CN104841010A - Novel PHBV modified induced type bone repair scaffold material - Google Patents
Novel PHBV modified induced type bone repair scaffold material Download PDFInfo
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Abstract
A novel PHBV modified induced type bone repair scaffold material is prepared by the steps: dissolving amino acetic acid in a sodium bicarbonate solution, to obtain a first solution; dissolving di-tert-butyl dicarbonate in dioxane, to obtain a second solution; dropwise adding the second solution into the first solution; adjusting the pH value to neutral, dissolving the solids, dropwise adding the filtrate into cold ethyl ether, washing and drying an insoluble substance, and thus obtaining dicarbonate t-butyloxycarboryl-glycine; carrying out condensation on icariin and dicarbonate t-butyloxycarboryl-glycine, adding a mixed liquid of trifluoroacetic acid and dichloromethane into the product, and after the reaction is stopped, evaporating; dissolving the remainder with ethyl acetate; removing the solvent, to obtain aminated icariin; carrying out a condensation reaction; pouring aminated icariin-PHBV into a chitosan solution, injecting into an annular mold, freezing with liquid nitrogen, demoulding, drying, and thus obtaining the novel PHBV modified induced type bone repair scaffold material. The bone repair scaffold material can promote directional differentiation of stem cells into osteoblasts, accelerates bone growth and functional recovery, and can effectively repair bone defects.
Description
Technical field
The invention belongs to field of medical technology, relate to a series of poly butyric ester altogether hydroxyl valerate (PHBV) modification induction type bone renovating bracket material, made by methods such as the chemical modifications of the structure of material, Physical Processing and Chinese medicine monomer icariin.
Background technology
Bone injury is disease common clinically, and bone is a kind of natural inorganic matter with complicated hierarchy and organic composite.Fracture, Cranial defect and bone that the reasons such as war, vehicle accident, industrial injury, athletic injury, disease and natural disaster cause lack
lose woundthe trouble number whole world reaches millions of every year, and the bone of damage can with endogenous bone or external source Bone Defect Repari.Endogenous bone is easily accepted by patient, and repairing effect is better, but its quantity is very limited, and needs second operation, likely brings other to damage; And although it is easy to adopt external source bone to draw materials, repairing effect can not show a candle to endogenous bone, and current external source Bone Defect Repari means exist immunological rejection, difficult degradation, may with potential pathogen and other side reactions etc.So adopt the material of artificial preparation as bone renovating material clinically more and more widely.
Research shows, bone renovating material itself must possess: enough mechanical strengths, the material degradation speed matched with Bone Ingrowth, good biocompatibility and certain organize inducibility.
Bone renovating material is that a class can repair body tissue, substitute and regeneration, has the material of specific function effect.Such material, in clinical application, provides possibility for wound (trouble) person recovers normal physiological function, also can avoid the problem adopting endogenous bone or external source bone to bring simultaneously.
In view of the shortcoming of traditional osseous tissue biomaterial, the third generation bioactive materials with " initiatively repair function " and " controllable biological response " characteristic becomes current study hotspot and following developing direction.
Summary of the invention
The shortcoming that the object of the invention is to overcome existing bone renovating material, with not enough, provide a series of and possesses suitable mechanical property and effectively can induce bone formation again, thus realizes the bone tissue engineering stent material repaired effectively fast Cranial defect.
