CN110487858A - A kind of electrochemistry ratio sensor detecting peroxynitrite and its preparation and application - Google Patents
A kind of electrochemistry ratio sensor detecting peroxynitrite and its preparation and application Download PDFInfo
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Abstract
The present invention provides a kind of electrochemistry ratio sensors for detecting peroxynitrite, are to disperse the organic poromerics Pt-CMP of platinum/conjugation in the acetum of chitosan, and being made into Pt-CMP concentration is 0.2 ~ 10 mgmL‑1Dispersion liquid；And dispersant liquid drop is coated in pretreated bare glassy carbon electrode, it dries at room temperature, modified electrode Pt-CMP-CTS/GCE obtained is electrochemistry ratio sensor.Using modified electrode Pt-CMP-CTS/GCE electrode as sensing platform, with peroxynitrite and the potassium ferricyanide for two kinds of active materials.In the case where operating potential is -0.2 ~ 1.0V, it is scanned with differential pulse voltametry；In specific ONOO‑Concentration range in, ONOO‑Response current (IONOO ‑) and K3[Fe(CN)6] response current (IFe) the ratio between and ONOO‑Concentration is in good linear relationship.
Description
Technical field
The present invention relates to a kind of preparation methods of electrochemistry ratio sensor for detecting peroxynitrite.
Background technique
Peroxynitrite (ONOO-) it is by nitric oxide free radical (NO•) and superoxide anion (O2 •-) quickly anti-
It should be formed, show extensive biochemical reaction, include (a) nitroprotein (tyrosine residue), carbohydrate and nucleic acid,
(b) lipid oxide, sulfydryl, the center Fe/S and Zn/S, (c) convert ferrihemoglobin for oxyhemoglobin.Since it is wide
General reactivity, ONOO-Related with several pathological conditions, such as acute ischemia reperfusion injury, arthritis is carcinogenic, alzheimer '
Mos disease and Parkinson's disease and AIDS etc..Therefore, more and more scholars are dedicated to the inspection of peroxynitrite
It surveys and studies.The reported detection method about peroxynitrite includes the oxidation of fluorescence probe, chemiluminescence and
Immunohistochemistry.However, due to its intrinsic complexity, these methods are all dfficult to apply to real-time quantization.
For the traditional sensors exported with mono signal, signal strength is difficult to measure, because it is highly susceptible to inherence
Or the influence of external factor, such as sensor concentration, environmental condition and instrument efficiency.Usage rate dual signal strategy can be effective
Ground overcomes this limitation.Electrochemistry ratio sensor containing different electroactive materials is exported with dual signal, and can be led to
It crosses and measures the ratio of the current peak intensity of the dual oxide under different oxidation-reduction potentials to determine analyte, improve electrochemistry biography
The accuracy and sensitivity of sense.Therefore, sensitive, convenient and fast ONOO is developed-Electrochemistry ratio sensor is just particularly important.
With the development of nanotechnology, nano material, especially noble metal nano particles, in terms of improving sensor performance
It plays a crucial role.Since it is with special surface nature, have more compared with the noble metal electrode of stock size
Good catalysis characteristics.It is conjugated organic poromerics or is conjugated microporous polymer (CMP) as a kind of by the three of conjugate unit building
Multi-dimensional polymer network skeleton presents excellent porosity, stable skeleton structure and diversified function, it is shown that in crowd
Multi-field application value and bright prospects.But poor dispersibility significantly limits its application in water.
Summary of the invention
The object of the present invention is to provide a kind of preparation sides of electrochemistry ratio sensor for detecting peroxynitrite
Method.
One, the preparation of peroxynitrite electrochemistry ratio sensor
(1) preparation of the organic poromerics of platinum/conjugation (Pt-CMP): by reactant H2PtCl6, surfactant Qula it is logical-
100, hydrochloric acid is mixed and is dissolved under ultrasound, is added reducing agent ascorbic acid (AA), uses water-bath by solution under ultrasonic treatment
Mixture is heated to 30 ~ 90 DEG C and keeps 3 ~ 8h；It is cooled to room temperature after reaction, centrifuge washing 3 ~ 7 times；Product through drying,
Grinding, obtains precious metals pt；Then precious metals pt is mixed with the organic poromerics (CMP) of conjugation organic micro- to get platinum/conjugation
Porous materials (Pt-CMP).
