CN110407933A - Graft product and method for the production thereof - Google Patents
Graft product and method for the production thereof Download PDFInfo
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- CN110407933A CN110407933A CN201910565915.XA CN201910565915A CN110407933A CN 110407933 A CN110407933 A CN 110407933A CN 201910565915 A CN201910565915 A CN 201910565915A CN 110407933 A CN110407933 A CN 110407933A
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- 238000000034 method Methods 0.000 title claims description 10
- 238000004519 manufacturing process Methods 0.000 title description 4
- 238000002360 preparation method Methods 0.000 claims abstract description 19
- 239000000047 product Substances 0.000 claims description 51
- 239000000243 solution Substances 0.000 claims description 51
- 102000008186 Collagen Human genes 0.000 claims description 40
- 108010035532 Collagen Proteins 0.000 claims description 40
- 229920001436 collagen Polymers 0.000 claims description 40
- 238000006243 chemical reaction Methods 0.000 claims description 30
- 239000006228 supernatant Substances 0.000 claims description 14
- 238000006467 substitution reaction Methods 0.000 claims description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 13
- -1 methacrylate acid anhydride Chemical class 0.000 claims description 11
- 238000013019 agitation Methods 0.000 claims description 8
- 238000000502 dialysis Methods 0.000 claims description 7
- 238000010790 dilution Methods 0.000 claims description 6
- 239000012895 dilution Substances 0.000 claims description 6
- 108090000623 proteins and genes Proteins 0.000 claims description 6
- 238000004108 freeze drying Methods 0.000 claims description 5
- 239000008367 deionised water Substances 0.000 claims description 4
- 229910021641 deionized water Inorganic materials 0.000 claims description 4
- 102000004169 proteins and genes Human genes 0.000 claims description 4
- 239000012634 fragment Substances 0.000 claims description 3
- 230000002068 genetic effect Effects 0.000 claims description 3
- 239000003292 glue Substances 0.000 claims description 3
- 239000007788 liquid Substances 0.000 claims description 3
- FQPSGWSUVKBHSU-UHFFFAOYSA-N methacrylamide Chemical compound CC(=C)C(N)=O FQPSGWSUVKBHSU-UHFFFAOYSA-N 0.000 claims description 3
- 238000005215 recombination Methods 0.000 claims description 3
- 238000003756 stirring Methods 0.000 claims description 3
- 230000006798 recombination Effects 0.000 claims 2
- QQONPFPTGQHPMA-UHFFFAOYSA-N propylene Natural products CC=C QQONPFPTGQHPMA-UHFFFAOYSA-N 0.000 claims 1
- 239000000126 substance Substances 0.000 abstract description 4
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 7
- 241001465754 Metazoa Species 0.000 description 4
- 238000001228 spectrum Methods 0.000 description 4
- 241000894006 Bacteria Species 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 229920002994 synthetic fiber Polymers 0.000 description 3
- NIXOWILDQLNWCW-UHFFFAOYSA-N 2-Propenoic acid Natural products OC(=O)C=C NIXOWILDQLNWCW-UHFFFAOYSA-N 0.000 description 2
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 238000005033 Fourier transform infrared spectroscopy Methods 0.000 description 2
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 2
- 239000004472 Lysine Substances 0.000 description 2
- 239000002202 Polyethylene glycol Substances 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 150000001408 amides Chemical class 0.000 description 2
- 238000005452 bending Methods 0.000 description 2
- 230000005847 immunogenicity Effects 0.000 description 2
- DCUFMVPCXCSVNP-UHFFFAOYSA-N methacrylic anhydride Chemical compound CC(=C)C(=O)OC(=O)C(C)=C DCUFMVPCXCSVNP-UHFFFAOYSA-N 0.000 description 2
- 239000005445 natural material Substances 0.000 description 2
- 229920001983 poloxamer Polymers 0.000 description 2
- 229920001223 polyethylene glycol Polymers 0.000 description 2
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- SMZOUWXMTYCWNB-UHFFFAOYSA-N 2-(2-methoxy-5-methylphenyl)ethanamine Chemical compound COC1=CC=C(C)C=C1CCN SMZOUWXMTYCWNB-UHFFFAOYSA-N 0.000 description 1
- HRPVXLWXLXDGHG-UHFFFAOYSA-N Acrylamide Chemical compound NC(=O)C=C HRPVXLWXLXDGHG-UHFFFAOYSA-N 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 238000004566 IR spectroscopy Methods 0.000 description 1
- CERQOIWHTDAKMF-UHFFFAOYSA-N Methacrylic acid Chemical compound CC(=C)C(O)=O CERQOIWHTDAKMF-UHFFFAOYSA-N 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 206010034960 Photophobia Diseases 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 150000008065 acid anhydrides Chemical class 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 229910021529 ammonia Inorganic materials 0.000 description 1
- 238000006065 biodegradation reaction Methods 0.000 description 1
- 239000012620 biological material Substances 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000007385 chemical modification Methods 0.000 description 1
- 238000009833 condensation Methods 0.000 description 1
- 230000005494 condensation Effects 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 125000004185 ester group Chemical group 0.000 description 1
- 229930182470 glycoside Natural products 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000009434 installation Methods 0.000 description 1