For achieving the above object, the technical solution adopted in the present invention is: a kind of
novelpHBV modification induction type bone renovating bracket material, it is characterized in that, this bone renovating bracket material obtains according to the following steps:
Step 1: be dissolved in by glycine in sodium bicarbonate solution, stirs, obtains the first solution; Bis(tert-butoxycarbonyl)oxide is dissolved in dioxane completely, after cooling, forms the second solution;
Step 2: in ice-water bath, under the condition stirred, the second solution is instilled the first solution, after dropwising, continue stirring reaction 20 ~ 24h under room temperature;
Then, regulate the pH value of the rear liquid of reaction to neutral with 0.01M hydrochloric acid solution, except desolventizing, residual solids dichloromethane dissolves, filter, filtrate is dropped in cold diethyl ether, separate out solid, collected by suction insoluble matter, with washed with diethylether, vacuum drying, obtain two carbonic acid tertbutyloxycarbonyl-glycine;
Step 3: be dehydrant with dicyclohexylcarbodiimide, take DMAP as catalyst, icariin and two carbonic acid tertbutyloxycarbonyl-glycine are carried out condensation reaction by 1 ︰ 1 in mass ratio, obtain icariin-glycine-two carbonic acid tertbutyloxycarbonyl;
Step 4: trifluoroacetic acid and dichloromethane are mixed into mixed solution, adds in icariin-glycine-two carbonic acid tertbutyloxycarbonyl, stirred at ambient temperature by this mixed solution, reaction stops, after rotary evaporation; Surplus materials is dissolved in ethyl acetate completely, wash to pH value be 8 ~ 9; Except desolventizing, obtain amination icariin;
Step 5: by condensation reaction, couples together the carboxyl of carboxymethyl on the amino in amination icariin and chitosan, obtains amination icariin-PHBV;
Step 6: by mass percentage, pours in the chitosan solution of 20% by the amination icariin-PHBV of 80%, and after stirring, annular stainless steel mould is injected in extruding, be placed in the freezing 10 ~ 20min of liquid nitrogen, take out rapidly, put into cold water 5 ~ 10s that temperature is 4 DEG C, the demoulding, dry, obtained
novelpHBV modification induction type bone renovating bracket material.
The present invention has and organizes the bone renovating bracket material of inductivity function to be a kind of PHBV and Chitosan Composite Scaffold Materials of compound icariin medicine, and tool has the following advantages:
1) icariin and the covalently bound preparation technology of material reach designing requirement, achieve the slow releasing function of medicine, and medicine is to the continuous action of tissue induction;
2) material has the effect that good biomechanical strength can play support rib growth;
3) effect of icariin and the covalently bound slow release of material promotes stem cell to osteoblast directed differentiation, accelerated bone growth and functional rehabilitation;
4) there is good degradability and biocompatibility, meet the requirement of tissue engineering bracket material;
5) can effective repairing bone defect.
Accompanying drawing explanation
fig. 1it is the infrared contrast of the amino modified front and back of icariin
figure.
fig. 2that icariin is combined with PHBV material and amino modified infrared contrast
figure.
fig. 3it is the amino modified front particle diameter of icariin
figure.
fig. 4be icariin amino modified after particle diameter
figure.
fig. 5it is X-ray diffraction (XRD) contrast of the amino modified front and back of icariin
figure.
fig. 6it is differential thermal-thermogravimetric analysis (TG/DTA) contrast of the amino modified front and back of icariin
figure.
fig. 7it is the cross-sectional scans Electronic Speculum of bone renovating bracket material of the present invention
figure.
fig. 8it is the scanning electron microscope of bone renovating bracket material pattern of the present invention
figure.
fig. 9it is the scanning electron microscope of bone renovating bracket material superficial cell of the present invention growth
figure.
fig. 10 is histological stain (HE dyeing) optical microscope of icariin-PHBV bone renovating bracket material when carrying out zoopery January
figure.
fig. 11 is histological stain (HE dyeing) optical microscope of amination icariin-PHBV bone renovating bracket material of the present invention when carrying out zoopery January
figure.
fig. 12 is icariin-PHBV bone renovating bracket material histological stain (HE dyeing) optical microscopes when carrying out zoopery March
figure.
fig. 13 is amination icariin-PHBV bone renovating bracket material of the present invention histological stain (HE dyeing) optical microscopes when carrying out zoopery March
figure.
fig. 14 is icariin-PHBV bone renovating bracket material histological stain (HE dyeing) optical microscopes when carrying out zoopery June
figure.
fig. 15 is amination icariin-PHBV bone renovating bracket material of the present invention histological stain (HE dyeing) optical microscopes when carrying out zoopery June
figure.
fig. 16 histological stain (Masson dyeing) optical microscopes when being icariin-PHBV timbering material zooperal June
figure.
fig. 17 histological stain (Masson dyeing) optical microscopes when being amination icariin-PHBV timbering material zoopery of the present invention June
figure.
Detailed description of the invention
Below in conjunction with
accompanying drawingthe present invention is described in detail with detailed description of the invention.
The invention provides and a kind of there is the PHBV bone renovating bracket material organizing inductivity function, using the PHBV Chitosan Composites of compound icariin medicine as bone renovating bracket material, have and organize inductivity, for bone defect healing, can effectively improve bone injury reparation and functional rehabilitation thereof, reduce disability rate.
