CN109202098A - A kind of preparation method of keratin compound silver nanometer cluster - Google Patents
A kind of preparation method of keratin compound silver nanometer cluster Download PDFInfo
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Abstract
The present invention relates to a kind of preparation methods of keratin compound silver nanometer cluster, using following steps: (1) mixing keratin solution with silver nitrate solution and adjust pH value；(2) sodium borohydride solution is added and stands reaction 30min；(3) it is terminated and is reacted by purification process, fluorescence silver nanoclusters are prepared.Compared with prior art, raw material of the present invention is easy to get and inexpensively, and system is simple, and generated time is short, favorable reproducibility.Obtained material has good biocompatibility and stronger and stable photoluminescent property.
Description
Technical field
The invention belongs to fluorescence nano cluster preparation fields, more particularly, to a kind of preparation side of keratin compound silver nanometer cluster
Method.
Background technique
Silver nanoclusters are due to its unique physical, electrical and optical properties, in recent years in catalysis, chemical sensor, electronics
The fields such as device and bio-imaging are got the attention.Relative to gold nanoclusters, silver nanoclusters have lower production cost.
Silver nanoclusters with high quantum production rate, near infrared emission wavelength and high-biocompatibility possess huge in bio-imaging field
Application prospect.Biomineralization method is a kind of environmentally protective synthetic method that last decade is risen, and reaction principle is by egg
The groups such as amine, carboxyl and thiol group in white matter or nucleic acid sequence are as stabilizer in synthetic reaction and reducing agent substitution tradition
Chemical reagent.Nucleic acid sequence and bovine serum albumin(BSA) are common two kinds of biological raw materials at present, but these raw materials all have cost
Higher problem, bovine serum albumin(BSA) also need to pre-process in the synthesis process, and step is complex；And existing method is logical
Crossing reduces reaction speed using the means of low concentration reducing agent in the reaction, in order to need the side of purifying for a long time using dialysis etc.
Method terminates reaction, at 24 hours or more the time required to entire reaction.
Keratin is prevalent in the hair of humans and animals, using soxhlet extraction can laboratory voluntarily in animal
Extract keratin in hair, raw material is easy to get and inexpensively；Keratin polypeptide have complete segmented structure and hydroxyl, carboxyl, amino,
Disulfide bond etc. can form the functional group of strong effect power with metal cation；It is big with molecular weight but for silver-colored cluster
For albumen as templated synthesis, current document is all seldom, and it is insufficient to be primarily due to sulfhydryl content in general albumen, separately
Outer NaBH4The time for participating in reaction system is difficult to control.
Chinese patent CN108031857A discloses the preparation method of the gold nano cluster of hair red fluorescence, and chlorine is respectively configured
Chlorauric acid solution, is slowly added into keratin solution by auric acid solution, keratin solution at room temperature, is then sufficiently stirred, and makes
Solution is uniformly mixed, and obtains yellow solution；The pH value of solution is adjusted to 8~12；Solution is placed under the conditions of 25~60 DEG C and is incubated
It educates, obtains the gold nano cluster solution of brown；It is to dialyse in bag filter that gold nano cluster solution, which is poured into molecular cut off, then
Freeze-drying obtains gold nanoclusters particle.Silver nanoclusters and gold nanoclusters belong to noble-metal nanoclusters, send out the gold of red fluorescence
Belonging to cluster can be used as fluorescence probe use.Relative to gold nanoclusters, need to close using reducing agent in the synthesis process of silver nanoclusters
It is relative complex at process；But the synthesis cost of silver nanoclusters is lower.
Summary of the invention
It is an object of the present invention to overcome the above-mentioned drawbacks of the prior art and provide one kind to have life well
The preparation method of the keratin compound silver nanometer cluster of object compatibility and stronger and stable photoluminescent property.
The purpose of the present invention can be achieved through the following technical solutions:
A kind of preparation method of keratin compound silver nanometer cluster, using following steps:
(1) keratin solution and silver nitrate solution are stirred 5 minutes and the reaction was continued after adjusting pH value 15 points
Clock；
(2) sodium borohydride solution is added and stands reaction 30 minutes；
(3) it is terminated and is reacted by purification process, fluorescence silver nanoclusters are prepared.
In step (1):
Controlling reaction temperature is 0-40 DEG C, and 25 DEG C are preferably used in 0 DEG C, 25 DEG C, 40 DEG C.
The mass ratio of silver nitrate and the keratin is 1:6.59-1:39.54, and preferred proportionate relationship is 1:13.18.
Addition concentration is 0.1-10mol/L sodium hydroxide solution, and preferably 1mol/L sodium hydroxide adjusts pH value to 12.4.