H2PtCl6, surfactant Qula is logical -100, molar ratio of hydrochloric acid are as follows: 1:2:203 ~1:8:104。
The dosage of reducing agent ascorbic acid (AA) is reactant (H2PtCl6) 4 ~ 9 times of mole.
Precious metals pt and the mass ratio for being conjugated organic poromerics (CMP) are 1:1 ~ 1:8.
(2) preparation of electrochemistry ratio sensor: Pt-CMP prepared by step (1) is dispersed in the acetum of chitosan
In, being made into Pt-CMP concentration is 0.2 ~ 10 mgmL-1Dispersion liquid；And dispersant liquid drop is coated in pretreated naked glass carbon electricity
It on extremely, dries at room temperature, modified electrode Pt-CMP-CTS/GCE obtained is electrochemistry ratio sensor.
In the acetum of chitosan, the percentage by volume of chitosan is 0.01 ~ 0.3%, and the percentage by volume of acetic acid is 0.
1~1%。
On modified electrode (Pt-CMP-CTS/GCE), organic poromerics/chitosan (Pt-CMP-CTS) coating of platinum/conjugation
With a thickness of 150 ~ 1500 nm.
Two, the performance of electrochemistry ratio sensor
Fig. 1 be Different electrodes (bare electrode GCE(a), modified electrode CMP/GCE(b), CTS/GCE(c), CMP-CTS/GCE(d),
Pt/GCE(e) and Pt-CMP-CTS/GCE(f) contain 6.11 × 10-6 The ONOO of M- With 0.5mM K3[Fe(CN)6]
PBS(pH=7.68 0.2M) in differential pulse volt-ampere curve.It will be noted from fig. 1 that occurring two oxygen on six curves
Change peak, wherein 0.8V or so is ONOO-Oxidation peak, 0.2V or so correspond to K3[Fe(CN)6] oxidation.This explanation is above
Two electrochemical signals can be generated on electrode.In comparison, ONOO-It is bent in modified electrode Pt-CMP-CTS/GCE(Fig. 1 f
Line) on the obvious highest of electrochemical response signal, and it is most negative to aoxidize spike potential, illustrates that the organic poromerics/shell of platinum/conjugation is poly-
Sugared (Pt-CMP-CTS) is to ONOO-Oxidation there is significant promote and catalytic action.
Fig. 2 is ONOO of the modified electrode Pt-CMP-CTS/GCE to various concentration- The differential pulse figure (A, C) of detection and
ONOO- Concentration and ONOO-Response current and K3[Fe(CN)6] the ratio between response current between linear relationship chart (B, D).
With ONOO it can be seen from figure A and C-The increase of concentration, ONOO-Oxidation peak current can constantly increase, and K3[Fe(CN)6]
Oxidation peak current can constantly reduce.By figure B, D it is found that ONOO-The range of linearity be divided into two sections: work as ONOO-Concentration 3.09 ×
10-10~8.32×10-8When within the scope of M, ONOO-Response current and K3[Fe(CN)6] the ratio between response current (IONOO -/IFe)
With ONOO-Concentration is in good linear relationship, equation of linear regression are as follows: IONOO -/ IFe =6.81 [ONOO-](μM)+
0.0899, R2=0.9990.And work as ONOO-Concentration is 2.19 × 10-7~1.20×10-4When within the scope of M, ONOO-Response current
And K3[Fe(CN)6] the ratio between response current (IONOO -/IFe) and ONOO-Concentration is also in good linear relationship, linear regression
Equation are as follows: IONOO -/IFe =0.00538 [ONOO-] (μM)+0.673, R2=0.9969.It follows that ONOO-That detects is linear
Range is 3.09 × 10-10~1.20×10-4 M, detection are limited to 1.03 × 10-10M。
In conclusion the present invention has the advantage that compared with prior art
1, the present invention utilizes ONOO-With K3[Fe(CN)6] dual signal ratiometric must detect ONOO- Electrochemistry ratio sensing
Device, the sensor have the characteristics that detection range is wide, detection limit is low, detection process is simple, high sensitivity；
2, electrochemistry ratio sensor prepared by the present invention is to ONOO-There are sensitive electrochemical response, strong anti-interference performance, stabilization
Property and favorable reproducibility, can be measured in real time；
3, preparation process of the present invention is simple, low in cost, easy to operate, can be used for a long time.
