CN107376032A - A kind of vesicoureteric reflux injection treatment filler and preparation method thereof - Google Patents
A kind of vesicoureteric reflux injection treatment filler and preparation method thereof Download PDFInfo
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- CN107376032A CN107376032A CN201710702758.3A CN201710702758A CN107376032A CN 107376032 A CN107376032 A CN 107376032A CN 201710702758 A CN201710702758 A CN 201710702758A CN 107376032 A CN107376032 A CN 107376032A
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- 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
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- 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
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- 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
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- 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
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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
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Abstract
The invention belongs to functional material and biological technical field, specially a kind of vesicoureteric reflux injection treatment filler and preparation method thereof.The present invention's is a kind of gel that agarose microbeads and cross-linking hyaluronic acid prepared by inverse suspension method is formed, or a kind of agarose microbeads and the gel of cross-linking hyaluronic acid formation with fluorophor prepared by inverse suspension method.Agarose microbeads are prepared by inverse suspension method, it is crosslinked in the basic conditions again, Sepharose is mixed to form gel filling agent with hyaluronic acid, and can be in agarose microbeads surface grafting fluorophor, efficient and noninvasive living imaging can be achieved in later stage zoopery, to observe the reservation situation of gel filling agent in vivo.Preparation time of the present invention is short, and process is simply efficient, and product heat endurance is good, high mechanical strength.
Description
Technical field
The invention belongs to functional material and biological technical field, and in particular to a kind of vesicoureteric reflux injection treatment is used
Filler and preparation method thereof.
Background technology
Vesicoureteric reflux (Vesicoureteral reflux, VUR) is most common Congenital Renal and urinary tract
One of deformity.VUR infants have the excessive risk of urinary system infection contamination, can cause the formation of pyelonephritis and kidney scar
（Pediatrics, 1999, 103(4): 843）, kidney scar and subsequent Chronic Renal Impairment are reflux type nephrosis（reflux
nephropathy, RN）Adulthood hypertension, albuminuria and progressive renal failure can be caused.Research represents, there is 5% ~ 10%
VUR infants are finally because RN progresses to ESRD（The Journal of urology, 2003, 169(1):309；
Pediatric Nephrology. 2012, 27(4):551）.Therefore, it is early to find, correctly diagnose and carry out to children VUR
Canonical management is always the focus of medical field concern research.
Microendoscopic injection treatment VU, R is because its is simple to operate quick, and several no postoperative pains are minimally invasive attractive in appearance, only in outpatient service hand
It can complete, Small side effects, and be widely studied under the conditions of art, its principle is that all kinds of fillers are injected in into urine output using scope
The submucosa of section between pipe the bladder wall, filler can raise distal ureteral and aperture position, can reach anti-reflux purpose.
The packing material reported at present has：Teflon（Polytetrafluoroethylene (PTFE)）（Br Med J (Clin Res Ed).
1984, 289(6436):7）, animal protein（J. Urology. 2000, 55(5):759）, DIMETHYLPOLYSILOXANE particle
（Urology. 2002, 60(5):894）, Autologous Chondrocyte（The Journal of urology. 1999, 162
(3):1185）, polyacrylate-polyol copolymer（Journal of pediatric urology. 2011,7(6):
654）Deng.But due to the possibility for moving to other organs, it is impossible to it is biodegradable, there is sensitization possibility, can be by human body
Absorb, it is impossible to exist long-term and stably the affected part the shortcomings of, above-mentioned material above receives certain limitation in application.
Any treatment VUR Microendoscopics injection filling medicine of domestic not yet official approval at present.The present invention is by simple
Method prepares agarose microbeads/hyaluronic acid gel, and the VUR gel filling agents of injectable are made.This invention address that prepare
The VUR Microendoscopic injection fillers of domestic first approval, this will fill up domestic current blank, be domestic VUR infants band
Carry out Gospel.
