CN107158485A - Antibiont, extracellular matrix stick coating and preparation method and application - Google Patents
Antibiont, extracellular matrix stick coating and preparation method and application Download PDFInfo
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- CN107158485A CN107158485A CN201710381228.3A CN201710381228A CN107158485A CN 107158485 A CN107158485 A CN 107158485A CN 201710381228 A CN201710381228 A CN 201710381228A CN 107158485 A CN107158485 A CN 107158485A
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- antibiont
- extracellular matrix
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- stick
- protein
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Classifications
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- A—HUMAN NECESSITIES
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- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
- A61L31/14—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L31/00—Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
- A61L31/08—Materials for coatings
- A61L31/10—Macromolecular materials
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
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Abstract
Stick coating the present invention relates to a kind of antibiont, extracellular matrix, it is that protein plastifies layer, and the protein plasticizing layer includes albumin and hydrophilicity condiment.Above-mentioned antibiont, extracellular matrix stick coating, good biocompatibility, and then can clinically use safely；Raw material is easy to get and quality-high and inexpensive.Its is applied widely, goes for high polymer material matrix, inorganic material matrix, on metal material matrix；And the antibiont, extracellular matrix stick coating and base material is firmly combined with, no degraded and diffusion, its good hydrophily of internal stability is high, infiltration is fast, so as to facilitate medical care precess and routine use.Importantly, its can the most of bacteriums of impedance, cell and extracellular matrix sticking and deposit on its surface, with wide spectrum antibiont, extracellular matrix adhesion effect.Present invention also offers the preparation method and applications that a kind of antibiont, extracellular matrix stick coating.
Description
Technical field
The present invention relates to medical instruments field, more particularly to a kind of antibiont, extracellular matrix stick coating and its preparation
Method and application.
Background technology
The organisms such as bacterium, cell and extracellular matrix stick always biology, medical science, anti-in the harmful of material surface
One problem in the fields such as pollution, water filtration.It is above-mentioned it is harmful stick, by the breeding of material surface, diffusion, accumulate, and then
There may be bacterium infection, blood vessel embolism, surface contamination, toxic breakdown thing etc., influence human health and reduce materials'use effect
Really.
At present, antibiont, the research of extracellular matrix pasting material achieve certain progress, wherein with micro-nano bionic structural wood
Material, heparin, PEO-like structural material (PEG/PEO) and betaine type amphoteric ion composite this four classes material
Antibiont, extracellular matrix Adhesion property are protruded the most, and the research concern being subject to is also most.But these materials, which have, significantly to be lacked
Fall into, such as synthesis technique is complicated, extraction costs dearly, stability is poor, anti-Mechanism of Adhesion is indefinite, this also make their application by
Certain limitation.
Therefore, it is badly in need of a kind of new antibiont, extracellular matrix and sticks coating.
The content of the invention
Based on this, it is necessary to provide a kind of new antibiont, extracellular matrix and stick coating.
A kind of antibiont, extracellular matrix stick coating, and it is that protein plastifies layer, and the protein plasticizing layer includes clearly
Albumen and hydrophilicity condiment.
Above-mentioned antibiont, extracellular matrix stick coating, due to its material of main part be albumin, good biocompatibility, and then
It can clinically use safely；Albumin raw material is easy to get and quality-high and inexpensive.Above-mentioned antibiont, extracellular matrix stick coating, are applicable
Scope is wide, goes for high polymer material matrix, inorganic material matrix, on metal material matrix；And the coating and base material
It is firmly combined with, no degraded and diffusion, internal stability are good.Above-mentioned antibiont, extracellular matrix stick coating, and its hydrophily is high, leaching
Profit is fast, so as to facilitate medical care precess and routine use.
Importantly, above-mentioned antibiont, extracellular matrix stick coating, can the most of bacteriums of impedance (such as Escherichia coli,
Staphylococcus aureus, Candida albicans, Pseudomonas aeruginosa, streptococcus fecalis etc.), cell (such as fibroblast, blood platelet) and born of the same parents
Epimatrix (such as polysaccharide, plasma fibrinogen, seralbumin) is sticked and deposited its surface, collection antibacterial, anti-thin
Born of the same parents are sticked, antiplatelet sticks multiple function, the effect sticked with wide spectrum antibiont, extracellular matrix.
In one of the embodiments, the albumin is white selected from seralbumin, lactalbumin, leucosin, egg white
One or more in albumen and soy whey.
In one of the embodiments, the hydrophilicity condiment is selected from polar hydrophilic protein, amino acid, polysaccharide and its spread out
One or more in biology.
In one of the embodiments, the hydrophilicity condiment be selected from gelatin, sericin, serine, aspartic acid,
One or more in Sodium Hyaluronate, sodium carboxymethylcellulose and Arabic gum.
In one of the embodiments, fibrous protein is also included in the albumen plasticizing layer.
In one of the embodiments, on the basis of the albuminised quality, the mass fraction of the hydrophilicity condiment
For 0.5wt%~10wt%.
In one of the embodiments, the isoelectric point of the protein in the protein plasticizing layer is between 3.5~5.5.
Present invention also offers the preparation method that a kind of antibiont, extracellular matrix stick coating.
