CN104384523B - Preparing method of three-dimensional chiral silver nanometer material - Google Patents
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Abstract
The invention belongs to the field of nanometer materials and optics, and provides a preparing method of a three-dimensional chiral silver nanometer material using DNA (deoxyribonucleic acid) frameworks as templates. According to the method, the DNA frameworks are used as templates for absorbing silver nanometer ions, and the silver nanometer ions are reduced into silver nanometer particles through reducing agents. The method concretely comprises the following steps that (1) DNA three-dimensional frameworks are prepared; (2) the DNA frameworks absorb the silver nanometer particles; (3) the silver nanometer ions which are not adsorbed are removed through ultrafiltration; (4) the silver nanometer particles are reduced through reducing agents. The problems of low assembling yield, nonuniform assembling body structure and the like during the preparation of chiral nanometer materials by using chiral molecules in the prior art are solved, and the preparing method of the three-dimensional chiral silver nanometer material using the DNA frameworks as the templates is provided. The method has the advantages that the growth dimension of the silver nanometer particles can be effectively controlled through regulating the concentration of the silver nanometer ions and the reducing agents in the system, and a transmission electron microscope result shows that the four silver nanometer particles are in spiral arrangement, so that the chiral signal resources are explained.
Description
Technical field
The invention belongs to nano material and optical field, it is related to a kind of three-dimensional chirality silver nanoparticle with DNA skeleton as template
The preparation method of material.
Background technology
Nanometer is to weigh a kind of unit of test-material yardstick, and 1nm is equal to 10-9Rice, is equivalent to the size of 10 atoms.Nanometer skill
Art is of the number, species and the geometric configuration that accurately identify, observe and control material on single atom or molecular scale
The investigative technique of multiple subject crossing.The appearance of nanotechnology has brought the mankind into a uncharted field " be situated between and see ".It is deposited
It is between microcosmic and macroscopic view, spatial dimension arrives 100nm 1, the material being in this narrow and small world has much peculiar light
The properties such as, magnetics, electricity and mechanics, this kind of material and so that nanometer is referred to as by the material that they form as individual element
Material.
The preparation of chiral nanomaterial is the focus direction of Recent study, is made using the special optically-active of chiral nanomaterial
With can be used to screen chiral biomolecule, build chiral sensor and be used for the important Hazard factor of food, biomacromolecule, with
And the key areas such as disease early diagnosiss.Assemble the chiral nanometer with asymmetric space conformation using single nanoparticle from group
Assembling structure is the important application of DNA self-assembling technique, is also the important channel of the multi-functional chiral material of preparation.Three-dimensional manometer
Grain spirillum is one of most typical chiral structure, can successfully prepare homogeneous, high yield nanometer spiral using DNA paper folding technology
Body, and can be current with the configuration of the distance between the spirochetal physical dimension of precise control, nano-particle and enantiomer
The technological means of comparative maturity.But the preparation of chiral nanomaterial, at present still in the primary stage, develops novel chiral nanometer
Assembled material novel preparation method and new way, and mechanism that its chirality is produced to be studied and explored be following research side
To.
At present, the preparation of chiral nanomaterial mainly will with multiple chiral molecules (e.g., protein, polypeptide, DNA etc.)
Nanomaterial assembly becomes different spaces configuration.In the presence of linearly polarized light, there is plasma in chiral molecule and nanoparticle
Resonance, the chiral signal (200-350nm) of chiral molecule itself can transfer to nanoparticle surface, and circular dichroism spectra shows as receiving
There is obvious circular dichroism signal at the plasma resonance wavelength of rice material.However, the chiral nanomaterial using the method preparation
Usually there is, thus having a strong impact on its optical property and practical application the problems such as assembling low yield, assembly structure heterogeneity.
Therefore, exploitation chiral nanomaterial novel preparation method, new way, thus obtain homogeneous, high yield, stable chirality receive
Rice material is a kind of technological innovation developing direction.