The content of the invention
It is an object of the invention to propose it is a kind of prepare conveniently, heat endurance is good, high mechanical strength vesicoureteral is anti-
Streamer penetrates treatment fluorescence filler and preparation method thereof.Sepharose microballoon with fluorophor/bright matter acid gel is filled out
Fill the preparation method of agent and its application in the injection treatment of the diseases such as vesicoureteric reflux.
Vesicoureteric reflux injection treatment fluorescence filler proposed by the present invention is one kind by anti-phase suspension legal system
The gel that standby agarose microbeads are formed with cross-linking hyaluronic acid, or a kind of carry fluorescence by prepared by inverse suspension method
The gel that the agarose microbeads of group are formed with cross-linking hyaluronic acid.The latter can make later stage zoopery to carry out convenience and high-efficiency
And the holding situation of noninvasive living imaging observation gel filling agent in vivo.
The preparation method of Sepharose microballoon/hyaluronic acid derivatives proposed by the present invention, is concretely comprised the following steps：
（1）Agarose solution is configured to as raw material using agarose powder and deionized water；
（2）Using inverse suspension method by step（1）Agarose solution be prepared into agarose microbeads；
（3）In step（2）Crosslinking agent and alkali are added in the system of preparation, carries out cross-linking reaction, forms Sepharose microballoon；
（4）By step（3）The system of preparation is washed by separating for several times, obtains having the Sepharose of certain grain size distribution micro-
Ball；
（5）Step（4）Obtained Sepharose microballoon is configured to gel with hyaluronic acid, as injection filler；Or
By step（4）Obtained Sepharose microballoon grafting fluorescence molecule, is then configured to gel, as injection with hyaluronic acid again
With filler, to meet that the observation of later stage living imaging retains the needs of situation in vivo.
Step of the present invention（1）In, the concentration of agarose solution is 1-10%（w/w）.
Step of the present invention（2）In, the inverse suspension method, normal octane is used as liquid phase, emulsification is used as using Span-80
Agent, at a temperature of high speed machine stirring, 50-85 DEG C, agarose solution is added, it is reacted to obtain agarose microbeads.
Step of the present invention（3）In, the cross-linking reaction is carried out under 150-1000rpm mixing speed, cross-linking reaction temperature
Spend for 35-80 DEG C, reaction time 0.5-6h.The crosslinking agent is selected from epoxychloropropane, the bromo- 2- propyl alcohol of 1,3- bis- etc., described
The one kind of alkali in sodium hydroxide, potassium hydroxide, ammoniacal liquor, sodium acid carbonate, sodium carbonate, or it is a variety of.
Step of the present invention（4）In, the separating, washing is ethanol, 20% ethanol-water solution and deionized water with solution；
Separation can be used and centrifuged（Centrifugal rotational speed 1000-5000rpm, 1-10min）, can also use standing separation (time of repose
0.5-5h)；Then sub-sieve (65-190 mesh) filtration washing is used, obtains certain particle diameter（80-250 microns）The Cross-linked Agar of distribution
Sugared microballoon.
Step of the present invention（5）In, the fluorescence molecule of the grafting can be fluorescein isothiocynate（Isomers I）, indoles cyanines
Green, rhodamine one kind；Sepharose microballoon is directly configured to gel with hyaluronic acid, and its concentration is 1-100 mg/ml, hands over
The gel being configured to again with hyaluronic acid after connection agarose microbeads grafting fluorescence molecule, its concentration is 1-50 mg/ml.
It is subcutaneous that above-mentioned Sepharose microballoon hyaluronic acid derivatives are injected at mouse, observe reservation situation.
The preparation method of Sepharose microballoon hyaluronic acid derivatives proposed by the present invention, the Sepharose microballoon are
Agarose is crosslinked in the basic conditions by inverse suspension method and is prepared, by Sepharose surface grafting fluorescent base
Group, can efficiently and non-invasively observe the reservation situation of gel in vivo.
