CN109254044A - A kind of preparation method and application of the macrolide antibiotics sensor based on nitrided iron - Google Patents
A kind of preparation method and application of the macrolide antibiotics sensor based on nitrided iron Download PDFInfo
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Abstract
The preparation method of the invention discloses a kind of macrolide antibiotics sensor based on nitrided iron.Belong to Nano-function thin films and chemical biosensor technical field.The present invention is prepared for nitrided iron nano-chip arrays can disposably throw first on electrode, using its big specific surface area and to the high adsorption activity of amino, using the method for growth in situ, directly be prepared in succession on nitrided iron nano-chip arrays in succession poly-dopamine film and in-stiu coating luminol using macrolide antibiotics molecule as the molecularly imprinted polymer of template molecule, 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 macrolide antibiotics sensor based on nitrided iron just prepares completion.
Description
Technical field
The present invention relates to a kind of preparation method and applications of Electrochemiluminescsensor sensor.Belong to Nano-function thin films
With chemical biosensor technical field.
Background technique
Macrolide antibiotics (macrolides antibiotics, MA) is that have 12-16 in molecule structure
The general name of the antibacterials of carbon lactone ring inhibits bacterio protein by blocking the activity of peptidy transeferace in 50s ribosomes
Synthesis, belongs to quick bacteriostatic agent.It is mainly used for treating aerobic gram-positive cocci and negative cocci, certain anaerobic bacterias and legion
The infection such as bacterium, mycoplasma, Chlamydia.After people are edible remains the animal food of such antibiotic, animal is generated resistance to
Pharmacological property can also be transferred to the mankind, to endanger human health.Currently, the method for detection macrolide antibiotics molecule mainly has
Enzyme-linked immunization, high performance liquid chromatography, mass spectrography etc..Such method instrument is valuable, complicated for operation, and laboratory personnel needs profession
It just can be carried out detection after training.Therefore, it develops as early as possible a kind of quick, highly selective and Sensitive Detection macrolide antibiotics
Method is extremely important to publilc health, and has wide market application prospect.
Electroanalytical Chemistry sensor includes electrochemical sensor, Electrochemiluminescsensor sensor, optical electro-chemistry sensor etc.,
Such sensor has high specific selectivity, excellent stability, excellent reproducibility, wide detection range and floor detection
Limit.Due to the sensor prepare simple, easy to detect, high sensitivity, it is at low cost the advantages that be widely used in chromatographic isolation, film
Point, the fields such as Solid Phase Extraction, medicine controlled releasing, chemical sensitisation.Molecularly imprinted polymer (MIP), also referred to as " plastics antibody ", can
Specific recognition and the specific target molecule of selective absorption (i.e. template molecule).Since molecular imprinting technology has many advantages,
Such as organic reagent corrosion resistance, good stability, heat-resisting quantity and preparation are simple.Therefore, in the past few years, it is based on
The MIP Electroanalytical Chemistry sensor (MIP-ECS) that MIP is combined with Electroanalytical Chemistry sensor causes Electroanalytical Chemistry field
Focus, the especially detection of small molecule contaminants.However, having template point in the preparation process of traditional MIP-ECS
The disadvantages of sub- hardly possible elutes, the thickness of blotting membrane is difficult to control, reproducibility is poor, limits molecular engram film in Electroanalytical Chemistry sensor
In application.These problems, especially molecular engram film thickness are not easy to control cause electrochemical sensor sensitivity decrease and
Molecular engram film easily falls off from electrode surface during elution leads to the technical problem of stability and reproducibility reduction, limits
Therefore the application of MIP_ECS finds the modification side of new molecularly imprinted polymer synthetic method, new molecular engram film electrode
The combination method of method and molecular engram film and base material has important grind to solve preparation and the application problem of MIP-ECS
Study carefully meaning 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 macrolide antibiotics sensor based on nitrided iron, prepared Electrochemiluminescsensor sensor, preparation
Simply, favorable reproducibility, stability are strong, can be used for quick, the Sensitive Detection of macrolide antibiotics molecule.Based on this purpose,
The present invention is prepared for nitrided iron nano-chip arrays can disposably throw first on electrode, using its big specific surface area and to amino
High adsorption activity poly- DOPA is directly prepared on nitrided iron nano-chip arrays in succession using the method for growth in situ in succession
Amine film and in-stiu coating luminol using macrolide antibiotics molecule as the molecularly imprinted polymer of template molecule, will
After template molecule elution, the position of template molecule originally has become hole, i.e. the molecular engram polymerization of eluted template molecule
Object, a kind of macrolide antibiotics sensor based on nitrided iron just prepares completion as a result,.When for anti-to macrolides
When raw element molecule is detected, the macrolide antibiotics sensor based on nitrided iron is inserted into solution to be measured, it is to be measured molten
Macrolide antibiotics molecule in liquid can be adsorbed onto the hole of NIP.Macrolide antibiotics point in solution to be measured
Sub- concentration is bigger, and it is more to be adsorbed onto macrolide antibiotics molecule in the hole of NIP.When progress electrochemiluminescdetection detection
When, it can be become smaller with increasing for macrolide antibiotics molecule in the hole of NIP is adsorbed by the current strength of electrode,
Corresponding electrochemiluminescence signal can also become smaller therewith, thus the journey reduced according to the light signal strength of electrogenerated chemiluminescence
Degree is capable of the concentration of macrolide antibiotics molecule in qualitative, quantitative solution to be measured.
