaced polyethylene glycol polymeric list
Body and pyridine lead to nitrogen deoxygenation, 30 minutes, are rapidly added CuCl under ice-water bath, react 5 hours at 50 DEG C, later in pure water
Middle dialysis, freeze-drying obtain block copolymer PEG-b-PDPA.
8. the method that block copolymer according to claim 1 carries insulin control release, which is characterized in that above-mentioned second
In step, the process of cladding are as follows: 3 will be stirred in organic solvent that above-mentioned block copolymer, insulin, glucose oxidase are dissolved in
Hour, it reacts 12 hours, freezing is protected from light drying, must carry insulin micella.
9. the method that block copolymer according to claim 2 carries insulin control release, which is characterized in that above-mentioned third
In step, the release feelings of insulin under different time after being loaded with the block copolymer of insulin are added by fluophotometer scanning
Condition, specifically: the block copolymer for being loaded with insulin is placed in test tube, the PBS with the back-off of dialysis membrane sealing end in pH7.4 is buffered
In liquid, and keeping temperature is 37 DEG C, and the rate of release and burst size of insulin are calculated using fluophotometer.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN115317437A (en) * | 2021-05-11 | 2022-11-11 | 中国科学院上海硅酸盐研究所 | Insulin delivery microneedle based on bilirubin nanomaterial and preparation method thereof |
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