CN109254053A - A kind of preparation method and application of environmental estrogens electro-chemical analyzing sensor - Google Patents
A kind of preparation method and application of environmental estrogens electro-chemical analyzing sensor Download PDFInfo
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Abstract
The invention discloses a kind of preparation methods of environmental estrogens electro-chemical analyzing sensor.Belong to Nano-function thin films and biosensory analysis technology field.The present invention is prepared for cobalt hydroxide nano-chip arrays can disposably throw first on electrode, utilize its big specific surface area and high activity hydroxy functional group, and the amido functional group of poly-dopamine, using the method for growth in situ, directly it is prepared for the poly-dopamine film containing electron mediator in succession and using environmental estrogens as the molecularly imprinted polymer of template molecule on cobalt hydroxide nano-chip arrays in succession, after by template molecule elution, the position of template molecule originally has become hole, that is the molecularly imprinted polymer of eluted template molecule, thus, a kind of environmental estrogens electro-chemical analyzing sensor just prepares completion.
Description
Technical field
The present invention relates to a kind of preparation method and applications of electro-chemical analyzing sensor.Belong to Nano-function thin films with
Biosensory analysis technology field.
Background technique
Environmental estrogens are a kind of chemical substances existed in the environment, have the activity of similar organism inner estrogen,
Have the function of simulating estrogen after into human body, by upsetting the normal function of the systems such as human endocrine, immune, nerve
Such as carcinogenic, damage reproductive development system, immune system, a variety of diseases of nervous system are generated, is that a kind of serious harm mankind are strong
The environmental toxin of health.Common environmental estrogens source includes if pesticide (such as organo-chlorine pesticide), secondhand smoke, exhaust gas are (as burnt
The toxic gas that plastic garbage generates), food additives etc..Detection method for environmental estrogens mainly has high-efficient liquid phase color
Spectrometry, gas chromatography, chromatograph-mass spectrometer coupling method, enzyme-linked immunization, radioimmunology etc..But most of detection method instruments
It is valuable, complicated for operation, reviewer has to pass through screw up discipline after, can be detected.The method such as electrochemistry of some simplicity
Analytic approach, though easy to operate, sensitive, quick, the poor specificity of detection.Therefore, it develops as early as possible a kind of quick, highly selective and clever
The method of quick detection environmental estrogens is extremely important to publilc health, and has wide market application prospect.
Molecular imprinting electrochemical sensor has high specific selectivity, excellent stability, excellent reproducibility, wide inspection
Survey range and floor detection limit.Due to the sensor prepare simple, easy to detect, high sensitivity, it is at low cost the advantages that it is extensive
Applied to the fields such as chromatographic isolation, film point, Solid Phase Extraction, medicine controlled releasing, chemical sensitisation.Molecularly imprinted polymer (MIP), also referred to as
It, being capable of specific recognition and the specific target molecule of selective absorption (i.e. template molecule) for " plastics antibody ".Due to molecular engram
Technology has many advantages that, such as organic reagent corrosion resistance, good stability, heat-resisting quantity and preparation are simple.Therefore, in mistake
In several years gone, electroanalysis is caused based on the MIP electrochemical sensor (MIP-ECS) that MIP is combined with electrochemical sensor
The detection of the focus of chemical field, especially small molecule contaminants.However, having in the preparation process of traditional MIP-ECS
The elution of template molecule difficulty, the disadvantages of thickness of blotting membrane is difficult to control, reproducibility is poor, limit molecular engram film and passed in electrochemistry
Application in sensor.These problems, especially molecular engram film thickness are not easy to control to lead to electrochemical sensor sensitivity decrease
And molecular engram film easily falls off from electrode surface the technical problem for causing stability and reproducibility to reduce during elution, limit
The application of MIP_ECS has been made, therefore, has found new molecularly imprinted polymer synthetic method, new molecular engram film electrode is repaired
The combination method of decorations method and molecular engram film and base material, to solve the preparation of MIP-ECS and have using problem important
Research significance and market value.