Content of the invention
Present invention aim to address utilizing the chiral nanomaterial of chiral molecule preparation that assembling product occurs in prior art
Rate is low, assembly structure heterogeneity the problems such as, a kind of preparation of the three-dimensional chirality silver nano material with DNA skeleton as template is provided
Method.
1. the present invention provides a kind of preparation method of three-dimensional chirality silver nano material, and this silver nano material with DNA skeleton is
Template Adsorption For Ag nanoparticle, by reducing agent by silver nanoparticle ion reduction be silver nano-grain, thus obtaining with rival's property
Silver nanoparticle assembled material；
Wherein, this DNA skeleton is formed by 4 single stranded DNA phase mutual crosses；
Article 4, DNA is single-stranded is respectively：L1, sequence is as shown in SEQ ID NO.1；L2, sequence is as shown in SEQ ID NO.2；
L3, sequence is as shown in SEQ ID NO.3；L4, sequence is as shown in SEQ ID NO.4.
2. the preparation method of the three-dimensional chirality silver nano material of above-mentioned 1 offer, wherein, this DNA is single-stranded to pass through transformation, passes through
The pyramidal spatial skeleton of base pair complementarity one DNA of formation, pyramid totally 6 sides, four edges wherein are embedded with
10 base C sequences；
DNA is single-stranded can be synthesized with engineer, and on DNA pyramid four edges, embedded base C sequence is single-stranded in itself by DNA
Sequence provides.Four silver ions growing out are on pyramidal four sides.
3. the preparation method of the three-dimensional chirality silver nano material of above-mentioned 2 offers, comprises the steps：
1) preparation of DNA three dimensions skeleton：Four single strand dna equal-volumes are mixed, between base, passes through complementation
It is DNA three dimensions skeleton that pairing hybridization formation has the pyramidal spatial skeleton of DNA；
2) DNA skeleton Adsorption For Ag nanoparticle：To step 1) add AgNO in the three dimensional DNA skeleton prepared3Solution, Ag+Can
With specific absorption in base C Surface；
3) ultrafiltration removes unadsorbed silver nanoparticle ion：To step 2) solution prepared carries out ultrafiltration, removes in solution not
Adsorb the Ag on sub- DNA surface+；
4) reducing agent reduction silver nano-grain：To step 3) add sodium borohydride, by Ag in the solution prepared+It is reduced to Ag
Nano-particle, three-dimensional chirality silver nano material preparation completes.
4. the preparation method of the three-dimensional chirality silver nano material of above-mentioned 3 offers, specifically includes following steps：
1) preparation of DNA three dimensions skeleton：
4 kinds of different single stranded DNAs：L1, L2, L3, L4, with buffer resuspended to 100-500nM, after water-bath, quick cool down
Be cooled to room temperature to 4 DEG C or gradually, during cooling, four single stranded DNA phase mutual crosses are formed has the pyramidal sky of DNA
Between skeleton be DNA three dimensions skeleton；
2) DNA skeleton Adsorption For Ag nanoparticle：
Take step 1) the DNA three dimensions skeleton prepared, add AgNO3Solution, DNA and AgNO3Mol ratio be 1: 50-1
: 200, lucifuge reaction concussion is overnight；
3) reducing agent reduction silver nano-grain：
Take step 3) reaction solution 100ul, add final concentration of 200-500mM sodium borohydride solution, solution colour by
Gradual change is yellow, finally becomes glassy yellow by colourless, then silver nano-grain preparation completes.
5. the preparation method of the three-dimensional chirality silver nano material of above-mentioned 4 offers, specifically includes following steps：
1) preparation of DNA three dimensions skeleton：
Concentration is 4 kinds of different single stranded DNAs of 100mM：L1, L2, L3, L4, resuspended extremely with 10mM tris-HCl buffer
Final concentration of 500nM, 4 kinds of single strand dnas, often pipe take 50uL equal-volume to be mixed in 1.5mL centrifuge tube, 5 points of 95 DEG C of water-baths
Zhong Hou, is cooled fast to 4 DEG C, and during cooling, four single stranded DNA phase mutual cross formation 500nM have DNA pyramid
Spatial skeleton be DNA three dimensions skeleton；
2) DNA skeleton Adsorption For Ag nanoparticle：
Take step 1) the DNA three dimensions skeleton 200uL for preparing, add AgNO3Solution, DNA and AgNO3Mol ratio be 1
: 200, lucifuge reaction concussion is overnight.