Detailed description of the invention
Fig. 1 is that different modifying electrode is containing 6.11 × 10-6 The ONOO of M- With 0.5mM K3[Fe(CN)6] 0.2M
PBS(pH=7.68) in differential pulse volt-ampere curve.
Fig. 2 is ONOO of the Pt-CMP-CTS/GCE to various concentration- The differential pulse figure and ONOO of detection- Concentration with
ONOO-Response current and K3[Fe(CN)6] the ratio between response current between linear relationship chart.
Specific embodiment
Below by specific embodiment to the system of the electrochemistry ratio sensor of present invention detection peroxynitrite
It is standby to be described further.
The preparation of embodiment 1, electrochemistry ratio sensor
(1) preparation of the organic poromerics of platinum/conjugation (Pt-CMP): by reactant H2PtCl6, surfactant Qula it is logical-
100, hydrochloric acid is mixed and is dissolved under ultrasound with the molar ratio of 1:3:5000；0.0013 times of reactant integral molar quantity anti-bad is added
Solution mixture is heated to 45 DEG C using water-bath under ultrasonic treatment and keeps 5h by hematic acid (AA)；It is cooled to after reaction
Room temperature, centrifuge washing 4 times；Product is mixed after drying, grinding with the organic poromerics (CMP) of conjugation with the mass ratio of 1:3, is obtained
The organic poromerics of platinum/conjugation (Pt-CMP).
(2) glass-carbon electrode successively the pretreatment of glass-carbon electrode: is used to the aluminum oxide suspension throwing of 0.30 μm, 0.05 μm
Light is at mirror surface, then successively after ethyl alcohol that volume fraction is 95 %, secondary distilled water ultrasonic cleaning, the glass carbon electricity that obtains that treated
Pole；It then is three-electrode system to electrode, saturated calomel electrode for reference electrode by working electrode, platinum column of glass-carbon electrode,
In the 0.1M potassium chloride electrolyte solution containing 1.0mM potassium ferricyanide probe molecule, carrying out cyclic voltammetry scan, (sweeping speed is
50mV/s), finally electrode is taken out and is rinsed and dried up with secondary distilled water；
(3) it the preparation of electrochemistry ratio sensor: disperses Pt-CMP prepared by step (1) in chitosan-acetic acid solution and is made into
Concentration is 1 mgmL-1Dispersion liquid, and drop coating is dried at room temperature in pretreated bare glassy carbon electrode, and modified electrode is made
Pt-CMP-CTS/GCE, i.e. electrochemistry ratio sensor.In chitosan-acetic acid solution, the percentage by volume of chitosan is 0.1%,
The percentage by volume of acetic acid is 0.5%.On modified electrode (Pt-CMP-CTS/GCE), the organic poromerics/chitosan of platinum/conjugation
(Pt-CMP-CTS) coating with a thickness of 1000 nm.
ONOO in embodiment 2, electrochemistry ratio sensor (Pt-CMP-CTS/GCE) detection solution-Concentration
(1) preparation of sample solution: by 1.3M H under conditions of ice bath and stirring2O21.0M is sequentially added with 0.1M HCl
NaNO2In.Secondly the pH of solution is adjusted to 3, continues stirring 5 minutes.Then same volume 7M NaOH is added.Next,
0.70g MnO is added2Powder is to remove excessive H2O2.Product is stored at 4 DEG C, is existed by ultraviolet-visible absorption spectroscopy
302.0 nm measure its concentration.By the ONOO of the known concentration-Solution is diluted to 3.51 × 10-9 M。
(2) ONOO in solution-The detection of concentration: using the modified electrode Pt-CMP-CTS/GCE of embodiment preparation as work electricity
It is that reference electrode forms three-electrode system that pole, platinum column, which are used as to electrode, saturated calomel electrode, and the sample solution of above-mentioned preparation is added
In the phosphate buffer solution for entering pH=7.68 0.2M containing the potassium ferricyanide, and with this as electrolyte, wherein ONOO-For inspection
Survey object, the potassium ferricyanide is reference element (concentration is 0.1 ~ 3mM)；In the case where operating potential is -0.2 ~ 1.0V, with differential pulse volt-ampere
Method is scanned.According to ONOO-Response current and K3[Fe(CN)6] the ratio between response current (IONOO -/ IFe) and ONOO-Concentration
Equation of linear regression are as follows: IONOO -/ IFe =6.81 [ONOO-] (μM)+0.0899 calculate ONOO-Concentration.Testing result:
ONOO in sample solution-Concentration is 3.48 × 10-9 M.