Keratin extraction purification from feather, using following steps: feather thoroughly being cleaned with water, and is taken off with acetone
Rouge operation, the smashed feather of extracting degreasing, which immerses, contains 8mol/L urea, 0.2mol/L lauryl sodium sulfate and 0.5mol/L
In the mixed solution of sodium pyrosulfite, after keratin degrading liquid is filtered removal insoluble matter after heat treatment, it is placed in obstruction size
To dialyse in the bag filter of 8000-14000Da, keratin solution is finally lyophilized into powder.
The molar ratio of silver nitrate and sodium borohydride is 1:0.1-1:1 in step (2), and preferred proportionate relationship is 1:1.
Step (3) as the chromatographic column of filler carried out column purification using Sephadex G25 gel, using ultraviolet lamp into
Row judges position of the fluorescence silver nanoclusters in chromatographic column, the process time-consuming about 10 minutes.
The size for the silver nanoclusters being prepared is 2.5nm, there is stronger fluorescent emission at 710nm, and fluorescent quantum produces
Rate is 1.7% (being control with rhodamine B).Obtained material has good biocompatibility and stronger and stable fluorescence
Property.
Compared with prior art, the present invention is low in cost and has good biocompatibility；Present invention employs gels
It crosses column method and quickly removes reducing agent small molecule in reaction system, the termination time of accurate regulation reaction, thus in the reaction
Use the NaBH of higher concentration4Accelerate reaction speed, this method generated time only needs 1 hour, easy to operate and reproducible；
Based on sulfydryl rich in keratin, the silver nanoclusters of this method synthesis have launch wavelength, the higher amount of near-infrared
Sub- yield and in aqueous solution have stable fluorescent intensity, in terms of bio-imaging have huge application prospect.
The present invention by keratin there are the particular advantages of abundant disulfide bond and gel to cross column method can be with Quick stop
The characteristics of reaction, obtain using keratin as template have biocompatibility, fluorescence property is excellent, stability is good, synthetic method
Simple and quick, the lower silver nanoclusters synthetic method of cost.
Detailed description of the invention
Fig. 1 is the transmission electron microscope schematic diagram and particle diameter distribution for the silver nanoclusters that the embodiment of the present invention 1 synthesizes.
Fig. 2 is the abosrption spectrogram for the silver nanoclusters that the embodiment of the present invention 1 synthesizes.
Fig. 3 is the fluorescent stability phenogram for the silver nanoclusters that the embodiment of the present invention 1 synthesizes.
Specific embodiment
The present invention is described in detail combined with specific embodiments below.Following embodiment will be helpful to the technology of this field
Personnel further understand the present invention, but the invention is not limited in any way.It should be pointed out that the ordinary skill of this field
For personnel, without departing from the inventive concept of the premise, various modifications and improvements can be made.These belong to the present invention
Protection scope.
Reagent raw material
Solvent used in reaction is ultrapure water；Sodium hydroxide, sodium borohydride (96%), nitric acid, phosphoric acid (95%), peroxide
Change hydrogen (30%) and is purchased from Sinopharm Chemical Reagent Co., Ltd.；Silver nitrate (99.8%) is limited purchased from Shanghai Ling Feng chemical reagent
Company；Keratin is voluntarily extracted from feather.
Fluorescence spectrum is recorded by Fluorescence Spectrometer (Edinburgh, Britain FS-5)；UV-vis absorption spectrum is to use UV-
2450 spectrophotometers (Japanese Shimadzu) measurement；Transmission electron microscope (TEM) is in JEOLJEM-2100 ultrahigh resolution
It is carried out under transmission electron microscope.
The preparation of solution used:
(1) preparation of keratin:
Keratin voluntarily extraction purification from feather by laboratory, specific purification step: feather is thoroughly cleaned with water, and
Degreasing operation is carried out with acetone.The smashed feather 10g of extracting degreasing immerses 100mL and contains 8mol/L urea, 0.2mol/L 12
In the solution of sodium alkyl sulfate and 0.5mol/L sodium pyrosulfite.Above-mentioned solution is heated 70 DEG C, 1 hour.Then by keratin
Degradation solution filtering removal insoluble matter is placed on to stop dialyses in the bag filter that size is 8000-14000Da.Finally by angle egg
It is spare that white solution is lyophilized into powder.
(2) preparation of the solution used in:
It weighs 0.0849g silver nitrate solid and prepares 5mL silver nitrate solution (0.1mol/L), keratin is prepared with ultrapure water
5 parts of concentration are 5~30mg/mL；Weigh 0.4000g sodium hydrate solid prepare 1mL sodium hydroxide (10mol/L) solution and by its
Another two parts of 1mL sodium hydroxides (1mol/L) of dilution and 1mL sodium hydroxide (0.1mol/L)；Weigh 0.1892g sodium borohydride solids
5mL sodium borohydride solution (1mol/L), then the hydrogen for being 0.1mol/L with concentration by it are configured to 1mol/L sodium hydroxide solution
The sodium hydroxide solution that sodium hydroxide solution and concentration are 0.01mol/L be diluted to respectively 0.1mol/L sodium borohydride solution and
0.01mol/L sodium borohydride solution.