The technical solution adopted by the invention is as follows:
1. a kind of preparation method of the macrolide antibiotics sensor based on nitrided iron, described based on the big of nitrided iron
Cyclic lactone class antibiotic sensor is by growth in situ on nitrided iron nano-chip arrays electrode FeN-nanoarray without template molecule point
What sub- imprinted polymer NIP was obtained；The molecule that template molecule is free from without template molecule molecularly imprinted polymer NIP
Imprinted polymer；The molecularly imprinted polymer without containing template molecule is by molecularly imprinted polymer containing template molecule
What MIP was obtained by eluted template molecule；The MIP of molecularly imprinted polymer containing template molecule is point containing template molecule
Sub- imprinted polymer；The template molecule is macrolide antibiotics molecule；
2. the preparation method of nitrided iron nano-chip arrays electrode FeN-nanoarray described in technical solution 1 includes following system
Standby 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, six nitric hydrate iron Fe (NO is weighed3)3·6H2O and 3 ~ 9 mmol urea CO (NH2)2, it is put into 50
In mL beaker, 30 mL deionized waters are added and stir to clarify, are 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, iron hydroxide nano-chip arrays presoma electrode is prepared；
(4) the iron hydroxide nano-chip arrays presoma electrode that step (3) obtains is inserted into ammonium hydroxide and is taken out after 5 ~ 30 seconds, in ammonia
Under compression ring border, after being heated to 340 ~ 400 DEG C and being kept for 4 ~ 8 hours, continuation naturally rings to room temperature under ammonia environment, then by it
Be inserted into the phosphate buffer solution PBS containing dopamine and Ammonium Persulfate 98.5 in, 20 ~ 40 DEG C at a temperature of react 4 ~ 6 hours
Afterwards, it takes out and is embathed 2 ~ 4 times with deionized water, nitrided iron nano-chip arrays electrode FeN-nanoarray is prepared；
The described disposable electrode of throwing is selected from one of cycle unit: foamed iron, foam copper, pure iron piece, pure copper sheet, pure iron piece,
Pure silicon piece, conductive carbon cloth；
In the phosphate buffer solution PBS containing dopamine and Ammonium Persulfate 98.5: dopamine concentration is 2 ~ 5 mg/mL, mistake
The concentration of amine sulfate is 3 ~ 8 mg/mL, and the concentration of phosphate buffer solution PBS is 0.1 mol/L, and pH value is 7.2 ~ 8.5；
3. the MIP's of molecularly imprinted polymer containing template molecule of FeN-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) FeN-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
1 ~ 3 mL of luminol solution of 1 mmol/L of dropwise addition and 1 mmol azo two are different simultaneously into mixed solution for ~ 20 drops/sec of speed
Butyronitrile AIBN carries out initiation polymerization, and the molecularly imprinted polymer containing template molecule of growth in situ is obtained on FeN-nanoarray
MIP；
4. FeN-nanoarray growth in situ described in technical solution 1 without template molecule molecularly imprinted polymer NIP's
Preparation step are as follows: by the molecular engram containing template molecule of growth in situ gathers on FeN-nanoarray obtained in technical solution 3