A kind of antibiont, extracellular matrix stick the preparation method of coating, comprise the following steps：
Albumin, hydrophilicity condiment and water are configured to glue；
The glue is coated in substrate surface formation film layer；
The film layer is subjected to plastics processing, antibiont, extracellular matrix is obtained and sticks coating.
Above-mentioned antibiont, extracellular matrix stick the preparation method of coating, environmentally friendly, simple, convenient and easy.
Present invention also offers the apparatus that a kind of antibiont, extracellular matrix stick.
The apparatus that a kind of antibiont, extracellular matrix stick, including antibiont provided by the present invention, extracellular matrix stick painting
Layer.
The apparatus that above-mentioned antibiont, extracellular matrix stick, because it has antibiont provided by the present invention, extracellular matrix
Stick coating, so good biocompatibility, and then can clinically use safely.And antibiont, extracellular matrix stick coating
It is firmly combined with base material, no degraded and diffusion, internal stability are good.In addition, its hydrophily is high, infiltration is fast, so that convenient medical treatment
Operation and routine use.Importantly, integrating antibacterial, anti-cellular adhesion, antiplatelet sticks multiple function.
In one of the embodiments, the apparatus that the antibiont, extracellular matrix stick is medical catheter, nail, painstaking effort
Pipe holder, operating theater instruments, biological culture product, glass, quartz or ceramic utensil.
Brief description of the drawings
Fig. 1 for institute's adherent bacteria bacterium colony on catheter plate count comparison diagram --- (upper row is catheter to Escherichia coli
A1, lower row is catheter AC1).
Fig. 2 for institute's adherent bacteria bacterium colony on catheter plate count comparison diagram --- (upper row is catheter to Pseudomonas aeruginosa
A1, lower row is catheter AC1).
Fig. 3 for institute's adherent bacteria bacterium colony on catheter plate count comparison diagram --- (upper row is catheter to streptococcus fecalis
A1, lower row is catheter AC1).
Fig. 4 is that (left side is slide A3 to E. coli clones violet staining effect contrast figure, and right side is slide
AC3)。
Fig. 5 is that (left side is slide A3 to S. aureus colonies violet staining effect contrast figure, and right side is load glass
Piece AC3).
Fig. 6 is that (left side is slide A3, and right side is carries glass for the fluorescence contrast figure that sticks of human plasma fibrinogen (HFg)
Piece AC3).
Fig. 7 is the fluorescence intensity comparison diagram of human plasma fibrinogen (HFg) and human serum albumins (HSA).
Fig. 8 is that (left side is slide A3, and right side is slide for the fluorescence contrast figure that sticks of human serum albumins (HSA)
AC3)。
Fig. 9 is the adhesion effect comparison diagram of rabbit platelet (left side is slide A3, and right side is slide AC3).
Figure 10 is the aspect graph of rabbit platelet rich plasma under an optical microscope.
Figure 11 is the OD value comparison diagrams sticked of L929 cells.
Embodiment
In order to make the purpose , technical scheme and advantage of the present invention be clearer, it is right below in conjunction with embodiment
The present invention is further elaborated.It should be appreciated that embodiment described herein is only to explain the present invention,
It is not intended to limit the present invention.
Unless otherwise defined, all of technologies and scientific terms used here by the article is with belonging to technical field of the invention
The implication that technical staff is generally understood that is identical.Term used in the description of the invention herein is intended merely to description tool
The purpose of the embodiment of body, it is not intended that in the limitation present invention.Term as used herein " and/or " including one or more
The arbitrary and all combination of related Listed Items.
A kind of antibiont, extracellular matrix stick coating, its be protein plastify layer, protein plasticizing layer include albumin,
And hydrophilicity condiment.
Wherein, albumin is the material of main part that antibiont, extracellular matrix stick coating.Albumin, also known as albumin, English
Title albumin, is abbreviated as Alb.Albumin is widely present in natural animal-plant or microorganism, typically by chemically or physically
Means are extracted in vivo from natural animal-plant or microorganism or regenerate and obtain.
Albumin is a kind of water-soluble, electronegative, globular protein, its easy plastic, and with most of water-soluble biologicals
Macromolecular material has preferable compatibility.Importantly, albumin has special electric charge and unit structure, and preferential suction
Attached the characteristics of, to most of bacteriums, cell, protein, the non-specific adsorption of nucleic acid or stick and have certain hindrance function.
Preferably, albumin of the invention is selected from seralbumin, lactalbumin, leucosin, oralbumin, soybean
One or more in albumin.
It is, of course, understood that the albumin of the present invention is not limited to above-mentioned albumin, other clear eggs are can also be
In vain；Such as albumin can also be the soy whey being modified by biological or chemical means or lactalbumin, also or manually
The albumin of synthesis.
In a preferred embodiment, the fibrous protein of humidification has also been included in protein plasticizing layer.In clear egg
In white object, fibrous protein is added, albumin can be overcome to be difficult film forming, the problem of film forming is difficult can be very good to improve
The filming performance of overall protein, and the mechanical strength after film forming.
Preferably, one or more of the fibrous protein in fibroin albumen, collagen and lysozyme.
