CN104820003B - A kind of papery microfluidic system and method for Detecting Pesticide - Google Patents
A kind of papery microfluidic system and method for Detecting Pesticide Download PDFInfo
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Abstract
The invention discloses a kind of papery microfluidic system and method for Detecting Pesticide.Papery microfluidic system is made up of papery micro-fluidic chip, three-electrode system, data acquisition device.Substrate and agricultural chemicals extract solution injection papery micro-fluidic chip, are transmitted to data acquisition device by three-electrode system detection signal, and Detecting Pesticide result is quickly obtained based on enzyme level principle.The present invention is easy to operate, improves the automaticity of inhibiting AChE detection process, while the use of paper base material greatly reduces cost, can be applied to fruits and vegetables, soil, water body equal samples Pesticide Residues fast and automatically, Site Detection.
Description
Technical field
The present invention relates to the technical field that papery micro-fluidic chip analytical technology and the Detection Technologies of Pesticide Residues are combined, specifically
It is related to a kind of papery microfluidic system and method for Detecting Pesticide.
Background technology
Agricultural chemicals is played an important role in agricultural production, and in China's agricultural production, the pesticide species for using are various, bag
Include several major classes such as Insecticides (tech) ＆ Herbicides (tech), bactericide, rat poison kind more than totally 600.Organophosphorus and carbamate pesticides class agricultural chemicals is China
The maximum agricultural chemicals of current usage amount, and more be used on fruit trees and vegetables.Therefore, at present water fruits and vegetables, aquatic products,
Residues of pesticides in the agricultural product such as soil are protruded the most so that organophosphorus pesticide and carbamate pesticide residue are exceeded.For having
Machine Phosphorus and carbamate chemicals for agriculture monitoring and management, prevent the exceeded agricultural product list marketing of residues of pesticides as agricultural production
The most important thing of quality trouble free service.
Constantly strengthen the detection work of residues of pesticides both at home and abroad, the method for detection is also more and more.The detection of residues of pesticides
Method is varied, has following a few classes by Cleaning Principle：1）Chromatography, including gas chromatography, liquid chromatography and capillary
Electrophoresis tube method 2）Spectroscopic methodology 3）Immunoassay 4）Inhibiting AChE 5）Chemical fast determining method 6) living body biological method.Inhibiting AChE is research
A kind of Fast Determination of Pesticide Residue technology of most and relative maturity, the method determines Organophosphorus and carbamate pesticides pesticide residue
It is deep to be welcome by vast grass-roots supervision department, production base and market with quick, easy, sensitive and low cost and other advantages.
The Fast Determination of Pesticide Residue means based on inhibiting AChE are roughly divided into enzyme piece method in the prior art（Quick measuring card method）,
PH methods, AAS（Colorimetric method）With hexavalent chrome bio-removal etc..These residues of pesticides for being based on inhibiting AChE principle are made a general survey of to examine
Survey means present situation it can be found that enzyme level principle requires the priority hybrid reaction and blank of plurality of reagents in itself, thus
Bring the low problem of complex operation step, automaticity, and the detection means reported outside Current Domestic all fails to have
Effect ground solves this problem.
Therefore the research heat that a kind of the Detection Technologies of Pesticide Residues fast and automatically, portable, inexpensive turns into current is developed
Point.Using papery micro-fluidic chip, making channel design and the analysis skill that it is merged with biosensor technology on its surface
Art can effectively solve the above problems, while greatly reduce cost, but at present the technology in Detecting Pesticide field not yet
There is substantial breakthrough.
The content of the invention
It is an object of the invention to be directed at present the Detecting Pesticide means generally existing based on inhibiting AChE both at home and abroad
A kind of low problem of complex operation step, automaticity, there is provided Detecting Pesticide skill fast and automatically, portable, inexpensive
Art.
In order to solve the above technical problems, the present invention is based on papery micro-fluidic chip technology and electrochemica biological sensor skill
The analytical technology of art fusion.Concrete technical scheme is as follows：
A kind of papery microfluidic system for Detecting Pesticide, it is characterised in that：Described system is by papery miniflow
Control chip, three-electrode system A, three-electrode system B and data acquisition device composition；Three-electrode system A and three-electrode system B are integrated
In papery micro-fluidic chip, data acquisition device connects three-electrode system A and three-electrode system B by data wire.
