CN102824654A - Double-bioenzyme modified blending biological material containing gelatin and chitosan and preparation method and application thereof - Google Patents
Double-bioenzyme modified blending biological material containing gelatin and chitosan and preparation method and application thereof Download PDFInfo
- Publication number
- CN102824654A CN102824654A CN2012103328038A CN201210332803A CN102824654A CN 102824654 A CN102824654 A CN 102824654A CN 2012103328038 A CN2012103328038 A CN 2012103328038A CN 201210332803 A CN201210332803 A CN 201210332803A CN 102824654 A CN102824654 A CN 102824654A
- Authority
- CN
- China
- Prior art keywords
- gelatin
- solution
- chitosan
- tryrosinase
- blend
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Abstract
The invention provides a double-bioenzyme modified blending biological material containing gelatin and chitosan and a preparation method and application of the double-bioenzyme modified blending biological material. The blending biological material comprises gelatin and chitosan which are used a blending main body and microbial transglutaminase and tyrosinase which are used as a modifier. The stability of the blending biological material is enhanced, and the mechanical property thereof is improved; furthermore, the material has antimicrobial activity, and has good application prospect and excessively wide application in the fields of biomedical materials and a biomedicine, such as the reduction of infection and inflammatory response, antibiosis, wound protection, a biologically degradable skin dressing, a wound healing agent, artificial skin, and the like; and meanwhile, the material is simple in preparation technology, easy to control, free from side and toxic effects and temperate in preparation condition.
Description
Technical field
The invention belongs to bio-medical material, polymer chemistry and biochemical field, particularly relate to a kind of gelatin that adopts twin thing enzyme modification and chitosan blend biomaterial and preparation method thereof and application.
Background technology
Gelatin is as the natural polymer and the biological macromolecule material of nature source rich in protein class; It has excellent biological compatibility, nontoxic, biodegradable, and advantage such as can be absorbed by the body and cheap is widely used in biomedical materials field, and especially the aminoacid composition of gelatin and ratio and human body skin are approaching; Therefore at biological dressing; Skin tissue engineering such as Wound-healing agent and artificial skin aspect has good application prospects [functional material, 2004,35:2310-2313]; Yet gelatin film fragility is big, it is broken to be prone to, its application that has been prone to grow drawbacks limit such as bacterial infection.Therefore, the mechanical performance and the anti-microbial property that improve gelatin have crucial meaning to expand its application at biomedical sector.Chitosan is the deacetylated product of the N-of chitin; Also be the natural polymer and the biological macromolecule material of the abundant biological polyoses class in a kind of occurring in nature source; Have good film property, biocompatibility, degradability; Wide material sources property also has unique broad spectrum antibacterial performance [International Journal of Food Microbiology, 2000,62 (1-2): 139-148].Therefore, gelatin and chitosan blend can be formed the mutual supplement with each other's advantages between each component of material, not only help improving the mechanical performance of gelatin-based material, particularly help improving the gelatin-based material and be prone to grow the shortcoming with bacterial infection.
In the preparation process of gelatin and chitosan blend material, often need carry out crosslinked mechanical performance and the stability [Food Chemistry, 2007,103:295 – 300] that improves intermingling material.Yet; Though traditional chemical crosslinking can effectively improve the mechanical performance and the stability of material; But because common chemical cross-linking agent aldehydes (formaldehyde, glutaraldehyde and glyceraldehyde) for example, epoxide component and carbodiimides etc. often have cytotoxicity; This biomaterial that will cause the Chemical Crosslinking Methods preparation influences the growth of receptor normal structure after implanting.In view of the foregoing, in the preparation process of material, use natural and avirulent dressing agent, have crucial meaning for the good biomaterial of preparation biocompatibility.Enzyme has a series of advantage as biocatalyzer: enzyme is a kind of biocatalyzer, does not have side reaction, can the toxigenicity problem; Reaction condition is gentle, and reactions step is simple; Catalytic efficiency is high, and specificity is strong, and is widely used in industries such as food, medical inspection, biological manufacturing.Therefore, whether can find suitable enzyme that gelatin and chitosan blend material are modified and become the research of this area focus.
Summary of the invention
The technical problem that the present invention will solve is: a kind of gelatin that adopts twin thing enzyme modification and chitosan blend biomaterial and preparation method thereof and application are provided; Gelatin and chitosan blend material are through behind the twin thing enzyme modification; The stability of material and mechanics mechanical performance improve, and this material has good antibacterial activity.
