CN109701029A - Nanocrystal self assembly aggregation of protein mediation and preparation method thereof - Google Patents
Nanocrystal self assembly aggregation of protein mediation and preparation method thereof Download PDFInfo
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Abstract
The invention discloses the nanocrystal self assembly aggregations of protein mediation, belong to technical field of nano materials in biomedicine, the nanocrystal self assembly aggregation is that oil-soluble nano particles are encapsulated in formation nanocrystal self assembly aggregation in carrier material using protein as carrier material.The invention also discloses preparation methods, are that oil solubility nanometer crystal is wrapped in the nano-spherical structure that protein is self-assembled by main reaction mechanism using protein as carrier with the hydrophobic interaction between protein and oil solubility nanometer crystal.The preparation method is easy to operate, is easily enlarged metaplasia production, obtained self assembly aggregation have many advantages, such as high nanocrystal loading density, nanocrystal utilization rate high (more than 90%), self assembly aggregation partial size can regulate and control in a wide range of, be easy to be surface modified, with preferable colloidal stability, high-biocompatibility and low bio-toxicity；Meanwhile multi-functional nanometer material can be very easily prepared using the method.
Description
Technical field
The invention belongs to technical field of nano materials in biomedicine, and in particular to the nanocrystal self assembly of protein mediation
Aggregation and preparation method thereof.
Background technique
In the past twenty years, people develop a variety of nanocrystals such as Superparamagnetic Iron Oxide nanoparticle, quantum
Point, gold nano grain, carbon nanotube etc..These nanocrystals have the function of fluorescence, superparamagnetism or plasma effect etc., this
Them are made to be widely used in field of biomedicine.Nanocrystal, which is assembled into bigger colloidal solid, to be helped to improve nanometer
The property of crystal, such as its dissolubility and stability in water, offer signal more stronger than single nanocrystal etc. are provided.More
Importantly, multifunctional nanoparticle can be constructed by assembling two or more with nanocrystal of different nature.And
And a large amount of result of study shows 100-300 nanometers of nano particle, is more suitably applied to drug delivery and tumor imaging system
System.
In order to realize the assembling of nanocrystal, researcher develops a variety of timbering materials, including micella, silicon meso-porous nano
Material etc., to construct nanocrystal self assembly aggregation to meet the needs of biosystem.However, with natural material such as albumen
Matter, nucleic acid are compared, these artificial synthesized materials have the shortcomings that toxicity height, poor biocompatibility, this manually to synthesize
Material be timbering material building nanocrystal self-assembly system cannot be really converted into drug or diagnostic reagent.Moreover,
In most cases, these nanocrystal self-assembly systems are there is also nanocrystal load efficiency is lower, self assembly aggregation
The disadvantages such as interior crystalline content is low.
In recent years, protein is as natural material nontoxic, with good biocompatibility, in some researchs
It is designed to nano-spherical structure for delivering hydrophobic small molecules drug.Drug is loaded into protein nano ball by researcher
In shape structure, to enhance drug targeting and reduce the toxicity of Chemotherapeutic Drugs On Normal tissue.And using protein as carrier most at
The example of function is exactly albumin mating type taxol (Abraxane), it is ratified by food and drug administration (FDA)
Treatment for kinds cancer.
In some researchs, protein is used for the treatment of tumour as the timbering material of certain hydrophilic nano grains, this
The load density that nanocrystal self assembly aggregation in a little researchs all has nanocrystal is low, product structure controls poor, size
The disadvantages of shape uniformity is poor affects the preparation of composite nano materials.
Summary of the invention
Goal of the invention: to solve problems of the prior art, an object of the present invention is to provide protein mediation
Nanocrystal self assembly aggregation, utilize the hydrophobic phase between the protein with hydrophobic domains and oil solubility nanometer crystal
Interaction successfully constructs that nanocrystal load efficiency is high, it is big to carry intracorporal nanocrystal density, the continuous controllable, bio-compatible of partial size
The good nanocrystal self-assembly system of property；Another object of the present invention is to provide preparation methods.