ONOO in embodiment 3, electrochemistry ratio sensor (Pt-CMP-CTS/GCE) detection solution-Concentration
(1) preparation of sample solution: by 1.3M H under conditions of ice bath and stirring2O21.0M is sequentially added with 0.1M HCl
NaNO2In.Secondly the pH of solution is adjusted to 3, continues stirring 5 minutes.Then same volume 7M NaOH is added.Next, plus
Enter 0.70g MnO2Powder is to remove excessive H2O2.Product is stored at 4 DEG C, by ultraviolet-visible absorption spectroscopy 302.0
Nm measures its concentration.By the ONOO of the known concentration-Solution is diluted to 5.68 × 10-5M。
(2) ONOO in solution-The detection of concentration: using the modified electrode Pt-CMP-CTS/GCE of embodiment preparation as work electricity
It is that reference electrode forms three-electrode system that pole, platinum column, which are used as to electrode, saturated calomel electrode, and the sample solution of above-mentioned preparation is added
In the phosphate buffer solution for entering pH=7.68 0.2M containing the potassium ferricyanide, and with this as electrolyte, wherein ONOO-For inspection
Survey object, the potassium ferricyanide is reference element (concentration is 0.1 ~ 3mM)；In the case where operating potential is -0.2 ~ 1.0V, with differential pulse volt-ampere
Method is scanned.According to ONOO-Response current and K3[Fe(CN)6] the ratio between response current (IONOO -/ IFe) and ONOO-Concentration
Equation of linear regression are as follows: IONOO -/ IFe =0.00538 [ONOO-] (μM)+0.673 calculate ONOO-Concentration.Detection
As a result: ONOO in sample solution-Concentration is 5.70 × 10-5M。
Instrument used in implementation process of the present invention and drug:
CHI 660C electrochemical workstation (Shanghai Chen Hua instrument company) is used to carry out the experiment of differential pulse voltametry, quartz ampoule
Automatically dual pure water distiller (1810B, Asian-Pacific Glass-Tech. Co., Shanghai City) is used to steam ultrapure water heated type.Electronic balance (north
Jing Saiduolisi Instrument Ltd.), for weighing drug.Ultrasonic cleaner (Kunshan Ultrasonic Instruments Co., Ltd.).Three
(0.30 μm, 0.05 μm, Shanghai Chen Hua instrument reagent company) of Al 2 O polishing powder for handling glass-carbon electrode.It is saturated calomel ginseng
Than electrode, platinum to electrode, potassium chloride, sodium dihydrogen phosphate, disodium hydrogen phosphate (Xi'an chemical reagent factory)；(Shanghai Ji is extremely for chloroplatinic acid
Biochemical technology Co., Ltd)；Qula leads to -100(Aladdin)；Ascorbic acid (upper marine Qin's chemical reagent Co., Ltd)；Iron cyanogen
Change potassium (development in science and technology Co., Ltd is recovered in Tianjin).
Claims (9)
1. a kind of preparation method for the electrochemistry ratio sensor for detecting peroxynitrite, comprises the following steps that:
(1) preparation of the organic poromerics Pt-CMP of platinum/conjugation: by reactant H2PtCl6, surfactant Qula lead to -100, salt
Acid is mixed and is dissolved under ultrasound, and reducing agent ascorbic acid is added, and is heated solution mixture using water-bath under ultrasonic treatment
To 30 ~ 90 DEG C and keep 3 ~ 8h；It is cooled to room temperature after reaction, centrifuge washing 3 ~ 7 times；Product obtains expensive through drying, grinding
Pt metal；Then precious metals pt is mixed with organic poromerics CMP is conjugated to get the organic poromerics Pt-CMP of platinum/conjugation；
(2) preparation of electrochemistry ratio sensor: dispersing Pt-CMP prepared by step (1) in the acetum of chitosan,
Being made into Pt-CMP concentration is 0.2 ~ 10 mgmL-1Dispersion liquid；And dispersant liquid drop is coated in pretreated bare glassy carbon electrode
On, it dries at room temperature, modified electrode Pt-CMP-CTS/GCE obtained is electrochemistry ratio sensor.
2. such as a kind of preparation method of the electrochemistry ratio sensor of detection peroxynitrite of claim 1, feature
Be: in step (1), reducing agent ascorbic acid dosage is reactant H2PtCl64 ~ 9 times of mole.