Preferably, on the basis of albuminised quality, the mass fraction of fibrous protein is 5wt%~30wt%.
It is, of course, understood that the fibrous protein of the present invention is not limited to above-mentioned albumen, it can also be that other are fine
Tie up shape albumen；Such as myosin, keratin.
Preferably, protein is overall in protein plasticizing layer still has strong elecrtonegativity.
It is highly preferred that in protein plasticizing layer the isoelectric point of protein between 3.5~5.5, more preferably 3.5~
4.8 between.It so can further improve the anti-Adhesion property that antibiont, extracellular matrix stick coating.
Wherein, the main function of hydrophilicity condiment is to improve the surface hydrophilicity that antibiont, extracellular matrix stick coating
Energy.The inventors found that protein is after plastics processing, a part of pendant hydrophilic base on protein molecule can be caused
Group internally overturns, so that the hydrophily reduction of protein, and then reduce anti-Adhesion property；Add after hydrophilicity condiment,
The change that hydrophily caused by can offsetting after protein plastics processing declines, so as to effectively increase whole antibiont, born of the same parents
Epimatrix sticks the anti-Adhesion property of coating.
Preferably, the one kind or several of hydrophilicity condiment in polar hydrophilic protein, amino acid, many carbohydrates and their derivatives
Kind.
It is highly preferred that hydrophilicity condiment is selected from gelatin, sericin, serine, aspartic acid, Sodium Hyaluronate, carboxylic
One or more in sodium carboxymethylcellulose pyce and Arabic gum.
Preferably, on the basis of albuminised quality, the mass fraction of hydrophilicity condiment is 0.5wt%~10wt%.
If it is highly preferred that the one kind or several of hydrophilicity condiment in gelatin, sericin, serine, aspartic acid
When planting, then the addition of hydrophilicity condiment is albuminised 3wt%~10wt%.If hydrophilicity condiment is selected from Sodium Hyaluronate
Or/and during sodium carboxymethylcellulose, then the addition of hydrophilicity condiment is albuminised 0.5wt%~1.5wt%.If hydrophily
When auxiliary material is selected from Arabic gum, then the addition of Arabic gum is albuminised 2wt%~5wt%.
Preferably, the thickness that antibiont sticks coating is 10nm~500nm.Pliability so to matrix does not almost have shadow
Ring.
Preferably, antibiont sticks the r.m.s. roughness no more than 100nm of coating.So antibiont sticks the table of coating
Face is more smooth, and extraordinary lubricity can be shown in water environment, facilitates surgical operation, and patient's use feeling is comfortable.
Above-mentioned antibiont, extracellular matrix stick coating, are native protein because its material of main part is albumin, biological
Compatibility is good, and then can clinically use safely；Natural protein source is easy to get and quality-high and inexpensive.It is above-mentioned antibiont, extracellular
Matrix adhesion coating, it is applied widely, go for high polymer material matrix, inorganic material matrix, on metal material matrix；
And the coating is firmly combined with base material, no degraded and diffusion, internal stability are good.Above-mentioned antibiont, extracellular matrix stick painting
Layer, its hydrophily is high, infiltration is fast, so as to facilitate medical care precess and routine use.
Importantly, above-mentioned antibiont, extracellular matrix stick coating, can the most of bacteriums of impedance (such as Escherichia coli,
Staphylococcus aureus, Candida albicans, Pseudomonas aeruginosa, streptococcus fecalis etc.), cell (such as fibroblast, blood platelet) and born of the same parents
Epimatrix (such as polysaccharide, plasma fibrinogen, seralbumin) is sticked and deposited its surface, collection antibacterial, anti-thin
Born of the same parents are sticked, antiplatelet sticks multiple function, the effect sticked with wide spectrum antibiont, extracellular matrix.
Above-mentioned antibiont, extracellular matrix stick coating, can apply to medical and civilian antibacterial.
Above-mentioned antibiont, extracellular matrix stick coating, the work for sticking, colonizing in coating surface with the most of bacteriums of impedance
With the formation of bacterial clump can be prevented, but does not kill bacterium.The antibiont of the present invention, extracellular matrix sticks coating to have
Effect prevents gram-positive bacteria, negative bacterium, fungi and its bacterium colony of formation sticking in material surface, and anti-adherence rate is up to 80%
More than.
The present inventor thinks that the mechanism that the antibiont of the present invention, extracellular matrix stick is as follows：
Learned according to scientists such as Van Oss, Fowkes on " Li Fushici-Van der Waals-Lewis Acids and Bases " of surface energy
Say (Lifshitz-Van der Waals-Lewis acid-base theory), material and bacterium, cell, extracellular matrix etc.
Between adhesion depend on material surface energy among lewis base property component (γ _ S^-) and lewis acidity component
The relative size of (γ _ S^+), γ _ S^- is bigger, and material more tends to electron donor, is less susceptible to electronegative bacterium, carefully
Born of the same parents, protein etc., which are produced, to be sticked.It by strong elecrtonegativity protein is main former that antibiont, the extracellular matrix of the present invention, which sticks coating to be,
Prepared by material, from material properties for be lewis base, i.e. electron donor, so there is stronger impedance to bio-adhesive
Effect.