- 208000013469 light sensitivity Diseases 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 230000005311 nuclear magnetism Effects 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- ARJOQCYCJMAIFR-UHFFFAOYSA-N prop-2-enoyl prop-2-enoate Chemical compound C=CC(=O)OC(=O)C=C ARJOQCYCJMAIFR-UHFFFAOYSA-N 0.000 description 1
- 239000008213 purified water Substances 0.000 description 1
- 239000000376 reactant Substances 0.000 description 1
- 230000001172 regenerating effect Effects 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
Landscapes
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Biochemistry (AREA)
- Gastroenterology & Hepatology (AREA)
- Zoology (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Toxicology (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention provides a grafting product and a preparation method thereof, wherein the molecular formula of the grafting product is as follows:the grafting product prepared according to the embodiment of the invention has the advantages of simple molecular formula, good physical and chemical properties and mechanical properties and wide application.
Description
Technical field
The present invention relates to field of biotechnology more particularly to a kind of graft product and preparation method thereof.
Background technique
Collagen raw material a part used in existing technology is from the natural materials such as animal, another part source
In synthetic material, natural material is the collagen obtained by enzymolytic tissue animal, organ, and immunogenicity is poor, mechanical strength compared with
Difference, and Costco Wholesale is higher.Synthetic material refers mainly to pluronic (Pluronic) and polyethylene glycol (PEG) etc., synthetic material
Although improve mechanical performance, printability, in terms of there is good prospect compared to natural material promote it is thin
Born of the same parents' attachment and proliferation aspect performance want poor.Both raw materials are not easy to prepare graft product, and application field is limited.
Summary of the invention
The present invention is directed at least solve one of the technical problems existing in the prior art.
For this purpose, the present invention proposes that a kind of graft product and preparation method thereof, the graft product are widely used convenient for preparation.
According to the molecular formula of the graft product of invention first aspect embodiment are as follows:
Graft product molecular formula according to an embodiment of the present invention is simple, and physicochemical property and better mechanical property are widely used,
It is easy to implement engineering bacterium expression collagen.
Graft product according to an embodiment of the present invention can also have following additional technical feature:
According to one embodiment of present invention, the graft product is modified by GelMA method by recombined collagen
It is made.
Embodiment also provides a kind of preparation method of graft product according to a second aspect of the present invention, and the preparation method includes
Following steps: S1, recombined collagen is dissolved into PBS solution；S2, methacrylate acid anhydride is added to dissolved with described heavy
In the PBS solution of group collagen, reaction solution is formed；The PBS solution is poured into S3, Xiang Suoshu reaction solution to dilute
It states reaction solution and terminates reaction；S4, the reaction solution after dilution is dialysed, and by the solution after dialysis carry out from
The heart collects supernatant；S5, the supernatant is subjected to frozen dried, obtains graft product.
According to one embodiment of present invention, in step sl, the recombined collagen is by that can translate into people's collagen
The recombinant vector of the genetic fragment of albumen, which is transferred to, to be obtained in the host cell of express express target protein through everfermentation, purifying and freeze-drying
.
According to one embodiment of present invention, in step sl, the recombined collagen is in the PBS solution, In
By magnetic agitation to being completely dissolved under 45 DEG C of -55 DEG C of environment.
According to one embodiment of present invention, step S2 includes: S21, the methacrylate acid anhydride is passed through to dropwise addition addition
Into the PBS solution dissolved with the recombined collagen, time for adding 8min-12min；S22, by step S21's
Solution magnetic agitation in 45 DEG C -55 DEG C of water bath with thermostatic control reacts 2h-4h.
According to one embodiment of present invention, in step s3, when pouring into the PBS solution in Xiang Suoshu reaction solution, no
The disconnected stirring reaction solution is to terminate reaction.
According to one embodiment of present invention, step S4 include: S41, by after dilution the reaction solution be placed in bag filter
In, it dialyses -8 days 6 days and is stirred continuously in deionized water at room temperature；S42, the solution after dialysis is poured into centrifuge tube
In, and 10min-20min is centrifuged at 2000rpm-3000rpm, collect supernatant.
