CN102600510A - Absorbable bone induction polypeptide-carried microsphere and preparation method thereof - Google Patents
Absorbable bone induction polypeptide-carried microsphere and preparation method thereof Download PDFInfo
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- CN102600510A CN102600510A CN2011100215368A CN201110021536A CN102600510A CN 102600510 A CN102600510 A CN 102600510A CN 2011100215368 A CN2011100215368 A CN 2011100215368A CN 201110021536 A CN201110021536 A CN 201110021536A CN 102600510 A CN102600510 A CN 102600510A
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Abstract
The invention relates to an absorbable bone induction polypeptide-carried microsphere, comprising a water-soluble chitosan carrier and BMP-2 (bone morphogenetic protein-2) active polypeptide, wherein the weight ratio of the water-soluble chitosan carrier to the BMP-2 active polypeptide is (0.1-1000): 1. The absorbable bone induction polypeptide-carried microsphere provided by the invention can be prepared within 24 hours, thus a preparation method is simple; the absorbable bone induction polypeptide-carried microsphere has good biocompatibility and has the characteristics of low temperature, low vacuum, low energy consumption and low pollution, and the requirement that material is synthesized by adopting a 'soft processing' manner can be met. The microsphere prepared by the preparation method provided by the invention is spherical, particle size distribution is 10-100 mu m, an in-vitro degradation experiment shows that a growth factor is persistently and slowly released, and a preparation technology has no obvious influence to biological function of the active polypeptide.
Description
Technical field
The invention belongs to the bio-medical material technical field, it is simple especially to relate to a kind of preparation, good biocompatibility, absorbable microsphere with bone-inducting active polypeptide slow-release function and preparation method thereof.
Background technology
Bone is damaged to be the difficult problem that orthopaedics faces always.In recent years, more and more receive various countries scholar's concern, become one of research focus of current biomaterial through compound skeletal growth factor on bone renovating material.
(bone morphogenetic proteins BMP) belongs to the transforming growth factor member to bone morphogenetic protein, is a kind of multi-functional cell growth factor, and BMP has the ability that promotes union of fracture, has shortened the healing cycle of fracture.At present given very high evaluation to BMP at clinical bone inductive effect both at home and abroad.Wherein BMP-2 is one of topmost bone formation regulatory factor.But natural B MP-2 limited amount, and the life activity is difficult to performance.By the synthetic small peptide that contains 1 24 nucleotide of BMP-2 Core Feature region amino acid sequence---BMP-2 active polypeptide, correlational study show that the BMP-2 active polypeptide has good induced osteogenesis ability.Yet the easy in vivo diffusion dilution of BMP-2 active polypeptide, availability is low.
In order better to bring into play the biological function of BMP-2 active polypeptide; Realize it and repair the slowly release that continues at position at bone; The present invention is carrier material with the water-soluble chitosan; Preparation has the chitosan microball of BMP-2 bioactive peptide control-release function, thereby for bone reparation field the carrier system with somatomedin control-release function is provided.
Summary of the invention
The object of the present invention is to provide a kind of preparation simple, good biocompatibility, obvious results
Bone was induced in absorbable yearThe polypeptide microsphere.
For realizing above-mentioned purpose, the present invention adopts following technical scheme:
A kind of
Bone was induced in absorbable yearThe polypeptide microsphere comprises water-soluble chitosan carrier and BMP-2 active polypeptide.The weight ratio of water-soluble chitosan carrier and BMP-2 active polypeptide is 0.1～1000:1.
More excellent scheme is: described water-soluble chitosan is a kind of of carboxymethyl chitosan, Pyrusussuriensis acyl chitosan, p-benzoyl chitosan etc.
Another object of the present invention is to provide a kind of
Bone was induced in absorbable yearThe preparation technology of polypeptide microsphere.
For realizing above-mentioned purpose, the present invention adopts following technical scheme:
Step 1: 0.1g～10g water-soluble chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.01g～5g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 5ml ~ 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To sodium tripolyphosphate (TPP) aqueous solution of Dropwise 5 ml ~ 50mL 5% (mass fraction) wherein, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
This method for preparing prescription is simple, and preparation technology is prone to row.In 24 hours, can prepare, preparation is simple, good biocompatibility.And have low temperature, coarse vacuum, low energy consumption, oligosaprobic characteristics.Meet " soft processing " (Soft Processing) mode and carry out the synthetic requirement of material.Be regular spherical on the microsphere form of the inventive method preparation, particle size distribution is at 10～100 μ m, and the external degradation experiment finds that somatomedin is lasting and slowly discharges, and preparation technology does not have appreciable impact to the biological function of bioactive peptide.