For bacterium, cell, the angle of extracellular matrix, antibiont, the extracellular matrix Adhering capacity of material are depended on carefully
Bacterium, cell, extracellular matrix stick free energy (Δ G_adh) to material, and Δ G_adh values are bigger, stick and are more difficult to occur.And it is general
The material such as 304 stainless steels, medical grade silicon rubber, glass for, its Δ G_adh values are all negative value, bacterium, cell, extracellular matrix
Easily stick these material surfaces are spontaneous；And the present invention antibiont, extracellular matrix stick coating, its Δ G_adh be on the occasion of,
So compared with other materials, bacterium, cell, extracellular matrix are spontaneous to stick relative difficulty；So as to more prominent, broader spectrum of
Antibacterium, cell, extracellular matrix Adhesion property.
People and protein and cell in animal body, most of is all to be presented electronegative, and antibiosis of the present invention
Thing, extracellular matrix, which stick coating, has strong negative electricity characteristic and strong hydrophilicity effect, therefore also has impedance cell and its secreted born of the same parents
Epimatrix is in its surface adhesion, the effect of absorption.
Present invention also offers the preparation method that a kind of antibiont, extracellular matrix stick coating.
A kind of antibiont, extracellular matrix stick the preparation method of coating, comprise the following steps：
S1, albumin, hydrophilicity condiment and water is configured to glue；
S2, the glue is coated in substrate surface formation film layer；
S3, by the film layer carry out plastics processing, obtain antibiont, extracellular matrix and stick coating.
In step sl, the main function of water is that protein material is uniformly dispersed with hydrophilicity condiment, forms aqueous point
Granular media system.Preferably, water can be selected from purified water, deionized water, distilled water, distilled water.It is, of course, understood that can also
The additive formation water system buffer solution in water, such as PBS, MES-Tris, with water system buffer solution dispersed protein material and parent
Aqueous auxiliary material.
Preferably, first by albumin, fibrous protein (selectively), with water be configured to initial glue, then again to
Hydrophilicity condiment is added in primary glue and is configured to secondary glue liquid.
Further, step S1 also includes：Secondary glue liquid is filtered, centrifuged, after de-bubble, the secondary glue clarified
Liquid.
In step sl, when preparing glue, when adding hydrophilicity condiment, other auxiliary materials can also be added, other are auxiliary
The one or more that material includes but is not limited in crosslinking agent, thickener, dispersant.Crosslinking agent can greatly improve the plasticizing of coating
Performance, makes antibiont, extracellular matrix stick coating more stable, firm.If using crosslinking agent, typically in normal temperature after well mixed
Lower standing 5min~30min.
It is highly preferred that crosslinking agent is selected from glyoxal, glutaraldehyde, carbodiimide, n-hydroxysuccinimide, Geniposide, original
One or more in anthocyanidin.The addition of crosslinking agent is the 0.1%~2% of protein material gross mass.Thickener is selected from
The one or more of xanthans or sodium alginate.The addition of thickener be preferably initial glue gross weight 0.5%~
1.5%.One or more of the dispersant in sodium tripolyphosphate, polyethylene glycol 400 or polysorbate60.The addition of dispersant
The 0.5%~1.5% of preferably initial glue gross weight.
In step s 2, it is preferable that base material before coating, may be pretreated, such as ultrasonic cleaning, pickling, alkali
Wash, also or other he washing of organic/inorganic solvent, corona treatment, LBL self-assembly, Modification of Photo-grafting Copolymerization, supramolecular chemistry
The modes such as adsorption modification.
Glue painting method can be infusion process, brushing method, rotation spray coating or ultrasonic spraying process.Glue is coated
Single application, can also be multiple coating.
In step s3, plastics processing refers to material becoming stable solid-state, in the process, material by solution or melt
Expect that great change occurs for molecular chain configuration and chain spacing, and there may be physically or chemically crosslinked.
Preferably, plastics processing can be using spontaneous curing, heat cure, microwave curing, ultraviolet light solidification or chemical crosslinking
Plasticizing, Radiation etc..
In step s3, it is highly preferred that using thermosetting, light wave or Radiation, the plastics processing of chemical crosslinking mode.
In the present embodiment, plastics processing mode is thermosetting.Preferably, plastics processing condition is：Temperature be 60 DEG C~
180 DEG C, pressure be 0.1MPa~1.5MPa, time be 10min~45min.It is highly preferred that plastics processing condition is：Temperature is
80 DEG C~150 DEG C, pressure be 1MPa~1.3MPa, time be 20min.
Preferably, can also use plasticizer during plastics processing, plasticizer be selected from glycerine, sorbierite, urea or
One or more in triethyl citrate.
Above-mentioned antibiont, extracellular matrix stick the preparation method of coating, environmentally friendly, simple, convenient and easy.
Present invention also offers the apparatus that a kind of antibiont, extracellular matrix stick.
The apparatus that a kind of antibiont, extracellular matrix stick, including antibiont provided by the present invention, extracellular matrix stick painting
Layer.
That is, the apparatus that antibiont, extracellular matrix stick includes instrument body, and it is at least partially coated in apparatus
Antibiont, extracellular matrix on body surface stick coating.