During reaction, positively charged Ag+Specifically adsorb in base C in DNA skeleton until reaching
Saturation, unadsorbed Ag+Dispersion is in the solution；
3) ultrafiltration removes unadsorbed silver nanoparticle ion：
Take step 2) the solution 200uL for preparing, it is diluted to 600uL with ultra-pure water, be added separately to 3 0.5mL 30kDa
Super filter tube in, after often pipe 200uL, 13000r/min are centrifuged 10 minutes, recovered to 200uL with ultra-pure water, repeat ultrafiltration once
Afterwards, the Polyvinylpyrrolidone being 55000 then with mass volume ratio 1%, molecular weight is resuspended, and the cumulative volume of final solution is
200uL；
4) reducing agent reduction silver nano-grain：
Step 3) solution after the ultrafiltration prepared takes in 100uL to 1 PCR pipe, adds the boron hydrogen of final concentration of 500mM
Change sodium solution, after 10 seconds, solution colour gradually turns yellow, and finally becomes glassy yellow by colourless, then silver nano-grain preparation completes；
Glassy yellow is observed result, as long as yellow occurs all illustrate that reduction is successful, the degree of color and particle diameter
Size has relation.
Beneficial effects of the present invention：
1. the present invention utilizes base C to Ag+Specific adsorption effect, by Ag+It is fixed on DNA skeleton, by reducing agent
By Ag+It is reduced to silver nano-grain, after effectively prevent nano-particle, assemble the unstable and inhomogeneity of product.
2. pair solution adopts 2 ultrafiltration, effectively eliminates unadsorbed Ag in system+It is ensured that only absorption is in DNA skeleton
On Ag+It is reduced to Ag nano-particle, improve the specificity of this method.
3. pass through appropriate design DNA sequence, controlled is embedded on DNA skeleton by Ag nano-particle, four silver nanoparticles
Particle shape becomes helical structure so as to have stronger circular dichroism, develops a kind of new way of chiral nanomaterial preparation with newly
Method.
4. pass through the concentration of silver nanoparticle ion and reducing agent in regulation system, before not up to saturation value, the silver of addition
The quantity of ion is more, and the particle diameter of the silver nano-grain of growth is bigger；Concentration one timing of silver ion, the amount of reducing agent is bigger,
Growth silver nano-grain particle diameter less, can effective control silver nano-grain growth size.
Brief description
Fig. 1 three-dimensional chirality silver nanoparticle assembly high-resolution-ration transmission electric-lens figure, sodium borohydride concentration is 500mM.
Fig. 2 three-dimensional chirality silver nanoparticle assembly low power transmission electron microscope picture, sodium borohydride concentration is 500mM.
Circular dichroism spectrogram before and after Fig. 3 three-dimensional chirality silver nanoparticle assembly preparation.
Ultraviolet spectrogram before and after Fig. 4 three-dimensional chirality silver nanoparticle assembly preparation.
Fig. 5 three-dimensional chirality silver nanoparticle assembly pyramid space structure schematic diagram.
Specific embodiment
With reference to embodiment, the present invention will be further described, the experiment side of unreceipted actual conditions in the following example
Method, generally according to the known approaches of this area.