This bone renovating bracket material specifically obtains according to the following steps:
Step 1: 2mmo1 glycine is dissolved in the NaHCO that 20mL mass percent concentration is 1.68%
3in solution, stirring at room temperature 1 ~ 2h, obtains the first solution; 0.45g Bis(tert-butoxycarbonyl)oxide (Boc) is dissolved in dioxane completely, after cooling, forms the second solution;
Step 2: in ice-water bath, under the condition stirred, 2m,mo1 second solution 2 mL is slowly instilled the first solution, after dropwising, continue stirring reaction 20 ~ 24h under room temperature; Reaction equation is:
Then, extremely neutral with the pH value of liquid after 0.01M hydrochloric acid solution adjustment reaction, rotary evaporation removes desolventizing, residual solids dichloromethane (CH
2cCl
2) dissolve, cross and filter insoluble matter (NaC1), filtrate is dropped in cold diethyl ether, separate out faint yellow solid, collected by suction insoluble matter, with washed with diethylether 3 ~ 5 times, be placed in vacuum drying oven dry, obtain two carbonic acid tertbutyloxycarbonyl-glycine (Boc-GLY);
Step 3: with dicyclohexylcarbodiimide (DCC) for dehydrant, with DMAP (DMAP) for catalyst, in mass ratio 1 ︰ 1 by icariin (Icariin) and Boc-GLY(Boc-GLY little over amount) carry out condensation reaction, obtain icariin-glycine-two carbonic acid tertbutyloxycarbonyl (Icariin-GLy-Boc), reaction equation is:
Step 4: 9mL trifluoroacetic acid and 10mL dichloromethane are mixed into mixed solution, then joins in 5g Icariin-GLy-Boc by this mixed solution, stirred at ambient temperature 1 ~ 2h, and reaction stops, after rotary evaporation; Surplus materials being put into appropriate ethyl acetate to dissolve completely, then, is the NaCO of 5% with mass percent concentration
3solution washing to pH value is 8 ~ 9; Except desolventizing, obtain the product after sloughing Boc protecting group, this product is amination icariin (Icariin-NH
2), reaction equation is:
Step 5:Icariin-NH
2be connected with PHBV:
Utilize carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) method (list of references: Wang Yingjun, Yang Chunrong, Wang Lingyun. EDC/NHS is cross-linked the impact on collagen physical and chemical performance, South China Science & Engineering University's journal (natural science edition), 2007,35(12): 66-70), by condensation reaction, by Icariin-NH
2in amino and chitosan on the carboxyl of carboxymethyl couple together, obtain amination icariin-PHBV(Icariin-NH
2-PHBV), reaction equation is:
Step 6: by mass percentage, by the Icariin-NH of 80%
2-PHBV pours in the chitosan solution of 20%, after refiner high speed (2500 revs/min) stirs, annular stainless steel mould (the length 10 ~ 12cm of this mould, internal diameter 1 ~ 1.2mm, external diameter 4 ~ 4.2mm) is injected in extruding, then the freezing 10 ~ 20min of liquid nitrogen is together placed in together with mould, rapid taking-up, puts into cold water 5 ~ 10s that temperature is 4 DEG C, the demoulding, drying, obtained
novelpHBV modification induction type bone renovating bracket material.
Icariin effectively can promote the propagation of mesenchymal stem cells MSCs (MSCs); And promote that MSCs is to osteoblast differentiation, thus can be considered a kind of good bone-inducting active factor by the expression of inducible alkaline phosphatase, Bone Gla protein and the formation of calcium scoring.The present invention utilizes chemical modification method to make icariin modified with PHBV material covalent bond, and the degraded along with PHBV material makes icariin realize slow release, promotes that the effect of proliferation and differentiation of osteoblasts realizes the function of tissue repair.
