CN103768038B - Fibroin albumen one pack system microcapsule, fibroin albumen-nanometer gold hybridized microcapsule and preparation method thereof - Google Patents
Fibroin albumen one pack system microcapsule, fibroin albumen-nanometer gold hybridized microcapsule and preparation method thereof Download PDFInfo
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Abstract
Disclosure of the invention fibroin albumen one pack system microcapsule, fibroin albumen-nanometer gold hybridized microcapsule and preparation method thereof, the preparation process of fibroin albumen one pack system microcapsule is: fibroin albumen, Poly(D,L-lactide-co-glycolide microgranule and carbonate buffer solution are mixed to get mixed liquor; Drip the poor solvent mixing of albumen, protein-crosslinking, adds acetone and N-Methyl pyrrolidone mixed liquor for removing Poly(D,L-lactide-co-glycolide, and washing obtains fibroin albumen one pack system microcapsule suspension; With the nanometer gold of DNA functionalization, modification is carried out to fibroin albumen one pack system microcapsule again and obtain fibroin albumen-nanometer gold hybridized microcapsule, it is a kind of hollow microcapsule, both can load biomacromolecule class medicine, decorated by nano-gold gives its long-range release regulation and control and Raman strengthens characteristic simultaneously.Preparation technology is simple, favorable reproducibility, and universality is strong, be suitable for large-scale production, and material therefor has good biocompatibility, degradable, non-immunogenicity, cheap and easy to get.
Description
Technical field
The invention belongs to pharmaceutical carrier field, relate to a kind of fibroin albumen one pack system microcapsule and preparation method thereof particularly, a kind of fibroin albumen-nanometer gold hybridized microcapsule and preparation method thereof
Background technology
In drug delivery system (drug delivery systems), the organic or inorganic carrier of hollow structure receives extensive concern due to its special encapsulating, protection and transmission characteristic.At present, common hollow micro-/ nano particle mainly comprises polyelectrolyte microcapsule, polymeric hollow micro-/ nano ball, liposome, carbon pipe and silica-based microgranule etc., and its preparation method is also different.Wherein, LBL self-assembly (Layer-by-layer, LBL) technology due to advantages such as it are simple and easy to do, the suitability is strong hollow microcapsule preparation in attention.But LBL method also has himself deficiency, the potential source biomolecule toxicity that the polyelectrolyte as lotus positive electricity is introduced when preparing Medical hollow microcapsule; Inorganic template is removed and severe corrosive acid often must be used as Fluohydric acid., hydrochloric acid etc.; Non-degradable polymer remains as A Polystyrene Spheres Template.Based on above-mentioned deficiency, 2011, Tsukruk group utilized albuminous degeneration principle; prepare one-component LBL fibroin albumen microcapsule; by controlling the number of plies regulation and control microcapsule permeability of fibroin albumen deposition, avoid bio-toxicity (Shchepelina, O. that lotus positive electrical polyelectrolyte brings; Drachuk, I.; Gupta, M.K.; Lin, J.; Tsukruk, V.V., Adv.Mater.2011,23,4655-4660.).And remove condition based on the template that the LBL microcapsule of polylactic acid (PLA) or Poly(D,L-lactide-co-glycolide (PLGA) microsphere template overcomes mineral acid harshness undoubtedly; simultaneously because the biocompatibility that PLA and PLGA is good also solves template residue problem (Shenoy, D.B.; Antipov, A.A.; Sukhorukov G.B.;
h.Biomacromolecules2003,4,265-272.).But, when LBL method still cannot overcome its operating cost and the difficult problem that reduces with the increase of the shell number of plies of output, greatly limit traditional LBL method practical application in industry and be worth (Richardson, J.J.; Liang, K.; Kempe, K.; Ejima, H.; Cui, J.; Caruso, F.Adv.Mater.2013,25,6874-6878.Zhu, Y.; Tong, W.; Gao, C.; Mohwald, H.J.Mater.Chem.2008,18,1153-1158).Thus, one-step synthesis method polyelectrolyte microcapsule obtains desk study, as with mesopore silicon oxide or MnCO
3microgranule is templated synthesis polymer or native protein one pack system microcapsule, and has regulated and controled the permeability of microcapsule by condition optimizing.（Wang,Y.;Bansal,V.;Zelikin,A.N.;Caruso,F.,Nano Lett.2008,8,1741-1745.Zhu,Y.;Tong,W.;Gao,C.;Mohwald,H.,J.Mater.Chem.2008,18,1153-1158.）。But microcapsule class nano-device that gentle for method easy universality, the template biological compatibility, microcapsule permeability and multifunctionality are integrated in one and preparation method thereof have not been reported.