Summary of the invention
The purpose of the present invention is to provide a kind of high specificity, prepare simple, easy to detect, high sensitivity, at low cost
The preparation method of environmental estrogens electro-chemical analyzing sensor, prepared electrochemical sensing electrode prepare simple, reproducibility
Well, stability is strong, can be used as quick, Sensitive Detection of the electrochemical sensor for environmental estrogens.Based on this purpose, the present invention
It is prepared for cobalt hydroxide nano-chip arrays on electrode can disposably throw first, utilizes its big specific surface area and high activity hydrogen-oxygen
The amido functional group of base functional group and poly-dopamine, using the method for growth in situ, in succession in cobalt hydroxide nano-chip arrays
It is upper to be directly prepared for the poly-dopamine film containing electron mediator in succession and printed by the molecule of template molecule of environmental estrogens
Mark polymer, after by template molecule elution, the position of template molecule originally has become hole, i.e. eluted template molecule
Molecularly imprinted polymer, a kind of environmental estrogens electro-chemical analyzing sensor just prepares completion as a result,.Swash when for female to environment
When element is detected, environmental estrogens electro-chemical analyzing sensor is inserted into solution to be measured, the environment in solution to be measured is female to swash
Element can be adsorbed onto the hole of NIP.Environmental estrogens concentration in solution to be measured is bigger, and it is female to be adsorbed onto environment in the hole of NIP
Hormone is more.When carrying out Electrochemical Detection, the intensity for detecting electric current can be with environmental estrogens in the hole for being adsorbed onto NIP
Increase and become smaller, thus according to the degree that current strength reduces, is capable of the dense of environmental estrogens in qualitative, quantitative solution to be measured
Degree.
The technical solution adopted by the invention is as follows:
1. a kind of preparation method of environmental estrogens electro-chemical analyzing sensor, the environmental estrogens electrochemical analysis is passed
Sensor is by growth in situ on cobalt hydroxide nano-chip arrays electrode Co-nanoarray without template molecule molecularly imprinted polymer NIP
It obtains；The molecularly imprinted polymer that template molecule is free from without template molecule molecularly imprinted polymer NIP；It is described
The molecularly imprinted polymer without containing template molecule be by the MIP of molecularly imprinted polymer containing template molecule by eluted template point
What son obtained；The MIP of molecularly imprinted polymer containing template molecule is the molecularly imprinted polymer containing template molecule；It is described
Template molecule be environmental estrogens；
2. the preparation method of cobalt hydroxide nano-chip arrays electrode Co-nanoarray described in technical solution 1 includes following
Preparation step:
(1) it will can disposably throw electrode and carry out ultrasonic cleaning processing using dilute hydrochloric acid, dehydrated alcohol and deionization respectively, to go
Except the oxide layer and surface impurity that can disposably throw electrode；
(2) 1 ~ 3 mmol Co (NO is weighed3)2With 3 ~ 9 mmol urea CO (NH2)2, it is put into 50 mL beakers, is added 30
ML deionized water stirs to clarify, and is then transferred into 50 mL ptfe autoclaves；
(3) in the disposable solution thrown in the reaction kettle that electrode is put into step (2) for handling step (1) well, 100 ~
It is reacted 9 ~ 12 hours at a temperature of 130 DEG C, cobalt hydroxide nano-chip arrays presoma electrode is prepared；
(4) the cobalt hydroxide nano-chip arrays presoma electrode insertion obtained step (3) contains dopamine, Ammonium Persulfate 98.5 and nitre
In the phosphate buffer solution PBS of sour cobalt, 20 ~ 40 DEG C at a temperature of reaction 4 ~ 6 hours after, take out simultaneously soaked with deionized water
It washes 2 ~ 4 times, cobalt hydroxide nano-chip arrays electrode Co-nanoarray is prepared；
The described disposable electrode of throwing is selected from one of cycle unit: nickel foam, foam copper, pure nickel piece, pure copper sheet, pure cobalt piece,
Pure silicon piece, conductive carbon cloth；
In the phosphate buffer solution PBS containing dopamine, Ammonium Persulfate 98.5 and cobalt nitrate: dopamine concentration is 2 ~ 5