the evaluation of PHBV modification induction type bone renovating bracket material of the present invention:
1) determination of infrared spectroscopy: potassium bromide grinds, film-making, utilizes the Fourier transform infrared spectroscopy (Fourier transform infrared spectrometer, FTIR) of day island proper rule C8-20 type Fourier sample.Icariin before modified after infrared contrast
figure,
as Fig. 1shown in,
in figuredisplay, amination icariin is at 3371cm
-1and 3328cm
-1the N-H stretching vibration peak that place's appearance 2 moderate strengths, shape are sharp-pointed, at 1689cm
-1there is N-H angle vibration peak, show NH
2existence; At 1708cm
-1, 1625cm
-1occurring C=O stretching vibration peak, is the characteristic peak of ester group; 2926cm
-1, the corresponding C-H stretching vibration peaks in 2 peaks that occur of 2850cm-1 place, show the appearance of methylene; Above result table
bright excessivesheep icariin is by successfully amination modified.Utilize the infrared signature absworption peak of DCC judge reaction carry out degree and whether raw material processes completely, icariin is combined and amino modified infrared contrast with PHBV material
figure,
as Fig. 2shown in, from
fig. 2can find out in chitosan microball process ,-the NH on chitosan
2schiff alkali cross-linking reaction is there occurs, 1714cm with the-CHO on glutaraldehyde
-1, 1560 cm
-1there is significantly new absworption peak in place; With the INFRARED ABSORPTION of chitosan
figurespectrum is compared, amination icariin-chitosan (Icariin-NH
2-CSM) INFRARED ABSORPTION
figurethere is not new characteristic peak in spectrum, shows Icariin-NH
2and there is no the generation of new key between chitosan, but all there is a certain amount of red shift, Icariin-NH in its principal character peak
2mainly with the mode of physically trapping embed by chitosan, Icariin-NH is described
2may with covalently cross-linked mode fix by chitosan.Intermediate product Icariin-Gly-Boc is at 2117cm
-1the characteristic peak occurred is contain C=N in DCC structure, shows end product Icariin-NH
2peak is not had to occur herein.
2) particle size distribution: utilize the LS-POP sound state multi-angle laser light of the American-European gram Instrument Ltd. in Zhuhai instrument that falls apart to measure particle size distribution.The grain size curve figure of Fig. 3 to be the grain size curve figure of the amination modified front microsphere of icariin, Fig. 4 be amination modified rear microsphere of icariin, show in figure, the particle diameter of two kinds of microspheres mainly concentrates on 3 ~ 12
, 3 ~ 9
in scope, compared with SEM observed result, its particle size range all has increase to a certain degree, and mainly part microsphere aggregation causes.
3) X-ray diffraction: utilize Rigaku XG control RINT2500 X-ray diffraction (X-ray diffraction, XRD) instrument to analyze the thing phase of icariin and amination icariin respectively.
as Fig. 5shown in,
in figuredisplay, before modified, sample crystallinity is all better; After amination modified, due to the existence of amino, the crystallinity of sample strengthens to some extent, and the main diffraction peak of sample all presents low-angle skew, table
bright excessiveafter the amination of sheep icariin, its crystallinity strengthens, and contributes to material crystalline and is shaped.
4) thermal weight loss and differential thermal analysis: utilize the SDT-Q600 type Re Chong – differential thermal analyzer (thermogravimetry-differential thermal analysis, TG – DTA) of TA company of the U.S. to analyze thermal weight loss and the differential thermal analysis of icariin sample and amination icariin sample respectively.As shown in Figure 6, show in figure, after amination modified, by amino impact, Ica-NH
2heat stability obviously reduce.The obvious weightlessness of Ica appears near 190 DEG C, and Ica-NH
2then occur near 160 DEG C significantly weightless.There are 2 obvious exothermic peaks respectively in corresponding DTA curve display icariin near 57 DEG C and near 196 DEG C, and Ica-NH
2then near 51 DEG C, near 108 DEG C and near 222 DEG C, there are 3 obvious exothermic peaks, after showing icariin amination, heat stability obviously reduces than before amination, heat stability reduction shows that material mixability is higher, contributes to homogeneous and effective release of medicine during material degradation.
5) scanning electron microscope (SEM) is observed: after bone renovating bracket material section metal spraying of the present invention, utilizes Dutch PHILIPS company XL-30 environmental scanning electron microscope to observe the cross-section morphology of material; This bone renovating bracket material cross-sectional scans Electronic Speculum figure, as shown in Figure 7, show this bone renovating bracket material external diameter about 3.4 ~ 3.5mm in figure, hollow aperture is about 740 ~ 760
, and structure is comparatively fine and close.The topography scan Electronic Speculum figure of this bone renovating bracket material, as shown in Figure 8, chitosan material is as the network-like distribution of base material as can be seen from Figure, PHBV is spherical in link, between the network being evenly distributed on chitosan polymer, chitosan network and PHBV sphere network form inierpeneirating network structure, have certain micropore distribution, micropore size 3 ~ 10 between network
.Illustrate that this material has certain pore structure, contribute to the formation of mass exchange and tissue.
