CN109884023A - Zinc ion detection method and zinc ion the detection preparation method of Illuminant nanometer probe - Google Patents
Zinc ion detection method and zinc ion the detection preparation method of Illuminant nanometer probe Download PDFInfo
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Abstract
The present invention relates to a kind of zinc ion detection method and the preparation methods of zinc ion detection Illuminant nanometer probe.The Illuminant nanometer probe is prepared by cysteine, copper nanocluster, ethoxy chitosan nanogel composite material.The Illuminant nanometer probe in detecting zinc ion of aggregation-induced emission effect based on CuNCs@GC solution, with fast and convenient, technical requirements are low, high sensitivity, detection limit is low, and the range of linearity is wide, good selective, and under experiment condition after optimization, the imaging of zinc ion in the detection and living biological cell of Trace Elements in Food zinc can be successfully applied to.
Description
Technical field
The present invention relates to a kind of zinc ion detection method and the zinc ion detection preparation methods of Illuminant nanometer probe, belong to
Luminescent detection techniques field.
Background technique
It is well known that zinc (Zn) is one of microelement necessary to organism, the various aspects of wide participation vital movement,
Such as growth and development, reproduction heredity, immune, endocrine.National Science research committee has formulated zinc intake diet and has referred to
South, the reason is that human body is easy to cause hypoimmunity in the case where lacking Zn, wound is difficult to heal and growth and development is slow
Slowly.Studies have shown that with age, the cognitive function of brain can be gradually reduced, this may be related with the Zn content in cell,
That is the adjustable brain cynapse transmitting of Zn, plays an important role in terms of protecting brain normal operation.Food is people except other
The mode of most directly most safe most natural intake zinc other than nutriment, drug often eats the food rich in zinc in diet
Object is just able to satisfy itself needs substantially.Therefore, a kind of method for establishing sensitive, quick, easy, accurate measurement zinc has important
Meaning.
The method of detection Zn has very much, such as high performance liquid chromatography, gas chromatography, capillary electrophoresis, ultraviolet spectrometry
Photometry, atomic absorption spectrography (AAS), inductively coupled plasma atomic emission spectrometry, Electrochemical Detection etc..These methods or
Expensive equipment or pre-treatment are cumbersome, are not suitable for realizing quickly detection.
Summary of the invention
To solve above-mentioned the deficiencies in the prior art, the present invention provides a kind of preparation of zinc ion detection Illuminant nanometer probe
Method.The probe is to Zn2+Highly sensitive and highly selective response is carried out to detect.
The present invention provides a kind of preparation method of zinc ion detection Illuminant nanometer probe, and step includes:
(1) ethoxy chitosan and ultrapure water are subjected to water-bath, until ethoxy chitosan is completely dissolved,
Ethoxy chitosan nanogel stock solution is obtained, placement is cooled to room temperature；The deacetylated shell of ethoxy after cooling is more
The ethoxy chitosan nanogel that sugared nanogel stock solution is diluted to 0.5-2.0 mg/mL with ultrapure water is molten
Liquid；
(2) the ethoxy chitosan nanogel solution for obtaining step (1) and cysteine ultrasonic mixing extremely dissolve,
Add CuSO4·5H2O solution, 30 min of gentle agitation obtain zinc ion detection Illuminant nanometer probe CuNCs@GC solution.
Further, the ratio of ethoxy chitosan described in step (1) and ultrapure water is (10-25) mg:
(1-10) mL。
Further, ethoxy chitosan nanogel solution described in step (2), cysteine and
CuSO4·5H2The ratio of O solution is (2.5-4.5) mL: (0.0121-0.0605) g:(0.5-2.5) mL.
The present invention also provides a kind of zinc ion detection method, it is 7.4 as above-mentioned that step, which includes: to the pH value of certain volume,
CuNCs@GC solution in be added same volume various concentration Zn2+Solution, then zinc ion content is carried out after being diluted with ultrapure water
Detection.