Concrete operations flow of the present invention is as follows：
（1）0.5-3g agarose powders are weighed with 50g deionized waters, being heated under the conditions of 90-100 DEG C, magnetic force or machinery stir
Mix 10-30mins all to dissolve to agarose, it is 1-10% to form concentration（w/w）Agarose solution；The agarose solution
Viscosity is larger；
（2）（With liquid-transfering gun or pipette）50ml normal octanes are pipetted, add 0.1-5g emulsifying agent Span-80, high speed machine stirs
Mix, mixing speed 500-1500rpm, in whipping process, while be warming up to 50-85 DEG C, by step（1）Obtained agarose
The aqueous solution（Viscosity is larger）Pour into the normal octane of high-speed stirred, height continues speed stirring 0.5-2h；
（3）By step（2）Obtained temperature of reaction system is adjusted to 35-80 DEG C, adds 0.1-2M alkaline solution 2ml, crosslinking agent
0.1-4g, mixing speed 150-1000rpm, cross-linking reaction 0.5-6h；
（4）Stop heating, solution is layered after standing, and lower floor is agarose microbeads；Supernatant liquid is poured out, lower floor's solid is used successively
For several times, washing separation can be used and centrifuged for ethanol, 20% ethanol-water solution and deionized water washing separation（1000-
5000rpm, 1-10min）Form, standing separation (0.5-5h) can be also used, afterwards with sub-sieve (65-190 mesh) filtration washing
Obtain the Sepharose microballoon of certain grain size distribution, product be stored in 20% ethanol-water solution or deionized water in it is standby
With；
（5）Sepharose microballoon is grafted fluorescence molecule, takes step（4）Obtained product 1-5g（Weight in wet base）In diamine solution,
35-50 DEG C of reaction 0.5-5h under magnetic agitation, diamine solution is to be added to by 0.1-2g diamines in 50ml 0.1M alkaline solutions
It is formulated；Heating is removed after the completion of reaction, the microballoon that reaction obtains is placed in sub-sieve（65-190 mesh）On, with it is a large amount of go from
Sub- water washing is to filtrate in neutrality；1-10 mg fluoresceins are dispersed in 100 ml deionized waters, 15-30min of ultrasound make it
Dissolving, add 0.1-2g（Weight in wet base）The agarose microbeads of surface grafting hexamethylene diamine, magnetic agitation reaction 1-18 h under normal temperature；Will be anti-
The microballoon that should be obtained later is placed in sub-sieve（65-190 mesh）On, washed with a large amount of deionized waters, remembered until cleaning solution is colourless
To the Sepharose microballoon of grafting fluorophor；
（6）Take step（4）Or step（5）Obtained Sepharose microballoon is made into gel with hyaluronic acid, and concentration is respectively 1-
100 mg/ml and 1-50 mg/ml, are stirred overnight, and form it into the uniform gel of system.
It is subcutaneously monitored in mouse preserve situation in vivo with syringe injection 0.1ml-2ml gels；
In the present invention, step（5）Described in diamines can be ethylenediamine, propane diamine, butanediamine, pentanediamine, one in hexamethylene diamine
Kind；
In the present invention, step（5）Described in fluorescein can be fluorescein isothiocynate（Isomers I）, indocyanine green, rhodamine
One kind；
The Sepharose microballoon obtained using preparation method of the present invention, Size Distribution is controllable, and heat endurance is good and mechanicalness
Can be strong；
Sepharose fluorescent microsphere is prepared in the present invention, and fluorescent effect is good, and hypodermic injection can be convenient by living imaging
Efficiently observe it and retain situation in vivo, without obvious nocuity, be a kind of preferable and there is wide variety of injection fillers material
Material；
Sepharose microballoon hyaluronic acid derivatives has the characteristics that prepared by the present invention：1. prepared by inverse suspension method
Journey is simple, efficient, and with the potentiality of industrialization large-scale production；2. Sepharose microballoon, heat endurance is good and can pass through
Crosslinking degree is regulated and controled, high mechanical strength, and size is controllable；3. fluorescence molecule can be grafted, follow-up zoopery can be facilitated simple
Efficiently observation, and do not have nocuity；4. gel biological compatibility is good, no cytotoxicity, no obvious adverse reaction is subcutaneously injected；
5. gel can preserve in vivo at least two weeks and more than.It is a kind of to illustrate prepared Sepharose microballoon hyaluronic acid derivatives
It is excellent and can wide variety of ideal injection packing material.