It closes object MIP to be immersed in eluant, eluent, template molecule is subjected to 5 ~ 20 min of elution at room temperature, is then taken out, no mould is obtained
Plate molecular imprinted polymer NIP；The eluant, eluent is the mixed liquor of formic acid and methanol, wherein the volume of formic acid and methanol
Than for 9:(1 ~ 5)；
5. the preparation step of the macrolide antibiotics sensor described in technical solution 1 based on nitrided iron are as follows: by technology
In scheme 2 ~ 4 the obtained growth in situ on FeN-nanoarray without template molecule molecularly imprinted polymer NIP, spend from
Sub- water logging is washed 2 ~ 4 times, is dried at room temperature, and the macrolide antibiotics sensor based on nitrided iron is obtained；
6. using the macrolide antibiotics sensing prepared by technical solution described in technical solution 1 ~ 5 based on nitrided iron
Device, applied to the detection of macrolide antibiotics molecule, including following applying step:
(1) standard solution is prepared: preparing the macrolide antibiotics molecule of one group of various concentration including blank standard specimen
Standard solution；
(2) working electrode is modified: being working electrode, inserting step by the macrolide antibiotics sensor based on nitrided iron
(1) the macrolide antibiotics molecular criteria solution for the various concentration prepared in takes out after hatching 10 min, uses deionized water
It embathes 3 times；
(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, is connected in electrochemiluminescdetection detection equipment；In a cell first
Be added afterwards 15 mL phosphate buffer solution PBS and 1mL 2 mmol/L hydrogen peroxide (H2O2) solution；With double rank Pulse Voltammetries
Method applies cyclical voltage to the working electrode of assembling, detects the light signal strength of electrogenerated chemiluminescence；The response light of blank standard specimen
Signal strength is denoted asA 0, the response light signal strength of the macrolide antibiotics standard solution containing various concentration is denoted asA i, ring
The difference for answering light signal strength to reduce is ΔA = A 0-A i, ΔAWith the mass concentration of macrolide antibiotics standard solutionCIt
Between it is linear, draw ΔA?CWorking curve；The phosphate buffer solution PBS concentration is 10 mmol/L, pH value
It is 7.4；Parameter setting when double rank pulse voltammetries detect are as follows: initial potential is 0 V, and pulse potential is 0.9 V, arteries and veins
Rushing the time is 0.1 s, and the pulse period is 30 s；
(4) in sample to be tested macrolide antibiotics detection: replace the macrolides in step (1) anti-with sample to be tested
Raw element standard solution, is detected, the difference DELTA that light signal strength reduces according to response according to the method in step (2) and (3)A
And working curve, obtain the content of macrolide antibiotics in sample to be tested；
7. macrolide antibiotics molecule described in technical solution 1 ~ 6 is one of following macrolide antibiotics molecule: red
Mycin, medecamycin, azithromycin, clarithromycin, roxithromycin, tylosin, ivermectin.