Embodiment 1
In the state of room temperature and stirring, by 5mL keratin solution (10mg/mL) and 0.2mL silver nitrate solution
(0.1mol/L) sufficiently reacts after five minutes, after addition 0.15mL sodium hydroxide solution (0.1mol/L) is stirred to react 15 minutes, to
0.23mL sodium borohydride solution (0.1mol/L) is added in above-mentioned solution, after mixing, stands 30 minutes, passes through
Sephadex-G25 gel chromatography Column methods terminate reaction and carry out preparing 1mg/ to get stable product silver nanoclusters are arrived
It is most strong to measure fluorescence at 710nm with 400nm excitation for mL silver nanoclusters.
Embodiment 2
In the state of room temperature and stirring, by 5mL keratin solution (5mg/mL, 7.5mg/mL, 15mg/mL, 30mg/mL)
It is sufficiently reacted after five minutes with 0.2mL silver nitrate solution (0.1mol/L), 0.15mL sodium hydroxide solution (1mol/L) stirring is added
After reaction 15 minutes, 0.23mL sodium borohydride solution (0.1mol/L) is added to above-mentioned solution, after mixing, stands 30 points
Clock terminates reaction by Sephadex-G25 gel chromatography Column methods and carries out preparing 1mg/ to get product silver nanoclusters are arrived
ML silver nanoclusters measure 0.12,0.65,0.70,0.49 times that light intensity is respectively 1 product of example with 400nm excitation.
Embodiment 3
In the state of room temperature and stirring, by 5mL keratin solution (10mg/mL) and 0.2mL silver nitrate solution
(0.1mol/L) sufficiently reaction after five minutes, is added 0.15mL sodium hydroxide solution (0.1mol/L, 10mol/L) and is stirred to react 15
After minute, 0.23mL sodium borohydride solution (0.1mol/L) is added to above-mentioned solution, after mixing, stands 30 minutes, leads to
It crosses Sephadex-G25 gel chromatography Column methods and terminates reaction progress to get to stable product silver nanoclusters, prepare
1mg/mL silver nanoclusters measure 0.00,0.19 times that light intensity is respectively 1 product of example with 400nm excitation.
Embodiment 4
In the state of room temperature and stirring, by 5mL keratin solution (10mg/mL) and 0.2mL silver nitrate solution
(0.1mol/L) sufficiently reacts after five minutes, after addition 0.15mL sodium hydroxide solution (0.1mol/L) is stirred to react 15 minutes, to
0.23mL sodium borohydride solution (0.01mol/L) is added in above-mentioned solution, after mixing, stands 30 minutes, passes through
Sephadex-G25 gel chromatography Column methods terminate reaction and carry out preparing 1mg/ to get stable product silver nanoclusters are arrived
ML silver nanoclusters measure 0.70 times that light intensity is respectively 1 product of example with 400nm excitation.
Embodiment 5
In the state of (0 DEG C, 40 DEG C) and stirring, by 5mL keratin solution (10mg/mL) and 0.2mL silver nitrate solution
(0.1mol/L) sufficiently reacts after five minutes, after addition 0.15mL sodium hydroxide solution (1mol/L) is stirred to react 15 minutes, upwards
0.23mL sodium borohydride solution (0.1mol/L) is added in the solution stated, and after mixing, stands 30 minutes, passes through Sephadex-
G25 gel chromatography Column methods terminate reaction and carry out preparing 1mg/mL silver nanoparticle to get stable product silver nanoclusters are arrived
Cluster measures 0.36,0.23 times that light intensity is respectively 1 product of example with 400nm excitation.
Embodiment 6
In the state of room temperature and stirring, by 5mL keratin solution (10mg/mL) and 0.2mL silver nitrate solution
(0.1mol/L) sufficiently reacts after five minutes, after addition 0.15mL sodium hydroxide solution (1mol/L) is stirred to react 15 minutes, upwards
0.23mL sodium borohydride solution (0.1mol/L) is added in the solution stated, and after mixing, stands 30 minutes, passes through (ultrapure water, PH
=6 BR caching solution) to get stable product silver nanoclusters are arrived, preparation 1mg/mL silver is received for purification process termination reaction progress
Rice cluster measures 0.50,0.67 times that light intensity is respectively 1 product of example with 400nm excitation.