Further, step includes: the Zn that (1) takes the 200 μ L of at least five kinds various concentrations2+Standard solution is added immediately
It is in CuNCs@GC solution that 400 μ L, pH are 7.4 to volume, being finally diluted to volume with ultrapure water is 1 mL, is sufficiently mixed
It is the measurement that 387 nm carry out photoluminescence emission spectrum to fix excitation wavelength, and be recorded in this and swash after 5 min of homogeneous reaction
It sends out and various concentration Zn is added under wavelength2+The emissive porwer spectrogram of CuNCs@GC solution afterwards；(2) molten to 400 μ L CuNCs@GC
It is separately added into the 200 μ L of sample liquid of above-mentioned non-mark-on and different scalar quantities in liquid, 1 mL is finally diluted to by ultrapure water, 387
Fluorescence intensity is measured under nm excitation wavelength；Last linear equation calculation Zn content and the rate of recovery.
Further, the detection for Zinc in Foods ion, step include: (1) using micro-wave digestion to milk, beer or
Tealeaves carries out pre-treatment, obtains sample liquid；(2) sample liquid is according to non-mark-on, 0.05 mM of mark-on, 0.15 mM of mark-on, mark-on
0.45 mM carries out the measurement of Zn content, specifically: the sample of non-mark-on and different scalar quantities is separately added into CuNCs@GC solution
200 μ L of liquid, is finally diluted to 1 mL by ultrapure water, measures fluorescence intensity under 387 nm excitation wavelengths；Last linear equation
Calculate Zn content and the rate of recovery.
The utility model has the advantages that
The present invention is unmarked type Illuminant nanometer probe, can be directly to Zn2+Highly sensitive and highly selective response is carried out to examine
It surveys.Illuminant nanometer probe is to Zn prepared by the present invention2+Feature with quick response, and detection limit is low, the range of linearity is wider,
Sensitivity and selectivity are high, and biocompatibility is excellent.Based on Zn2+The CuNCs for causing GC nanogel package generates aggregation inducing
(AIE) effect that shines is simple and quick to prepared by the nano-probe of novel " luminous " detection pattern of building, low toxicity is pollution-free,
With Zn2+Relevant drinking water safety and food analysis field have potential application prospect.
Detailed description of the invention
Fig. 1 is the CuNCs@GC of embodiment 1 and Zeta potential figure of the CuNCs when pH is 7.4 of comparative example 1.
Fig. 2 is that various concentration Zn is added in embodiment 12+The Zeta potential variation diagram of CuNCs@GC is measured later.
Fig. 3 be the embodiment of the present invention 1, comparative example 1-4 product transmission electron microscope picture (TEM)；
Wherein Fig. 3 A is the transmission electron microscope picture of the CuNCs of the product of comparative example 1；
Fig. 3 B is the transmission electron microscope picture of the CuNCs@GC (0.5 mg/mL) of the product of comparative example 2；
Fig. 3 C is the transmission electron microscope picture of the CuNCs@GC (1.0 mg/mL) of the product of embodiment 1；
Fig. 3 D is the transmission electron microscope picture of the CuNCs@GC (1.5 mg/mL) of the product of comparative example 3；
Fig. 3 E is the transmission electron microscope picture of the CuNCs@GC (2.0 mg/mL) of the product of comparative example 4.
Fig. 4 is the x-ray photoelectron spectroscopy figure (XPS) of CuNCs@GC and the Cu2p electronics of the embodiment of the present invention 1.
Fig. 5 is comparative example 1 (a) of the present invention, the fluorescence intensity spectrogram of embodiment 1 (b).
Fig. 6 is the fluorescence intensity spectrogram after the GC nanogel composite Cu NCs of various concentration.
Fig. 7 is the synthesis step and Zn of CuNCs@GC of the invention2+Testing principle.
Fig. 8 is that 30 μM of Zn is being added in nano-probe2+After common metal ion is added in fluorescence intensity difference comparison afterwards
Fluorescence intensity difference.