Mg/mL, the concentration of Ammonium Persulfate 98.5 are 3 ~ 8 mg/mL, and the concentration of cobalt nitrate is 0.1 ~ 0.5 mg/mL, phosphate buffer solution PBS
Concentration be 0.1 mol/L, pH value be 7.2 ~ 8.5；
3. the system of the MIP of molecularly imprinted polymer containing template molecule of Co-nanoarray growth in situ described in technical solution 1
Preparation Method includes following preparation step:
(1) 0.25 ~ 0.45mmol template molecule and 3 ~ 5 mmol 2- methacrylic acid MAA are weighed respectively in peace times bottle, are added
Enter 8 ~ 15 mL acetonitriles, 30 min of ultrasound to whole dissolutions；
(2) 15 ~ 25 mmol ethylene glycol dimethacrylate EDMA are added in the solution of step (1), 30 min of ultrasound
To being uniformly mixed, precursor mixed solution is obtained；
(3) Co-nanoarray prepared in technical solution 2 is clipped on Stirring device, the forerunner being inserted into step (2)
In body mixed solution, in N2At a temperature of environment and 20 ~ 40 DEG C of water-bath, with 5 ~ 200 revolutions per seconds of speed Stirring, while with 1
~ 20 drops/sec of speed carries out initiation polymerization to 1 mmol azodiisobutyronitrile AIBN of mixed solution and dripping, in Co-
The MIP of molecularly imprinted polymer containing template molecule of growth in situ is obtained on nanoarray；
4. the system without template molecule molecularly imprinted polymer NIP of Co-nanoarray growth in situ described in technical solution 1
Standby step are as follows: by the molecular engram containing template molecule of growth in situ polymerize on Co-nanoarray obtained in technical solution 3
Object MIP is immersed in eluant, eluent, and template molecule is carried out 5 ~ 20 min of elution at room temperature, is then taken out, no template is obtained
Molecular imprinted polymer NIP；The eluant, eluent is the mixed liquor of formic acid and methanol, wherein the volume ratio of formic acid and methanol
For 9:(1 ~ 5)；
5. the preparation step of environmental estrogens electro-chemical analyzing sensor described in technical solution 1 are as follows: by technical solution 2 ~ 4
In the obtained growth in situ on Co-nanoarray without template molecule molecularly imprinted polymer NIP, embathe 2 with deionized water
It ~ 4 times, dries at room temperature, obtains environmental estrogens electro-chemical analyzing sensor；
6. being applied to using environmental estrogens electro-chemical analyzing sensor prepared by technical solution described in technical solution 1 ~ 5
The detection of environmental estrogens, including following applying step:
(1) standard solution is prepared: preparing the environmental estrogens standard solution of one group of various concentration including blank standard specimen；
(2) working electrode is modified: being working electrode by environmental estrogens electro-chemical analyzing sensor, is prepared in inserting step (1)
Various concentration environmental estrogens standard solution, hatch 10 min after take out, embathed 3 times with deionized water；
(3) working curve is drawn: using saturated calomel electrode electrode as reference electrode, platinum electrode is used as to electrode, with step
(2) the working electrode composition three-electrode system modified, connects electrochemical workstation, 15 mL is successively added in a cell
Phosphate buffer solution PBS；Pass through the current-responsive of the working electrode of Differential Pulse Voltammetry DPV detection assembling；Blank standard specimen
Response current intensity be denoted asI 0, the response current intensity of the environmental estrogens standard solution containing various concentration is denoted asI i, response
The difference that current strength reduces is ΔI = I 0-I i, ΔIWith the mass concentration of environmental estrogens standard solutionCBetween linear pass
Δ is drawn by systemI?CWorking curve；The phosphate buffer solution PBS concentration is 10 mmol/L, pH value 7.4；Described
Parameter setting when DPV is detected are as follows: range and direction are 0 ~ 1 V, and stride is 0.05 V, and the burst length is 0.05 s, when sampling
Between be 0.016 s, the pulse period be 0.5 s；
(4) in sample to be tested environmental estrogens detection: replace the environmental estrogens standard in step (1) molten with sample to be tested
Liquid is detected according to the method in step (2) and (3), the difference DELTA that current strength reduces according to responseIAnd working curve,
Obtain the content of environmental estrogens in sample to be tested；
7. environmental estrogens described in technical solution 1 ~ 6 are one of following environmental estrogens: estradiol, estriol, hexene are female
Phenol, bisphenol-A, nonyl phenol.