The present invention solves the problems of the technologies described above the technical scheme of being taked to be:
A kind of gelatin and chitosan blend biomaterial that adopts twin thing enzyme modification is characterized in that said pair of enzyme is respectively microbial transglutaminase and tryrosinase.
Adopt the method for preparing of the gelatin and the chitosan blend biomaterial of twin thing enzyme modification, it is characterized in that it comprises following concrete steps:
1) preparation of solution:
Press gelatin: deionized water=2～6g:50～100mL, choose gelatin and deionized water, subsequent use; Gelatin is joined in the deionized water, after gelatin fully dissolves, obtained gelatin solution in 1～2 hour 40～60 ℃ stirred in water bath;
Press chitosan: 1% (percent by volume) acetic acid solution=1～2g:50～100mL, choose chitosan and acetic acid solution, subsequent use; Chitosan is joined in the acetic acid solution, stir under the room temperature and fully dissolved to chitosan in 3～5 hours, in solution, drip 0.5～1.0mol/L sodium hydroxide solution then, the pH of regulator solution is 5.0～6.0, obtains chitosan solution;
Press microbial transglutaminase: deionized water=10～20U:5～10mL, choose microbial transglutaminase and deionized water, subsequent use; Microbial transglutaminase is joined in the deionized water, stir fully dissolving in 15～30 minutes under the room temperature, refilter and obtain microbial transglutaminase solution;
By tryrosinase: pH is PBS buffer solution=5000～10000U:5～10mL of 7.0, chooses tryrosinase and pH and be 7.0 PBS buffer solution, subsequent use; It is in 7.0 the PBS buffer solution that tryrosinase is dissolved in said pH, stirs under the room temperature and after tryrosinase fully dissolves, obtains tryrosinase solution in 2～5 minutes;
2) blend of gelatin and chitosan:
Press gelatin solution: chitosan solution=50～100mL:50～100mL, choose the gelatin solution and the chitosan solution of above-mentioned preparation, subsequent use; Gelatin solution and chitosan solution equal-volume are mixed, and 40～60 ℃ were stirred 15～30 minutes down, made the solution mix homogeneously, obtained gelatin and chitosan blend solution;
3) preparation of the gelatin of twin thing enzyme modification and chitosan blend material:
Press gelatin and chitosan blend solution: microbial transglutaminase: tryrosinase=50～100mL:3～8U:3000～7000U, choose above-mentioned gelatin and chitosan blend solution, microbial transglutaminase and tryrosinase, subsequent use; In gelatin and chitosan blend solution, add mentioned microorganism T-5398 and tryrosinase; 28～30 ℃ of following stirring reactions 10～20 minutes; Reactant liquor is poured in the culture dish, and aeration-drying at room temperature 48～72 hours obtains the gelatin and the chitosan blend material diaphragm of twin thing enzyme modification; Then diaphragm was soaked in 0.1～0.5mol/L sodium hydroxide solution 0.5～1 hour; Reuse deionized water rinsing diaphragm, 30～40 ℃ were descended dry 12～24 hours, obtained the gelatin and the chitosan blend biomaterial of the twin thing enzyme modification of said employing.
The gelatin and the chitosan blend biomaterial that adopt twin thing enzyme modification are at dressing for skin, Wound-healing agent, or the application of artificial skin aspect.
The invention has the beneficial effects as follows:
1) the present invention adopts two enzyme (microbial transglutaminase and tryrosinase) that gelatin and chitosan blend material are modified; The advantage that not only has the enzyme method of modifying; And because this material adopts the synergistically modified effect of two kinds of enzymes; Compare as dressing agent with a kind of enzyme of independent use and also to have following advantage: thus microbial transglutaminase can make and between hydroxyl amide groups and the epsilon-amino on the lysine on the gamma-glutamic acid in the gelatin molecule association reaction takes place and form cross-linked network; And tryrosinase is through being oxidized to the tyrosine residue in the gelatin molecule quinones residue, then with chitosan molecule in the amino reaction form covalently bound.Therefore,, not only can gelatin molecule and chitosan molecule be formed stable covalently boundly, also form stable cross-linked network simultaneously, thereby improve the stability of material greatly through the synergism of two kinds of enzyme.In addition; This covalently bound and cross-linked network that forms through the synergistically modified effect of two enzyme; Mechanics mechanical performance that can also reinforcing material; And the stability of material and mechanics mechanical performance improve, just greatly expansion material in the purposes and the range of application of bio-medical material and biomedical sector.And; Because microbial transglutaminase is cheap and easy to get; Therefore adopt two enzyme (microbial transglutaminase and tryrosinase) can when improving material settling out performance and mechanical property, can also effectively reduce the production cost of material as dressing agent.