According to one embodiment of present invention, in step s 5, the supernatant is put into frozen dried 2 in freeze drier
It -3 days, obtain the graft product.
According to one embodiment of present invention, different methyl can be obtained using the methacrylate acid anhydride of different volumes
The graft product of acrylamide degree of substitution.
Detailed description of the invention
Fig. 1 is the nuclear magnetic spectrum according to different degree of substitution graft product in the embodiment of the present invention；
Fig. 2 is the preparation method flow chart according to graft product in the embodiment of the present invention.
Fig. 3 is the Water quality analysis curve graph according to different degree of substitution graft product in the embodiment of the present invention；
Fig. 4 is the FT-IR analysis graph according to different degree of substitution graft product in the embodiment of the present invention；
Fig. 5 is according to the graft product outside drawing in the embodiment of the present invention.
Appended drawing reference:
The preparation method 100 of graft product.
Specific embodiment
The embodiment of the present invention is described below in detail, examples of the embodiments are shown in the accompanying drawings, wherein from beginning to end
Same or similar label indicates same or similar element or element with the same or similar functions.Below with reference to attached
The embodiment of figure description is exemplary, and for explaining only the invention, and is not considered as limiting the invention.
In the description of the present invention, it is to be understood that, term " center ", " longitudinal direction ", " transverse direction ", " length ", " width ",
" thickness ", "upper", "lower", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom" "inner", "outside", " up time
The orientation or positional relationship of the instructions such as needle ", " counterclockwise ", " axial direction ", " radial direction ", " circumferential direction " be orientation based on the figure or
Positional relationship is merely for convenience of description of the present invention and simplification of the description, rather than the device or element of indication or suggestion meaning must
There must be specific orientation, be constructed and operated in a specific orientation, therefore be not considered as limiting the invention.In addition, limit
There is the feature of " first ", " second " to can explicitly or implicitly include one or more of the features surely.Of the invention
In description, unless otherwise indicated, the meaning of " plurality " is two or more.
In the description of the present invention, it should be noted that unless otherwise clearly defined and limited, term " installation ", " phase
Even ", " connection " shall be understood in a broad sense, for example, it may be being fixedly connected, may be a detachable connection, or be integrally connected；It can
To be mechanical connection, it is also possible to be electrically connected；It can be directly connected, can also can be indirectly connected through an intermediary
Connection inside two elements.For the ordinary skill in the art, above-mentioned term can be understood at this with concrete condition
Concrete meaning in invention.
Graft product according to an embodiment of the present invention is specifically described in conjunction with attached drawing first below.
Graft product according to the present invention, the molecular formula of graft product are as follows:
It should be noted that for white solid and being soluble in as shown in figure 5, the molecular weight of graft product is more stable
Water.
Graft product molecular formula according to an embodiment of the present invention is simple as a result, physicochemical property and better mechanical property, application
Extensively, it is easy to implement engineering bacterium expression collagen.
According to one embodiment of present invention, graft product is made by recombined collagen by the way that GelMA method is modified.
That is, carry out chemical modification to recombined collagen using GelMA method, light sensitivity and certain is made it have
Mechanical performance, and biodegradation rate is controllable, becomes a kind of excellent biomaterial for medical purpose as a result, can be used for tissue
In engineering and regenerative medicine research.
The preparation method 100 of the graft product of embodiment according to a second aspect of the present invention, as shown in Figure 1, this method is main
The following steps are included:
S1, recombined collagen is dissolved into PBS solution；
S2, methacrylate acid anhydride is added in the PBS solution dissolved with recombined collagen, forms reaction solution；
S3, PBS solution is poured into reaction solution with dilute reaction solution and terminates reaction；
S4, the reaction solution after dilution is dialysed, and the solution after dialysis is centrifuged, collect supernatant；
S5, supernatant is subjected to frozen dried, obtains graft product.
Specifically, graft product is mainly obtained by following formulas:
In one embodiment of the invention, in step sl, recombined collagen is by that can translate into human collagen
The recombinant vector of genetic fragment be transferred to and can be obtained through everfermentation, purifying and freeze-drying in the host cell of express express target protein.
Specifically, recombined collagen uses gene recombination technology, the gene piece of human collagen will can be translated into
The recombinant vector of section, which is transferred to, to express people's glue on a large scale in vitro with the host cell of express express target protein so as to realize
Former albumen not only has the characteristics that the low immunogenicity, biodegradable, good biocompatibility of collagen, and
And it is lacked that animal virus hidden danger, rejection existing for animal sources collagen, unstable quality, molecular weight are not uncertain etc.