Beneficial achievement of the invention
(1) environmental estrogens electro-chemical analyzing sensor preparation of the present invention is simple, easy to operate, realizes to sample
Quickly, sensitive, highly selective detection, and it is at low cost, it can be applied to portable inspectiont, there is market development prospect；
(2) growth in situ molecular engram polymerize the present invention on cobalt hydroxide nano-chip arrays electrode Co-nanoarray for the first time
On the one hand object can grow more, molecularly imprinted polymer more evenly using the big specific surface area of Co-nanoarray, and
Co-nanoarray has excellent electron transmission ability, to greatly improve detection sensitivity；On the other hand, the present invention will be more
When on bar amine in-situ polymerization to cobalt hydroxide nano-chip arrays, creative is doped into cobalt ions as electron mediator, is examining
Electrochemical response electric current is directly generated when survey, allows sensor straight in the buffer solution it is not necessary that other media substance is added
Capable detection is tapped into, to greatly reduce testing cost simultaneously while further decreasing signal background, improving detection sensitivity
It reduces environmental pollution；
(3) present invention is rich in using cobalt hydroxide nano-chip arrays high activity hydroxy functional group and big specific surface area with
Dopamine combines, sufficiently thin poly- more being formed so that dopamine is in cobalt hydroxide nano-chip arrays in situ Polymerization
While bar amine film, on uniform fold to cobalt hydroxide nano-chip arrays, thus for more better polymerizable moleculars in next step
Imprinted polymer carries out place mat；Later using poly-dopamine to the amino being rich on hydroxy functional group and molecularly imprinted polymer
Strong connection function, then dexterously use Co-nanoarray as blender, in molecular engram precursor mixed solution
Immersion stirring is carried out, by control mixing speed, the rate of addition and polymeric reaction temperature of initiators for polymerization, in Co-
The surface nanoarray direct in-situ grows the molecularly imprinted polymer that can control film thickness, on the one hand makes Co-
Nanoarray can secured supporting molecular imprinted polymer, to significantly improve the stability of prepared electrochemical sensor
And reproducibility；On the other hand molecularly imprinted polymer can effectively be controlled in the film forming thickness of electrode surface, solving can not control
Molecular engram film processed is unable to control the technical problem so as to cause poor reproducibility in electrode surface film forming thickness；In addition, more due to
Preparation method of the invention can sufficiently improve molecule print to the in-stiu coating of the effective control and electron mediator of film forming thickness
The sensitivity of electrochemical sensor based on mark and detection limit have important scientific meaning and application value.
Specific embodiment
The preparation of 1 Co-nanoarray of embodiment
(1) it will can disposably throw electrode and carry out ultrasonic cleaning processing using dilute hydrochloric acid, dehydrated alcohol and deionization respectively, to go
Except the oxide layer and surface impurity that can disposably throw electrode；
(2) 1 mmol Co (NO is weighed3)2With 3 mmol urea CO (NH2)2, it is put into 50 mL beakers, 30 mL is added and go
Ionized water stirs to clarify, and is then transferred into 50 mL ptfe autoclaves；
(3) in the disposable solution thrown in the reaction kettle that electrode is put into step (2) for handling step (1) well, 100
It is reacted 12 hours at a temperature of DEG C, cobalt hydroxide nano-chip arrays presoma electrode is prepared；
(4) the cobalt hydroxide nano-chip arrays presoma electrode insertion obtained step (3) contains dopamine, Ammonium Persulfate 98.5 and nitre
In the phosphate buffer solution PBS of sour cobalt, 20 DEG C at a temperature of reaction 4 hours after, take out simultaneously embathed 2 times with deionized water,
Cobalt hydroxide nano-chip arrays electrode Co-nanoarray is prepared；
The electrode therein that can disposably throw is nickel foam；Dopamine concentration is 2 mg/mL, and the concentration of Ammonium Persulfate 98.5 is 3 mg/mL,
The concentration of cobalt nitrate is 0.1 mg/mL, and the concentration of phosphate buffer solution PBS is 0.1 mol/L, pH value 7.2.
The preparation of 2 Co-nanoarray of embodiment
(1) it will can disposably throw electrode and carry out ultrasonic cleaning processing using dilute hydrochloric acid, dehydrated alcohol and deionization respectively, to go
Except the oxide layer and surface impurity that can disposably throw electrode；
(2) 2 mmol Co (NO are weighed3)2With 6 mmol urea CO (NH2)2, it is put into 50 mL beakers, 30 mL is added and go
Ionized water stirs to clarify, and is then transferred into 50 mL ptfe autoclaves；
(3) in the disposable solution thrown in the reaction kettle that electrode is put into step (2) for handling step (1) well, 110
It is reacted 11 hours at a temperature of DEG C, cobalt hydroxide nano-chip arrays presoma electrode is prepared；
(4) the cobalt hydroxide nano-chip arrays presoma electrode insertion obtained step (3) contains dopamine, Ammonium Persulfate 98.5 and nitre
In the phosphate buffer solution PBS of sour cobalt, 30 DEG C at a temperature of reaction 5 hours after, take out simultaneously embathed 3 times with deionized water,
Cobalt hydroxide nano-chip arrays electrode Co-nanoarray is prepared；
The electrode therein that can disposably throw is pure copper sheet；Dopamine concentration is 3.5 mg/mL, and the concentration of Ammonium Persulfate 98.5 is 6.2
Mg/mL, the concentration of cobalt nitrate are 0.3 mg/mL, and the concentration of phosphate buffer solution PBS is 0.1 mol/L, pH value 8.0.