Preferably, the apparatus that antibiont, extracellular matrix stick be medical catheter, nail, angiocarpy bracket, operating theater instruments,
Biological culture product, glass, quartz or ceramic utensil.
The apparatus that above-mentioned antibiont, extracellular matrix stick, because it has antibiont provided by the present invention, extracellular matrix
Stick coating, so good biocompatibility, and then can clinically use safely.And antibiont, extracellular matrix stick coating
It is firmly combined with base material, no degraded and diffusion, internal stability are good.In addition, its hydrophily is high, infiltration is fast, so that convenient medical treatment
Operation and routine use.Importantly, integrating antibacterial, anti-cellular adhesion, antiplatelet sticks multiple function.
Below in conjunction with specific embodiment, the invention will be further elaborated.
Embodiment 1
By the catheter of uncoated silicon rubber material, 30min and drying are cleaned by ultrasonic in purified water, then through ultraviolet light
Irradiation 15min is sterilized.
By the catheter after sterilizing, it is put into vacuum plasma treatment equipment and carries out surface activation process.Plasma
It is NH to handle the atmosphere used3-O2- Ar three-element mixed gas bodies, vacuum is 50Pa, and discharge power is 30W, and handling duration is
10min。
Lactalbumin, soy whey, fibroin albumen, sericin are pressed 7:10:1.5:1.5 mass ratio mixing, plus
Enter purified water, obtain the initial glue that total mass fraction is 7wt%.Then according to the volume of initial glue with 50:1 (v/v's)
Ratio, adds dispersant (the 5mg/ml PEG-400 aqueous solution), well mixed to form secondary glue liquid.Then by secondary glue liquid again
Through steps such as centrifugation, filterings, bubble removing and suspended particulate are removed, the secondary glue liquid clarified.
By the secondary glue liquid of clarification, using the mode of ultrasound spraying to the whole outer surface of catheter after surface activation process
The accurate coating of single, individual layer is carried out, film layer is formed.
Film layer is subjected to plastics processing, the condition of wherein plastics processing is：Temperature is 70 DEG C, pressure is 1.5MPa, duration
For 20min.
Obtained catheter, is denoted as A1.
Embodiment 2
By the cylindrical nail of uncoated Nitinol, be cut into 2cm or so nail section, successively in acetone, ethanol, go
It is cleaned by ultrasonic 30min in ionized water, then uses N2Air-blowing dry doubling 75vol% alcohol disinfecting.
By the nail section after sterilization, it is put into vacuum plasma treatment equipment and carries out surface activation process.Plasma
It is acrylic acid-O to handle the atmosphere used2Binary mixture, vacuum is 30Pa, and discharge power is 50W, and handling duration is
15min。
By the nail section after surface activation process, 0.5mg/ml aq. polyethyleneimine (branched PEI, MW=is immersed
10000), dip-coating keeps 5min, so that the adsorption of nail section is enriched with the PEI coatings of positive electricity.
Soy whey, collagen, sodium carboxymethylcellulose are pressed 16:3:1 mass ratio mixing, adds purified water,
Stirring in 37 DEG C of water bath with thermostatic control is placed in until being completely dissolved, the initial glue that total mass fraction is 10wt% is obtained.Then
According to the volume of initial glue with 50:1 (v/v) ratio, adds dispersant (the 5mg/ml PEG-400 aqueous solution), and mixing is equal
Even formation secondary glue liquid.Then secondary glue liquid is removed bubble removing and suspended particulate, clarified again through steps such as centrifugation, filterings
Secondary glue liquid.
In mass ratio 50:1 ratio adds 5wt% carbodiimide/N- hydroxysuccinimidyl acyls into the secondary glue liquid of clarification
Imines (mass ratio 1:1) crosslinking agent, carries out precrosslink, obtains coating liquid.
The multiple coating of albumen glue is carried out with the pattern of LBL self-assembly.Nail section after positive charge is taken out, after
With deionized water rinsing 3 times, coating liquid dip-coating 10min is then put into again；Taking-up afterwards is gently rinsed 3 times with deionized water, then
Immerse 0.5mg/ml aq. polyethyleneimine (branched PEI, MW=10000), dip-coating keep 5min, then take out spend from
Sub- water is rinsed 3 times, then puts into coating liquid dip-coating 10min, is gently rinsed with deionized water 3 times after taking-up.If repeating above step
Dry time, outermost layer is set to coat coating liquid.
Film layer is subjected to plastics processing, the condition of wherein plastics processing is：Temperature is 70 DEG C, pressure is 1.5MPa, duration
For 45min.
Obtained nail, is denoted as A2.
Embodiment 3
Take uncoated slide is each in acetone, ethanol, deionized water to be successively cleaned by ultrasonic 30min.
By slide immersion Piranha washing lotions (the dense H after ultrasonic cleaning2SO4:30%H2O2(v/v)=3:1) 70 in
Water-bath 30min at DEG C, to remove the organic matter remained on slide and make slide negative electrical charge.
After water-bath terminates, deionized water rinsing slide is used, and slide is immersed to 0.5mg/ml polyethyleneimine
The aqueous solution (branched PEI, MW=10000), dip-coating keeps 10min, makes the PEI coatings of slide surface adsorption and enrichment positive electricity.So
Slide is taken out afterwards, with deionized water rinsing 3 times, dried naturally standby.