Embodiment 1
(1) preparation of DNA three dimensions skeleton
4 kinds of different single stranded DNAs are by Sangon Biotech (Shanghai) Co., Ltd.：L1 sequence such as SEQ ID NO.1
Shown；L2 sequence is as shown in SEQ ID NO.2；L3 sequence is as shown in SEQ ID NO.3；L4 sequence as shown in SEQ ID NO.4,
Table 1 shows the sequence of 4 single stranded DNAs used, and black italic is base C sequence, positively charged Ag+Specifically adsorb
In base C in DNA skeleton.Article 4, the initial concentration of single stranded DNA is 100mM, uses 10mM tris-HCl buffer weight respectively
Hang to final concentration of 500nM, 4 kinds of DNA moleculars, often pipe takes 50uL equal-volume to be mixed in 1.5mL centrifuge tube, 5 points of 95 DEG C of water-baths
Zhong Hou, is cooled fast to 4 DEG C, and during cooling, four single stranded DNA phase mutual crosses are formed has three dimensions configuration
DNA assembly.It is 4 kinds of single stranded sequences used in table 1, wherein black italic is base C sequence.
(2) DNA skeleton Adsorption For Ag nanoparticle
Take DNA three dimensions assembly 200uL prepared by step (1), add a certain amount of AgNO3Solution, DNA and AgNO3
Mol ratio be 1: 200, lucifuge reaction concussion overnight.During reaction, positively charged Ag+Specifically adsorb
Until reaching saturation in base C in DNA skeleton, unadsorbed Ag+Dispersion is in the solution.
(3) ultrafiltration removes unadsorbed silver nanoparticle ion
The solution 200uL taking step (2) to prepare, is diluted to 600uL with ultra-pure water, is added separately to 3 0.5mL 30kDa
Super filter tube in, after often pipe 200uL, 13000r/min are centrifuged 10 minutes, recovered to original volume with ultra-pure water, repeat ultrafiltration once
Afterwards, more resuspended with mass volume ratio 1% Polyvinylpyrrolidone (molecular weight 55000), the cumulative volume of final solution is 200uL.
(4) reducing agent reduction silver nano-grain
Solution after ultrafiltration prepared by step (3) respectively takes in 100uL to 1 PCR pipe, adds final concentration of 500mM's
Sodium borohydride solution, after 10 seconds, solution colour gradually turns yellow, and finally becomes glassy yellow by colourless, then prepared by silver nano-grain
Become.
Embodiment 2
(1) preparation of DNA three dimensions skeleton
4 kinds of different single stranded DNAs, L1 sequence is as shown in SEQ ID NO.1；L2 sequence is as shown in SEQ ID NO.2；L3 sequence
Row are as shown in SEQ ID NO.3；As shown in SEQ ID NO.4, table 1 shows the sequence of 4 single stranded DNAs used, black to L4 sequence
Italic is base C sequence, positively charged Ag+Specifically adsorb in base C in DNA skeleton.Article 4, single stranded DNA is first
Beginning concentration is 100mM, respectively use resuspended to the final concentration of 100nM of 10mMtris-HCl buffer, 4 kinds of DNA moleculars, often pipe take
50uL equal-volume is mixed in 1.5mL centrifuge tube, after 95 DEG C of water-baths 5 minutes, naturally cools to room temperature, during cooling,
Article four, the mutual cross of single stranded DNA phase forms and has the pyramidal spatial skeleton of DNA.
(2) DNA skeleton Adsorption For Ag nanoparticle
Take DNA three dimensions assembly 200uL prepared by step (1), add a certain amount of AgNO3Solution, DNA and AgNO3
Mol ratio be 1: 50, lucifuge reaction concussion overnight.During reaction, positively charged Ag+Specifically adsorb in DNA
Until reaching saturation in base C in skeleton, unadsorbed Ag+Dispersion is in the solution.
(3) ultrafiltration removes unadsorbed silver nanoparticle ion
The solution 200uL taking step (2) to prepare, is diluted to 600uL with ultra-pure water, is added separately to 3 0.5mL 30kDa
Super filter tube in, after often pipe 200uL, 13000r/min are centrifuged 10 minutes, recovered to original volume with ultra-pure water, repeat ultrafiltration once
Afterwards, more resuspended with 1% Polyvinylpyrrolidone (molecular weight 55000), the cumulative volume of final solution is 200uL.