The channel design of described papery micro-fluidic chip is in Y types；Described Y types channel design is by sample inlet pool A and sample introduction
Pond B, sample intake passage A and sample intake passage B, detection cell A and detection cell B, hybrid channel, passive mixed structure are constituted；Sample intake passage A
It is connected with hybrid channel after converging with sample intake passage B, sample inlet pool A and sample inlet pool B is located at sample intake passage A and sample intake passage B respectively
Upper end, detection cell A is located at the middle part of sample intake passage A, and detection cell B is located at hybrid channel end, and passive mixed structure is located at
The centre position of hybrid channel.
The making material of described papery micro-fluidic chip is any one in filter paper, chromatographic paper or nitrocellulose filter.
Described three-electrode system A and three-electrode system B is integrated in papery micro-fluidic chip；The three-electrode system
A and three-electrode system B include working electrode, reference electrode, are located at working electrode respectively to electrode, reference electrode and to electrode
Both sides；Three-electrode system A and three-electrode system B are respectively positioned at the center of detection cell A and detection cell B.
Enzyme is secured on described three-electrode system A and two working electrodes of three-electrode system B.
The fixing means of the fixation is any one in absorption method, investment, cross-linking method or covalent coupling method；Described
Enzyme is any one in acetylcholinesterase, butyrylcholine esterase, plant esterase.
Described working electrode and the material to electrode are any one in platinum, gold, carbon；The reference electrode material is
It is covered with the argent of silver chloride layer and is immersed in any one in composite, silver, the carbon formed in potassium chloride or hydrochloric acid solution.
Described data acquisition device is that the data acquisition circuit of core and the program of microcontroller are constituted by microcontroller；
Described microcontroller is any one in single-chip microcomputer, DSP, ARM；Described data acquisition circuit is by potentiostat circuit, amplification
Circuit, filter circuit, A/D conversion circuits, interface circuit, microcontroller peripheral circuit are constituted.
A kind of pesticide residue detection method, it is characterised in that including following characteristics step：
Step one, pre-treatment：Pre-treatment to testing sample, including take testing sample be put into beaker in, add extractant,
Soak extraction goes out agricultural chemicals extract solution；
Step 2, injection：By substrate and agricultural chemicals extract solution simultaneously from sample introduction by way of syringe pump or hand injection
Pond A and sample inlet pool B injection papery micro-fluidic chips；
Step 3, hybrid reaction：After substrate and agricultural chemicals extract solution inject from sample inlet pool A and sample inlet pool B respectively, bottom first
The detection cell A that logistics is crossed in the middle part of sample intake passage A, the consolidating on three-electrode system A working electrodes in the case of without agricultural chemicals extract solution
Surely changing enzyme A carries out catalytic reaction to substrate, produces electroactive material, and at the same time agricultural chemicals extract solution flows through sample intake passage B, then
Substrate and agricultural chemicals extract solution converge after commonly through hybrid channel, under the passive mixed structure effect in hybrid channel centre position
It is well mixed, the last detection cell B for flowing into hybrid channel end simultaneously, three electrode bodies in the case where there is agricultural chemicals extract solution to suppress
It is that immobilised enzymes B on B working electrodes carries out catalytic reaction to substrate, produces electroactive material；
Step 4, Electrochemical Detection：Electroactive material redox is detected by three-electrode system A and three-electrode system B
Micro-current signal produced by reaction, so as to form blank；
Step 5, signal acquisition：Micro- electricity is gathered by the data acquisition device of connection three-electrode system A and three-electrode system B
Stream signal, transmitting to host computer carries out data processing, and control calculates the residues of pesticides concentration of testing sample.
Described testing sample is any one in fruits and vegetables, soil, water body；Described substrate is acetylcholine, thioacetyl
In choline, acetonaphthone ester any one；Described agricultural chemicals extract solution is organophosphorus pesticide or carbamate chemicals for agriculture.