Lactalbumin, leucosin, Arabic gum are pressed 3:16:1 mass ratio mixing, adds deionized water and obtains total matter
Fraction 12wt% initial glue is measured, then according to the volume of initial glue with 50:1 (v/v) ratio, adds dispersant
(the 5mg/ml PEG-400 aqueous solution), then removes bubble removing and suspended particulate, is clarified through steps such as centrifugation, filterings again
Secondary glue liquid.
By 100:The OPC aqueous solution that 1 (v/v) ratio adds 2wt% into the secondary glue liquid of clarification is pre-payed
Connection, obtains coating liquid.
The slide dried is immersed into 10min in coating liquid, then takes out and uses deionized water rinsing slide 3 times.
Slide after flushing is subjected to plastics processing, the condition of wherein plastics processing is：Temperature is 95 DEG C, pressure is
1.5MPa, when a length of 45min.
Obtained slide, is denoted as A3.
Comparative example 1
By the catheter of uncoated silicon rubber material, 30min and drying are cleaned by ultrasonic in purified water, then through ultraviolet light
Irradiation 15min is sterilized.
Obtained catheter, is denoted as AC1.
Comparative example 2
By the cylindrical nail of uncoated Nitinol, be cut into 2cm or so nail section, successively in acetone, ethanol, go
It is each in ionized water to be cleaned by ultrasonic 30min, then use N2Air-blowing dry doubling 75vol% alcohol disinfecting.
Obtained nail, is denoted as AC2.
Comparative example 3
Take uncoated slide is each in acetone, ethanol, deionized water to be successively cleaned by ultrasonic 30min.
Obtained slide, is denoted as AC3.
Performance test：
Catheter antibacterium sticks test：
By three groups of catheters (every group is an a catheter A1 and catheter AC1), it is cut at urination cavity
7cm segments, are put into 15ml centrifuge tube, are then respectively adding the 1 × 10 of 6ml3~5 × 103The bacteria suspension of cfu/ml concentration
(first group be Escherichia coli, second group be pseudomonas aeruginosa, the 3rd group be streptococcus fecalis), be subsequently placed in 35 ± 2 DEG C of perseverance
Cultivated in warm incubator.Escherichia coli and pseudomonas aeruginosa are cultivated 17~19 hours, and streptococcus fecalis is cultivated 45~48 hours, training
Catheter after supporting is gone out with sterile tweezer respectively, in input 50ml centrifuge tubes, and injection 30ml sterile salines turn upside down 10
Secondary, rinse urethral catheterization pipeline section surfaces externally and internally planktonic bacteria repeats this step 3 time.
Each urethral catheterization pipeline section after rinse is put into respectively ultrasonic in the 15ml centrifuge tubes of a physiological saline containing 10ml
The bacterium colony (40kHz, 100%) that 10min elution catheter surfaces stick.
Appropriate dilution, takes 0.5ml to be cleaned by ultrasonic liquid and is placed in plate, with cool to 40~50 DEG C nutrient agars respectively
15ml~20ml is poured into, and rotates plate, flat board is overturn after making its full and uniform agar solidification, and (35 ± 2) DEG C culture 24~
48h。
The flat board that bacterium colony grows in the form of sheets should not be used, and the naked eyes obtained on different extension rate flat boards be recorded respectively visible
Clump count.
Antibacterium adherence rate is calculated by formula：X is antibacterium adherence rate (%) in X=(A-B)/A × 100%, formula, and A is
Comparative example group clump count average value, B is embodiment group clump count average value.
By above-mentioned steps, final calculating obtains catheter A1 and Escherichia coli, pseudomonas aeruginosa and streptococcus fecalis is resisted
Adherence rate is respectively 89%, 86% and 83%.The anti-result of sticking of Escherichia coli, pseudomonas aeruginosa and streptococcus fecalis is shown in respectively
Fig. 1, Fig. 2 and Fig. 3.
It can be seen that from Fig. 1, Fig. 2, Fig. 3：Escherichia coli, pseudomonas aeruginosa and streptococcus fecalis are respectively in catheter AC1
Surface is largely sticked, and bacterium is not countable on flat board；And bacterial adhesion amount is fewer on catheter A1, the two has quantity
Level gap.The catheter of this explanation present invention has obvious anti-adhesion to bacterium.
Nail antibacterium sticks test：
Method is sticked using above-mentioned antibacterium bacterial adhesion quantitative experiment is carried out to nail A2 and AC2, finally calculated
It is respectively 99%, 93% and 87% to the anti-adherence rate of Escherichia coli, pseudomonas aeruginosa and streptococcus fecalis to nail A2.
The antibacterium of slide sticks test：
Bacteria Culture：It is each on each slide after cooling by slide A3 and slide AC3 through autoclave sterilization
It is 1 × 10 that concentration, which is added dropwise,5~9 × 105The bacteria suspension (Escherichia coli, staphylococcus aureus) of cfu/ml concentration, slide is put into
Disposable sterilized plate is placed in starting to cultivate bacterium in (35 ± 2) DEG C constant incubator.