Point.
According to one embodiment of present invention, in step sl, recombined collagen is in PBS solution, at 45 DEG C -55 DEG C
By magnetic agitation to being completely dissolved under environment.
Specifically, respectively weigh recombined collagen (5g) be added to containing 50mL PBS solution (pH=7.5) A, B,
C, in tetra- containers of D, magnetic agitation is carried out until being completely dissolved at 50 DEG C.
Further, step S2 includes:
S21, by methacrylate acid anhydride by being added in the PBS solution dissolved with recombined collagen, when dropwise addition
Between be 8min-12min；
S22, by the solution of step S21, magnetic agitation reacts 2h-4h in 45 DEG C -55 DEG C of water bath with thermostatic control.
Specifically, the methacrylate acid anhydride for measuring 50uL, 100uL, 200uL, 600uL respectively is added drop-wise to aforementioned four difference
In container, and controlling time for adding is 10min, and then magnetic agitation reacts 3h in 50 DEG C of thermostat water bath.
In one embodiment of the invention, in step s3, it when pouring into PBS solution into reaction solution, is stirred continuously anti-
Liquid is answered to terminate reaction.Specifically, using 50 DEG C of 200mL PBS dilute reaction solution, and be stirred continuously to terminate reaction.
According to still another embodiment of the invention, step S4 includes:
S41, the reaction solution after dilution is placed in bag filter, is dialysed in deionized water at room temperature -8 days 6 days and continuous
Stirring；
S42, the solution after dialysis is poured into centrifuge tube, and is centrifuged 10min-20min at 2000rpm-3000rpm,
Collect supernatant.
Specifically, first the solution in tetra- containers of A, B, C, D is placed in bag filter (12kDa-14kDa), at room temperature
It dialyses 7 days, and constantly whisks, then the solution after dialysis is poured into centrifuge tube in deionized water, be centrifuged at 2500rpm
15min collects supernatant.
Further, in step s 5, supernatant is put into frozen dried -3 days 2 days in freeze drier, obtains grafting and produces
Object.Specifically, supernatant is put into lyophilized preparation, it is lyophilized 3 days, obtains the sample of different Methacrylamide degree of substitution,
And respectively marked as A, B, C, D.
It should be noted that different Methacrylamide degree of substitution can be obtained using the methacrylate acid anhydride of different volumes
Graft product.
The identification method of graft product (MARHC) degree of substitution is specifically described below with reference to embodiment:
Nuclear magnetic resonance pop method: it is measured whether be grafted success with 1H NMR.RHC and MARHC are dissolved in D2In O, sample
Concentration is 50mg/mL.Chemical shift peak type at 5.5ppm and 1.8ppm changes, and shows to be grafted successfully, this is because first
After amino in base acrylic acid and recombinant collagen and hydroxyl reaction, wherein methyl and acrylic acid proton can be because on nucleus magnetic hydrogen spectrum
Grafting degree is different and peak area is caused to change, and can increase with the increase of metering system acid concentration, meanwhile, chemical shift
What is indicated at 2.9ppm is the variation of lysine methyl signal, and peak area is smaller, the bad ammonia to dissociate on recombined collagen
Acid is fewer.Nuclear magnetic spectrum is as shown in Figure 2.
The table that nuclear-magnetism pop method calculates recombined collagen methacrylic acid degree of substitution is as follows:
| Sample ID | Blank control group | A | B | C | D |
| Area(lysine) | 0.78 | 0.69 | 0.67 | 0.60 | 0.32 |
| Degree of substitution (%) | / | 11 | 14 | 23 | 59 |
Conclusion: with the increase of methyl-prop acid anhydrides amount, the degree of substitution of sample increases to 59% from 11% respectively, shows methyl
Dehydration condensation occurs for the amino on the carboxyl and recombined collagen in acrylic anhydride, so that degree of substitution be made to gradually increase.
Hydrophobicity analysis: analyzing the hydrophobicity of modified sample using C18 column, after blank control group, graft reaction
A, B, C sample difference it is soluble in water, HPLC analysis after map as shown in figure 3, with sample degree of substitution increase, sample appearance
Time is more late, shows that graft product hydrophobicity gradually increases, that is to say, that as methacrylic anhydride is to recombined collagen
Modifying and decorating degree increases, and produces influence to the hydrophobicity of recombinant collagen.