The preparation of 3 Co-nanoarray of embodiment
(1) it will can disposably throw electrode and carry out ultrasonic cleaning processing using dilute hydrochloric acid, dehydrated alcohol and deionization respectively, to go
Except the oxide layer and surface impurity that can disposably throw electrode；
(2) 3 mmol Co (NO are weighed3)2With 9 mmol urea CO (NH2)2, it is put into 50 mL beakers, 30 mL is added and go
Ionized water stirs to clarify, and is then transferred into 50 mL ptfe autoclaves；
(3) in the disposable solution thrown in the reaction kettle that electrode is put into step (2) for handling step (1) well, 130
It is reacted 9 hours at a temperature of DEG C, cobalt hydroxide nano-chip arrays presoma electrode is prepared；
(4) the cobalt hydroxide nano-chip arrays presoma electrode insertion obtained step (3) contains dopamine, Ammonium Persulfate 98.5 and nitre
In the phosphate buffer solution PBS of sour cobalt, 40 DEG C at a temperature of reaction 6 hours after, take out simultaneously embathed 4 times with deionized water,
Cobalt hydroxide nano-chip arrays electrode Co-nanoarray is prepared；
The electrode therein that can disposably throw is conductive carbon cloth；Dopamine concentration is 5 mg/mL, and the concentration of Ammonium Persulfate 98.5 is 8 mg/
ML, the concentration of cobalt nitrate are 0.5 mg/mL, and the concentration of phosphate buffer solution PBS is 0.1 mol/L, pH value 8.5.
The preparation method of 4 environmental estrogens electro-chemical analyzing sensor of embodiment
(1) 0.25 mmol template molecule and 3 mmol 2- methacrylic acid MAA are weighed respectively in peace times bottle, and 8 mL second are added
Nitrile, 30 min of ultrasound to whole dissolutions；
(2) 15 mmol ethylene glycol dimethacrylate EDMA are added in the solution of step (1), 30 min of ultrasound are to mixed
It closes uniformly, obtains precursor mixed solution；
(3) Co-nanoarray prepared in embodiment 1 is clipped on Stirring device, the presoma being inserted into step (2)
In mixed solution, in N2At a temperature of environment and 20 DEG C of water-bath, with 200 revolutions per seconds of speed Stirring, while with 1 drop/sec
Speed carry out initiation polymerization to 1 mmol azodiisobutyronitrile AIBN of mixed solution and dripping, obtained on Co-nanoarray
The MIP of molecularly imprinted polymer containing template molecule of growth in situ；
(4) MIP of molecularly imprinted polymer containing template molecule of growth in situ on Co-nanoarray for obtaining step (3) soaks
Not in eluant, eluent, template molecule is subjected to 5 min of elution at room temperature, is then taken out, obtains no template molecule molecule print
Mark polymer NIP；Continue to be embathed 2 times with deionized water, dry at room temperature, obtains environmental estrogens electrochemical analysis sensing
Device；
Eluant, eluent therein is the mixed liquor of formic acid and methanol, and wherein the volume ratio of formic acid and methanol is 9:1.