(4) reducing agent reduction silver nano-grain
Solution after ultrafiltration prepared by step (3) respectively takes in 100uL to 1 PCR pipe, adds final concentration of 200mM's
Sodium borohydride solution, after 10 seconds, solution colour gradually turns yellow, and finally becomes glassy yellow by colourless, then prepared by silver nano-grain
Become.
The sequence of DNA used in table 1 experiment
Embodiment 3
Optics and structural characterization are carried out to the three-dimensional chirality silver nano-grain assembly of preparation
The product that embodiment 1 is prepared carries out 15000r/min and is centrifuged 20 minutes, removes supernatant, with weight in ultra-pure water
Outstanding, ultrapure washing is once.Electronic Speculum characterizes：The above-mentioned sample processing of 10 μ L is added drop-wise on the copper mesh of carbon film support, infrared
It is dried under lamp.Electronic Speculum adopts the Electronic Speculum of JEOLJEM-2100 model thoroughly, and its accelerating potential is 200kV, as Fig. 1 and Fig. 2 institute
Show, Fig. 1 be the high power transmission electron microscope body of silver nano-grain helical structure it can be seen that the concrete configuration of silver nano-grain, four
Granule not on same horizontal plane, in clockwise helical structure；Fig. 2 is three-dimensional chirality silver nanoparticle assembly low power transmission electricity
Mirror figure, does not have the aggregation of sheet, the good dispersion of grains, assembly is more in electron microscope.
Circular dichroism spectra characterizes：Take the system 100 μ L assembling in cuvette, do blank with ultra-pure water.Circular dichroism
Spectrometer adopts French Bio-Logic MOS-450+SMF-300, as shown in figure 3, before silver nano-grain does not reduce, system is
There is no chiral signal, thus in 400nm about there is no obvious absworption peak；After silver nano-grain reduces, four of growth
Silver nano-grain helically structural arrangement, presents chiral structure, so obvious absworption peak near 400nm, thus
Prove that chiral nanomaterial is successfully prepared.
Ultraviolet spectra characterizes：Take the system 100 μ L assembling in cuvette, do blank with ultra-pure water.Ultraviolet light
Spectrometer adopts U.S. UNICO 2100 PC UV/Vis/NIR, as shown in figure 4, before silver nano-grain does not reduce, in system
There is no silver nano-grain, so being not in the ultraviolet absorption peak of silver nano-grain near 400nm；When silver nano-grain reduction
Afterwards, contain silver nano-grain in system, so the ultraviolet absorption peak of silver nano-grain occurs near 400nm, thus prove silver
The reduction of nano-particle is successful.
It is recognised that the illustrative embodiments that above-described embodiment adopts only for explanation inventive principle, but this
Bright be not limited only to this, those skilled in the art without departing under real situation of the present invention, can make various improve and change, this
A little improvement and change fall within protection scope of the present invention.
Claims (4)
1. a kind of preparation method of three-dimensional chirality silver nano material it is characterised in that：This silver nano material is with DNA skeleton as template
Adsorption For Ag nanoparticle, by reducing agent by silver nanoparticle ion reduction be silver nano-grain, thus obtaining the silver with rival's property
Nanometer assembled material；
Wherein, this DNA skeleton is formed by 4 single stranded DNA phase mutual crosses；
Wherein, 4 DNA are single-stranded respectively：L1, sequence is as shown in SEQ ID NO.1；L2, sequence is as shown in SEQ ID NO.2；
L3, sequence is as shown in SEQ ID NO.3；L4, sequence is as shown in SEQ ID NO.4；
DNA is single-stranded to pass through transformation, by the pyramidal spatial skeleton of base pair complementarity one DNA of formation, totally 6, pyramid
Side, four edges wherein are embedded with the sequence of 10 bases C.