Beneficial achievement of the invention
(1) the macrolide antibiotics sensor preparation of the present invention based on nitrided iron is simple, easy to operate, realizes
Quick, sensitive, highly selective detection to sample, and it is at low cost, it can be applied to portable inspectiont, before there is market development
Scape；
(2) the growth in situ molecularly imprinted polymer on nitrided iron nano-chip arrays electrode FeN-nanoarray for the first time of the invention,
On the one hand more, molecularly imprinted polymer more evenly can be grown using the big specific surface area of FeN-nanoarray, and
FeN-nanoarray has excellent electron transmission ability, to greatly improve detection sensitivity；On the other hand, FeN-
Nanoarray has electro catalytic activity to hydrogen peroxide, and may not need addition horseradish peroxidase can be realized luminol-mistake
The stabilization of hydrogen oxide electrogenerated chemiluminescence system, highly effective reaction, so that prepared sensor is without considering that biological enzyme inactivation is asked
Topic, so that the use and storage of sensor can be more stable loose with condition, thus further decreasing signal back
While scape, raising detection sensitivity, greatly reduces testing cost and reduce environmental pollution；
(3) present invention specific surface area and dopamine big using high adsorption activity and nano-array electrode of the nitride to amino
It combines, so that dopamine in nitrided iron nano-chip arrays in situ Polymerization, is forming sufficiently thin poly-dopamine film
While, on uniform fold to nitrided iron nano-chip arrays, thus for more preferably polymerizable molecular imprinted polymers in next step
Carry out place mat；Later using poly-dopamine to the strong absorption connection function for the amino being rich on molecularly imprinted polymer, then ingeniously
It uses FeN-nanoarray as blender wonderfully, immersion stirring is carried out in molecular engram precursor mixed solution, pass through control
The rate of addition and polymeric reaction temperature of mixing speed processed, initiators for polymerization, in the surface FeN-nanoarray direct in-situ
Growth can control the molecularly imprinted polymer of film thickness, print the secured supporting molecular of FeN-nanoarray
Mark polymer and luminol, to significantly improve the stability and reproducibility of prepared Electrochemiluminescsensor sensor；Separately
On the one hand molecularly imprinted polymer can be effectively controlled in the film forming thickness of electrode surface, solve and be unable to control molecular engram film
The technical problem so as to cause poor reproducibility is unable to control in electrode surface film forming thickness；In addition, more due to preparation of the invention
Method coats effective control of film forming thickness and the in situ quantitation of luminol, can sufficiently improve the electricity based on molecular engram
The sensitivity and detection limit for causing chemiluminescence sensor have important scientific meaning and application value.
Specific embodiment
The preparation of 1 FeN-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, six nitric hydrate iron Fe (NO is weighed3)3·6H2O and 3 mmol urea CO (NH2)2, it is put into 50 mL
In beaker, 30 mL deionized waters are added and stir to clarify, are 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, iron hydroxide nano-chip arrays presoma electrode is prepared；
(4) the iron hydroxide nano-chip arrays presoma electrode that step (3) obtains is inserted into ammonium hydroxide and is taken out after 5 seconds, in ammonia
Under environment, be heated to 340 DEG C and keep 8 hours after, continuation naturally ring to room temperature under ammonia environment, be then inserted into containing
In the phosphate buffer solution PBS of dopamine and Ammonium Persulfate 98.5,20 DEG C at a temperature of reaction 4 hours after, take out and spend from
Sub- water logging is washed 2 times, and nitrided iron nano-chip arrays electrode FeN-nanoarray is prepared；
The electrode therein that can disposably throw is foamed iron；Dopamine concentration is 2 mg/mL, and the concentration of Ammonium Persulfate 98.5 is 3 mg/mL,
The concentration of phosphate buffer solution PBS is 0.1 mol/L, pH value 7.2.
The preparation of 2 FeN-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, six nitric hydrate iron Fe (NO is weighed3)3·6H2O and 6 mmol urea CO (NH2)2, it is put into 50 mL
In beaker, 30 mL deionized waters are added and stir to clarify, are 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, iron hydroxide nano-chip arrays presoma electrode is prepared；
(4) the iron hydroxide nano-chip arrays presoma electrode that step (3) obtains is inserted into ammonium hydroxide and is taken out after 15 seconds, in ammonia
Under environment, be heated to 370 DEG C and keep 6 hours after, continuation naturally ring to room temperature under ammonia environment, be then inserted into containing
In the phosphate buffer solution PBS of dopamine and Ammonium Persulfate 98.5,30 DEG C at a temperature of reaction 5 hours after, take out and spend from
Sub- water logging is washed 3 times, and nitrided iron nano-chip arrays electrode FeN-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
The concentration of mg/mL, phosphate buffer solution PBS are 0.1 mol/L, pH value 8.0.
The preparation of 3 FeN-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, six nitric hydrate iron Fe (NO is weighed3)3·6H2O and 9 mmol urea CO (NH2)2, it is put into 50 mL
In beaker, 30 mL deionized waters are added and stir to clarify, are 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, iron hydroxide nano-chip arrays presoma electrode is prepared；
(4) the iron hydroxide nano-chip arrays presoma electrode that step (3) obtains is inserted into ammonium hydroxide and is taken out after 30 seconds, in ammonia
Under environment, be heated to 400 DEG C and keep 4 hours after, continuation naturally ring to room temperature under ammonia environment, be then inserted into containing
In the phosphate buffer solution PBS of dopamine and Ammonium Persulfate 98.5,40 DEG C at a temperature of reaction 6 hours after, take out and spend from
Sub- water logging is washed 4 times, and nitrided iron nano-chip arrays electrode FeN-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/
The concentration of mL, phosphate buffer solution PBS are 0.1 mol/L, pH value 8.5.