Technical solution: to achieve the above object, the present invention adopts the following technical scheme:
The nanocrystal self assembly aggregation of protein mediation, the nanocrystal self assembly aggregation are with protein for load
Oil-soluble nano particles are encapsulated in formation nanocrystal self assembly aggregation in carrier material by body material.Self assembly aggregation
Body is self-assembly of by protein and oil solubility nanometer crystal.The driving force of assembling is hydrophobicity effect, and protein mediation is received
The hydration kinetics partial size of meter Jing Ti self assembly aggregation is continuously adjusted within the scope of 20-500nm.
Further, the protein is selected from bovine serum albumin(BSA), Hen egg-white lysozyme, cow's milk albumin etc. with thin
The protein of aqueous structural domain.
Further, the oil-soluble nano particles are quantum dot, Superparamagnetic Iron Oxide nanoparticle, silver nanoparticle crystalline substance
The combination of one or more of body, gold nano-crystal.
The preparation method of the nanocrystal self assembly aggregation of the protein mediation, by the oil solubility nanometer grain
Sub (one or more) are dissolved in organic solvent simultaneously, and mix with protein solution to get having a variety of nanocrystals special simultaneously
The multifunctional nanocrystals self assembly aggregation of property.The oil solubility nanometer crystal self assembly aggregation of the protein mediation,
Its synthetic method is to mix protein solution with oil solubility nanometer crystal.
Further, the preparation method of the nanocrystal self assembly aggregation of the protein mediation, including walk as follows
It is rapid:
1) protein solution is dissolved in protein buffer liquid, is configured to protein solution；Protein buffer liquid be water or
Person's phosphate buffer；
2) in organic solvent by the dissolution of one or more of oil-soluble nano particles, it is configured to nanocrystal solution；
3) nanocrystal solution is mixed with protein solution, concussion shakes up；
4) above-mentioned mixed liquor is incubated at room temperature, the mixed liquor after being incubated for；
5) mixed liquor after above-mentioned incubation is centrifuged, removes supernatant, the nanocrystal of precipitating as protein mediation is certainly
Assemble aggregation.
Further, the concentration of the protein solution is 0.1-10mg/mL, and the concentration of the nanocrystal solution is
The concentration of 0.05-20mg/mL, nanocrystal are higher, and the partial size of gained nanocrystal self assembly aggregation is bigger.
Further, in step 4), the time of the incubation is three hours or more.
Inventive principle: the variation of inducible protein matter recurring structure makes protein when the nanocrystal and protein contacts of small size
Hydrophobic domains are exposed, the hydrophobic surface of the hydrophobic domains in protein and oil solubility nanometer crystal passes through hydrophobic forces
It is connected with each other, and then is self-assembled into protein-nanocrystal self assembly aggregation.
The utility model has the advantages that compared with prior art, the nanocrystal self assembly aggregation of protein mediation of the invention has
Higher crystalline density loads more than 50 nanocrystals in each nanocrystal aggregation, is readily synthesized multifunctional nano
Particle；Nanocrystal packaging efficiency with higher, the nanocrystal more than 90% are encapsulated in BSA self assembly particle；It should
Self assembly aggregation has good biocompatibility and low bio-toxicity, is easy to surface modification, nanocrystal self assembly aggregation
The surface of body is protein, exposes a large amount of amino and carboxyl, is easy to be coupled with functional moleculars such as antibody, polypeptides；The self assembly
Aggregate particle size is uniform, and has preferable colloidal stability, can save for a long time.
Preparation, easy to operate, being easily enlarged are assembled in the nanocrystal self assembly of protein mediation of the invention
Production, meanwhile, multi-functional nanometer material, the hydration kinetics of nanocrystal aggregation can be very easily prepared using the method
Partial size can regulate and control between 20nm-500nm.Therefore, the oil solubility nanometer crystal self assembly aggregation of this protein mediation
The fields of biomedicine such as imaging, detection, treatment can be widely applied to.