PLGA is a kind of medical macromolecular materials with good biocompatibility and biological degradability, can be used as apparatus and the pharmaceutic adjuvants such as medical operation suture thread, microcapsule, microsphere through U.S. FDA approval.PLGA in vivo intermediate product is lactic acid, and be the normal metabolite of human organism, can not assemble at vitals, final metabolite is CO
2and H
2o.The template prepared using PLGA microgranule as microcapsule, not only can solve template residue problem, and removes the process also relative ease gentleness of PLGA template.Fibroin albumen has remarkable mechanical strength, good biocompatibility and machinability and not easily causes inflammatory reaction, is the ideal material of pharmaceutical carrier.Gold is the material of extremely inertia, and golden nanometer particle stable in physicochemical property, environmental friendliness, good biocompatibility, median lethal dose(LD 50) are high, and has excellent optical property, by physical absorption or-SH ,-S-SH ,-NH
2fix various biomolecules Deng covalent bonding Absorbable organic halogens, as nucleic acid, antibody and medicine etc., and keep its biological activity.
Therefore, with PLGA microgranule for template, utilize desolventizing method one-step synthesis fibroin albumen one-component microcapsule, and then utilize the nanometer gold of DNA functionalization to carry out microcapsule finishing, while its permeability of regulation and control, give the special optical character of microcapsule inorganic nano-particle introducing and the various biological function of DNA, the function making such hybridized microcapsule have drug delivery and diagnosis concurrently have not been reported.
Summary of the invention
The present invention seeks to overcome the deficiencies in the prior art, a kind of fibroin albumen one pack system microcapsule is provided.
Second object of the present invention is to provide a kind of preparation method of fibroin albumen one pack system microcapsule.
3rd object of the present invention is to provide a kind of fibroin albumen-nanometer gold hybridized microcapsule.
4th object of the present invention is to provide the preparation method of a kind of fibroin albumen-nanometer gold hybridized microcapsule.
5th object of the present invention is to provide the purposes of a kind of fibroin albumen-nanometer gold hybridized microcapsule.
Technical scheme of the present invention is summarized as follows:
A preparation method for fibroin albumen one pack system microcapsule, the step comprised:
Be that the Poly(D,L-lactide-co-glycolide microgranule of 5000-50000 and the carbonate buffer solution of pH=9-11 are mixed to get mixed liquor by fibroin albumen, molecular weight, make the concentration of fibroin albumen in mixed liquor be 2-5mg/mL, make the concentration of described Poly(D,L-lactide-co-glycolide microgranule be 1 × 10
8-3 × 10
8individual/mL; In mixed liquor, drip the poor solvent mixing of albumen, the volume ratio of the poor solvent of described mixed liquor and albumen is 1:6-10; Centrifugal, removing supernatant, obtains the first microgranule, adds glutaraldehyde water solution and carry out protein-crosslinking in the first microgranule, centrifugal, and washing, obtains the second microgranule; In the second microgranule after washing, add the acetone that volume ratio is 1:1 and the mixing of N-Methyl pyrrolidone mixed liquor, for removing Poly(D,L-lactide-co-glycolide, washing obtains fibroin albumen one pack system microcapsule suspension.
The poor solvent particular methanol of albumen or ethanol.
Fibroin albumen one pack system microcapsule prepared by said method.