2) the prepared biomaterial of the present invention all is made up of biomacromolecule; What wherein the blend material of main part adopted is gelatin and chitosan; What dressing agent adopted is enzyme, and therefore this biomaterial that all is made up of biomacromolecule has excellent biological compatibility, and no cytotoxicity.
3) gelatin is a nature source rich in protein class biological macromolecule material; Chitosan is the abundant polysaccharide biological macromolecule material in nature source; All have excellent biological compatibility, nontoxic, biodegradable, can be absorbed by the body and advantage such as cheap; Chitosan also has unique broad spectrum antibacterial performance, and this material by protein and biological polyoses blend preparation can form the mutual supplement with each other's advantages between each component of material; Not only help improving the mechanical performance of gelatin-based material, particularly help improving the gelatin-based material and be prone to grow the shortcoming with bacterial infection.
4) enzyme has avoided adopting traditional chemical dressing agent problem, the problem includes: the problem that biocompatibility is poor, toxicity is big as the bio-modification agent; Do not need complicated instrument and equipment, reaction condition is gentle, and reactions step is simple, and enzyme has high efficiency and specificity.
5) method for preparing of the present invention is simple, is easy to control, has no side effect, and preparation condition is gentle.
6) the present invention adopts the gelatin and the chitosan blend biomaterial of twin thing enzyme modification; Purposes at bio-medical material and biomedical sector; Particularly reducing infection and inflammatory reaction, antibiotic, protection wound, biodegradable dressing for skin and Wound-healing agent, and bio-medical material such as artificial skin has a good application prospect with biomedical sector and ten minutes purposes widely.
Description of drawings
Fig. 1 does not adopt enzyme to modify, adopt separately the comparison diagram of the hot strength of the employing microbial transglutaminase of a kind of enzyme modification and embodiment 1 gained and gelatin that tryrosinase is modified and chitosan blend material; Wherein the gelatin and the chitosan blend material of enzyme modification do not adopted in the G/CS representative; Gelatin and chitosan blend material that on behalf of microbial transglutaminase, m-G/CS modify; Gelatin and chitosan blend material that on behalf of tryrosinase, t-G/CS modify, gelatin and chitosan blend material that the two enzyme (microbial transglutaminase and tryrosinase) of mt-G/CS representative are modified.
Fig. 2 is a gelatin materials; And the gelatin of the twin thing enzyme modification of employing that obtains of embodiment 1 and chitosan blend material are to colibacillary fungistatic effect comparison diagram; Wherein the Control representative does not add the blank group of sample; G represents the gelatin materials of unmodified, gelatin and chitosan blend material that the two enzyme (microbial transglutaminase and tryrosinase) of mt-G/CS representative are modified.
Fig. 3 is a gelatin materials; And the gelatin of the twin thing enzyme modification of employing that obtains of embodiment 1 and chitosan blend material are to the fungistatic effect comparison diagram of staphylococcus aureus; Wherein the Control representative does not add the blank group of sample; G represents the gelatin materials of unmodified, gelatin and chitosan blend material that the two enzyme (microbial transglutaminase and tryrosinase) of mt-G/CS representative are modified.
The specific embodiment
Below in conjunction with accompanying drawing and embodiment the present invention is done further description, certain following embodiment should not be construed as limitation of the present invention.
The present invention provides a kind of gelatin and chitosan blend biomaterial that adopts twin thing enzyme modification; It adopts biological macromolecule material to constitute as intermingling material and dressing agent fully; What the blend material of main part adopted is gelatin and chitosan; Adopt two kinds of enzyme then simultaneously, promptly microbial transglutaminase and tryrosinase be as dressing agent, obtains after gelatin and chitosan blend biomaterial are modified.Below in conjunction with specific embodiment method for preparing of the present invention is described further, but does not limit the present invention.