The preparation method of 5 environmental estrogens electro-chemical analyzing sensor of embodiment
(1) 0.35mmol template molecule and 4 mmol 2- methacrylic acid MAA are weighed respectively in peace times bottle, and 12 mL second are added
Nitrile, 30 min of ultrasound to whole dissolutions；
(2) 18 mmol ethylene glycol dimethacrylate EDMA are added in the solution of step (1), 30 min of ultrasound are to mixed
It closes uniformly, obtains precursor mixed solution；
(3) Co-nanoarray prepared in technical solution 2 is clipped on Stirring device, the forerunner being inserted into step (2)
In body mixed solution, in N2At a temperature of environment and 30 DEG C of water-bath, with 60 revolutions per seconds of speed Stirring, while with 10 drops/sec
Speed carry out initiation polymerization to 1 mmol azodiisobutyronitrile AIBN of mixed solution and dripping, obtained on Co-nanoarray
The MIP of molecularly imprinted polymer containing template molecule of growth in situ；
(4) MIP of molecularly imprinted polymer containing template molecule of growth in situ on Co-nanoarray for obtaining step (3) soaks
Not in eluant, eluent, template molecule is subjected to 10 min of elution at room temperature, is then taken out, obtains no template molecule molecule print
Mark polymer NIP；Continue to be embathed 3 times with deionized water, dry at room temperature, obtains environmental estrogens electrochemical analysis sensing
Device；
Eluant, eluent therein is the mixed liquor of formic acid and methanol, and wherein the volume ratio of formic acid and methanol is 9:3.
The preparation method of 6 environmental estrogens electro-chemical analyzing sensor of embodiment
(1) 0.45mmol template molecule and 5 mmol 2- methacrylic acid MAA are weighed respectively in peace times bottle, and 15 mL second are added
Nitrile, 30 min of ultrasound to whole dissolutions；
(2) 25 mmol ethylene glycol dimethacrylate EDMA are added in the solution of step (1), 30 min of ultrasound are to mixed
It closes uniformly, obtains precursor mixed solution；
(3) Co-nanoarray prepared in technical solution 2 is clipped on Stirring device, the forerunner being inserted into step (2)
In body mixed solution, in N2At a temperature of environment and 40 DEG C of water-bath, with 5 revolutions per seconds of speed Stirring, while with 20 drops/sec
Speed carry out initiation polymerization to 1 mmol azodiisobutyronitrile AIBN of mixed solution and dripping, obtained on Co-nanoarray
The MIP of molecularly imprinted polymer containing template molecule of growth in situ；
(4) MIP of molecularly imprinted polymer containing template molecule of growth in situ on Co-nanoarray for obtaining step (3) soaks
Not in eluant, eluent, template molecule is subjected to 20 min of elution at room temperature, is then taken out, obtains no template molecule molecule print
Mark polymer NIP；Continue to be embathed 4 times with deionized water, dry at room temperature, obtains environmental estrogens electrochemical analysis sensing
Device；
Eluant, eluent therein is the mixed liquor of formic acid and methanol, and wherein the volume ratio of formic acid and methanol is 9:5.
The environmental estrogens sensor of 7 embodiment 1 ~ 6 of embodiment preparation, applied to the detection of environmental estrogens, step
It is as follows:
(1) standard solution is prepared: preparing the environmental estrogens standard solution of one group of various concentration including blank standard specimen；
(2) working electrode is modified: being working electrode by environmental estrogens electro-chemical analyzing sensor, is prepared in inserting step (1)
Various concentration environmental estrogens standard solution, hatch 10 min after take out, embathed 3 times with deionized water；
(3) working curve is drawn: using saturated calomel electrode electrode as reference electrode, platinum electrode is used as to electrode, with step
(2) the working electrode composition three-electrode system modified, connects electrochemical workstation, 15 mL is successively added in a cell
PBS；Pass through the current-responsive of the working electrode of Differential Pulse Voltammetry DPV detection assembling；The response current intensity of blank standard specimen
It is denoted asI 0, the response current intensity of the environmental estrogens standard solution containing various concentration is denoted asI i, response current strength reduction
Difference is ΔI = I 0-I i, ΔIWith the mass concentration of environmental estrogens standard solutionCBetween it is linear, draw ΔI?C
Working curve；The PBS is the phosphate buffer solution of 10 mmol/L, and the pH value of the phosphate buffer solution is
7.4；Parameter setting when the described DPV detection are as follows: range and direction are 0 ~ 1 V, and stride is 0.05 V, burst length 0.05
S, sampling time are 0.016 s, and the pulse period is 0.5 s；
(4) in sample to be tested environmental estrogens detection: replace the environmental estrogens standard in step (1) molten with sample to be tested
Liquid is detected according to the method in step (2) and (3), the difference DELTA that current strength reduces according to responseIAnd working curve,
Obtain the content of environmental estrogens in sample to be tested.