Fig. 9 is that 30 μM of Zn is being added in nano-probe2+The fluorescence after amino acids is added in fluorescence intensity difference comparison afterwards
Strength difference.
Figure 10 is that 30 μM of Zn is being added in nano-probe2+After other biological substance is added in fluorescence intensity difference comparison afterwards
Fluorescence intensity difference.
Figure 11 is that various concentration Zn is added to the CuNCs@GC solution in embodiment 22+Fluorescence intensity change spectrogram afterwards；
Wherein
Figure 11 A is the fluorescence intensity change for the CuNCs@GC solution in embodiment 2 being added the Zn2+ standard liquid that concentration is 0-750 μM
Spectrogram.
Figure 11 B is fluorescence intensity difference (I-I0) and various concentration Zn2+Gained is respectively in low strength range after standard liquid mapping
(I0 is CuNCs@GC to the good linear relational graph presented in (1.5 μM -100 μM) and high concentration range (100 μM -750 μM)
The fluorescence intensity of solution, I are that various concentration Zn is added in CuNCs@GC solution2+Fluorescence intensity after standard liquid).
Figure 12 is the fluorescence probe based on CuNCs@GC nanocomposite for detecting Zn in A549 living cells2+It is resulting
Confocal fluorescence image.
Specific embodiment
The present invention is explained further below in conjunction with specific embodiment, but embodiment does not do any type of limit to the present invention
It is fixed.
Embodiment 1
A kind of zinc ion detection preparation method of Illuminant nanometer probe, steps are as follows:
(1) preparation of ethoxy chitosan (hereinafter referred to as GC) nanogel solution: accurately weigh 50 mg GC fall in
In the centrifuge tube for filling 10 mL ultrapure waters, it is then ready for water-bath of the temperature at 80 DEG C or so, GC solution is put into water-bath
20 min, until the GC nanogel stock solution that it is 5 mg/mL to get concentration that GC, which is completely dissolved, placement are cooled to room temperature.
GC nanogel stock solution after cooling is diluted to the GC nanogel solution of 1.0mg/mL with ultrapure water.It places
In 4 DEG C of environment, used for subsequent experimental.
(2) preparation of CuNCs@GC solution: the GC nanogel that 4.5 mL steps (1) are prepared is pipetted using liquid-transfering gun
Solution is in the small beaker of 10 mL, and number 1, the cysteine (hereinafter referred to as Cys) for then accurately weighing 0.0605 g are added 1
In number small beaker, the CuSO of 60 mM of 0.5 mL is then added to dissolving in 5 min of ultrasound into No. 1 beaker4·5H2O solution,
The color of mixed solution becomes shallow yellow transparent solution from clear, colorless immediately, last 30 min of gentle agitation has synthesized zinc
Ion detection is with Illuminant nanometer probe (CuNCs@GC solution).The CuNCs@GC solution of synthesis, which is placed in 4 DEG C of environment, to be stored, with
It is used for subsequent step.
No. 1 CuNCs@GC solution is poured into centrifuge tube, is irradiated under the ultraviolet lantern of 365 nm, the feelings that shine are observed
Then condition records the fluorescence intensity of the solution combined CuNCs of various concentration GC nanogel under the excitation wavelength of 387 nm again.
Nanogel type used in the present invention is ethoxy deacetylated chitin.On the one hand, common chitosan passes through
The processing of ethoxy deacetylation, can dissolve well in water, not need the solubility of addition acetic acid increase in water.It is another
Aspect, ethoxy deacetylated chitin compared with common chitosan, stablize, general by aqueous solution property within the scope of the full pH of 1-14
Logical chitosan aqueous solution will appear flocculent deposit after pH=8, so the copper nanoclusters that ethoxy deacetylated chitin is compound
Cluster property is more excellent, stability is more preferable.
Copper nanocluster (CuNCs) due to its preparation cost is low, the simple and quick, excellent optical performance of synthesis, low toxicity and
The advantages that good biocompatibility, can match in excellence or beauty in nano material with gold, silver nanocluster, thus be concerned in illumination field.