Embodiment 1:
A kind of method for preparing that adopts twin thing enzyme modification gelatin and chitosan blend biomaterial, it comprises following concrete steps:
1) preparation of solution:
Take by weighing the 2g gelatin and join in the 50mL deionized water, dissolve fully to gelatin 50 ℃ stirred in water bath 1 hour and obtain the gelatin solution that concentration is 0.04g/mL; Taking by weighing the 1g chitosan joins in 50mL 1% (percent by volume v/v%) acetic acid solution; Stir under the room temperature and fully dissolved to chitosan in 3 hours; Obtain the chitosan solution that concentration is 0.02g/mL, the pH that in chitosan solution, slowly drips 1mol/L sodium hydroxide solution to solution is 5.5; Microbial transglutaminase joined in the deionized water under the room temperature stirred 10 minutes, refilter the microbial transglutaminase solution that obtains 2U/mL; Tryrosinase is joined in PBS (pH7.0) buffer solution, stirred 2 minutes under the room temperature, obtain the tryrosinase solution of 1000U/mL.
2) blend of gelatin and chitosan:
The gelatin solution and the chitosan solution equal-volume of measuring the above-mentioned preparation of 50 mL respectively mix, and 50 ℃ were stirred 30 minutes down, made the solution mix homogeneously, obtained gelatin and chitosan blend solution.
3) preparation of the gelatin of twin thing enzyme modification and chitosan blend material:
In the blend solution of 100mL gelatin and chitosan, add 4U microbial transglutaminase and 5000U tryrosinase; 28 ℃ of following stirring reactions 15 minutes; Reactant liquor is poured in the culture dish, and aeration-drying at room temperature 72 hours obtains the gelatin and the chitosan blend material diaphragm of twin thing enzyme modification; Then diaphragm was soaked 0.5 hour in the 0.1mol/L sodium hydroxide solution; The a large amount of deionized water rinsing diaphragms of reuse, 30 ℃ were descended dry 12 hours, obtained adopting the gelatin and the chitosan blend biomaterial of twin thing enzyme modification.
Fig. 1 does not adopt enzyme to modify, adopt separately the comparison diagram of the hot strength of the employing microbial transglutaminase of a kind of enzyme modification and embodiment 1 gained and gelatin that tryrosinase is modified and chitosan blend material.
Test process is: choose smooth, clean, flawless material film, be cut into the horizontal stripe of 70mm * 1Omm.MEASUREMENTS OF THIN thickness, each sample are measured six points in gauge length, get its meansigma methods, as the thickness of film.Place relative humidity to be about 50% exsiccator on sample, be positioned under 25 ℃ of conditions balance 24 hours.Adopt universal testing machine (CMT-6503, the material tests company limited is newly thought carefully in Shenzhen) to carry out mechanics performance determining then, initial folder is apart from being set at 30mm, and rate of extension is set at 40mm/min.The mechanical property of different samples is all parallel does 6 appearance, gets 6 results' meansigma methods.As can be seen from Figure 1; Not adopting the gelatin of enzyme modification and the hot strength of chitosan blend material G/CS is 30.75MPa; And adopt the gelatin of microbial transglutaminase enzyme modification and the hot strength of chitosan blend material m-G/CS separately is 43.09MPa; The gelatin of employing tryrosinase modification and the hot strength of chitosan blend material t-G/CS are 39.97MPa separately, and this explanation adopts the gelatin of a kind of enzyme (microbial transglutaminase or tryrosinase) modification and the hot strength of chitosan blend material to be improved separately.Adopting common gelatin of modifying of two enzyme (microbial transglutaminase and tryrosinase) and the hot strength of chitosan blend material mt-G/CS is 50.06MPa; With G/CS mutually specific tensile strength improved 62.80%; Above result shows that the twin thing enzyme modification method that the present invention adopts can obviously improve the mechanical property of gelatin and chitosan blend material.
Fig. 2 is a gelatin materials, and the gelatin of the twin thing enzyme modification of employing that obtains of embodiment 1 and chitosan blend material are to colibacillary fungistatic effect comparison diagram.
Its test process is with reference to People's Republic of China's light industry standard: QB/T 2591-2003 " antibiotic plastic-anti-microbial property method of testing and antibacterial effect ".Concrete test process is: with fresh bacterial cultures (transferring in 24 hours), add in the culture fluid, take turns doing 10 times and increase progressively dilution, selecting bacterial concentration is (5.0～10.0) * 10
5The diluent of CFU/mL is used bacterium liquid as test.In each culture tube, add 2mL bacterium liquid; Add 1 gelatin diaphragm and the gelatin and the chitosan blend diaphragm that adopt twin thing enzyme modification respectively; And do not add diaphragm-operated culture tube as the blank group, after 24 hours, calculate the bacterium colony number 37 ℃ of constant-temperature shaking culture with colony counting method.Every group of data are done twice parallel laboratory test, and each sample is done three parallel antibacterial tests.Wherein antibiotic rate (%)=(the blank sample average reclaims bacterium number-antibiotic sample average and reclaims the bacterium number)/blank sample average reclaims bacterium number * 100%.