The environmental estrogens sensor of 8 embodiment 1 ~ 6 of embodiment preparation, is applied to according to the detecting step of embodiment 7
The detection of varying environment estrogen, the range of linearity and detection limit are shown in Table 1:
The detection technique index of 1 environmental estrogens of table
The detection of environmental estrogens in 9 milk of embodiment
Milk sample is accurately pipetted, the environmental estrogens standard solution of certain mass concentration is added, environmental estrogens not to be added
Milk be blank, carry out recovery testu, with embodiment 1 ~ 6 prepare environmental estrogens sensor, according to embodiment 7
Step is detected, and the rate of recovery of environmental estrogens in milk sample is measured, and testing result is shown in Table 2:
The testing result of environmental estrogens in 2 milk of table
2 testing result of table it is found that the relative standard deviation (RSD) of result less than 3.0 %, average recovery rate is 98.0 ~
101.6%, show that the present invention can be used for the detection of a variety of environmental estrogens in milk, the high sensitivity of method, high specificity, knot
Fruit is accurate and reliable.
The detection of environmental estrogens in 10 water sample of embodiment
Certain water sample is accurately pipetted, the environmental estrogens standard solution of certain mass concentration is added, environmental estrogens not to be added
Water sample be blank, carry out recovery testu, with embodiment 1 ~ 6 prepare environmental estrogens sensor, according to embodiment 7
Step is detected, and the rate of recovery of environmental estrogens in water sample is measured, and testing result is shown in Table 3:
The testing result of environmental estrogens in 3 water sample of table
For 3 testing result of table it is found that the relative standard deviation (RSD) of result is less than 3.0 %, average recovery rate is 98.4 ~ 102%,
Show that the present invention can be used for the detection of a variety of environmental estrogens in water sample, as a result the high sensitivity of method, high specificity accurately may be used
It leans on.
Claims (7)
1. a kind of preparation method of environmental estrogens electro-chemical analyzing sensor, which is characterized in that the environmental estrogens electricity
Chemical analysis sensor is printed by growth in situ on cobalt hydroxide nano-chip arrays electrode Co-nanoarray without template molecule molecule
Mark polymer NIP is obtained；The molecular engram that template molecule is free from without template molecule molecularly imprinted polymer NIP
Polymer；The molecularly imprinted polymer without containing template molecule is passed through by the MIP of molecularly imprinted polymer containing template molecule
Cross what eluted template molecule obtained；The MIP of molecularly imprinted polymer containing template molecule is the molecule print containing template molecule
Mark polymer；The template molecule is environmental estrogens.
2. cobalt hydroxide nano-chip arrays electrode Co-nanoarray as described in claim 1 is it is characterized in that, the Co-
The preparation method of nanoarray includes following preparation step:
(1) it will can disposably throw electrode and carry out ultrasonic cleaning processing using dilute hydrochloric acid, dehydrated alcohol and deionization respectively, to go
Except the oxide layer and surface impurity that can disposably throw electrode；
(2) 1 ~ 3 mmol Co (NO is weighed3)2With 3 ~ 9 mmol urea CO (NH2)2, it is put into 50 mL beakers, is added 30
ML deionized water stirs to clarify, and is then transferred into 50 mL ptfe autoclaves；
(3) in the disposable solution thrown in the reaction kettle that electrode is put into step (2) for handling step (1) well, 100 ~
It is reacted 9 ~ 12 hours at a temperature of 130 DEG C, cobalt hydroxide nano-chip arrays presoma electrode is prepared；
(4) the cobalt hydroxide nano-chip arrays presoma electrode insertion obtained step (3) contains dopamine, Ammonium Persulfate 98.5 and nitre
In the phosphate buffer solution PBS of sour cobalt, 20 ~ 40 DEG C at a temperature of reaction 4 ~ 6 hours after, take out simultaneously soaked with deionized water
It washes 2 ~ 4 times, cobalt hydroxide nano-chip arrays electrode Co-nanoarray is prepared；
The described disposable electrode of throwing is selected from one of cycle unit: nickel foam, foam copper, pure nickel piece, pure copper sheet, pure cobalt piece,
Pure silicon piece, conductive carbon cloth；
In the phosphate buffer solution PBS containing dopamine, Ammonium Persulfate 98.5 and cobalt nitrate: dopamine concentration is 2 ~ 5
Mg/mL, the concentration of Ammonium Persulfate 98.5 are 3 ~ 8 mg/mL, and the concentration of cobalt nitrate is 0.1 ~ 0.5 mg/mL, phosphate buffer solution PBS
Concentration be 0.1 mol/L, pH value be 7.2 ~ 8.5.