FT-IR analysis: the sample after freeze-drying being placed in liquid nitrogen and freezes 5min, and taking-up is ground into powder, and KBr pressure is added
Piece carries out infrared test, compares recombinant collagen (RHC) and graft product (RHCMA) infrared spectroscopy difference, as shown in figure 4, difference spectrum
Figure is shown: being amide band at 3315cm-1,2956cm-1 is the stretching vibration peak of-CH3, it was demonstrated that there is the presence of methyl, and
It is the bending vibration of-CH (CH3) at 1447cm-1, further demonstrates the presence of methyl, 1651cm-1 is the flexible vibration of C=0
Dynamic peak, it was demonstrated that the presence of ester group is amide II band at 1539cm-1, is drawn by N-H bending vibration and C-N stretching vibration coupling
It rises, this is just illustrating the amido bond that recombined collagen and methacrylic acid anhydride reactant are newly formed, and also further illustrates weight
Formation of group collagen (RHC) through the new graft product (RHCMA) of methacrylic acid glycosides (MA) modified formative.
Physical and chemical performance analysis: as shown in figure 5, in spongy after the freeze-drying of recombined collagen graft product, color be white,
Soluble easily in water, graft product is dissolved in purified water, is configured to the solution of concentration 10mg/mL, weakly acidic (pH range 3-7), and takes
Lower for degree, sample pH value is higher.
To sum up, graft product molecular formula according to an embodiment of the present invention is simple, physicochemical property and better mechanical property,
It is easily prepared, it is widely used, is easy to implement engineering bacterium expression collagen.
The above is a preferred embodiment of the present invention, it is noted that for those skilled in the art
For, without departing from the principles of the present invention, it can also make several improvements and retouch, these improvements and modifications
It should be regarded as protection scope of the present invention.
Claims (10)
1. a kind of graft product, which is characterized in that the molecular formula of the graft product are as follows:
2. graft product according to claim 1, which is characterized in that the graft product is passed through by recombined collagen
GelMA method is modified to be made.
3. a kind of preparation method of graft product, which comprises the following steps:
S1, recombined collagen is dissolved into PBS solution；
S2, methacrylate acid anhydride is added in the PBS solution dissolved with the recombined collagen, forms reaction solution；
The PBS solution is poured into S3, Xiang Suoshu reaction solution to dilute the reaction solution and terminate reaction；
S4, the reaction solution after dilution is dialysed, and the solution after dialysis is centrifuged, collect supernatant；
S5, the supernatant is subjected to frozen dried, obtains graft product.
4. the preparation method of graft product according to claim 3, which is characterized in that in step sl, the recombination glue
Former albumen by by the recombinant vector that can translate into the genetic fragment of human collagen be transferred to can express express target protein host cell
It is interior, it is obtained through everfermentation, purifying and freeze-drying.
5. the preparation method of graft product according to claim 3, which is characterized in that in step sl, the recombination glue
Former albumen is in the PBS solution, by magnetic agitation to being completely dissolved under 45 DEG C of -55 DEG C of environment.
6. the preparation method of graft product according to claim 3, which is characterized in that step S2 includes:
S21, by the methacrylate acid anhydride by being added to the PBS solution dissolved with the recombined collagen
In, time for adding 8min-12min；
S22, by the solution of step S21, magnetic agitation reacts 2h-4h in 45 DEG C -55 DEG C of water bath with thermostatic control.
7. the preparation method of graft product according to claim 3, which is characterized in that in step s3, to the reaction
When pouring into the PBS solution in liquid, the reaction solution is stirred continuously to terminate reaction.
8. the preparation method of graft product according to claim 3, which is characterized in that step S4 includes:
S41, the reaction solution after dilution is placed in bag filter, is dialysed in deionized water at room temperature -8 days 6 days and continuous
Stirring；
S42, the solution after dialysis is poured into centrifuge tube, and is centrifuged 10min-20min at 2000rpm-3000rpm,
Collect supernatant.
9. the preparation method of graft product according to claim 3, which is characterized in that in step s 5, by the supernatant
Frozen dried -3 days 2 days in freeze drier are put into, the graft product is obtained.
10. the preparation method of graft product according to claim 3, which is characterized in that using the first of different volumes
Base propylene acid anhydrides can obtain the graft product of different Methacrylamide degree of substitution.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111962210A (en) * | 2020-06-22 | 2020-11-20 | 华南理工大学 | Polycaprolactone/methacryloylated elastin nanofiber composite membrane and preparation method and application thereof |
| CN114788899A (en) * | 2022-05-07 | 2022-07-26 | 西安德诺海思医疗科技有限公司 | Photocuring anti-adhesion gel and preparation method thereof |
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