The chemical name of cysteine (Cys) is 2- amino-3-mercaptopropionic acid, and wherein sulfydryl (- SH) is easy to form strong coordination with Cu
Key Cu-S key, therefore can be used as protective agent and reducing agent to prepare CuNCs.Ethoxy chitosan (GC) nanogel tool
There is big specific surface area, good biocompatibility is wrapped up CuNCs with GC nanogel, can prevent the nanometer of aggregation
There is a phenomenon where loose and structures to ossify for cluster, and significantly improves the luminous intensity of script CuNCs.Present invention combination Cys is repaired
The advantages of CuNCs, GC nanogel of decorations, is compound by the two, and CuNCs@GC solution is used to answer as Illuminant nanometer probe, and successfully
For in food zinc detection and living biological cell in zinc ion imaging.
Comparative example 1
A kind of zinc ion detection preparation method of Illuminant nanometer probe, prepares the CuSO of 20 mM first4·5H2O solution,
The CuSO of 5 mL preparation is pipetted with liquid-transfering gun in the small beaker of 7 10 mL4·5H2O solution, and number 1-7, it is then quasi-
Really claim the cysteine of 0.0121 g, 0.0204 g, 0.0363 g, 0.0484 g, 0.0605 g, 0.0726 g and 0.0847 g
(Cys), CuSO is filled by what weighed Cys successively poured into number 1-74·5H2In the small beaker of O solution, 5 min of ultrasound are extremely
It is completely dissolved, can observe that mixed solution becomes taupe by clear and fades to light yellow suspension again in this process,
The CuNCs most protected afterwards through 30 min of gentle agitation to get concentration for 100 mM Cys.By the CuNCs prepared in 365 nm
It is irradiated under ultraviolet lantern, observes luminous situation, then record various concentration Cys protection under the excitation wavelength of 405 nm again
The fluorescence intensity of CuNCs.
Fig. 1's the result shows that, simple CuNCs surface charge in comparative example 1 is negative electrical charge, the Zeta electricity in pH=7.4
Gesture is -17.8 mV, and the CuNCs GC surface charge in embodiment 1 is positive charge, and the Zeta electric potential in pH=7.4 is
28.9 mV, this is because containing a large amount of amido in GC nanogel, so the Zeta electric potential of the CuNCs after being wrapped up is positive
Value, and the Zeta electric potential absolute value than simple CuNCs is big, shows that the CuNCs structure of GC nanogel package is more stable, is not easy pine
It dissipates.
Fig. 2's the result shows that, by addition various concentration Zn2+After (0,10,100,150,250,500,750,1000 μM)
Zeta electric potential variation diagram can be seen that with Zn2+The Zeta electric potential value of the increase of concentration, CuNCs@GC is also increasing, reason
It is Zn2+It can be with the carboxylic acid ion (COO for the Cu-S composite surface being wrapped in GC nanogel-) reaction, then consume
Part COO is fallen-Afterwards, in the presence of the amido for being more electric positive charge, therefore, CuNCs@GC system in CuNCs@GC system
Zeta electric potential is with Zn2+Concentration increases the trend for presenting and increasing.
Comparative example 2
A kind of zinc ion detection preparation method of Illuminant nanometer probe, step is the same as embodiment 1.It the difference is that only: will be cold
But the GC nanogel stoste after is diluted to the GC nanogel solution of 0.5mg/mL with ultrapure water；It is prepared No. 2
CuNCs@GC。
Comparative example 3
A kind of zinc ion detection preparation method of Illuminant nanometer probe, step is the same as embodiment 1.It the difference is that only: will be cold
But the GC nanogel stoste after is diluted to the GC nanogel solution of 1.5mg/mL with ultrapure water；It is prepared No. 3
CuNCs@GC。
Comparative example 4
A kind of zinc ion detection preparation method of Illuminant nanometer probe, step is the same as embodiment 1.It the difference is that only: will be cold
But the GC nanogel stoste after is diluted to the GC nanogel solution of 2mg/mL with ultrapure water；No. 4 CuNCs@are prepared
GC。
Fig. 3 be the embodiment of the present invention 1, comparative example 1-4 product transmission electron microscope picture (TEM).