6) Evaluating Mechanical Properties: the comprcssive strength utilizing Instron5865 universal material test machine test material, balances 5 samples.Experimental result shows, the bending strength 45 ~ 50MPa of bi-material, and the bending strength being significantly greater than spongy bone requires (to be greater than 20 M P a).Show that such material possesses certain mechanical strength, can certain supporting role be played in bone defect healing process.
the evaluation of its biocompatibility of PHBV modification induction type bone renovating bracket material of the present invention
This bone renovating bracket material of the present invention is placed in DMEM/F12 culture medium, after inoculation rat bone marrow mesenchymal stem cells (MSCs), 37 DEG C of 5% CO
2incubator cultivates 3d and 7d respectively, the combining case of scanning electric mirror observing cell and material,
as Fig. 9shown in,
in figureshowed cell can grow at material surface, breed, and illustrates that this material is to cytotoxic side effect, has good biocompatibility.
the repairing effect evaluation of PHBV modification induction type bone renovating bracket material of the present invention
The preparation of matched group bone renovating bracket material: by mass percentage, the PHBV+Icarin of 80% is poured in 20% chitosan solution, after refiner high speed (2500 revs/min) stirs, annular stainless steel mould (the length 10 ~ 12cm of this mould, internal diameter 1 ~ 1.2mm, external diameter 4 ~ 4.2mm) is injected in extruding, then the freezing 10 ~ 20min of liquid nitrogen is together placed in together with mould, rapid taking-up, put into cold water 5 ~ 10s that temperature is 4 DEG C, the demoulding, drying, obtained icariin-PHBV material.
1) zoopery: select cleaning grade Chinchilla Rabbit 18, body weight 3 ~ 4 kg, male and female are regardless of, and are divided into 2 groups at random, i.e. experimental group (amination icariin-PHBV material group) and matched group (icariin-PHBV material group), often organize 9.Experimental rabbit auricular vein in giving 2 groups by the consumption of every kg body weight 30mg is injected 3% pentobarbital sodium and is anaesthetized, and left side forearm shaves hair, and 2.5% iodine disinfection also spreads aseptic hole-towel.In Radial Forearm stringer otch, successively appear radius backbone, excision left front leg radius 1.0 ~ 1.2cm bone section, experimental group implants amination icariin-PHBV, and matched group implants icariin-PHBV material.Raise under postoperative Clean Facility.
2) Bone Defect Repari imaging evaluation: respectively at postoperative January, March, do June three three dimensional CT and X-ray iconography detect, observe its Bone Defect Repari situation.Result shows: during postoperative January, experimental group and matched group Cranial defect place Image detection zero difference.During postoperative March, experimental group defect place bone bridge is set up, and area of new bone is along Material growth; When 6 months, area of new bone growth significantly increases, and defect place bone density obviously increases.During matched group 3 months, defect place bone bridge is not yet set up, area of new bone negligible amounts when 6 months, and defect place bone density is starkly lower than experimental group.
3) histology's (HE and Masson dyeing) analyze: respectively at month after operation, 3 months and 6 months time, intravenous injection air method puts to death rabbit, draw materials and total length Bone Defect Repari place draw materials, routine paraffin wax bag quilt, HE and Masson dyes, the characteristics of organizational structure that under optical microscope, the biocompatibility of observation analysis material and host, inflammatory reaction degree and host/material interface are formed.During month after operation, experimental group and matched group Cranial defect place host/material interface invade profit without inflammatory cell, mainly based on fibroblast;
as Fig. 10 He
fig. 1shown in 1.During 3 months after operation, matched group surrounding materials mainly based on fibrous tissue,
as Fig. 1shown in 2; During 3 months after operation, experimental group defect place has area of new bone to be formed,
as Fig. 1shown in 3.When postoperative 6 months, matched group has a small amount of area of new bone to be formed, surrounding materials to become fibrous tissue in the majority,
as Fig. 14
figureshown in 16; Experimental group defect place has a large amount of area of new bone to be formed, and part area of new bone enters in edge of materials,
as Fig. 1shown in 5 and 17; Two pack support material component are degraded.
Confirmed: PHBV inductivity bone renovating material of the present invention, by amination icariin induction of bone growth, achieves the tissue engineering bracket material having and organize inductivity Bone Defect Repari, to being formed, there is independent intellectual by above-mentioned experiment
property rightbiomaterial and for clinical treatment.