Brief description of the drawings
Fig. 1 is that ungarbled size is about shone in the light microscope of the Sepharose of 10-700 microns in embodiment 1
Piece；
Fig. 2 be embodiment 4 in after screening size about in the optical microscope photograph of 80-250 micron Sepharoses；
Fig. 3 is the optical microscope photograph of heated post-crosslinking agarose overnight in embodiment 5, it is seen that microballoon form keep compared with
Good, stability is good in heating process；
Fig. 4 is the agarose microbeads photo of surface modification fluorescein isothiocynate FITC in embodiment 8；
Fig. 5 is the fluorescent microscopy images that FITC is grafted agarose microbeads in embodiment 8, and fluorescent effect is apparent；
Fig. 6 is Sepharose in embodiment 10（FITC is not met）/ hyaluronic acid derivatives injection material photo；
Fig. 7 is the relation that agarose/hyaluronic acid injectable packing material modulus of shearing varies with temperature in embodiment 11, different
Hyaluronic acid（HA）And agarose microbeads（AG）Proportioning, it is shown that the ratio of hyaluronic acid is higher in system, gel viscosity
It is bigger.The modulus of shearing of gel will not be decreased obviously because temperature raises, and maintained preferable viscosity, illustrated what is obtained with such a method
Gel is relatively stable, there is the potentiality for retaining the long period in vivo；
Fig. 8 is the small animal living body images of mouse in embodiment 12, and before left figure is injection, after right figure is injection, green is glimmering
Light part is injected gel position, and imaging effect is obvious.
Embodiment
Embodiment 1：The preparation of Sepharose microballoon
1. weigh 3g agarose powders and in 50ml deionized waters, heating stirring all dissolves to agarose under the conditions of 95 DEG C, shape
Into 6%（w/w）Agarose solution, solution viscosity is larger after dissolving；
2. pipetting 50ml normal octanes with liquid-transfering gun, 2g emulsifying agent Span-80 are added, stirring and dissolving, then use mechanical stirring device
High-speed stirred is carried out, mixing speed 800rpm, is warming up to 65 DEG C simultaneously in this course；
3. complete agarose solution will be dissolved（Viscosity is larger）Pour into while hot in the normal octane of high-speed stirred, maintain 65 DEG C,
High-speed stirred 30min；
4. temperature of reaction system is down into 50 DEG C, 2M NaOH solution 2ml, crosslinking agent epoxychloropropane 2g are added, reduction of speed stirs
Mix, mixing speed 250rpm, cross-linking reaction 1h；
5. removing heating, solution is layered after standing, and lower floor is agarose microbeads.Supernatant liquid is poured out, lower floor's solid is used successively
Ethanol, 20% ethanol-water solution and deionized water washing, the form centrifuged can be used by washing, and also can be used to stand and be divided
From or with sub-sieve filtration washing；
6. product should be stored in 20% ethanol-water solution or deionized water, interim slide is made, passes through light microscope
Observe the form and size of agarose microbeads.
Embodiment 2：The preparation of Sepharose microballoon
1. weigh 2g agarose powders and in 50g deionized waters, heating stirring all dissolves to agarose under the conditions of 100 DEG C, shape
Into 4%（w/w）Agarose solution, solution viscosity is larger after dissolving；
2. pipetting 50ml normal octanes with liquid-transfering gun, 2g emulsifying agent Span-80 are added, stirring and dissolving, then use mechanical stirring device
High-speed stirred is carried out, mixing speed 1000rpm, is warming up to 65 DEG C simultaneously in this course；
3. complete agarose solution will be dissolved（Viscosity is larger）Pour into while hot in the normal octane of high-speed stirred, maintain 65 DEG C,
High-speed stirred 30min；
4. temperature of reaction system is down into 50 DEG C, 2M KOH solution 2ml, crosslinking agent epoxychloropropane 2g are added, reduction of speed stirs,
Mixing speed 250rpm, cross-linking reaction 1h；
5. removing heating, solution is layered after standing, and lower floor is agarose microbeads.Supernatant liquid is poured out, lower floor's solid is used successively
Ethanol, 20% ethanol-water solution and deionized water washing, the form centrifuged can be used by washing, and also can be used to stand and be divided
From or with sub-sieve filtration washing；
6. product should be stored in 20% ethanol-water solution or deionized water, interim slide is made, passes through light microscope
Observe the form and size of agarose microbeads.