As can beappreciated from fig. 2, the blank culture medium that does not add sample of Control representative is 6.48 * 10 through colibacillary concentration after the incubated overnight
7CFU/mL, and the gelatin materials of the adding unmodified of G representative is 8.22 * 10 through the e. coli concentration after the incubated overnight
7CFU/mL, the gelatin-based material of this explanation unmodified does not only have colibacillary antibacterial activity, and it can promote colibacillary growth and breeding on the contrary.The gelatin and the chitosan blend material of the twin thing enzyme modification of adding of mt-G/CS representative are 5.70 * 10 through the e. coli concentration after the incubated overnight
7CFU/mL compares with the gelatin materials G of unmodified, and antibiotic rate has improved 30.66%.Above result shows that the gelatin and the chitosan blend material of the resulting twin thing enzyme modification of the present invention have antibacterial activity to escherichia coli.
Fig. 3 is a gelatin materials, and the gelatin of the twin thing enzyme modification of employing that obtains of embodiment 1 and chitosan blend material are to the fungistatic effect comparison diagram of staphylococcus aureus.
Its test process is with reference to People's Republic of China's light industry standard: QB/T 2591-2003 " antibiotic plastic-anti-microbial property method of testing and antibacterial effect ".Concrete test process is: with fresh bacterial cultures (transferring in 12 hours), add in the culture fluid, take turns doing 10 times and increase progressively dilution, selecting bacterial concentration is (1.0～5.0) * 10
9The diluent of CFU/mL is used bacterium liquid as test.In each culture tube, add 3mL bacterium liquid; Add 1 gelatin diaphragm and the gelatin and the chitosan blend diaphragm that adopt twin thing enzyme modification respectively; And do not add diaphragm-operated culture tube as the blank group, after 12 hours, calculate the bacterium colony number 37 ℃ of constant-temperature shaking culture with colony counting method.Every group of data are done twice parallel laboratory test, and each sample is done three parallel antibacterial tests.Wherein antibiotic rate (%)=(the blank sample average reclaims bacterium number-antibiotic sample average and reclaims the bacterium number)/blank sample average reclaims bacterium number * 100%.
As can beappreciated from fig. 3, the blank culture medium that does not add sample of Control representative is 2.79 * 10 through the concentration of staphylococcus aureus after the incubated overnight
11CFU/mL, and the gelatin materials of the adding unmodified of G representative is 3.71 * 10 through the staphylococcus aureus concentration after the incubated overnight
11CFU/mL, the gelatin-based material of this explanation unmodified does not only have the antibacterial activity to staphylococcus aureus, and it can promote growth and the breeding of staphylococcus aureus on the contrary.The gelatin and the chitosan blend material of the twin thing enzyme modification of adding of mt-G/CS representative are 2.26 * 10 through the staphylococcus aureus concentration after the incubated overnight
11CFU/mL compares with the gelatin materials G of unmodified, and antibiotic rate has improved 39.08%.Above result shows that the gelatin and the chitosan blend material of the resulting twin thing enzyme modification of the present invention have antibacterial activity to staphylococcus aureus.
Embodiment 2:
A kind of method for preparing that adopts twin thing enzyme modification gelatin and chitosan blend biomaterial, it comprises following concrete steps:
1) preparation of solution:
Take by weighing the 3g gelatin and join in the 50mL deionized water, dissolve fully to gelatin 40 ℃ stirred in water bath 1.5 hours and obtain the gelatin solution that concentration is 0.06g/mL; Take by weighing the 1.5g chitosan and join 50mL 1% (in (percent by volume v/v%) acetic acid solution; Stir under the room temperature and fully dissolved to chitosan in 4 hours; Obtain the chitosan solution that concentration is 0.03g/mL, the pH that in chitosan solution, slowly drips 0.5mol/L sodium hydroxide solution to solution is 5.5; Microbial transglutaminase is added in the deionized water relaxing the bowels with purgatives of warm nature stirred 15 minutes, refilter the microbial transglutaminase solution that obtains 2U/mL; Tryrosinase is added in PBS (pH7.0) buffer solution, stirred 3 minutes under the room temperature, obtain the tryrosinase solution of 1000U/mL.