A preparation method for fibroin albumen-nanometer gold hybridized microcapsule, comprises the steps:
1) be that the Poly(D,L-lactide-co-glycolide microgranule of 5000-50000 and the carbonate buffer solution of pH=9-11 are mixed to get mixed liquor by fibroin albumen, molecular weight, make the concentration of fibroin albumen in mixed liquor be 2-5mg/mL, make the concentration of described Poly(D,L-lactide-co-glycolide microgranule be 1 × 10
8-3 × 10
8individual/mL; In mixed liquor, drip the poor solvent mixing of albumen, the volume ratio of the poor solvent of described mixed liquor and albumen is 1:6-10; Centrifugal, removing supernatant, obtains the first microgranule, adds glutaraldehyde water solution and carry out protein-crosslinking in the first microgranule, centrifugal, and washing, obtains the second microgranule; In the second microgranule after washing, add the acetone that volume ratio is 1:1 and the mixing of N-Methyl pyrrolidone mixed liquor, for removing Poly(D,L-lactide-co-glycolide, washing obtains fibroin albumen one pack system microcapsule suspension;
2) be the ratio of 1:10000-100000 in the mol ratio of the golden nanometer particle of fibroin albumen one pack system microcapsule and DNA functionalization, fibroin albumen one pack system microcapsule suspension is mixed homogeneously with the gold nanoparticle colloid solution of DNA functionalization, hatch, obtain fibroin albumen-nanometer gold hybridized microcapsule, described DNA is the sequence that end is modified with 5-100 base of a sulfydryl;
The poor solvent of albumen is preferably methanol or ethanol.
A kind of fibroin albumen-nanometer gold hybridized microcapsule prepared by said method.
Fibroin albumen-nanometer gold hybridized microcapsule is preparing the application in fibroin albumen-nanometer gold medicine carrying hybridized microcapsule, and medicine is preferably albumen or polysaccharide.
A kind of fibroin albumen-nanometer gold hybridized microcapsule is preparing the application in Raman image contrast agent.
Fibroin albumen of the present invention-nanometer gold hybridized microcapsule is a kind of hollow microcapsule, both can load biomacromolecule class medicine, and decorated by nano-gold gives its long-range release regulation and control and Raman strengthens characteristic simultaneously.Fibroin albumen one pack system microcapsule and fibroin albumen-nanometer gold hybridized microcapsule preparation technology are simple, favorable reproducibility, and universality is strong, be suitable for large-scale production, and material therefor has good biocompatibility, degradable, non-immunogenicity, cheap and easy to get.
Accompanying drawing explanation
Figure 1A, B and C is respectively the obtained PLGA microgranule of fibroin albumen bag quilt of PLGA microgranule (molecular weight 50,000) prepared by embodiment 2, embodiment 5 and the stereoscan photograph of fibroin albumen one pack system microcapsule, and D is the transmission electron microscope photo of fibroin albumen one pack system microcapsule.
Fig. 2 is that embodiment 2 prepares PLGA microgranule (molecular weight 1.5 ten thousand).
Fig. 3 A, B are respectively the obtained PLGA microgranule of fibroin albumen bag quilt of embodiment 6 and the stereoscan photograph of fibroin albumen one pack system microcapsule.
The transmission electron microscope photo of the fibroin albumen one pack system microcapsule that Fig. 4 DNA functionalization gold nanosphere (A), gold nanorods (C) and gold nanorods and ball (E) are modified; B, D, F are the stereoscan photograph that A, C, E are corresponding.
The surface-enhanced Raman collection of illustrative plates of Fig. 5 hybridized microcapsule.Solid line is focus on the Raman signal that microcapsule gathers, and dotted line is focus on the Raman signal gathered in microcapsule surrounding medium.
Fig. 6 load FITC-BSA(A) and hybridized microcapsule laser co-focusing photo FITC-dextran250kDa(B).