Colony IFA technique：After 18h, the plate for filling slide is taken out, slide surface bacterium is absorbed with disposable sterilized dropper
Suspension, is then rinsed with purified water, washes away and the 0.5ml crystal violet aqueous solution is added dropwise after planktonic bacteria on every slide
(1wt%), is dyed to the bacterium colony remained on slide.
Bacterium colony is observed：Dye after 30min, every slide is rinsed with purified water 5 times, dyeing liquor is washed away, in optical microphotograph
Bacterium colony is observed with 100 times of oil mirrors under mirror, Fig. 4 and Fig. 5 is as a result seen.
It can be seen that from Fig. 4, Fig. 5：Intensive royal purple color dot or sheet area are presented after the crystallized purple dyeing of slide AC3
Domain, illustrates that Escherichia coli and staphylococcus aureus a large amount of tactophilies and form biomembrane on slide AC3 respectively, and carry
Slide A3 only has few several stains.This explanation present invention can substantially suppress the formation of bacterial biof iotalm.
Human plasma fibrinogen sticks test：
Prepare human fibrinogen (Human Fibrinogen, HFg) dilution：By human fibrinogen
(HumanFibrinogen, HFg) is dissolved in 0.2M, pH 4.5 NaAc_HAc buffer solution, and HFg ultimate densities are
0.5mg/ml；
Prepare bovine serum albumin(BSA) (Bovine Serum Albumin, BSA) confining liquid：Bovine serum albumin(BSA) is dissolved in
In 0.1M, pH7.4 PBS, BSA ultimate densities are 10mg/ml；
Prepare goat-anti HFg-FITC working solutions：By the goat-anti people of concentration 5mg/ml marked by fluorescein isothiocyanate (FITC)
Plasma fibrinogen stoste is used as working solution after diluting 30 times with PBS.
To addition HFg dilutions in slide A3 and slide AC3 culture plate are placed with, material is totally submerged,
Put in people's constant temperature water tank, 37 DEG C of incubation 1h；Incubation terminate after by dilution all suction out, with PBS rinse 3 times, every time
10min；Rinsing adds BSA confining liquids after terminating, and 1h is closed under the conditions of 37 DEG C；After closing terminates, confining liquid is suctioned out, again
Rinsed with PBS, 10min × 3 time；Rinsing adds goat-anti HFg-FITC working solutions after terminating, under the conditions of 37 DEG C
It is incubated 1h；Incubation is rinsed after terminating with deionized water, and then 10min × 3 time are entered with fluorescence microscope and sepectrophotofluorometer
Row is characterized, and its result difference is as shown in Figure 6 and Figure 7.
It can be seen that from Fig. 6, Fig. 7：There is a more FITC characteristic fluorescences point on slide AC3 surfaces, and the spy on slide A3
Levy phosphor dot seldom.The result explanation of goat-anti HFg-FITC spikes, HFg has more non-specificity to stick on slide AC3 surfaces,
And slide A3 HFg non-specificity stick it is less；Full wafer fluorescence intensity test result shows, slide AC3 FITC fluorescence
Intensity is higher by 4~5 times of slide A3.This explanation present invention is sticked to the non-specificity of human plasma fibrinogen (HFg) to be had
Notable hindrance function.
Human serum albumins sticks test：
Prepare HSA-FITC working solutions：The human serum albumins (Human SerumAlbumin, HSA) that FITC is marked is former
Liquid is used as working solution after diluting 30 times with 0.1M, pH7.4 PBS.
It is to addition HSA-FITC working solutions in slide A3 and slide AC3 culture plate are placed with, material is complete
Submergence, puts in people's constant temperature water tank, 37 DEG C of incubation 1h；Incubation all suctions out working solution after terminating, and is rinsed 3 times with deionized water,
Each 10min, is then characterized with sepectrophotofluorometer and fluorescence microscope, as a result respectively such as Fig. 7 and Fig. 8 institutes respectively
Show.
It can be seen that from Fig. 7, Fig. 8：There is more FITC characteristic fluorescences point on slide AC3 surfaces, and several on slide A3
There is no characteristic fluorescence point；The result explanation of HSA-FITC spikes, HSA has a large amount of non-specificity to stick on slide AC3 surfaces,
And slide A3 is seldom；Full wafer fluorescence intensity test result shows that slide AC3 FITC fluorescence intensities are higher by slide A3's
8~9 times.This explanation present invention is sticked to the non-specificity of human serum albumins (HSA) notable hindrance function.
Rabbit platelet sticks test：
Slide A3 and slide AC3 are put into super-clean bench ultraviolet light wave or Radiation sterilizing 1h, then again will
The two is respectively put into culture plate, and is added rabbit platelet rich plasma (PRP) and be totally submerged to material, is put into constant temperature water tank
In, 37 DEG C of incubation 1h；Incubation all suctions out rabbit PRP after terminating, and is rinsed 3 times with deionized water, each 10min, Ran Houyong
Observation by light microscope, is as a result shown in Fig. 9.
Figure 10 is the form contrast that rabbit PRP is capped slide observation under an optical microscope.