Detailed description of the invention
Fig. 1 is the Superparamagnetic Iron Oxide self-assembly aggregation that the bovine serum albumin(BSA) in embodiment 1 mediates
The transmission electron microscope photo (Transmission Electron Microscope, TEM) of (SPIONs@BSA)；
Fig. 2 is to control the Superparamagnetic Iron Oxide that bovine serum albumin(BSA) mediates by control crystal concentration in embodiment 1 to receive
The curve graph of rice corpuscles self assembly aggregation (SPIONs@BSA) partial size；
Fig. 3 is the transmission electricity for the quantum dot self assembly aggregation (QDs@BSA) that the bovine serum albumin(BSA) in embodiment 2 mediates
Sub- microscope photo (Transmission Electron Microscope, TEM)；
Fig. 4 is that the hydration for the quantum dot self assembly aggregation (QDs@BSA) that the bovine serum albumin(BSA) in embodiment 2 mediates is dynamic
Mechanics grading curve；
Fig. 5 is the hydration power for the quantum dot self assembly aggregation (QDs@BSA) that bovine serum albumin(BSA) mediates in embodiment 2
Learn the curve that partial size changes over time；
Fig. 6 is the multifunctional nanocrystals self assembly aggregation (SPIONs& that the bovine serum albumin(BSA) in embodiment 3 mediates
QDs@BSA) fluorescence and magnet attract photo；
Fig. 7 is multifunctional nanocrystals self assembly aggregation (the gQDs&rQDs@that bovine serum albumin(BSA) mediates in embodiment 4
BSA) the ratio control analysis result of interior different crystal；
Fig. 8 is the transmission for the quantum dot self assembly aggregation (QDs@CEWL) that the Hen egg-white lysozyme in embodiment 5 mediates
Electron micrograph (Transmission Electron Microscope, TEM)；
Fig. 9 is the high score for the quantum dot self assembly aggregation (QDs@CEWL) that the Hen egg-white lysozyme in embodiment 5 mediates
Distinguish transmission electron microscope photo (Transmission Electron Microscope, TEM).
Specific embodiment
In order to further illustrate the present invention, technical method provided by the invention is done further below in conjunction with example and attached drawing
Detailed description.It should be understood that these examples are only for illustrating the present invention and are not intended to limit the scope of the present invention.In addition, answering
Understand, after reading the content taught by the present invention, those skilled in the art can make various modifications or changes to the present invention,
These equivalent forms also fall within the scope of the appended claims of the present application.
The nanocrystal self assembly aggregation of protein mediation, the nanocrystal self assembly aggregation are with protein for load
Oil-soluble nano particles are encapsulated in formation nanocrystal self assembly aggregation in carrier material by body material.Self assembly aggregation
Body is self-assembly of by protein and oil solubility nanometer crystal.The driving force of assembling is hydrophobicity effect, and protein mediation is received
The hydration kinetics partial size of meter Jing Ti self assembly aggregation is continuously adjusted within the scope of 20-500nm.
Protein is selected from the egg with hydrophobic domain such as bovine serum albumin(BSA), Hen egg-white lysozyme, cow's milk albumin
White matter.Oil-soluble nano particles are quantum dot, in Superparamagnetic Iron Oxide nanoparticle, silver nanoparticle crystal, gold nano-crystal
One or more of combinations.
The preparation method of the nanocrystal self assembly aggregation of protein mediation, oil-soluble nano particles are (a kind of or several
Kind) it is dissolved in organic solvent simultaneously, and mix with protein solution to get having a variety of the multi-functional of nanocrystal characteristic to receive simultaneously
Meter Jing Ti self assembly aggregation.The oil solubility nanometer crystal self assembly aggregation of protein mediation, synthetic method are egg
White matter solution is mixed with oil solubility nanometer crystal.Include the following steps:
1) protein solution is dissolved in protein buffer liquid, is configured to protein solution；Protein buffer liquid be water or
Person's phosphate buffer；
2) in organic solvent by the dissolution of one or more of oil-soluble nano particles, it is configured to nanocrystal solution；
3) nanocrystal solution is mixed with protein solution, concussion shakes up；
4) above-mentioned mixed liquor is incubated at room temperature, the mixed liquor after being incubated for；
5) mixed liquor after above-mentioned incubation is centrifuged, removes supernatant, the nanocrystal of precipitating as protein mediation is certainly
Assemble aggregation.