As shown in Fig. 3 A, prepared CuNCs is spherical substantially, and size is similar, non-uniform to be distributed in aqueous solution,
Show successfully to be synthesized by cysteine as the CuNCs of protective agent and reducing agent；But nanocluster is unevenly distributed, and will lead to
CuNCs stability relative mistake.
Fig. 3 B is that the TEM of CuNCs@GC (0.5 mg/mL) schemes, the nanocluster distribution phase as can be seen from the figure dispersed
To uniform, but GC negligible amounts are not enough to completely wrap up nanocluster and fetter, and can only show and hand over to part nanocluster
Fork chain connects effect.
Fig. 3 C is that the TEM of CuNCs GC (1.0 mg/mL) schemes, and nanocluster shown in figure is wrapped up by GC agglomerating substantially
Shape, distribution is more uniform, and relative to pure CuNCs, fluorescence intensity improves about 3 times.
Fig. 3 D and 3E are CuNCs@GC (1.5 mg/mL) and the TEM figure of CuNCs@GC (2.0 mg/mL), above-mentioned TEM figure
The Severe aggregation of GC itself is clearly illustrated, and then causes the fluorescence intensity of CuNCs@GC that can be lower than CuNCs@GC (1.0
Mg/mL fluorescence intensity).
Therefore, an appropriate number of GC is conducive to obtain a highly organized self-assembled structures.
Fig. 4 is the CuNCs@GC (1.0 mg/mL) of embodiment 1 of the present invention and the X-ray photoelectricity of Cu2p electronics
Sub- energy spectrum diagram (XPS).From the XPS of CuNCs GC (1.0 mg/mL) can be seen that the member for including in CuNCs GC be known as C, O,
N, S, Cu, this is consistent with the element that the XPS of CuNCs is shown, illustrates that there is no chemically react compound GC with CuNCs
And change the script structure of CuNCs, CuNCs is wrapped up simply by electrostatic interaction, increases its stability.Pass through Cu2p electronics
The peak value that XPS can be seen that at 932.4 eV and 952.5 eV is respectively Cu2p3/2And Cu2p1/2Combination energy, the two peaks
It belongs to as Cu0Characteristic peak, and illustrate in CuNCs@GC system without characteristic peak almost without Cu in 943.7 eV2+It deposits
?.
Fig. 5 is comparative example 1 (a) of the present invention, and the fluorescence intensity spectrogram of embodiment 1 (b), CuNCs excitation wavelength is
405 nm, launch wavelength are 605 nm, and the excitation of CuNCs@GC and launch wavelength are respectively 387 nm, 615 nm.It can from figure
With the fluorescence intensity spectrogram both found out there are notable difference, the fluorescence intensity level of CuNCs@GC (1.0 mg/mL) is almost pure
3 times of CuNCs illustrate that GC nanogel can greatly enhance the luminous intensity of system.
Fig. 6 is the fluorescence intensity spectrogram after the GC nanogel composite Cu NCs of various concentration of the present invention, Cong Tuzhong
As can be seen that with the increase of GC nanogel concentration, the fluorescence intensity of CuNCs@GC system, which is presented, first increases becoming of reducing afterwards
Gesture, when GC nanogel concentration is 1 mg/mL, luminous intensity is maximum, schemes in conjunction with the TEM of Fig. 3, select 1.0 mg/mL as
Optium concentration.
Embodiment 2
A kind of zinc ion detection method, step include: identical to be added in 7.4 Illuminant nanometer probe to the pH value of certain volume
The Zn of volume various concentration2+Solution, then zinc ion content detection is carried out after being diluted with ultrapure water.