2) blend of gelatin and chitosan:
The gelatin solution and the chitosan solution equal-volume of measuring the above-mentioned preparation of 50 mL respectively mix, and 40 ℃ were stirred 30 minutes down, made the solution mix homogeneously, obtained gelatin and chitosan blend solution.
3) preparation of the gelatin of twin thing enzyme modification and chitosan blend material:
In the blend solution of 100mL gelatin and chitosan, add 6U microbial transglutaminase and 3000U tryrosinase; 28 ℃ of following stirring reactions 20 minutes; Reactant liquor is poured in the culture dish, and aeration-drying at room temperature 48 hours obtains the gelatin and the chitosan blend material diaphragm of twin thing enzyme modification; Then diaphragm was soaked 45 minutes in the 0.2mol/L sodium hydroxide solution; The a large amount of deionized water rinsing diaphragms of reuse, 35 ℃ were descended dry 18 hours, and obtained a kind of gelatin and chitosan blend biomaterial that adopts twin thing enzyme modification.
Embodiment 3:
A kind of method for preparing that adopts twin thing enzyme modification gelatin and chitosan blend biomaterial, it comprises following concrete steps:
1) preparation of solution:
Take by weighing the 6g gelatin and join in the 100mL deionized water, dissolve fully to gelatin 60 ℃ stirred in water bath 1.5 hours and obtain the gelatin solution that concentration is 0.06g/mL; Take by weighing the 2g chitosan and join 100mL 1% (in (percent by volume v/v%) acetic acid solution; Stir under the room temperature and fully dissolved to chitosan in 5 hours; Obtain the chitosan solution that concentration is 0.02g/mL, the pH that in chitosan solution, slowly drips 0.8mol/L sodium hydroxide solution to solution is 5.5; Microbial transglutaminase added under the deionized water room temperature stirred 20 minutes, refilter the microbial transglutaminase solution that obtains 2U/mL; Tryrosinase is added in PBS (pH7.0) buffer solution, stirred 5 minutes under the room temperature, obtain 1000U/mL tryrosinase solution.
2) blend of gelatin and chitosan:
The gelatin solution and the chitosan solution equal-volume of measuring the above-mentioned preparation of 100mL respectively mix, and 60 ℃ were stirred 30 minutes down, made the solution mix homogeneously, obtained gelatin and chitosan blend solution.
3) preparation of the gelatin of twin thing enzyme modification and chitosan blend material:
In the blend solution of 100 mL gelatin and chitosan, add 8U microbial transglutaminase and 4000U tryrosinase; 30 ℃ of following stirring reactions 10 minutes; Reactant liquor is poured in the culture dish, and aeration-drying at room temperature 60 hours obtains the gelatin and the chitosan blend material diaphragm of twin thing enzyme modification; Then diaphragm was soaked 1 hour in the 0.5mol/L sodium hydroxide solution; The a large amount of deionized water rinsing diaphragms of reuse, 40 ℃ were descended dry 24 hours, obtained adopting the gelatin and the chitosan blend biomaterial of twin thing enzyme modification.
Need to prove that those of ordinary skill in the art should be appreciated that and can make amendment or be equal to replacement technical scheme of the present invention, and do not break away from the aim and the scope of technical scheme of the present invention, it all should be encompassed in the middle of the claim scope of the present invention.
Claims (3)
1. a gelatin and a chitosan blend biomaterial that adopts twin thing enzyme modification is characterized in that said pair of enzyme is respectively microbial transglutaminase and tryrosinase.