It can be seen that from Fig. 9, Figure 10：Former rabbit PRP activity and density are all relatively good, but pass through incubating under the same terms
After educating, the platelet adhesion reaction amount on slide A3 is considerably less than slide AC3 and platelet PLA2 is also without significant changes, that is, does not have
There is obvious activation.The obvious impedance blood platelet of this explanation present invention energy is in surface adhesion, and no stimulating platelet activation.
L929 cell adhesions are tested：
Slide A3 and slide AC3 are put into super-clean bench ultraviolet light wave or Radiation sterilizing 1h, then with sterile
PBS piping and druming rinse, 24 orifice plates are loaded after drying.The L929 cell pancreatin for growing to increased logarithmic phase is digested, used
It is 1 × 10 that 1640 fresh complete mediums, which terminate digestion and are resuspended to density,5/ml.0.5ml cell suspensions are added dropwise per hole, gently
CO is put into after jog is even2Incubator (37 DEG C, 5%CO2) it is incubated 24h.Slide is transferred in clean orifice plate, with sterile PBS
Solution adds fresh culture medium after flushing three times.The MTT solution that 50 μ l are added per hole is incubated 4h, and suction is abandoned solution in orifice plate and added
200 μ l DMSO, orifice plate, which is placed in shaking table and shaken up, takes 150 μ l solution to be transferred in 96 orifice plates after 10min, ELIASA determines absorbance
(OD) value, is as a result shown in Figure 11.
As can be seen from Figure 11：In 0~24h, cell OD values and slide AC3 on slide A3 have statistics always
Learn the L929 cell quantities sticked on difference, slide A3 substantially less.This explanation present invention also has impedance work to cell adhesion
With.
Each technical characteristic of embodiment described above can be combined arbitrarily, to make description succinct, not to above-mentioned reality
Apply all possible combination of each technical characteristic in example to be all described, as long as however, the combination of these technical characteristics is not deposited
In contradiction, the scope of this specification record is all considered to be.
Embodiment described above only expresses the several embodiments of the present invention, and it describes more specific and detailed, but simultaneously
Can not therefore it be construed as limiting the scope of the patent.It should be pointed out that coming for one of ordinary skill in the art
Say, without departing from the inventive concept of the premise, various modifications and improvements can be made, these belong to the protection of the present invention
Scope.Therefore, the protection domain of patent of the present invention should be determined by the appended claims.
Claims (10)
1. the coating that a kind of antibiont, extracellular matrix stick, it is characterised in that the antibiont, extracellular matrix stick coating and be
Protein plastifies layer, and the protein plasticizing layer includes albumin and hydrophilicity condiment.
2. the coating that antibiont according to claim 1, extracellular matrix stick, it is characterised in that the albumin is selected from
One or more in seralbumin, lactalbumin, leucosin, oralbumin and soy whey.
3. the coating that antibiont according to claim 1, extracellular matrix stick, it is characterised in that the hydrophilicity condiment
One or more in polar hydrophilic protein, amino acid, many carbohydrates and their derivatives.
4. the coating that antibiont according to claim 3, extracellular matrix stick, it is characterised in that the hydrophilicity condiment
In gelatin, sericin, serine, aspartic acid, Sodium Hyaluronate, sodium carboxymethylcellulose and Arabic gum
It is one or more of.
5. the coating that antibiont according to claim 1, extracellular matrix stick, it is characterised in that the albumen plastifies layer
In also include fibrous protein.
6. the coating that antibiont according to claim 1, extracellular matrix stick, it is characterised in that with described albuminised
On the basis of quality, the mass fraction of the hydrophilicity condiment is 0.5wt%~10wt%.
7. the coating that the antibiont, extracellular matrix according to any one of claim 1~6 stick, it is characterised in that the egg
The isoelectric point of protein in white matter plasticizing layer is between 3.5~5.5.
8. the preparation method for the coating that a kind of antibiont, extracellular matrix stick, it is characterised in that comprise the following steps：
Albumin, hydrophilicity condiment and water are configured to glue；
The glue is coated in substrate surface formation film layer；
The film layer is subjected to plastics processing, antibiont, extracellular matrix is obtained and sticks coating.
9. the apparatus that a kind of antibiont, extracellular matrix stick, it is characterised in that including anti-described in any one of claim 1~7
Biological, extracellular matrix sticks coating.
10. the apparatus that antibiont according to claim 9, extracellular matrix stick, it is characterised in that the antibiont, born of the same parents
The apparatus that epimatrix is sticked be medical catheter, nail, angiocarpy bracket, operating theater instruments, biological culture product, glass, quartz or
Ceramic utensil.
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| CN113499479A (en) * | 2021-07-19 | 2021-10-15 | 科凯(南通)生命科学有限公司 | Preparation method of modified biological material and obtained modified biological material |
| US11168287B2 (en) | 2016-05-26 | 2021-11-09 | Kimberly-Clark Worldwide, Inc. | Anti-adherent compositions and methods of inhibiting the adherence of microbes to a surface |
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| CN115418031B (en) * | 2022-09-01 | 2023-10-20 | 北京化工大学 | Preparation method and application of a negatively charged small molecule/cationic polymer composite protein differential adhesion material |
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