Detailed description of the invention
Below in conjunction with specific embodiment, the present invention is further illustrated, and embodiments of the invention understand the present invention better to enable those skilled in the art to, but do not impose any restrictions the present invention.
The preparation of embodiment 1 silk fibroin water solution:
Fibroin powder is purchased from: Huzhou Xintiansi Biotechnology Co. Ltd., gets 1g fibroin powder, is placed in 5mL LiBr(9.3M) solution, 60 DEG C of slow magnetic agitation 4h, produce the solution of 20%, gained solution is proceeded to bag filter (MWCO3500), the every 4h of distill water dialysis 3-4d(changes water once), except desalting, the centrifugal 20min of gained solution 8,000rpm, repeats 3 times, removing insoluble matter, then gained solution is crossed 0.45 μm of filter membrane.The silk fibroin water solution concentration finally obtained is about 6%, and concentration determination is by Weight computation gained after the lyophilizing of mensuration certain volume silk fibroin water solution.Storage liquid deposit in 4 DEG C for subsequent use.
The preparation of embodiment 2PLGA microsphere:
Classical emulsion-solvent evaporation method is adopted to prepare PLGA microsphere:
1) by the PLGA(50:50 of 150mg, molecular weight 50,000) be dissolved in 10mL dichloromethane, dropwise instill to 100mL concentration be in the polyvinyl alcohol of 1%, the homogeneous formation emulsion of high speed, be transferred on magnetic stirring apparatus and continue to stir 18h, dichloromethane is made to volatilize completely, the centrifugal 15min of 10,000rpm, add water ultrasonic heavy dispersion, repeat this process three times, lyophilizing, low temperature storage (Figure 1A).
2) by the PLGA(50:50 of 150mg, molecular weight 1.5 ten thousand) be dissolved in 10mL dichloromethane, dropwise instill to 100mL concentration be in the polyvinyl alcohol of 1%, the homogeneous formation emulsion of high speed, be transferred on magnetic stirring apparatus and continue to stir 18h, with the dichloromethane that volatilizees, the centrifugal 15min of 10,000rpm, add water ultrasonic heavy dispersion, repeat this process three times, lyophilizing, low temperature storage (Fig. 2).
The preparation of the gold nanosphere of embodiment 3DNA functionalization:
Classical trisodium citrate reduction method is adopted to prepare ball shaped nano gold grain (AuNSs), by the mass concentration of 20mL be 0.01% aqueous solution of chloraurate add in round-bottomed flask, backflow is heated with stirring in oil bath pan, add the trisodium citrate aqueous solution that 1mL mass concentration is 1%, continue to stir 15min, be cooled to room temperature, obtain the colloid solution of gold nanosphere.
In gold nanosphere and sulfhydrylation DNA(sulfhydrylation oligonucleotide 5'HS-C6-TTTTT) mol ratio be the ratio of 1:10000, by the aqueous solution 40h of the colloid solution of gold nanosphere and sulfhydrylation DNA, centrifugal, add water in precipitation, obtain the gold nanosphere colloid solution of DNA functionalization.