The concentration of protein solution is 0.1-10mg/mL, and the concentration of nanocrystal solution is 0.05-20mg/mL, nanocrystal
Concentration it is higher, the partial size of gained nanocrystal self assembly aggregation is bigger.In step 4), time of incubation be three hours with
On.
Superparamagnetic Iron Oxide self-assembly aggregation (the SPIONs@that 1 bovine serum albumin(BSA) of embodiment mediates
BSA preparation and its size controlling)
Concrete operations are as follows:
1) the oil-soluble Superparamagnetic Iron Oxide nanoparticle that synthesis partial size is 6nm.
2) bovine serum albumin solution is dissolved in water, is configured to the protein solution that concentration is 2mg/mL.
3) by oil-soluble Superparamagnetic Iron Oxide nanoparticle dissolution in tetrahydrofuran, it is 0.2mg/mL that concentration, which is made,
The nanocrystal solution of 0.4mg/mL, 0.6mg/mL, 0.8mg/mL, 1mg/mL, 1.5mg/mL, 2mg/mL.
4) 800 μ L nanocrystal solutions are injected in 3mL protein solution, quickly concussion shakes up；
5) above-mentioned mixed liquor is incubated at room temperature three hours or more；
6) mixed liquor after above-mentioned incubation is centrifuged, removes supernatant, precipitating is gained.
Fig. 1 and Fig. 2 is the oil-soluble Superparamagnetic Iron Oxide nanoparticle that prepared bovine serum albumin(BSA) mediates respectively
The TEM figure of self assembly aggregation (SPIONs@BSA) and its hydration kinetics partial size are with Superparamagnetic Iron Oxide nanoparticle concentration
The curve of variation.As can be seen from Figure 1: prepared Superparamagnetic Iron Oxide self-assembly is at cluster structure, partial size
Uniform, nanocrystal density is high, there is 50 or more nanocrystals in the same cluster structure.Figure it is seen that oil-soluble
The concentration of nanoparticle is higher, and the partial size of gained nanocrystal self assembly aggregation is bigger.
The preparation for the quantum dot self assembly aggregation (QDs@BSA) that 2 bovine serum albumin(BSA) of embodiment mediates
Concrete operations are as follows:
1) oil-soluble quantum dot is synthesized.
2) bovine serum albumin solution is dissolved in phosphate buffer, it is molten is configured to the albumen that concentration is 2mg/mL
Liquid.
3) oil-soluble quantum dot is dissolved in tetrahydrofuran, the nanocrystal solution that concentration is 1mg/mL is made.
4) 100 μ L nanocrystal solutions are injected in 3mL protein solution, quickly concussion shakes up；
5) above-mentioned mixed liquor is incubated at room temperature three hours or more；
6) mixed liquor after above-mentioned incubation is centrifuged, removes supernatant, precipitating is gained.
Fig. 3 and Fig. 4 is the quantum dot self assembly aggregation (QDs@BSA) that prepared bovine serum albumin(BSA) mediates respectively
TEM figure and its hydration kinetics grain size curve.As can be seen from Figure 3: prepared quantum dot is self-assembled into cluster structure, nanometer
Crystalline density is high, there is 50 or more nanocrystals in the same cluster structure.From fig. 4, it can be seen that prepared quantum dot
Self assembly aggregate particle size is uniform.
Fig. 5 shows the quantum dot self assembly aggregation (QDs@BSA) of prepared bovine serum albumin(BSA) mediation in water
In colloidal stability.The quantum dot self assembly aggregation (QDs@BSA) that bovine serum albumin(BSA) mediates is dissolved in water, is placed
Under 4 degrees Celsius of environment, the partial size of QDs@BSA does not change within 1 month, shows good colloidal stability.