2. the gelatin of the twin thing enzyme modification of employing as claimed in claim 1 and the method for preparing of chitosan blend biomaterial is characterized in that it comprises following concrete steps:
1) preparation of solution:
Press gelatin: deionized water=2～6g:50～100mL, choose gelatin and deionized water, subsequent use; Gelatin is joined in the deionized water, after gelatin fully dissolves, obtained gelatin solution in 1～2 hour 40～60 ℃ stirred in water bath;
Press chitosan: 1% (percent by volume) acetic acid solution=1～2g:50～100mL, choose chitosan and acetic acid solution, subsequent use; Chitosan is joined in the acetic acid solution, stir under the room temperature and fully dissolved to chitosan in 3～5 hours, in solution, drip 0.5～1.0mol/L sodium hydroxide solution then, the pH of regulator solution is 5.0～6.0, obtains chitosan solution;
Press microbial transglutaminase: deionized water=10～20U:5～10mL, choose microbial transglutaminase and deionized water, subsequent use; Microbial transglutaminase is joined in the deionized water, stir fully dissolving in 15～30 minutes under the room temperature, refilter and obtain microbial transglutaminase solution;
By tryrosinase: pH is PBS buffer solution=5000～10000U:5～10mL of 7.0, chooses tryrosinase and pH and be 7.0 PBS buffer solution, subsequent use; It is in 7.0 the PBS buffer solution that tryrosinase is dissolved in said pH, stirs under the room temperature and after tryrosinase fully dissolves, obtains tryrosinase solution in 2～5 minutes;
2) blend of gelatin and chitosan:
Press gelatin solution: chitosan solution=50～100mL:50～100mL, choose the gelatin solution and the chitosan solution of above-mentioned preparation, subsequent use; Gelatin solution and chitosan solution equal-volume are mixed, and 40～60 ℃ were stirred 15～30 minutes down, made the solution mix homogeneously, obtained gelatin and chitosan blend solution;
3) preparation of the gelatin of twin thing enzyme modification and chitosan blend material:
Press gelatin and chitosan blend solution: microbial transglutaminase: tryrosinase=50～100mL:3～8U:3000～7000U, choose above-mentioned gelatin and chitosan blend solution, microbial transglutaminase and tryrosinase, subsequent use; In gelatin and chitosan blend solution, add mentioned microorganism T-5398 and tryrosinase; 28～30 ℃ of following stirring reactions 10～20 minutes; Reactant liquor is poured in the culture dish, and aeration-drying at room temperature 48～72 hours obtains the gelatin and the chitosan blend material diaphragm of twin thing enzyme modification; Then diaphragm was soaked in 0.1～0.5mol/L sodium hydroxide solution 0.5～1 hour; Reuse deionized water rinsing diaphragm, 30～40 ℃ were descended dry 12～24 hours, obtained the gelatin and the chitosan blend biomaterial of the twin thing enzyme modification of said employing.
3. the gelatin of the twin thing enzyme modification of employing as claimed in claim 1 and chitosan blend biomaterial are at dressing for skin, Wound-healing agent, or the application of artificial skin aspect.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210332803.8A CN102824654B (en) | 2012-09-11 | 2012-09-11 | Double-bioenzyme modified blending biological material containing gelatin and chitosan and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210332803.8A CN102824654B (en) | 2012-09-11 | 2012-09-11 | Double-bioenzyme modified blending biological material containing gelatin and chitosan and preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102824654A true CN102824654A (en) | 2012-12-19 |
| CN102824654B CN102824654B (en) | 2014-07-02 |
Family
ID=47328099
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201210332803.8A Expired - Fee Related CN102824654B (en) | 2012-09-11 | 2012-09-11 | Double-bioenzyme modified blending biological material containing gelatin and chitosan and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102824654B (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103980500A (en) * | 2014-04-23 | 2014-08-13 | 湖南尔康北山明胶有限公司 | Protein-grafted natural polysaccharide, preparation method and application thereof |
| CN104233436A (en) * | 2014-09-15 | 2014-12-24 | 武汉理工大学 | Chitosan/gelatin/nano-silver conductive antibacterial material and preparation method thereof |
| CN106519271A (en) * | 2016-11-29 | 2017-03-22 | 吉林大学 | Method for preparing antibacterial membrane by N-succinyl-chitosan immobilized lysozyme |
| CN109295137A (en) * | 2018-10-17 | 2019-02-01 | 四川大学 | A kind of method that the continuous enzymatic microfibrillar collagen of multistep is cross-linking modified |
| CN113773654A (en) * | 2021-09-17 | 2021-12-10 | 河南迪怡疗护科技开发有限公司 | Antibacterial material and preparation method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2011077388A1 (en) * | 2009-12-22 | 2011-06-30 | Lifebond Ltd | Modification of enzymatic crosslinkers for controlling properties of crosslinked matrices |