3. such as a kind of preparation method of the electrochemistry ratio sensor of detection peroxynitrite of claim 1, feature
Be: in step (1), precious metals pt and the mass ratio for being conjugated organic poromerics CMP are 1:1 ~ 1:8.
4. such as a kind of preparation method of the electrochemistry ratio sensor of detection peroxynitrite of claim 1, feature
Be: in the chitosan-acetic acid solution of step (2), the percentage by volume of chitosan is 0.01 ~ 0.3%, the percentage by volume of acetic acid
It is 0. 1 ~ 1%.
5. such as a kind of preparation method of the electrochemistry ratio sensor of detection peroxynitrite of claim 1, feature
Be: on the modified electrode Pt-CMP-CTS/GCE of step (2), the organic poromerics of platinum/conjugation/chitosan Pt-CMP-CTS is applied
Layer with a thickness of 150 ~ 1500 nm.
6. the electrochemistry ratio sensor of the detection peroxynitrite of method preparation as described in claim 1 is molten in detection
ONOO in liquid−The application of concentration.
7. the electrochemistry ratio sensor of the detection peroxynitrite of method preparation as claimed in claim 6 is molten in detection
ONOO in liquid−The application of concentration, it is characterised in that: using Pt-CMP-CTS/GCE as working electrode, platinum column be to electrode, saturation it is sweet
Mercury electrode is reference electrode, forms three-electrode system；To contain ONOO-It is slow with the phosphate of pH=7.68 0.2M of the potassium ferricyanide
Solution is rushed as electrolyte to be scanned in the case where operating potential is -0.2 ~ 1.0V with differential pulse voltametry；Specific
ONOO-Concentration range in, ONOO-Response current IONOO -And K3[Fe(CN)6] response current IFeThe ratio between IONOO -/IFeWith
ONOO-Concentration is in good linear relationship.
8. the electrochemistry ratio sensor of the detection peroxynitrite of method preparation as claimed in claim 7 is molten in detection
ONOO in liquid−The application of concentration, it is characterised in that: work as ONOO-Concentration 3.09 × 10-10~8.32×10-8When M, ONOO-'s
Response current and K3[Fe(CN)6] the ratio between response current IONOO -/IFeWith ONOO-Concentration is in good linear relationship, linear
Regression equation are as follows: IONOO -/ IFe =6.81 [ONOO-] (μM)+0.0899, R2=0.9990。
9. the electrochemistry ratio sensor of the detection peroxynitrite of method preparation as claimed in claim 7 is molten in detection
ONOO in liquid−The application of concentration, it is characterised in that: work as ONOO-Concentration is 2.19 × 10-7~1.20×10-4When within the scope of M,
ONOO-Response current and K3[Fe(CN)6] the ratio between response current IONOO -/IFeWith ONOO-Concentration is in good linear relationship,
Its equation of linear regression are as follows: IONOO -/ IFe =0.00538 [ONOO-] (μM)+0.673, R2=0.9969。
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109187687A (en) * | 2018-10-22 | 2019-01-11 | 西北师范大学 | It is conjugated the preparation of organic poromerics modified electrode and the application as peroxynitrite electrochemical sensor |
| CN110066399A (en) * | 2019-04-23 | 2019-07-30 | 西北师范大学 | A kind of preparation and its application of chitosan-modified microporous polymer nano material |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109187687A (en) * | 2018-10-22 | 2019-01-11 | 西北师范大学 | It is conjugated the preparation of organic poromerics modified electrode and the application as peroxynitrite electrochemical sensor |
| CN110066399A (en) * | 2019-04-23 | 2019-07-30 | 西北师范大学 | A kind of preparation and its application of chitosan-modified microporous polymer nano material |
Non-Patent Citations (3)
| Title |
|---|
| FEIYUE LIU等: "Real-time Monitoring of Peroxynitrite (ONOO-) in the Rat Brain by Developing a Ratiometric Electrochemical Biosensor", 《ANALYST》 * |
| JUJIE REN等: "A Novel Superoxide Anion Radical Nonenzymatic Electrochemical Sensor Based on PtRuCu Ternary Alloy Nanoparticles/Graphene Composite Modified Electrode", 《NANO》 * |
| 任聚杰等: "基于氯化血红素复合材料的过氧亚硝酸阴离子", 《分析化学研究报告》 * |
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