The preparation method of macrolide antibiotics sensor of the embodiment 4 based on nitrided iron
(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) FeN-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 into mixed solution simultaneously be added dropwise 1 mmol/L 1 mL of luminol solution and 1 mmol azodiisobutyronitrile AIBN into
Row causes polymerization, and the MIP of molecularly imprinted polymer containing template molecule of growth in situ is obtained on FeN-nanoarray；
(4) MIP of molecularly imprinted polymer containing template molecule of growth in situ on FeN-nanoarray for obtaining step (3)
It is immersed in eluant, eluent, template molecule is subjected to 5 min of elution at room temperature, then takes out, obtains no template molecule molecule
Imprinted polymer NIP；Continue to be embathed 2 times with deionized water, dry at room temperature, it is anti-to obtain the macrolides based on nitrided iron
Raw element sensor；
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 macrolide antibiotics sensor of the embodiment 5 based on nitrided iron
(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) FeN-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 into mixed solution simultaneously be added dropwise 1 mmol/L 2 mL of luminol solution and 1 mmol azodiisobutyronitrile AIBN into
Row causes polymerization, and the MIP of molecularly imprinted polymer containing template molecule of growth in situ is obtained on FeN-nanoarray；
(4) MIP of molecularly imprinted polymer containing template molecule of growth in situ on FeN-nanoarray for obtaining step (3)
It is immersed in eluant, eluent, template molecule is subjected to 10 min of elution at room temperature, then takes out, obtains no template molecule molecule
Imprinted polymer NIP；Continue to be embathed 3 times with deionized water, dry at room temperature, it is anti-to obtain the macrolides based on nitrided iron
Raw element sensor；
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 macrolide antibiotics sensor of the embodiment 6 based on nitrided iron
(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) FeN-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 into mixed solution simultaneously be added dropwise 1 mmol/L 3 mL of luminol solution and 1 mmol azodiisobutyronitrile AIBN into
Row causes polymerization, and the MIP of molecularly imprinted polymer containing template molecule of growth in situ is obtained on FeN-nanoarray；
(4) MIP of molecularly imprinted polymer containing template molecule of growth in situ on FeN-nanoarray for obtaining step (3)
It is immersed in eluant, eluent, template molecule is subjected to 20 min of elution at room temperature, then takes out, obtains no template molecule molecule
Imprinted polymer NIP；Continue to be embathed 4 times with deionized water, dry at room temperature, it is anti-to obtain the macrolides based on nitrided iron
Raw element sensor；
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 macrolide antibiotics sensor based on nitrided iron of 7 embodiment 1 ~ 6 of embodiment preparation, is applied to big ring
The detection of lactone antibiotic molecule, steps are as follows:
(1) standard solution is prepared: preparing the macrolide antibiotics molecule of one group of various concentration including blank standard specimen
Standard solution；
(2) working electrode is modified: being working electrode, inserting step by the macrolide antibiotics sensor based on nitrided iron
(1) the macrolide antibiotics standard solution for the various concentration prepared in takes out after hatching 10 min, is embathed with deionized water
3 times；
(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, is connected in electrochemiluminescdetection detection equipment；In a cell first
Be added afterwards 15 mL phosphate buffer solution PBS and 1mL 2 mmol/L hydrogen peroxide (H2O2) solution；With double rank Pulse Voltammetries
Method applies cyclical voltage to the working electrode of assembling, detects the light signal strength of electrogenerated chemiluminescence；The response light of blank standard specimen
Signal strength is denoted asA 0, the response light signal strength of the macrolide antibiotics standard solution containing various concentration is denoted asA i, ring
The difference for answering light signal strength to reduce is ΔA = A 0-A i, ΔAWith the mass concentration of macrolide antibiotics standard solutionCIt
Between it is linear, draw ΔA?CWorking curve；The phosphate buffer solution PBS concentration is 10 mmol/L, pH value
It is 7.4；Parameter setting when double rank pulse voltammetries detect are as follows: initial potential is 0 V, and pulse potential is 0.9 V, arteries and veins
Rushing the time is 0.1 s, and the pulse period is 30 s；
(4) in sample to be tested macrolide antibiotics detection: replace the macrolides in step (1) anti-with sample to be tested
Raw element standard solution, is detected, the difference DELTA that light signal strength reduces according to response according to the method in step (2) and (3)A
And working curve, obtain the content of macrolide antibiotics in sample to be tested.