The preparation of the gold nanorods of embodiment 4DNA functionalization:
1) classical seed mediated growth method synthesis gold nanorods (AuNRs) is adopted.Experimental procedure is as follows:
A) gold seeds solution is prepared:
Be the HAuCl of 0.01mol/L by 0.25mL concentration
4cetyl trimethyl ammonium bromide (CTAB) the aqueous solution Homogeneous phase mixing of aqueous solution and 7.5mL concentration 0.1mol/L obtains mixed liquor, by the NaBH of the concentration 0.01mol/L of fresh for 0.6mL preparation
4aqueous solution adds in mixed liquor under magnetic agitation condition, after continuing to stir 2min, leaves standstill 2h, obtains gold seeds solution;
B) gold nanorods growth-promoting media is prepared:
Get a beaker, adding 9.5mL concentration is successively 0.1mol/L CTAB aqueous solution, and 0.4mL concentration is the HAuCl of 0.01mol/L
4aqueous solution, the concentration of the fresh preparation of 0.06mL is the AgNO of 0.01mol/L
3aqueous solution, fully mixes; The concentration adding the fresh preparation of 0.064mL is the aqueous ascorbic acid stirring of 0.1mol/L, until solution transfers water white transparency to from faint yellow, obtains gold nanorods growth-promoting media;
C) gold nanorods is prepared:
Gold seeds solution 0.04mL step a) obtained adds in the obtained gold nanorods growth-promoting media of step b), to bring out the growth of gold nanorods, slowly stirs until solution colour transfers lavender to, leaves standstill 9h, keep constant temperature 28 DEG C gold nanorods colloid solution;
3) in gold nanorods and sulfhydrylation DNA(sulfhydrylation oligonucleotide 5'HS-C6-TTTTT) mol ratio be the ratio of 1:1000, by the aqueous solution 40h of the colloid solution of gold nanorods and sulfhydrylation DNA, centrifugal, the liquid of removing containing unconjugated sulfhydrylation DNA, is precipitated as the gold nanorods (AuNRs-T5) of DNA functionalization;
Experiment shows, the DNA sequence that end is modified with 5-100 base of a sulfydryl is all applicable to the present invention.
Embodiment 5
A preparation method for fibroin albumen one pack system microcapsule, the step comprised:
Be 2.4 μm by fibroin albumen, particle diameter, molecular weight is that the sodium carbonate-bicarbonate buffer of 50000 Poly(D,L-lactide-co-glycolide microgranules (Figure 1A) and pH=9 is mixed to get mixed liquor, make the concentration of fibroin albumen in mixed liquor be 2mg/mL, make the concentration of Poly(D,L-lactide-co-glycolide microgranule be 1 × 10
8individual/mL; In mixed liquor, drip dehydrated alcohol mixing, the volume ratio of mixed liquor and dehydrated alcohol is 1:6; Centrifugal, removing supernatant, obtains the first microgranule, the mass concentration adding 1 times of the volume of the supernatant just removed in the first microgranule be the glutaraldehyde water solution of 2% for crosslinking protein, stir 1h, centrifugal, washing, obtains the second microgranule (Figure 1B); In the second microgranule after washing, add the acetone and N-Methyl pyrrolidone mixed liquor mixing 16h that volume ratio is 1:1 for removing Poly(D,L-lactide-co-glycolide, washing obtains fibroin albumen one pack system microcapsule suspension.
Fig. 1 C, D be electron microscope observation to the fibroin albumen one pack system microcapsule will obtained after the drying of fibroin albumen one pack system microcapsule suspension.
This experiment utilizes hydrophobic interaction (Wang, the X. of fibroin albumen and PLGA; Wenk, E.; Hu, X.; Castro, G.R.; Meinel, L.; Wang, X.; Li, C.; Merkle, H.; Kaplan, D.L., Biomaterials2007,28 (28), 4161-4169) fibroin albumen of precipitation is deposited on PLGA microsphere surface, then cross-linking solidification, make it no longer depolymerization, dissolve template and finally form fibroin albumen one pack system microcapsule, therefore the PLGA microsphere of other yardsticks such as: any microgranule being 5000-50000 at molecular weight equally can as the fibroin albumen one pack system microcapsule of Template preparation different size.
Embodiment 6
A preparation method for fibroin albumen one pack system microcapsule, the step comprised:
Be 2.4 μm by fibroin albumen, particle diameter, molecular weight is that the sodium carbonate-bicarbonate buffer of 50000 Poly(D,L-lactide-co-glycolide microgranules (Figure 1A) and pH=11 is mixed to get mixed liquor, make the concentration of fibroin albumen in mixed liquor be 5mg/mL, make the concentration of Poly(D,L-lactide-co-glycolide microgranule be 3 × 10
8individual/mL; In mixed liquor, drip dehydrated alcohol mixing, the volume ratio of mixed liquor and dehydrated alcohol is 1:10; Centrifugal, removing supernatant, obtain the first microgranule, the mass concentration adding the same volume of the supernatant just removed in the first microgranule is the glutaraldehyde water solution of 2%, stirs 1h, centrifugal, and washing, obtains the second microgranule (Fig. 3 A); The acetone that volume ratio is 1:1 and N-Methyl pyrrolidone mixed liquor mixing 16h is added in the second microgranule, for removing Poly(D,L-lactide-co-glycolide, washing obtains fibroin albumen one pack system microcapsule suspension, drips after silicon chip drying, sem observation (Fig. 3 B).