Embodiment 3：The preparation of Sepharose microballoon
1. weigh 2g agarose powders and in 50g deionized waters, heating stirring all dissolves to agarose under the conditions of 100 DEG C, shape
Into 4%（w/w）Agarose solution, solution viscosity is larger after dissolving；
2. pipetting 50ml normal octanes with liquid-transfering gun, 2g emulsifying agent Span-80 are added, stirring and dissolving, then use mechanical stirring device
High-speed stirred is carried out, mixing speed 1000rpm, is warming up to 65 DEG C simultaneously in this course；
3. complete agarose solution will be dissolved（Viscosity is larger）Pour into while hot in the normal octane of high-speed stirred, maintain 65 DEG C,
High-speed stirred 30min；
4. temperature of reaction system is down into 50 DEG C, 2M KOH solution 2ml, crosslinking agent 1, the bromo- 2- propyl alcohol 2g of 3- bis-, reduction of speed are added
Stirring, mixing speed 250rpm, cross-linking reaction 1h；
5. removing heating, solution is layered after standing, and lower floor is agarose microbeads.Supernatant liquid is poured out, lower floor's solid is used successively
Ethanol, 20% ethanol-water solution and deionized water washing, the form centrifuged can be used by washing, and also can be used to stand and be divided
From or with sub-sieve filtration washing；
6. product should be stored in 20% ethanol-water solution or deionized water, interim slide is made, passes through light microscope
Observe the form and size of agarose microbeads.
Embodiment 4：The size screening of Sepharose microballoon
The agarose microbeads that will be dispersed in deionized water are placed on sub-sieve, and 69 mesh are corresponding 250 microns at the middle and upper levels for it, lower floor
190 mesh are corresponding 80 microns.By the flushing repeatedly of a large amount of deionized waters, meet Particle size requirements（80-250 microns）Microballoon can stay
In the intermediate layer of sieve, larger microballoon retains in the upper layer, and less microballoon can be washed off with deionized water.Among preserving
Layer microballoon, and partially desiccated is taken, calculate its dry weight.
Embodiment 5：Agarose microbeads cross-linking properties is verified
The microballoon for taking 1g to be crosslinked is dispersed in 5ml deionized waters, is placed in 95 DEG C of baking oven and is heated overnight, and observing microballoon, it is
No dissolving, and characterize the front and rear micro-sphere structure of heating with light microscope.
Embodiment 6：Sepharose microsphere surface is grafted 1,6- hexamethylene diamines
1. 2g hexamethylene diamines are taken to be dissolved in 50ml 0.1M NaHCO3In；
2. take 4g（Weight in wet base）Agarose microbeads are added in above-mentioned solution after activation, are reacted at 50 DEG C, magnetic agitation, react about 5h；
3. removing heating, the microballoon obtained after reaction is placed in sub-sieve（190 mesh）On, washed with a large amount of deionized waters to filter
Liquid illustrates that unreacted hexamethylene diamine has been removed in neutrality；Get on and verify to connect with elemental analysis method checking hexamethylene diamine grafting
Branch amount, the results are shown in Table 1, provable hexamethylene diamine is successfully grafted on agarose.
Embodiment 7：Sepharose microsphere surface is grafted ethylenediamine
1. 1g ethylenediamines are taken to be dissolved in 25ml 0.1M NaHCO3In；
2. take 2g（Weight in wet base）Agarose microbeads are added in above-mentioned solution after activation, are reacted at 65 DEG C, magnetic agitation, react about 4h；
3. removing heating, the microballoon obtained after reaction is placed in sub-sieve（190 mesh）On, washed with a large amount of deionized waters to filter
Liquid illustrates that unreacted ethylenediamine has been removed in neutrality；Get on and verify to connect with elemental analysis method checking hexamethylene diamine grafting
Branch amount.