The macrolide antibiotics sensor based on nitrided iron of 8 embodiment 1 ~ 6 of embodiment preparation, according to embodiment
7 detecting step is applied to the detection of different macrolide antibiotics, and the range of linearity and detection limit are shown in Table 1:
The detection technique index of 1 macrolide antibiotics of table
The detection of macrolide antibiotics in 9 pig urine samples of embodiment
Pig urine samples are accurately pipetted, the macrolide antibiotics standard solution of certain mass concentration are added, big ring not to be added
The pig urine samples of lactone antibiotic are blank, carry out recovery testu, with the preparation of embodiment 1 ~ 6 based on the big of nitrided iron
Cyclic lactone class antibiotic sensor, is detected according to the step of embodiment 7, measures macrolide antibiotics in pig urine samples
The rate of recovery, testing result is shown in Table 2:
The testing result of macrolide antibiotics in 2 pig urine samples of table
2 testing result of table it is found that the relative standard deviation (RSD) of result less than 3.5 %, average recovery rate is 98.4 ~
101.6%, show that the present invention can be used for the detection of a variety of macrolide antibiotics in pig urine, high sensitivity, the specificity of method
By force, as a result accurately and reliably.
The detection of macrolide antibiotics in 10 sheep urine samples of embodiment
Certain sheep urine samples are accurately pipetted, the macrolide antibiotics standard solution of certain mass concentration are added, not to be added
The sheep urine samples of macrolide antibiotics are blank, carry out recovery testu, with the preparation of embodiment 1 ~ 6 based on nitrided iron
Macrolide antibiotics sensor, detected according to the step of embodiment 7, it is anti-to measure macrolides in sheep urine samples
The rate of recovery of raw element, testing result are shown in Table 3:
The testing result of macrolide antibiotics in 3 sheep urine samples of table
For 3 testing result of table it is found that the relative standard deviation (RSD) of result is less than 3.2 %, average recovery rate is 98.6 ~ 101%,
Show the present invention can be used for sheep urine in a variety of macrolide antibiotics detection, the high sensitivity of method, high specificity, as a result
Accurately and reliably.
Claims (7)
1. a kind of preparation method of the macrolide antibiotics sensor based on nitrided iron, which is characterized in that it is described based on
The macrolide antibiotics sensor of nitrided iron by growth in situ on nitrided iron nano-chip arrays electrode FeN-nanoarray without
Template molecule molecularly imprinted polymer NIP is obtained；Described has been free from template without template molecule molecularly imprinted polymer NIP
The molecularly imprinted polymer of molecule；The molecularly imprinted polymer without containing template molecule is printed by molecule containing template molecule
What mark polymer MIP was obtained by eluted template molecule；The MIP of molecularly imprinted polymer containing template molecule is containing template
The molecularly imprinted polymer of molecule；The template molecule is macrolide antibiotics molecule.