Experiment proves, the poor solvent of albumen selects methanol or isopropyl alcohol also to may be used for the present invention.
Embodiment 7
A preparation method for fibroin albumen-gold nanosphere hybridized microcapsule, the step comprised:
1) preparation of fibroin albumen one pack system microcapsule suspension is with embodiment 5;
2) mol ratio of the gold nanosphere of the DNA functionalization prepared in fibroin albumen one pack system microcapsule and embodiment 3 is the ratio of 1:60000, fibroin albumen one pack system microcapsule suspension is mixed homogeneously with the gold nanosphere colloid solution of DNA functionalization, hatch 4h, obtain fibroin albumen-gold nanosphere hybridized microcapsule (Fig. 4 A, B).
PLGA microsphere only affects the size of fibroin albumen microcapsule as template, and does not change the physicochemical properties of microcapsule.Therefore, the fibroin albumen one pack system microcapsule of other yardsticks also produces corresponding fibroin albumen-gold nanosphere hybridized microcapsule by this process.
Embodiment 8
A preparation method for fibroin albumen-gold nanorods hybridized microcapsule, the step comprised:
1) preparation of fibroin albumen one pack system microcapsule is with embodiment 5;
2) mol ratio of the gold nanorods of the DNA functionalization prepared in fibroin albumen one pack system microcapsule and embodiment 4 is the ratio of 1:60000, fibroin albumen one pack system microcapsule suspension is mixed homogeneously with the gold nanorods colloid solution of DNA functionalization, hatch 4h, obtain fibroin albumen-gold nanorods hybridized microcapsule (Fig. 4 C, D).
The mol ratio of the gold nanorods of fibroin albumen one pack system microcapsule and DNA functionalization can also be selected such as in the scope of 1:10000-100000: 1:10000,1:20000,1:50000,1:80000,1:100000 etc., its objective is and allow the gold nanorods of DNA functionalization carry out shutoff to fibroin albumen one pack system microcapsule, to regulate its permeability, so that prepare fibroin albumen-nanometer gold medicine carrying hybridized microcapsule.
One pack system microcapsule is after gold nanorods modification obtains hybridized microcapsule, zeta-current potential becomes-4.22 ± 0.62mV from-9.44 ± 2.67mV and shows between gold nanorods and fibroin by electrostatic interaction and hydrogen bonded, with work before this and consistent (the Zhimin Zhou of other researcher work, A.C.Anselmo and S.Mitragotri, Adv.Mater., 2013, 25, 2723-2727, V.Kozlovskaya, E.Kharlampieva, I.Drachuk, D.Cheng and V.V.Tsukruk, Soft Matter2010, 6, 3596-9608.), PLGA microsphere only affects the size of fibroin albumen microcapsule as template, and do not change the physicochemical properties of microcapsule.Therefore, the fibroin albumen one pack system microcapsule of other yardsticks also produces corresponding fibroin albumen-gold nanorods hybridized microcapsule by this process.
Embodiment 9
A kind of fibroin albumen-nanometer gold hybridized microcapsule is preparing the application in Raman image contrast agent
Fibroin albumen embodiment 8 prepared-gold nanorods hybridized microcapsule makes the suspension containing 5 μMs of crystal violets, excitation wavelength 633nm, gathers Raman signal (Fig. 5).