Specific steps are as follows:
(1) Zn of the 200 μ L of at least five kinds various concentrations is taken2+It is molten that the CuNCs@GC for being 7.4 to pH is added in standard solution immediately
In liquid, being finally diluted to volume with ultrapure water is 1 mL, after being sufficiently mixed 5 min of homogeneous reaction, to fix excitation wavelength as 387
Nm carries out the measurement of photoluminescence emission spectrum, and is recorded in addition various concentration Zn under the excitation wavelength2+CuNCs@GC is molten afterwards
The emissive porwer spectrogram of liquid；
(2) the 200 μ L of sample liquid of above-mentioned non-mark-on and different scalar quantities is separately added into 400 μ L CuNCs@GC solution, finally
1 mL is diluted to by ultrapure water, measures fluorescence intensity under 387 nm excitation wavelengths；Last linear equation calculation Zn content with
And the rate of recovery.
Fig. 7 is the synthesis step and Zn of CuNCs@GC of the invention2+Testing principle, the CuNCs@GC in coherent condition
It can shine；Become colorless and transparent state when pH is adjusted to 7.4, CuNCs@GC solution, shows the nanometer in solution at this time
Dispersity is presented in cluster, cannot shine, Zn2+After addition, the nanocluster that will be dispersed in solution with a kind of effect of crosslinking agent
Reconnect gathers together, and solution shines again.
It carries out interference--free experiments: is ready to first containing common metal ion K+、Na+、Mg2+、Ca2+Aqueous solution, concentration
For 1.5 mM；Contain trace metal ion Fe3+、Cu2+、Cr3+、Mn2+、Ni2+Aqueous solution, concentration be 300 μM；Amino acids
Aqueous solution is 300 μM；Other biological substance DA, UA, AA concentration of aqueous solution is 1.5 mM, and GSH concentration of aqueous solution is 1 mM,
Glucose, lactate concentration of aqueous solution are 3 mM.Take 400 μ L CuNCs@GC solution in the small centrifuge tube of 2 mL, then to
Above-mentioned different 200 μ L of interfering substance is separately added into small centrifuge tube again, 1 mL is finally diluted to by ultrapure water, in 387 nm
Fluorescence intensity is measured under excitation wavelength.Finally be added 30 μ L Zn2+Compare fluorescence intensity.
Fig. 8,9,10 are that 30 μM of Zn is being added in nano-probe of the present invention2+Fluorescence intensity difference comparison afterwards adds
Enter to the fluorescence intensity difference after other interfering substances.It is to be separately added into common metal ion K first+、Na+、Mg2+、Ca2+Concentration
For 1.5 mM and trace meter Fe3+、Cu2+、Cr3+、Mn2+、Ni2+(gold is added in fluorescence intensity difference after concentration is 300 μM
Intensity when metal ion is not added for fluorescence intensity-after belonging to ion)；Followed by it is separately added into amino acids (1 glycine, 2
Alanine, 3 phenylalanines, 4 valines, 5 histidines, 6 isoleucines, 7 arginine, 8 aspartic acids, 9 proline, 10 ammonia
Acid, 11 leucines, 12 tyrosine, 13 tryptophans, 14 threonines, 15 asparagines, 16 glutamic acid, 17 methionines) concentration
It is the fluorescence intensity difference after 300 μM；It is finally to be separately added into other biological substance DA, UA, AA as 1.5 mM, GSH 1
MM, glucose, lactate are the fluorescence intensity difference after 3 mM.It can be seen that the nano-probe is to Zn from three figures2+Inspection
Survey has preferable selectivity.
The Illuminant nanometer probe that the present invention constructs is to detect in water and the zinc ion of food, biological field.Detection side
Method is the nano-probe based on " luminous " mode, and detection device is molecular fluorescence spectroscopy instrument (Cary Eclipse).