The specific embodiment
Below in conjunction with specific embodiment the present invention is explained further details:
Embodiment 1
Step 1: the 1g carboxymethyl chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.1g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 10mL 5% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Embodiment 2
Step 1: the 2g carboxymethyl chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.1g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 25mL 5% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Embodiment 3
Step 1: the 2g water-soluble chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.2g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 10ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 25mL 5% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Embodiment 4
Step 1: the 2g carboxymethyl chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.1g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 25mL 5% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Embodiment 5
Step 1: 1g Pyrusussuriensis acyl chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.4g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 25mL 5% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Embodiment 6
Step 1: 3g Pyrusussuriensis acyl chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.4g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 25mL 2% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Embodiment 7
Step 1: 2g p-benzoyl chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.2g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 15mL 5% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Embodiment 8
Step 1: 4g p-benzoyl chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.4g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 2h under the room temperature; To the sodium tripolyphosphate that wherein drips 25mL 5% (mass fraction) (TPP) aqueous solution, it is crosslinked that carboxymethyl chitosan is taken place then.Dropwise, continue to stir 2h.Deposition, the rinsing repeatedly of petroleum ether, isopropyl alcohol promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
Above content is to combine concrete preferred implementation to the further explain that the present invention did, and can not assert that practical implementation of the present invention is confined to these explanations.For the those of ordinary skill of technical field under the present invention, under the prerequisite that does not break away from the present invention's design, can also make some simple deduction or replace, all should be regarded as belonging to protection scope of the present invention.
Claims (3)
1. an absorbable year bone induced the polypeptide microsphere, it is characterized in that: comprise water-soluble chitosan carrier and BMP-2 active polypeptide, the weight ratio of water-soluble chitosan carrier and BMP-2 active polypeptide is 0.1～1000:1.
2. an absorbable according to claim 1 year bone is induced the polypeptide microsphere, it is characterized in that: described water-soluble chitosan is a kind of of carboxymethyl chitosan, Pyrusussuriensis acyl chitosan, p-benzoyl chitosan etc.
3. an absorbable year bone induced the preparation technology of polypeptide microsphere, and it is characterized in that: the present invention adopts following technical scheme:
Step 1: 0.1g～10g water-soluble chitosan is dissolved in the 20ml water to dissolving fully obtains solution A;
Step 2: 0.01g～5g BMP-2 active polypeptide is added in the solution A, and mechanical agitation to mix homogeneously obtains mixed solution B;
Step 3: 5ml ~ 20ml mixed solution B adding is contained among the liquid paraffin solution 100mL of 2% (mass fraction) emolsifier span-80 (Span80); Mechanical agitation 1 ~ 10h under the room temperature; To sodium tripolyphosphate (TPP) aqueous solution of Dropwise 5 ml ~ 50mL 5% (mass fraction) wherein, it is crosslinked that carboxymethyl chitosan is taken place, and dropwises then; Continue to stir 1 ~ 10h; Deposition with the rinsing repeatedly of petroleum ether, isopropyl alcohol, promptly obtains the required year bone that absorbs with precipitate lyophilization to constant weight at last and induces polypeptide (BMP-2 active polypeptide) microsphere.
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| CN2011100215368A CN102600510A (en) | 2011-01-19 | 2011-01-19 | Absorbable bone induction polypeptide-carried microsphere and preparation method thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104056304A (en) * | 2014-07-02 | 2014-09-24 | 昆明医科大学第一附属医院 | Growth-factor-chitosan-microsphere loaded DBM support joint cartilage repairing material |
| CN109700783A (en) * | 2019-03-04 | 2019-05-03 | 江西派尼生物药业有限公司 | A kind of preparation method of chitosan coating tilmicosin microspheres |
-
2011
- 2011-01-19 CN CN2011100215368A patent/CN102600510A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104056304A (en) * | 2014-07-02 | 2014-09-24 | 昆明医科大学第一附属医院 | Growth-factor-chitosan-microsphere loaded DBM support joint cartilage repairing material |
| CN109700783A (en) * | 2019-03-04 | 2019-05-03 | 江西派尼生物药业有限公司 | A kind of preparation method of chitosan coating tilmicosin microspheres |
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Application publication date: 20120725 |