2. three-dimensional chirality silver nano material according to claim 1 preparation method it is characterised in that：This preparation method bag
Include following steps：
1) preparation of DNA three dimensions skeleton：By four single strand dnas：L1, L2, L3, L4 equal-volume mixes, between base
Hybridize formation to have the pyramidal spatial skeleton of DNA by complementary pairing is DNA three dimensions skeleton；
2) DNA skeleton Adsorption For Ag nanoparticle：To step 1) add AgNO in the three dimensional DNA skeleton prepared3Solution, Ag+Can be special
The absorption of the opposite sex is in base C Surface；
3) ultrafiltration removes unadsorbed silver nanoparticle ion：To step 2) solution prepared carries out ultrafiltration, removes unadsorbed in solution
The Ag on sub- DNA surface+；
4) reducing agent reduction silver nano-grain：To step 3) add sodium borohydride, by Ag in the solution prepared+It is reduced to Ag nanometer
Granule, three-dimensional chirality silver nano material preparation completes.
3. three-dimensional chirality silver nano material according to claim 2 preparation method it is characterised in that：This preparation method has
Body comprises the steps：
1) preparation of DNA three dimensions skeleton：
4 kinds of different single stranded DNAs：L1, L2, L3, L4, use buffer resuspended to 100-500nM respectively, after water-bath, quick cooling
Be cooled to room temperature to 4 DEG C or gradually, during cooling, four single stranded DNA phase mutual crosses are formed has the pyramidal sky of DNA
Between skeleton be DNA three dimensions skeleton；
2) DNA skeleton Adsorption For Ag nanoparticle：
Take step 1) the DNA three dimensions skeleton prepared, add AgNO3Solution, DNA and AgNO3Mol ratio be 1:50-1:
200, lucifuge reaction concussion is overnight；
3) reducing agent reduction silver nano-grain：
Take step 3) reaction solution 100ul, add the sodium borohydride solution of final concentration of 200-500mM, solution colour gradually becomes
Huang, finally becomes glassy yellow by colourless, then silver nano-grain preparation completes.
4. three-dimensional chirality silver nano material according to claim 3 preparation method it is characterised in that：This preparation method has
Body comprises the steps：
1) preparation of DNA three dimensions skeleton：
Concentration is 4 kinds of different single stranded DNAs of 100mM：L1, L2, L3, L4, use 10mM tris-HCl buffer resuspended extremely respectively
Final concentration of 500nM, 4 kinds of single strand dnas, respectively take 50uL equal-volume to mix, after 95 DEG C of water-baths 5 minutes, be cooled fast to 4
DEG C, during cooling, four single stranded DNA phase mutual cross formation 500nM have the pyramidal spatial skeleton of DNA, i.e. DNA
Three dimensions skeleton；
2) DNA skeleton Adsorption For Ag nanoparticle：
Take step 1) the DNA three dimensions skeleton 200uL for preparing, add AgNO3Solution, DNA and AgNO3Mol ratio be 1:
200, lucifuge reaction concussion is overnight；
3) ultrafiltration removes unadsorbed silver nanoparticle ion：
Take step 2) the solution 200uL for preparing, it is diluted to 600uL with ultra-pure water, points 3 parts, after 13000r/min is centrifuged 10 minutes,
Recovered to 200uL with ultra-pure water, after repeating ultrafiltration once, then the polyethylene pyrrole being 55000 with mass volume ratio 1%, molecular weight
Pyrrolidone is resuspended, and the cumulative volume of final solution is 200uL；
4) reducing agent reduction silver nano-grain：
Step 3) solution after the ultrafiltration prepared takes 100uL, adds the sodium borohydride solution of final concentration of 500mM, and 10 seconds
Solution colour gradually turns yellow afterwards, finally becomes glassy yellow by colourless, then silver nano-grain preparation completes.
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| CN105967143B (en) * | 2016-05-06 | 2017-12-08 | 陕西师范大学 | A kind of chiral metal nanostructured for realizing circular dichroism and preparation method thereof |
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