Embodiment 10
A kind of fibroin albumen-nanometer gold hybridized microcapsule is preparing the application in fibroin albumen-nanometer gold medicine carrying (albumen medicine) hybridized microcapsule
1) preparation of fibroin albumen one pack system microcapsule is with embodiment 5;
2) preparation of the gold nanorods of DNA functionalization is with embodiment 4;
3) be that the bovine serum albumin (FITC-BSA) of FITC labelling of 2mg/mL is mixed to get mixed liquor by fibroin albumen one pack system microcapsule and mass concentration, after 30min, mixed liquor is mixed in the ratio that fibroin albumen one pack system microcapsule and the mol ratio of the gold nanorods of DNA functionalization are 1:60000 with the gold nanorods of DNA functionalization, hatch 4h, centrifugal washing obtains wrapping the fibroin albumen-nanometer gold medicine carrying hybridized microcapsule (Fig. 6 A) carrying model drug.
Due to known in this field, model drug bovine serum albumin is the representative of albumen, and bovine serum albumin can wrap and be loaded in fibroin albumen-gold nanorods hybridized microcapsule, and therefore, the medicine of other albumen also can wrap and be loaded in fibroin albumen-gold nanorods hybridized microcapsule.Embodiment 11
A kind of fibroin albumen-nanometer gold hybridized microcapsule is preparing the application in fibroin albumen-nanometer gold medicine carrying (polysaccharide medicine) hybridized microcapsule
1) preparation of fibroin albumen one pack system microcapsule is with embodiment 5;
2) preparation of the gold nanorods of DNA functionalization is with embodiment 4;
3) mass concentration being 250kDa by fibroin albumen one pack system microcapsule and molecular weight is that the glucosan (FITC-dextran) of the FITC labelling of 2mg/mL is mixed to get mixed liquor, after 30min, mixed liquor is mixed in the ratio that fibroin albumen one pack system microcapsule and the mol ratio of the gold nanorods of DNA functionalization are 1:60000 with the gold nanorods of DNA functionalization, hatch 4h, centrifugal washing obtains wrapping the fibroin albumen-nanometer gold medicine carrying hybridized microcapsule (Fig. 6 B) carrying model drug.
Due to known in this field, model drug glucosan is the representative of polysaccharide, and glucosan can wrap and be loaded in fibroin albumen-gold nanorods hybridized microcapsule, and therefore, other polysaccharide also can wrap and be loaded in fibroin albumen-gold nanorods hybridized microcapsule.
Embodiment 12
A preparation method for fibroin albumen-gold nanorods and ball hybridized microcapsule, the step comprised:
1) preparation of fibroin albumen one pack system microcapsule is with embodiment 5;
2) preparation of the gold nanosphere of DNA functionalization is with embodiment 3;
3) preparation of the gold nanorods of DNA functionalization is with embodiment 4;
4) be the ratio of 1:60000 in the mol ratio of the gold nanorods of fibroin albumen one pack system microcapsule and DNA functionalization, fibroin albumen one pack system microcapsule suspension is mixed homogeneously with the gold nanorods colloid solution of DNA functionalization, hatch 4h, mixed liquor is the ratio of 1:60000 with the gold nanosphere of DNA functionalization in the mol ratio of the gold nanosphere of fibroin albumen one pack system microcapsule and DNA functionalization again, mixing, hatch 4h, obtain fibroin albumen-gold nanorods and ball hybridized microcapsule (Fig. 4 E, F).
PLGA microsphere only affects the size of fibroin albumen microcapsule as template, and does not change the physicochemical properties of microcapsule.Therefore, the fibroin albumen one pack system microcapsule of other yardsticks also produces corresponding fibroin albumen-gold nanorods and ball hybridized microcapsule by this process.