Figure 11 is that various concentration Zn is added to the CuNCs@GC solution in embodiment 22+Fluorescence intensity change spectrogram afterwards,
Wherein Figure 11 A is the fluorescence intensity change for the CuNCs@GC solution in embodiment 2 being added the Zn2+ standard liquid that concentration is 0-750 μM
Spectrogram, Figure 11 B are fluorescence intensity difference (I-I0) and various concentration Zn2+Gained is respectively in low strength range after standard liquid mapping
(I0 is CuNCs@GC to the good linear relational graph presented in (1.5 μM -100 μM) and high concentration range (100 μM -750 μM)
The fluorescence intensity of solution, I are that various concentration Zn is added in CuNCs@GC solution2+Fluorescence intensity after standard liquid).It can from Figure 11 A
To find out, with Zn2+Concentration gradually increases in the range of 0.1 μM -750 μM, and the fluorescence intensity of CuNCs@GC solution is gradually
Increase, the data obtained is mapped, finds the Zn being added2+Concentration within the scope of 1.5 μM -100 μM and 100 μM -750 μM
With poor (the addition Zn of fluorescence intensity2+Fluorescence intensity-afterwards is without addition Zn2+When fluorescence intensity) linear relationship is presented respectively, from
As can be seen that corresponding linear equation is in Figure 11 BΔI 1 = 0.38c 1 + 9.79(R2=0.9943) andΔI 2 =
0.02c 2 + 46.79(R2=0.9859).Show that the nano-probe can delicately detect Zn2+Concentration.
Embodiment 3
A kind of Zinc in Foods ion detection method, step include:
Before detecting to food samples, pre-treatment is carried out to milk, beer and tealeaves using micro-wave digestion, specific practice is just
Be three kinds of food are taken it is a certain amount of be put into polyfluortetraethylene pipe, be added 5 mL nitric acid and 5 mL H2O2On electric boiling plate
30 min of pre- resolution, are then placed in microwave dissolver and are cleared up according to the program set, is finally settled to ultrapure water
100 mL.
Every kind of sample carries out Zn content all in accordance with non-mark-on, 0.05 mM of mark-on, 0.15 mM of mark-on, 0.45 mM of mark-on
Measurement.
The 200 μ L of sample liquid of above-mentioned non-mark-on and different scalar quantities is separately added into 400 μ L CuNCs@GC solution, most
1 mL is diluted to by ultrapure water afterwards, measures fluorescence intensity under 387 nm excitation wavelengths.Most afterwards through the linear equation in embodiment 2
Calculate Zn content and the rate of recovery.
Table 1 is that the zinc ion detection method of embodiment 3 and inductively coupled plasma body (ICP) method measure milk, beer, tea
The result of Zn content in leaf compares.
The result table of the Zn content of the different detection method detections of table 1
As can be seen from Table 1, the Zn content in food measured using the Illuminant nanometer probe is measured zinc with conventional method ICP and contained
Magnitude difference is little, illustrates that the Illuminant nanometer probe has good practicability, can substitute the traditional detection side of very complicated
Method.
Embodiment 4
Zn in a kind of living biological cell2+Imaging method, step include:
Carrying out living biological cell imaging experiment, to do is to cytotoxicity experiments before, are assessed using MTT experiment prepared
The cytotoxicity of nano-probe.Specific implementation process are as follows: A549 cell culture is in 96 porocyte culture plates first, final densities
For 8000 cell/plates, be then added into each tissue culture plate different volumes CuNCs@GC solution (1,2.5,5,
7.5,10,15 μ L) after in 37 DEG C, 5% CO2Under conditions of be incubated for 24 h；Then it is added again into each tissue culture plate
MTT(ultimate density is 5 mg/mL) 4 h are cultivated under similarity condition, there is Crystallization in the DMSO solution of 150 μ L, with thin
Born of the same parents' counter observes above-mentioned crystal and calculates cell survival rate in the absorbance (OD) of 570 nm.10 μ L are mixed with 500 μM
Zn2+CuNCs@GC solution be added to liquid-transfering gun in the cell culture medium of 1 mL, in confocal fluorescent after 30 min of cell incubation
Microscopically observation.