Embodiment 8：Microsphere surface is grafted fluorescein isothiocynate（FITC）
1. by 10 mg fluorescein isothiocynates（Isomers I）It is dispersed in 100 ml deionized waters, 15 min of ultrasound make its molten
Solution；
2. add 2g（Weight in wet base）The agarose microbeads of surface grafting hexamethylene diamine, the h of magnetic agitation 18 under normal temperature；
3. the microballoon that reaction is obtained later is placed in sub-sieve（190 mesh）On, microballoon is in orange-yellow, is washed with a large amount of deionizations
Wash, until cleaning solution is colourless, during the orange-yellow of microballoon do not take off.
Embodiment 9：Microsphere surface is grafted indocyanine green
1. 20 mg indocyanine greens are dispersed in 100 ml deionized waters, 15 min of ultrasound make its dissolving；
2. add 1g（Weight in wet base）The agarose microbeads of surface grafting hexamethylene diamine, the h of magnetic agitation 12 under normal temperature；
3. the microballoon that reaction is obtained later is placed in sub-sieve（190 mesh）On, washed with a large amount of deionized waters, until cleaning solution
It is colourless.
Embodiment 10：Injection fillers material is mixed with carrier hyaluronic acid
By Sepharose microballoon and hyaluronic acid powder with 50 mg/ml and 15 mg/ml concentration and proportioning and deionized water
Mixing, being stirred overnight makes system uniform.Obtained gel rubber system viscosity is very big.2 ml or so gel is taken to be fitted into 2 ml syringes
In case follow-up zoopery uses.
Embodiment 11：Dynamic rheology is tested
Be configured with three kinds of different mixing ratios gel go forward side by side Mobile state rheology test, as a result as shown in Figure 7.HA is represented in diagram
Hyaluronic acid（15mg/ml）, AG expression agarose microbeads（50mg/ml）, the two is with different volumes than mixing.
Embodiment 12：Small animal living body imaging test
By Sepharose fluorescent microsphere and hyaluronic acid powder with 50 mg/ml and 15 mg/ml concentration and proportioning with go from
Sub- water mixing, being stirred overnight makes system uniform.Obtained gel is subcutaneous in mouse with syringe injection 0.8ml, uses living imaging
Its state before and after injection is tested respectively.
Table 1 is to use elementary analysis test result after being grafted hexamethylene diamine in embodiment 6
Claims (10)
1. treatment gel is injected in a kind of vesicoureteric reflux, it is characterised in that is a kind of to be prepared by inverse suspension method
The gel that agarose microbeads and cross-linking hyaluronic acid are formed, or a kind of carry fluorophor by prepared by inverse suspension method
The gel that is formed of agarose microbeads and cross-linking hyaluronic acid.
2. a kind of preparation method of vesicoureteric reflux injection treatment gel, it is characterised in that concretely comprise the following steps：
（1）It is that raw material is configured to concentration as 1-10% using agarose powder and deionized water（w/w）Agarose solution；
（2）Using inverse suspension method by step（1）Agarose solution be prepared into agarose microbeads；
（3）In step（2）Crosslinking agent and alkali are added in the system of preparation, carries out cross-linking reaction, forms Sepharose microballoon；
（4）By step（3）The system of preparation is washed by separating for several times, obtains having the Sepharose of certain grain size distribution micro-
Ball；
（5）Step（4）Obtained Sepharose microballoon is configured to gel with hyaluronic acid, as injection filler；Or
By step（4）Obtained Sepharose microballoon grafting fluorescence molecule, is then configured to gel, as injection with hyaluronic acid again
Use filler.
3. preparation method according to claim 2, it is characterised in that step（2）Described in inverse suspension method, be using just
Octane is liquid phase, and using Span-80 as emulsifying agent, at a temperature of high speed machine stirring, 50-85 DEG C, it is water-soluble to add agarose
Liquid, it is reacted to obtain agarose microbeads.