Claims (8)
1. a preparation method for fibroin albumen one pack system microcapsule, is characterized in that comprising step:
Be that the Poly(D,L-lactide-co-glycolide microgranule of 5000-50000 and the carbonate buffer solution of pH=9-11 are mixed to get mixed liquor by fibroin albumen, molecular weight, make the concentration of fibroin albumen in mixed liquor be 2-5mg/mL, make the concentration of described Poly(D,L-lactide-co-glycolide microgranule be 1 × 10
8-3 × 10
8individual/mL; In mixed liquor, drip the poor solvent mixing of albumen, the volume ratio of the poor solvent of described mixed liquor and albumen is 1:6-10; Centrifugal, removing supernatant, obtains the first microgranule, adds glutaraldehyde water solution and carry out protein-crosslinking in the first microgranule, centrifugal, and washing, obtains the second microgranule; In the second microgranule after washing, add the acetone that volume ratio is 1:1 and the mixing of N-Methyl pyrrolidone mixed liquor, for removing Poly(D,L-lactide-co-glycolide, washing obtains fibroin albumen one pack system microcapsule suspension.
2. the preparation method of a kind of fibroin albumen one pack system microcapsule according to claim 1, is characterized in that the poor solvent of described albumen is methanol or ethanol.
3. the fibroin albumen one pack system microcapsule prepared of the method for claim 1 or 2.
4. a preparation method for fibroin albumen-nanometer gold hybridized microcapsule, is characterized in that comprising the steps:
1) be that the Poly(D,L-lactide-co-glycolide microgranule of 5000-50000 and the carbonate buffer solution of pH=9-11 are mixed to get mixed liquor by fibroin albumen, molecular weight, make the concentration of fibroin albumen in mixed liquor be 2-5mg/mL, make the concentration of described Poly(D,L-lactide-co-glycolide microgranule be 1 × 10
8-3 × 10
8individual/mL; In mixed liquor, drip the poor solvent mixing of albumen, the volume ratio of the poor solvent of described mixed liquor and albumen is 1:6-10; Centrifugal, removing supernatant, obtains the first microgranule, adds glutaraldehyde water solution and carry out protein-crosslinking in the first microgranule, centrifugal, and washing, obtains the second microgranule; In the second microgranule after washing, add the acetone that volume ratio is 1:1 and the mixing of N-Methyl pyrrolidone mixed liquor, for removing Poly(D,L-lactide-co-glycolide, washing obtains fibroin albumen one pack system microcapsule suspension;
2) be the ratio of 1:10000-100000 in the mol ratio of the golden nanometer particle of fibroin albumen one pack system microcapsule and DNA functionalization, fibroin albumen one pack system microcapsule suspension is mixed homogeneously with the gold nanoparticle colloid solution of DNA functionalization, hatch, obtain fibroin albumen-nanometer gold hybridized microcapsule, described DNA is the sequence that end is modified with 5-100 base of a sulfydryl.
5. the preparation method of a kind of fibroin albumen-nanometer gold hybridized microcapsule according to claim 4, is characterized in that the poor solvent of described albumen is methanol or ethanol.
6. a kind of fibroin albumen-nanometer gold hybridized microcapsule of preparing of the method for claim 4 or 5.
7. a kind of fibroin albumen-nanometer gold hybridized microcapsule of claim 6 is preparing the application in fibroin albumen-nanometer gold medicine carrying hybridized microcapsule, and described medicine is albumen or polysaccharide.
8. a kind of fibroin albumen-nanometer gold hybridized microcapsule of claim 6 is preparing the application in Raman image contrast agent.
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| CN106421800A (en) * | 2016-09-28 | 2017-02-22 | 天津医科大学口腔医院 | Silk fibroin modified depression structure lactic acid-based polymer drug-carrying microsphere and method for preparing same |
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| Fabrication of bovine serum albumin microcapsules by desolvation and destroyable cross-linking;Yi Zhu et.al;《Journal of Materials Chemistry》;20080129;第18卷;1153-1158 * |
| Silk Coatings on PLGA and Alginate Microspheres for Protein Delivery;Xiaoqin Wang et.al;《Biomaterials》;20071030;第28卷(第28期);4161-4169 * |
| Silk-on-Silk Layer-by-Layer Microcapsules;Olga Shchepelina et.al;《Adv. Mater》;20111231;第23卷;4655-4660 * |
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