3. the MIP of molecularly imprinted polymer containing template molecule as described in claim 1, which is characterized in that described containing template point
Sub- molecularly imprinted polymer MIP is that direct in-situ is grown on Co-nanoarray, and preparation method includes following preparation step:
(1) 0.25 ~ 0.45mmol template molecule and 3 ~ 5 mmol 2- methacrylic acid MAA are weighed respectively in peace times bottle, are added
Enter 8 ~ 15 mL acetonitriles, 30 min of ultrasound to whole dissolutions；
(2) 15 ~ 25 mmol ethylene glycol dimethacrylate EDMA are added in the solution of step (1), 30 min of ultrasound
To being uniformly mixed, precursor mixed solution is obtained；
(3) Co-nanoarray is clipped on Stirring device, is inserted into the precursor mixed solution in step (2), in N2Ring
At a temperature of border and 20 ~ 40 DEG C of water-bath, with 5 ~ 200 revolutions per seconds of speed Stirring, while with 1 ~ 20 drop/sec of speed to mixed
1 mmol azodiisobutyronitrile AIBN is added dropwise in conjunction solution and carries out initiation polymerization, growth in situ is obtained on Co-nanoarray
The MIP of molecularly imprinted polymer containing template molecule.
4. as described in claim 1 without template molecule molecularly imprinted polymer NIP, it is characterised in that the no template molecule
The preparation step of molecularly imprinted polymer NIP are as follows: by growth in situ contains on Co-nanoarray obtained in claim 3
Template molecule molecularly imprinted polymer MIP is immersed in eluant, eluent, template molecule is carried out 5 ~ 20 min of elution at room temperature, so
After take out, obtain no template molecule molecularly imprinted polymer NIP；The eluant, eluent is the mixed liquor of formic acid and methanol,
The volume ratio of middle formic acid and methanol is 9:(1 ~ 5).
5. the preparation step of environmental estrogens electro-chemical analyzing sensor as described in claim 1 are as follows: will be in claim 2 ~ 4
The growth in situ obtained on Co-nanoarray without template molecule molecularly imprinted polymer NIP, embathe 2 ~ 4 with deionized water
It is secondary, it dries at room temperature, obtains environmental estrogens electro-chemical analyzing sensor.
6. being applied to using environmental estrogens electro-chemical analyzing sensor prepared by preparation method described in claim 1 ~ 5
The detection of environmental estrogens, which is characterized in that the detecting step is as follows:
(1) standard solution is prepared: preparing the environmental estrogens standard solution of one group of various concentration including blank standard specimen；
(2) working electrode is modified: being working electrode by environmental estrogens electro-chemical analyzing sensor, is prepared in inserting step (1)
Various concentration environmental estrogens standard solution, hatch 10 min after take out, embathed 3 times with deionized water；
(3) working curve is drawn: using saturated calomel electrode electrode as reference electrode, platinum electrode is used as to electrode, with step
(2) the working electrode composition three-electrode system modified, connects electrochemical workstation, 15 mL is successively added in a cell
Phosphate buffer solution PBS；Pass through the current-responsive of the working electrode of Differential Pulse Voltammetry DPV detection assembling；Blank standard specimen
Response current intensity be denoted asI 0, the response current intensity of the environmental estrogens standard solution containing various concentration is denoted asI i, response
The difference that current strength reduces is ΔI = I 0-I i, ΔIWith the mass concentration of environmental estrogens standard solutionCBetween linear pass
Δ is drawn by systemI?CWorking curve；The phosphate buffer solution PBS concentration is 10 mmol/L, pH value 7.4；It is described
DPV detection when parameter setting are as follows: range and direction are 0 ~ 1 V, and stride is 0.05 V, and the burst length is 0.05 s, sampling
Time is 0.016 s, and the pulse period is 0.5 s；
(4) in sample to be tested environmental estrogens detection: replace the environmental estrogens standard in step (1) molten with sample to be tested
Liquid is detected according to the method in step (2) and (3), the difference DELTA that current strength reduces according to responseIAnd working curve,
Obtain the content of environmental estrogens in sample to be tested.
7. the environmental estrogens as described in claim 1 ~ 6 are one of following environmental estrogens: estradiol, estriol, hexene are female
Phenol, bisphenol-A, nonyl phenol.
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