Fig. 1 is the transmission electron microscope schematic diagram and particle diameter distribution (2.53 ± 0.54nm) for the silver nanoclusters that embodiment 1 synthesizes.
Fig. 2 is the abosrption spectrogram for the silver nanoclusters that embodiment 1 synthesizes, and middle line 1 is the purple of 1mg/mL keratin solution
Outer absorption spectrum；Line 2 is the ultra-violet absorption spectrum of 1mg/mL silver nanoclusters；Line 3 is the fluorescent absorption light of 1mg/mL silver nanoclusters
Spectrum.As a result understand the presence for not having large scale partial size in silver nanoclusters, and when exciting light is 400nm, maximum fluorescence emission peak
Position is 710nm.
Fig. 3 is the fluorescent stability phenogram for the silver nanoclusters that embodiment 1 synthesizes.Its middle line 1, line 2 are respectively to synthesize knot
Fluorescence intensity after Shu Shiyu preservation three months.The fluorescent intensity of silver nanoclusters can be also maintained at after three months 80% with
On, illustrate the stability of this method synthesis silver nanoclusters.
Specific embodiments of the present invention are described above.It is to be appreciated that the invention is not limited to above-mentioned
Particular implementation, those skilled in the art can make various deformations or amendments within the scope of the claims, this not shadow
Ring substantive content of the invention.
Claims (9)
1. a kind of preparation method of keratin compound silver nanometer cluster, which is characterized in that this method uses following steps:
(1) keratin solution is mixed with silver nitrate solution and adjusts pH value；
(2) sodium borohydride solution is added and stands reaction 30min；
(3) it is terminated and is reacted by purification process, fluorescence silver nanoclusters are prepared.
2. a kind of preparation method of keratin compound silver nanometer cluster according to claim 1, which is characterized in that step (1)
Middle control reaction temperature is 0-40 DEG C.
3. a kind of preparation method of keratin compound silver nanometer cluster according to claim 1, which is characterized in that step (1)
Described in the mass ratio of silver nitrate and the keratin be 1:6.59-1:39.54.
4. a kind of preparation method of keratin compound silver nanometer cluster according to claim 1, which is characterized in that step (1)
Middle addition sodium hydroxide solution adjusts pH value.
5. a kind of preparation method of keratin compound silver nanometer cluster according to claim 4, which is characterized in that the hydrogen-oxygen
The concentration for changing sodium solution is 0.1-10mol/L.
6. a kind of preparation method of keratin compound silver nanometer cluster according to claim 1 or 4 or 5, which is characterized in that step
Suddenly pH value is adjusted in (1) to 12.4.
7. a kind of preparation method of keratin compound silver nanometer cluster according to claim 1, which is characterized in that step (1)
Middle keratin extraction purification from feather, using following steps: feather thoroughly being cleaned with water, and carries out degreasing behaviour with acetone
Make, the smashed feather of extracting degreasing immerses in the mixed solution of urea, lauryl sodium sulfate and sodium pyrosulfite, heat treatment
After keratin degrading liquid is filtered removal insoluble matter afterwards, being placed in obstruction size is that progress is saturating in the bag filter of 8000-14000Da
Analysis, is finally lyophilized into powder for keratin solution.
8. a kind of preparation method of keratin compound silver nanometer cluster according to claim 1, which is characterized in that step (2)
The molar ratio of middle silver nitrate and sodium borohydride is 1:0.1-1:1.
9. a kind of preparation method of keratin compound silver nanometer cluster according to claim 1, which is characterized in that step (3)
Chromatographic column using Sephadex G25 gel as filler carried out column purification, carried out judging fluorescence silver nanoparticle using ultraviolet lamp
Position of the cluster in chromatographic column.
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Cited By (3)
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| CN110106706A (en) * | 2019-05-23 | 2019-08-09 | 叶盛 | A kind of preparation method of modified protein UV resistance finishing agent |
| CN110772432A (en) * | 2019-11-05 | 2020-02-11 | 北京科技大学 | Long-acting gold nanocluster fluorescent hair dye and hair dyeing method |
| CN114621755A (en) * | 2022-04-06 | 2022-06-14 | 广东省科学院生物与医学工程研究所 | Silver nanoparticle probe protected by cow hair keratin and preparation method and application thereof |
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| CN110772432A (en) * | 2019-11-05 | 2020-02-11 | 北京科技大学 | Long-acting gold nanocluster fluorescent hair dye and hair dyeing method |
| CN114621755A (en) * | 2022-04-06 | 2022-06-14 | 广东省科学院生物与医学工程研究所 | Silver nanoparticle probe protected by cow hair keratin and preparation method and application thereof |
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| CN109202098B (en) | 2022-04-01 |
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