The multifunctional nanocrystals self assembly aggregation (SPIONs&QDs@BSA) that 3 bovine serum albumin(BSA) of embodiment mediates
Preparation
Concrete operations are as follows:
1) oil-soluble quantum dot is synthesized.
2) the oil-soluble Superparamagnetic Iron Oxide nanoparticle that synthesis partial size is 6nm.
3) bovine serum albumin solution is dissolved in phosphate buffer, it is molten is configured to the albumen that concentration is 2mg/mL
Liquid.
4) oil-soluble quantum dot and oil-soluble Superparamagnetic Iron Oxide nanoparticle are dissolved in tetrahydrofuran simultaneously, are made
The nanocrystal solution for being 1mg/mL at concentration, the mass ratio of two kinds of nanocrystals are 1:1.
5) 200 μ L nanocrystal solutions are injected in 3mL protein solution, quickly concussion shakes up；
6) above-mentioned mixed liquor is incubated at room temperature three hours or more；
By the mixed liquor centrifugation after above-mentioned incubation, precipitating is gained.
Fig. 6 is prepared multifunctional nanocrystals self assembly aggregation (SPIONs&QDs@BSA) under magnet attraction
Fluorescent characteristic, indicate prepared multifunctional nanocrystals self assembly aggregation (SPIONs&QDs@BSA) have simultaneously it is magnetic with
Fluorescent characteristic.
The multifunctional nanocrystals self assembly aggregation (gQDs&rQDs@BSA) that 4 bovine serum albumin(BSA) of embodiment mediates is poly-
Collect the ratio control of internal different crystal
Concrete operations are as follows:
1) the red and green oil-soluble quantum dot of synthesis；
2) bovine serum albumin solution is dissolved in phosphate buffer, it is molten is configured to the albumen that concentration is 2mg/mL
Liquid；
4) the oil-soluble quantum dot of two kinds of colors is dissolved in tetrahydrofuran simultaneously, the nanometer that concentration is 1mg/mL is made
Crystalloid solution, the mass ratio of two kinds of nanocrystals are 1:1,3:1,5:1；
5) 200 μ L nanocrystal solutions are injected into 3mL protein solution, quickly concussion shakes up；
6) above-mentioned mixed liquor is incubated at room temperature three hours or more；By the mixed liquor centrifugation after above-mentioned incubation.
Fig. 7 is the ratio of prepared multifunctional nanocrystals self assembly aggregation (gQDs&rQDs@BSA) interior different crystal
Example control analysis result.
The quantum dot that 5 Hen egg-white lysozyme of embodiment (Chicken egg white lysozyme, CEWL) mediates is from group
Fill the preparation of aggregation (QDs@CEWL)
Concrete operations are as follows:
1) oil-soluble quantum dot is synthesized.
2) Hen egg-white lysozyme is dissolved in phosphate buffer, is configured to the protein solution that concentration is 5mg/mL.
3) oil-soluble quantum dot is dissolved in tetrahydrofuran, the nanocrystal solution that concentration is 0.2mg/mL is made.
4) 50 μ L nanocrystal solutions are injected in 3mL protein solution, quickly concussion shakes up；
5) above-mentioned mixed liquor is incubated at room temperature three hours or more；
6) mixed liquor after above-mentioned incubation is centrifuged, removes supernatant, precipitating is gained.
Fig. 8 and Fig. 9 is the TEM for the quantum dot self assembly aggregation (QDs@CEWL) that prepared Hen egg-white lysozyme mediates
Figure.As it can be observed in the picture that quantum dot is self-assembled into irregular spheric cluster.As can be seen from Figure 9, the quantum dot in each cluster structure
Packaging density it is high, can see 10 or so quantum dots in a microscopical level, the partial size of nano particle is 30 to receive
Rice is to 40 nanometers.
Claims (8)
1. the nanocrystal self assembly aggregation of protein mediation, it is characterised in that: the nanocrystal self assembly aggregation be with
Protein is carrier material, and oil-soluble nano particles are encapsulated in formation nanocrystal self assembly aggregation in carrier material.