By building the bone renovating material in conjunction with icariin medicine, making it have and organizing inductivity function, effectively solve bone injury reparation and functional rehabilitation thereof.The raw material of PHBV chitosan co-induction bone renovating material of the present invention can industrialization large-scale production, abundance, and tissue engineering bracket forming method is simply easy to operation, nontoxic, the easy degraded of material therefor and have good biocompatibility.Modification icariin and the covalently bound preparation technology of material reach designing requirement, can realize effective sustained drug release effect.
Claims (6)
1. a novel PHBV modification induction type bone renovating bracket material, it is characterized in that, this bone renovating bracket material obtains according to the following steps:
Step 1: be dissolved in by glycine in sodium bicarbonate solution, stirs, obtains the first solution; Bis(tert-butoxycarbonyl)oxide is dissolved in dioxane completely, after cooling, forms the second solution;
Step 2: in ice-water bath, under the condition stirred, the second solution is instilled the first solution, after dropwising, continue stirring reaction 20 ~ 24h under room temperature;
Then, regulate the pH value of the rear liquid of reaction to neutral with 0.01M hydrochloric acid solution, except desolventizing, residual solids dichloromethane dissolves, filter, filtrate is dropped in cold diethyl ether, separate out solid, collected by suction insoluble matter, with washed with diethylether, vacuum drying, obtain two carbonic acid tertbutyloxycarbonyl-glycine;
Step 3: be dehydrant with dicyclohexylcarbodiimide, take DMAP as catalyst, icariin and two carbonic acid tertbutyloxycarbonyl-glycine are carried out condensation reaction by 1 ︰ 1 in mass ratio, obtain icariin-glycine-two carbonic acid tertbutyloxycarbonyl;
Step 4: trifluoroacetic acid and dichloromethane are mixed into mixed solution, adds in icariin-glycine-two carbonic acid tertbutyloxycarbonyl, stirred at ambient temperature by this mixed solution, reaction stops, after rotary evaporation; Surplus materials is dissolved in ethyl acetate completely, wash to pH value be 8 ~ 9; Except desolventizing, obtain amination icariin;
Step 5: by condensation reaction, couples together the carboxyl of carboxymethyl on the amino in amination icariin and chitosan, obtains amination icariin-PHBV;
Step 6: by mass percentage, amination icariin-the PHBV of 80% is poured in the chitosan solution of 20%, after stirring, annular stainless steel mould is injected in extruding, is placed in the freezing 10 ~ 20min of liquid nitrogen, takes out rapidly, put into cold water 5 ~ 10s that temperature is 4 DEG C, the demoulding, dry, obtained novel PHBV modification induction type bone renovating bracket material.
2. novel PHBV modification induction type bone renovating bracket material according to claim 1, is characterized in that, in described step 1,2mmo1 glycine is dissolved in the NaHCO that 20mL mass percent concentration is 1.68%
3in solution, obtain the first solution; 0.45g Bis(tert-butoxycarbonyl)oxide is dissolved in dioxane completely, after cooling, forms the second solution.
3. novel PHBV modification induction type bone renovating bracket material according to claim 1, is characterized in that, in described step 3, two carbonic acid tertbutyloxycarbonyl-glycine are little over amount.
4. novel PHBV modification induction type bone renovating bracket material according to claim 1, it is characterized in that, in described step 4,9mL trifluoroacetic acid and 10mL dichloromethane are mixed into mixed solution, then this mixed solution are joined in 5g icariin-glycine-two carbonic acid tertbutyloxycarbonyl.
5. novel PHBV modification induction type bone renovating bracket material according to claim 1, it is characterized in that, in described step 4, be the NaCO of 5% with mass percent concentration
3solution washing to pH value is 8 ~ 9.
6. novel PHBV modification induction type bone renovating bracket material according to claim 1, is characterized in that, length 10 ~ 12cm, internal diameter 1 ~ 1.2mm, the external diameter 4 ~ 4.2mm of stainless steel mould used in step 6.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106905444A (en) * | 2017-03-06 | 2017-06-30 | 张君涛 | Conjugate of icariin and hyaluronic acid and its preparation method and application |
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| US20140141082A1 (en) * | 2012-11-16 | 2014-05-22 | Song Gao | Compositions Containing Enriched Natural Crocin and/or Crocetin, and Their Therapeutic or Nutraceutical Uses |
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| CN101928328A (en) * | 2009-06-17 | 2010-12-29 | 李毅林 | RGD peptide-epimedium flavone combination, preparation method and application thereof |
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