4. preparation method according to claim 2, it is characterised in that step（3）Described in cross-linking reaction in 150-
Carried out under 1000rpm mixing speed, cross-linking reaction temperature is 35-80 DEG C, reaction time 0.5-6h；The crosslinking agent is selected from
The bromo- 2- propyl alcohol of epoxychloropropane, 1,3- bis-, the alkali is in sodium hydroxide, potassium hydroxide, ammoniacal liquor, sodium acid carbonate, sodium carbonate
One kind, it is or a variety of.
5. preparation method according to claim 2, it is characterised in that step（4）Described in separating, washing with solution be second
Alcohol, 20% ethanol-water solution and deionized water；Separation, which uses, to be centrifuged, centrifugal rotational speed 1000-5000rpm, 1-10min,
Or using standing separation, time of repose 0.5-5h；Then 65-190 mesh sub-sieve filtration washings are used, it is 80-250 to obtain particle diameter
The equally distributed Sepharose microballoon of micron.
6. preparation method according to claim 2, it is characterised in that step（5）In, Sepharose microballoon directly with thoroughly
Bright matter acid is configured to gel, and its concentration is 1-100 mg/ml, after Sepharose microballoon grafting fluorescence molecule again with hyaluronic acid
The gel being configured to, its concentration are 1-50 mg/ml.
7. preparation method according to claim 2, it is characterised in that the Sepharose microballoon grafting fluorescence molecule
Flow is：Take step（4）Obtained product is calculated as 1-5g by weight in wet base and is put in diamine solution, 35-50 DEG C of reaction under magnetic agitation
0.5-5h, diamine solution are to be added in 50ml 0.1M alkaline solutions to be formulated by 0.1-2g diamines；Recession is completed in reaction
Go to heat, the microballoon that reaction obtains is placed on sub-sieve, washed with a large amount of deionized waters to filtrate in neutrality；By 1-10 mg
Fluorescein is dispersed in 100 ml deionized waters, and 15-30min of ultrasound make its dissolving, is added and is calculated as 0.1-2g surfaces by weight in wet base and connects
The agarose microbeads of branch hexamethylene diamine, at room temperature magnetic agitation reaction 1-18 h；The microballoon that reaction obtains later is placed in sub-sieve
On, washed with a large amount of deionized waters, until the colourless Sepharose microballoon for remembering to be grafted fluorophor of cleaning solution.
8. preparation method according to claim 7, it is characterised in that described diamines is ethylenediamine, propane diamine, fourth two
Amine, pentanediamine, one kind in hexamethylene diamine.
9. preparation method according to claim 7, it is characterised in that described fluorescein is fluorescein isothiocynate（It is different
Structure body I）, indocyanine green, one kind in rhodamine.