2. the nanocrystal self assembly aggregation of protein mediation according to claim 1, it is characterised in that: the egg
White matter is the protein with hydrophobic domain.
3. the nanocrystal self assembly aggregation of protein mediation according to claim 2, it is characterised in that: the tool
There is the protein of hydrophobic domain to be selected from bovine serum albumin(BSA), Hen egg-white lysozyme and cow's milk albumin.
4. the nanocrystal self assembly aggregation of protein mediation according to claim 1, it is characterised in that: the oil
Dissolubility nanoparticle is one of quantum dot, Superparamagnetic Iron Oxide nanoparticle, silver nanoparticle crystal, gold nano-crystal or several
The combination of kind.
5. the preparation method of the nanocrystal self assembly aggregation of protein mediation described in any one of claim 1-4,
It is characterized by: being mixed after the oil-soluble nano particles are dissolved in organic solvent with protein solution, shaking and shake up, i.e.,
Obtain the multifunctional nanocrystals self assembly aggregation simultaneously with a variety of nanocrystal characteristics.
6. the preparation method of the nanocrystal self assembly aggregation of protein mediation according to claim 5, feature exist
In: include the following steps:
1) protein solution is dissolved in protein buffer liquid, is configured to protein solution；
2) in organic solvent by the dissolution of one or more of oil-soluble nano particles, it is configured to nanocrystal solution；
3) nanocrystal solution is mixed with protein solution, concussion shakes up；
4) above-mentioned mixed liquor is incubated at room temperature, the mixed liquor after being incubated for；
5) mixed liquor after above-mentioned incubation is centrifuged, removes supernatant, precipitating is the nanocrystal self assembly of protein mediation
Aggregation.
7. the preparation method of the nanocrystal self assembly aggregation of protein mediation according to claim 6, feature exist
In: the concentration of the protein solution is 0.1-10mg/mL, and the concentration of the nanocrystal solution is 0.05-20mg/mL.
8. the preparation method of the nanocrystal self assembly aggregation of protein mediation according to claim 6, feature exist
In: in step 4), the time of the incubation is three hours or more.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111635539A (en) * | 2020-05-12 | 2020-09-08 | 南昌大学 | Production method of flavone-protein self-assembly reversible gel |
| CN114853882A (en) * | 2022-04-02 | 2022-08-05 | 西北工业大学 | Method for preparing protein nano crystal with uniform size |
Citations (3)
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| CN106124758A (en) * | 2016-06-12 | 2016-11-16 | 华中科技大学 | A kind of preparation method and applications of water soluble microsphere |
| CN106990250A (en) * | 2017-05-23 | 2017-07-28 | 华中科技大学 | A kind of preparation method and application for reducing self-assembled protein coated magnetic microballoon |
| CN107753435A (en) * | 2017-10-31 | 2018-03-06 | 聊城大学 | A kind of medicine phosphatide/albumin compound nano-particle and preparation technology |
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| CN106124758A (en) * | 2016-06-12 | 2016-11-16 | 华中科技大学 | A kind of preparation method and applications of water soluble microsphere |
| CN106990250A (en) * | 2017-05-23 | 2017-07-28 | 华中科技大学 | A kind of preparation method and application for reducing self-assembled protein coated magnetic microballoon |
| CN107753435A (en) * | 2017-10-31 | 2018-03-06 | 聊城大学 | A kind of medicine phosphatide/albumin compound nano-particle and preparation technology |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111635539A (en) * | 2020-05-12 | 2020-09-08 | 南昌大学 | Production method of flavone-protein self-assembly reversible gel |
| CN111635539B (en) * | 2020-05-12 | 2022-12-02 | 南昌大学 | Production method of flavone-protein self-assembly reversible gel |
| CN114853882A (en) * | 2022-04-02 | 2022-08-05 | 西北工业大学 | Method for preparing protein nano crystal with uniform size |
| CN114853882B (en) * | 2022-04-02 | 2024-04-05 | 西北工业大学 | Method for preparing protein nanocrystals with uniform size |
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