2. nitrided iron nano-chip arrays electrode FeN-nanoarray as described in claim 1 is it is characterized in that, the FeN-
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, six nitric hydrate iron Fe (NO is weighed3)3·6H2O and 3 ~ 9 mmol urea CO (NH2)2, it is put into 50
In mL beaker, 30 mL deionized waters are added and stir to clarify, are 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, iron hydroxide nano-chip arrays presoma electrode is prepared；
(4) the iron hydroxide nano-chip arrays presoma electrode that step (3) obtains is inserted into ammonium hydroxide and is taken out after 5 ~ 30 seconds, in ammonia
Under compression ring border, after being heated to 340 ~ 400 DEG C and being kept for 4 ~ 8 hours, continuation naturally rings to room temperature under ammonia environment, then by it
Be inserted into the phosphate buffer solution PBS containing dopamine and Ammonium Persulfate 98.5 in, 20 ~ 40 DEG C at a temperature of react 4 ~ 6 hours
Afterwards, it takes out and is embathed 2 ~ 4 times with deionized water, nitrided iron nano-chip arrays electrode FeN-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 iron piece,
Pure silicon piece, conductive carbon cloth；
In the phosphate buffer solution PBS containing dopamine and Ammonium Persulfate 98.5: dopamine concentration is 2 ~ 5 mg/mL, mistake
The concentration of amine sulfate is 3 ~ 8 mg/mL, and the concentration of phosphate buffer solution PBS is 0.1 mol/L, and pH value is 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 FeN-nanoarray, and preparation method includes following preparation step
It is rapid:
(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) FeN-nanoarray is clipped on Stirring device, is inserted into the precursor mixed solution in step (2), in N2
At 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 drop/sec of speed to
1 ~ 3 mL of luminol solution and 1 mmol azodiisobutyronitrile AIBN that 1 mmol/L is added dropwise simultaneously in mixed solution are caused
Polymerization, obtains the MIP of molecularly imprinted polymer containing template molecule of growth in situ on FeN-nanoarray.
4. as described in claim 1 without template molecule molecularly imprinted polymer NIP, it is characterised in that the no template point
The preparation step of sub- molecularly imprinted polymer NIP are as follows: by obtained in claim 3 on FeN-nanoarray growth in situ
The MIP of molecularly imprinted polymer containing template molecule be immersed in eluant, eluent, template molecule is subjected to elution 5 ~ 20 at room temperature
Min then takes out, and obtains no template molecule molecularly imprinted polymer NIP；The eluant, eluent is the mixing of formic acid and methanol
Liquid, wherein the volume ratio of formic acid and methanol is 9:(1 ~ 5).
5. the preparation step of the macrolide antibiotics sensor based on nitrided iron as described in claim 1 are as follows: by right
It is required that in 2 ~ 4 the obtained growth in situ on FeN-nanoarray without template molecule molecularly imprinted polymer NIP, spend from
Sub- water logging is washed 2 ~ 4 times, is dried at room temperature, and the macrolide antibiotics sensor based on nitrided iron is obtained.
6. using the macrolide antibiotics sensing prepared by preparation method described in claim 1 ~ 5 based on nitrided iron
Device, the detection applied to macrolide antibiotics, which is characterized in that the detecting step is as follows:
(1) standard solution is prepared: preparing the macrolide antibiotics standard of one group of various concentration including blank standard specimen
Solution；
(2) working electrode is modified: being working electrode, inserting step by the macrolide antibiotics sensor based on nitrided iron
(1) the macrolide antibiotics standard solution for the various concentration prepared in takes out after hatching 10 min, is embathed with deionized water
3 times；
(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, is connected in electrochemiluminescdetection detection equipment；In a cell first
Be added afterwards 15 mL phosphate buffer solution PBS and 1mL 2 mmol/L hydrogen peroxide (H2O2) solution；With double rank Pulse Voltammetries
Method applies cyclical voltage to the working electrode of assembling, detects the light signal strength of electrogenerated chemiluminescence；The response light of blank standard specimen
Signal strength is denoted asA 0, the response light signal strength of the macrolide antibiotics standard solution containing various concentration is denoted asA i, ring
The difference for answering light signal strength to reduce is ΔA = A 0-A i, ΔAWith the mass concentration of macrolide antibiotics standard solutionCIt
Between it is linear, draw ΔA?CWorking curve；The phosphate buffer solution PBS concentration is 10 mmol/L, pH value
It is 7.4；Parameter setting when double rank pulse voltammetries detect are as follows: initial potential is 0 V, and pulse potential is 0.9 V, arteries and veins
Rushing the time is 0.1 s, and the pulse period is 30 s；
(4) in sample to be tested macrolide antibiotics detection: replace the macrolides in step (1) anti-with sample to be tested
Raw element standard solution, is detected, the difference DELTA that light signal strength reduces according to response according to the method in step (2) and (3)A
And working curve, obtain the content of macrolide antibiotics in sample to be tested.
7. the macrolide antibiotics as described in claim 1 ~ 6 is one of following macrolide antibiotics: erythromycin, wheat
Enlightening mycin, azithromycin, clarithromycin, roxithromycin, tylosin, ivermectin.
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