| CN102552967A (en) * | 2006-12-15 | 2012-07-11 | 生命连结有限公司 | Gelatin-transglutaminase hemostatic dressings and sealants |
-
2012
- 2012-09-11 CN CN201210332803.8A patent/CN102824654B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102552967A (en) * | 2006-12-15 | 2012-07-11 | 生命连结有限公司 | Gelatin-transglutaminase hemostatic dressings and sealants |
| WO2011077388A1 (en) * | 2009-12-22 | 2011-06-30 | Lifebond Ltd | Modification of enzymatic crosslinkers for controlling properties of crosslinked matrices |
Non-Patent Citations (1)
| Title |
|---|
| 周小华: "壳聚糖-明胶共聚物的酶促合成及抑菌性质", 《应用化学》 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103980500A (en) * | 2014-04-23 | 2014-08-13 | 湖南尔康北山明胶有限公司 | Protein-grafted natural polysaccharide, preparation method and application thereof |
| CN103980500B (en) * | 2014-04-23 | 2016-08-24 | 湖南尔康北山明胶有限公司 | A kind of protein grafting natural polysaccharide as well as preparation method and application thereof |
| CN104233436A (en) * | 2014-09-15 | 2014-12-24 | 武汉理工大学 | Chitosan/gelatin/nano-silver conductive antibacterial material and preparation method thereof |
| CN106519271A (en) * | 2016-11-29 | 2017-03-22 | 吉林大学 | Method for preparing antibacterial membrane by N-succinyl-chitosan immobilized lysozyme |
| CN106519271B (en) * | 2016-11-29 | 2019-03-05 | 吉林大学 | A kind of method of the immobilized lysozyme preparation bacteriostatic film of N- succinyl-chitosan |
| CN109295137A (en) * | 2018-10-17 | 2019-02-01 | 四川大学 | A kind of method that the continuous enzymatic microfibrillar collagen of multistep is cross-linking modified |
| CN113773654A (en) * | 2021-09-17 | 2021-12-10 | 河南迪怡疗护科技开发有限公司 | Antibacterial material and preparation method thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102824654B (en) | 2014-07-02 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Zhang et al. | Zn2+-loaded TOBC nanofiber-reinforced biomimetic calcium alginate hydrogel for antibacterial wound dressing | |
| Guo et al. | Development of tannin-inspired antimicrobial bioadhesives | |
| Yu et al. | A self-healing and injectable oxidized quaternized guar gum/carboxymethyl chitosan hydrogel with efficient hemostatic and antibacterial properties for wound dressing | |
| Giano et al. | Injectable bioadhesive hydrogels with innate antibacterial properties | |
| CN102824654B (en) | Double-bioenzyme modified blending biological material containing gelatin and chitosan and preparation method and application thereof | |
| CN103360613B (en) | A kind of gelatin/nano silver composite material adopting microbial transglutaminase to modify and its preparation method and application | |
| CN101905034B (en) | Method for preparing biological polysaccharide self-assembly modificatory chitosan antibacterial biological material | |
| CN107602884B (en) | Silk fibroin/chitosan composite intelligent hydrogel and preparation method thereof | |
| CN111154149A (en) | Hydrogel and preparation method and dressing thereof | |
| CN104784741B (en) | chitosan functional hydrocolloid medical dressing | |
| CN105816903A (en) | Drug-loaded hyaluronic acid nanofiber composite dressing and preparation method thereof | |
| CN112999418B (en) | Medical hydrogel composition, medical hydrogel and preparation method thereof | |
| CN110747534A (en) | Antibacterial polysaccharide fiber material and preparation method thereof | |
| Fan et al. | Preparation and characterization of antibacterial polyvinyl alcohol/chitosan sponge and potential applied for wound dressing | |
| CN105288704A (en) | Polyamide-amine dendrimer modified chitosan wound repair hydrogel as well as preparation method and application thereof | |
| US20210015966A1 (en) | Antibacterial Cellulose Hydrogels and Preparation Method therefor | |
| CN113429589A (en) | Glycyrrhetinic acid-based pH-sensitive slow-release hydrogel material and preparation method and application thereof | |
| CN106822980B (en) | Plasticized nano bacterial cellulose functional dressing and preparation method thereof | |
| CN101810879B (en) | Bioactive polysaccharide self-assembly modified polyurethane material and preparation method thereof | |
| CN110152055A (en) | The functional drug that alginic acid amination derivative/bacteria cellulose nanocomposite gel is constructed is sustained medical dressing | |
| Zuo et al. | Preparation and characterization of tannin-maltodextrin-polyvinyl alcohol hydrogel based on hydrogen bonding for wound healing | |
| CN103751831B (en) | Chitosan-containing activated carbon fiber surgical dressing and preparation method thereof | |
| CN112480435B (en) | Injectable antibacterial hydrogel material and preparation method thereof | |
| CN114225097B (en) | Self-healing hydrogel wound dressing loaded with antibacterial peptide and preparation method thereof | |
| CN106693048A (en) | Dual-computer injectable in-situ functional hydrogel biomaterial and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20140702 Termination date: 20160911 |