Present invention employs cheap papery micro-fluidic chip, channel design and integrated two immobilized enzymes are made on its surface
Three-electrode system, then signal is gathered by data acquisition device, residues of pesticides inspection can be automatically performed under the driving of paper capillary force
The priority hybrid reaction and blank of plurality of reagents, easy to operate during survey, improves inhibiting AChE detection process
Automaticity, while the use of paper base material greatly reduces cost, is suitable for fruits and vegetables, soil, water body equal samples Pesticides
Residual fast and automatically, Site Detection.
Brief description of the drawings
Fig. 1 is the structural representation of papery microfluidic system in the present invention.
Fig. 2 is the hardware circuit composition schematic diagram of data acquisition device in the present invention.
In figure：1. sample inlet pool A, 2. sample intake passage A, 3. detection cell A, 4. sample inlet pool B, 5. sample intake passage B, 6. passive mixing
Structure, 7. hybrid channel, 8. detection cell B, 9. immobilised enzymes B, 10. three-electrode system B, 11. three-electrode system A, 12, immobilization
Enzyme A.
Specific embodiment
Below in conjunction with the accompanying drawings, by taking the organophosphorus pesticide detection of vegetables as an example, specific embodiment of the invention is entered
Row is described in detail.
In the present embodiment, for Detecting Pesticide papery microfluidic system include the micro-fluidic chip of papery and
One data acquisition device, the wherein structure of papery microfluidic system are as shown in figure 1, the hardware circuit composition of data acquisition device
As shown in Figure 2.
Specifically detection process is：
, first be placed in the vegetables sample of 5g in the containers such as beaker by step one, adds 2,000 one section of μ L extractants immersion treatments
Time, or extractant is added dropwise in vegetable surface, the agricultural chemicals extract solution for being extracted.
Step 2,2000 μ L substrate acetyls choline solutions are injected into chip from 1 sample inlet pool A, while by 2000 μ L agricultures
Medicament extracting solution injects chip from 4 sample inlet pool B.
Step 3, substrate acetyl choline solution flows through the 2 detection cell A of sample intake passage A to 3, and is fixed on 11 three-electrode system A
Working electrode on 12 immobilised enzymes A there is catalytic reaction, the electroactive material of generation forms oxidation also on the working electrode (s
Primary current signal.
Step 4, agricultural chemicals extract solution flows through 5 sample intake passage B, converges in 7 hybrid channels with substrate acetyl choline solution, and
It is sufficiently mixed in the presence of 6 passive mixed structures afterwards.
Step 5, the mixed solution of agricultural chemicals extract solution and substrate flow to 8 detection cell B, and is fixed on 10 three-electrode system B's
There is catalytic reaction in 9 immobilised enzymes B on working electrode, the electroactive material of generation forms redox electricity on the working electrode (s
Stream signal.
Step 6, the redox effect of the electroactive material on three-electrode system A and three-electrode system B detection working electrodes
Answer produced micro-current signal.
Step 7, data acquisition device gathers two detected current signals, blank is formed, by wireless telecommunications
Or the means such as bluetooth are transmitted to host computer, carry out data processing, obtain surveyed residue of organic phosphorus pesticides in vegetable amount.
Above-mentioned step two~seven are system automation and operate, the papery microfluidic system provided using this implementation,
Can be with the sxemiquantitative of acquisition Organophosphorus and carbamate pesticides pesticide residue or quantitative determination result in 10~15min.
Claims (8)
1. a kind of papery microfluidic system for Detecting Pesticide, it is characterised in that：Described system is micro-fluidic by papery
Chip, three-electrode system A, three-electrode system B and data acquisition device composition；Three-electrode system A and three-electrode system B are integrated in
In papery micro-fluidic chip, data acquisition device connects three-electrode system A and three-electrode system B by data wire；
The channel design of described papery micro-fluidic chip is in Y types；Described Y types channel design by sample inlet pool A and sample inlet pool B,
Sample intake passage A and sample intake passage B, detection cell A and detection cell B, hybrid channel, passive mixed structure are constituted；Sample intake passage A and enter
Sample passage B is connected after converging with hybrid channel, and sample inlet pool A and sample inlet pool B is respectively positioned at the upper of sample intake passage A and sample intake passage B
End, detection cell A is located at the middle part of sample intake passage A, and detection cell B is located at hybrid channel end, and passive mixed structure is located at mixing
The centre position of passage.