A kind of 10. application of gel as described in claim 1 in vesicoureteric reflux injection is prepared.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110448737A (en) * | 2019-07-09 | 2019-11-15 | 复旦大学附属儿科医院 | Treatment magnetic gel and preparation method thereof is injected in vesicoureteric reflux |
Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20080107744A1 (en) * | 2006-11-06 | 2008-05-08 | Jack Fa-De Chu | Injectable hollow tissue filler |
| CN101591441A (en) * | 2009-02-23 | 2009-12-02 | 协和生物制药（天津）有限公司 | The method that a kind of hyaluronic acid is connected with sepharose |
| CN101736087A (en) * | 2010-01-28 | 2010-06-16 | 华东医学生物技术研究所 | Method for digitally detecting micro-mutation by using micro-emulsion clone amplified bound water gel microsphere chip |
| CN101903408A (en) * | 2007-10-30 | 2010-12-01 | 粘凝胶股份公司 | Chitosan composition |
| CN102836465A (en) * | 2012-08-30 | 2012-12-26 | 浙江星月生物科技股份有限公司 | Silk-fibroi and hyaluronic-acid (HA) composite gel for injection and preparation and application thereof |
| WO2014072525A1 (en) * | 2012-11-12 | 2014-05-15 | MAX-PLANCK-Gesellschaft zur Förderung der Wissenschaften e.V. | Metal(iii)-polypeptide complexes |
| CN104774337A (en) * | 2014-08-20 | 2015-07-15 | 江阴市柏御天谷生物医药有限公司 | Agarose microsphere-containing cross-linked sodium hyaluronate gel for injection and preparation method thereof |
| CN104785222A (en) * | 2015-03-25 | 2015-07-22 | 安徽师范大学 | Preparation method and application of chitosan composite material |
| CN104826605A (en) * | 2015-04-24 | 2015-08-12 | 广州极泰生物科技有限公司 | Preparation of hyper-crosslinking uniform-particle-size agarose gel medium by using ultrasonic reversed phase suspension method |
-
2017
- 2017-08-16 CN CN201710702758.3A patent/CN107376032A/en active Pending
Patent Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20080107744A1 (en) * | 2006-11-06 | 2008-05-08 | Jack Fa-De Chu | Injectable hollow tissue filler |
| CN101903408A (en) * | 2007-10-30 | 2010-12-01 | 粘凝胶股份公司 | Chitosan composition |
| CN101591441A (en) * | 2009-02-23 | 2009-12-02 | 协和生物制药（天津）有限公司 | The method that a kind of hyaluronic acid is connected with sepharose |
| CN101736087A (en) * | 2010-01-28 | 2010-06-16 | 华东医学生物技术研究所 | Method for digitally detecting micro-mutation by using micro-emulsion clone amplified bound water gel microsphere chip |
| CN102836465A (en) * | 2012-08-30 | 2012-12-26 | 浙江星月生物科技股份有限公司 | Silk-fibroi and hyaluronic-acid (HA) composite gel for injection and preparation and application thereof |
| WO2014072525A1 (en) * | 2012-11-12 | 2014-05-15 | MAX-PLANCK-Gesellschaft zur Förderung der Wissenschaften e.V. | Metal(iii)-polypeptide complexes |
| CN104774337A (en) * | 2014-08-20 | 2015-07-15 | 江阴市柏御天谷生物医药有限公司 | Agarose microsphere-containing cross-linked sodium hyaluronate gel for injection and preparation method thereof |
| CN104785222A (en) * | 2015-03-25 | 2015-07-22 | 安徽师范大学 | Preparation method and application of chitosan composite material |
| CN104826605A (en) * | 2015-04-24 | 2015-08-12 | 广州极泰生物科技有限公司 | Preparation of hyper-crosslinking uniform-particle-size agarose gel medium by using ultrasonic reversed phase suspension method |
Non-Patent Citations (6)
| Title |
|---|
| ARNE STENBERG等: "A NEW BIOIMPLANT FOR THE ENDOSCOPIC TREATMENT OF VESICOURETERAL REFLUX: EXPERIMENTAL AND SHORT-TERM CLINICAL RESULTS", 《THE JOURNAL OF UROLOGY》 * |
| FERRUH SIMSEK等: "ORDINARY GLASS SPHERES AS AN ALTERNATIVE INJECTABLE MATERIAL FOR ENDOSCOPIC CORRECTION OF VESICOURETERAL REFLUX", 《THE JOURNAL OF UROLOGY》 * |
| LING-MIN ZHANG等: "Synthesis and characterization of a degradable composite agarose/HA hydrogel", 《CARBOHYDRATE POLYMERS》 * |
| 刘宏波等: "不同尺寸单分散 PMMA/GMA/DVB 聚合物荧光微球的制备", 《化学学报》 * |
| 潘丹卓等: "琼脂糖凝胶微球活化度的考察", 《山东化工》 * |
| 许加超主编: "《海藻化学与工艺学》", 30 September 2014, 中国海洋大学出版社 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110448737A (en) * | 2019-07-09 | 2019-11-15 | 复旦大学附属儿科医院 | Treatment magnetic gel and preparation method thereof is injected in vesicoureteric reflux |
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