Figure 12 is the fluorescence probe based on CuNCs@GC nanocomposite for detecting in A549 living cells obtained by Zn2+
Confocal fluorescence image, the nano-probe is quick on the draw as seen from the figure, can be realized organism living cells Zn2+Quick spirit
Quick detection.
The present invention can be summarized with others without prejudice to the concrete form of spirit or essential characteristics of the invention.Therefore, nothing
By from the point of view of which point, the embodiment above of the invention can only all be considered the description of the invention and cannot limit this hair
Bright, claims indicate the scope of the present invention, and above-mentioned explanation does not point out the scope of the present invention, therefore, with this
Any change in the comparable meaning and scope of claims of invention is all considered as including in claim of the invention
In the range of book.
Claims (6)
1. the preparation method that Illuminant nanometer probe is used in a kind of detection of zinc ion, which is characterized in that step includes:
(1) ethoxy chitosan and ultrapure water are subjected to water-bath, until ethoxy chitosan is completely dissolved,
Ethoxy chitosan nanogel stock solution is obtained, placement is cooled to room temperature；The deacetylated shell of ethoxy after cooling is more
The ethoxy chitosan nanogel that sugared nanogel stock solution is diluted to 0.5-2.0 mg/mL with ultrapure water is molten
Liquid；
(2) the ethoxy chitosan nanogel solution for obtaining step (1) and cysteine ultrasonic mixing extremely dissolve,
Add CuSO4·5H2O solution, 30 min of gentle agitation obtain zinc ion detection Illuminant nanometer probe CuNCs@GC solution.
2. preparation method as described in claim 1, which is characterized in that ethoxy chitosan described in step (1) and
The ratio of ultrapure water is (10-25) mg:(1-10) mL.
3. preparation method as described in claim 1, which is characterized in that ethoxy chitosan described in step (2) is received
Rice gel solution, cysteine and CuSO4·5H2The ratio of O solution is (2.5-4.5) mL: (0.0121-0.0605) g:
(0.5-2.5) mL.
4. a kind of zinc ion detection method, which is characterized in that it is 7.4 as right is wanted that step, which includes: to the pH value of certain volume,
The Zn of same volume various concentration is added in CuNCs@GC solution described in asking 12+Solution, then carried out after being diluted with ultrapure water zinc from
Sub- content detection.
5. detection method as claimed in claim 4, which is characterized in that step includes:
(1) Zn of the 200 μ L of at least five kinds various concentrations is taken2+Standard solution, being added to volume immediately is 400 μ L, 7.4 pH
CuNCs@GC solution in, finally with ultrapure water be diluted to volume be 1 mL, after being sufficiently mixed 5 min of homogeneous reaction, with fixation
Excitation wavelength is the measurement that 387 nm carry out photoluminescence emission spectrum, and is recorded under the excitation wavelength and various concentration is added
Zn2+The emissive porwer spectrogram of CuNCs@GC solution afterwards；
(2) the 200 μ L of sample liquid of above-mentioned non-mark-on and different scalar quantities is separately added into 400 μ L CuNCs@GC solution, finally
1 mL is diluted to by ultrapure water, measures fluorescence intensity under 387 nm excitation wavelengths；Last linear equation calculation Zn content with
And the rate of recovery.
6. detection method as claimed in claim 4, which is characterized in that for the detection of Zinc in Foods ion, step includes:
(1) pre-treatment is carried out to milk, beer or tealeaves using micro-wave digestion, obtains sample liquid；
(2) sample liquid carries out the survey of Zn content according to non-mark-on, 0.05 mM of mark-on, 0.15 mM of mark-on, 0.45 mM of mark-on
It is fixed, specifically: the 200 μ L of sample liquid of non-mark-on and different scalar quantities is separately added into CuNCs@GC solution, finally by ultrapure water
It is diluted to 1 mL, measures fluorescence intensity under 387 nm excitation wavelengths；Last linear equation calculation Zn content and the rate of recovery.
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