2. a kind of papery microfluidic system for Detecting Pesticide according to claim 1, it is characterised in that：It is described
The making material of papery micro-fluidic chip be any one in filter paper, chromatographic paper or nitrocellulose filter.
3. a kind of papery microfluidic system for Detecting Pesticide according to claim 1, it is characterised in that：It is described
Three-electrode system A and three-electrode system B be integrated in papery micro-fluidic chip；The three-electrode system A and three electrode bodies
It is that B includes working electrode, reference electrode, is located at the both sides of working electrode respectively to electrode, reference electrode and to electrode；Three electricity
Polar body system A and three-electrode system B are respectively positioned at the center of detection cell A and detection cell B.
4. a kind of papery microfluidic system for Detecting Pesticide according to claim 3, it is characterised in that：It is described
Three-electrode system A and three-electrode system B two working electrodes on secure enzyme.
5. a kind of papery microfluidic system for Detecting Pesticide according to claim 4, it is characterised in that：It is described
Fixed fixing means is any one in absorption method, investment, cross-linking method or covalent coupling method；Described enzyme is acetylcholine
In esterase, butyrylcholine esterase, plant esterase any one.
6. a kind of papery microfluidic system for Detecting Pesticide according to claim 1, it is characterised in that：It is described
Data acquisition device by microcontroller for the data acquisition circuit and the program of microcontroller of core are constituted；Described microcontroller
Device is any one in single-chip microcomputer, DSP, ARM；Described data acquisition circuit is by potentiostat circuit, amplifying circuit, filtered electrical
Road, A/D conversion circuits, interface circuit, microcontroller peripheral circuit are constituted.
7. the Detecting Pesticide side of a kind of papery microfluidic system for Detecting Pesticide according to claim 1
Method, it is characterised in that including following characteristics step：
Step one, pre-treatment：Pre-treatment to testing sample, including take testing sample be put into beaker in, add extractant, immersion
Extract agricultural chemicals extract solution；
Step 2, injection：By way of syringe pump or hand injection by substrate and agricultural chemicals extract solution simultaneously from sample inlet pool A and
Sample inlet pool B injects papery micro-fluidic chip；
Step 3, hybrid reaction：After substrate and agricultural chemicals extract solution inject from sample inlet pool A and sample inlet pool B respectively, bottoms stream first
The detection cell A crossed in the middle part of sample intake passage A, the immobilization in the case of without agricultural chemicals extract solution on three-electrode system A working electrodes
Enzyme A carries out catalytic reaction to substrate, produces electroactive material, and at the same time agricultural chemicals extract solution flows through sample intake passage B, then substrate
Commonly through hybrid channel after converging with agricultural chemicals extract solution, mix under the passive mixed structure effect in hybrid channel centre position
Uniformly, the last detection cell B for flowing into hybrid channel end simultaneously, the three-electrode system B in the case where there is agricultural chemicals extract solution to suppress
Immobilised enzymes B on working electrode carries out catalytic reaction to substrate, produces electroactive material；
Step 4, Electrochemical Detection：Electroactive material redox reaction is detected by three-electrode system A and three-electrode system B
Produced micro-current signal, so as to form blank；
Step 5, signal acquisition：Believed by the data acquisition device collection micro-current of connection three-electrode system A and three-electrode system B
Number, transmitting to host computer carries out data processing, and control calculates the residues of pesticides concentration of testing sample.
8. a kind of pesticide residue detection method according to claim 7, it is characterised in that：Described testing sample is fruit
In vegetable, soil, water body any one；Described substrate is any one in acetylcholine, acetylthiocholine, acetonaphthone ester；It is described
Agricultural chemicals extract solution be organophosphorus pesticide or carbamate chemicals for agriculture.
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