CN115290719A - Electrochemical test paper for uric acid detection and preparation method and application thereof - Google Patents
Electrochemical test paper for uric acid detection and preparation method and application thereof Download PDFInfo
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
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Abstract
The invention provides electrochemical test paper for uric acid detection and a preparation method and application thereof, belonging to the technical field of medical equipment. The electrochemical test paper comprises a working electrode and a counter electrode, the preparation method comprises the steps of at least respectively soaking the working electrode in a first polymer solution and a second polymer solution to obtain a polymer modified working electrode, and depositing flower-shaped nano gold particles on the surface of the polymer modified working electrode to obtain the electrochemical test paper. The electrochemical test paper prepared by the preparation method has strong anti-pollution capability, good stability and high repeatability, and can be applied to the detection of uric acid.
Description
Technical Field
The invention belongs to the technical field of medical equipment, relates to in-vitro diagnosis and detection test paper, and particularly relates to electrochemical test paper for uric acid detection, a preparation method thereof, and application of the electrochemical test paper in uric acid detection.
Background
Uric Acid (UA) is a final product of purine metabolism in the human body, and abnormality in its concentration may cause diseases such as gout, hyperuricemia, and the like. Clinically normal uric acid level, 149-416 μ M for male and 89-357 μ M for female. However, it is far from sufficient to control uric acid to normal value for gout patients and kidney disease patients with high uric acid, and if the deposition of a large amount of uric acid on organs such as joints, kidneys and the like is to be avoided and the kidney function is to be protected, the uric acid needs to be controlled to be within a normal range by diet. Therefore, there is a need for a real-time uric acid test for patients with hyperuricemic conditions, which can help them control the uric acid level in the body.
At present, methods for detecting uric acid mainly comprise uricase detection method, colorimetric method, spectroscopic method, chromatographic analysis method and the like. However, the existing methods have some defects, such as high detection conditions, high manufacturing cost, complex detection environment and the like, while the electrochemical method has the advantages of rapidness, convenience, small equipment volume and the like, and particularly, the occurrence of a screen printing electrode enables a patient to monitor the content level of uric acid of the patient at home in real time. However, the current electrochemical detection has a bottleneck which is difficult to break through, because the electrochemical detection is very dependent on the electron transfer rate of the detected solution phase, when the screen printing test paper is used for detecting substances in a whole blood environment, the sensitivity and the accuracy of the screen printing test paper are obviously reduced, and the anti-pollution capability of the test paper is insufficient. Although the difficulty that the industrial blood glucose test strip widely used for home detection breaks through and the problem that the property of enzyme catalysis is perfectly avoided is overcome, the activity of the enzyme is greatly influenced by the environment, the stability of the test strip is not good, and the quality guarantee period is only three months. Therefore, the provision of the uric acid electrochemical test paper which can monitor the uric acid content level of a patient at home in real time, has strong pollution resistance, good stability and high repeatability becomes an important problem to be solved by technical personnel in the field.
Disclosure of Invention
Aiming at the defects in the prior art, in combination with the tests and findings of the inventor, one of the purposes of the invention is to provide the electrochemical test paper for detecting uric acid, which has high sensitivity, fast response time, good repeatability and high stability, and the preparation method thereof.
According to one aspect of the present application, the present invention provides a preparation method of an electrochemical test paper for uric acid detection, comprising the following steps:
at least soaking the working electrode in a first polymer solution and a second polymer solution respectively to obtain a polymer modified working electrode, wherein one of the first polymer solution and the second polymer solution is a polymer solution containing primary amine and/or secondary amine, and the other is a polymer solution containing carboxyl;
and depositing flower-shaped nano gold particles on the surface of the working electrode modified by the polymer to obtain the electrochemical test paper.
According to the method, the polymer film is modified firstly, the polymer solution changes the microstructure of the electrode surface, the nucleation sites of the nano-gold particles are increased, and the nano-gold particles have the tendency of growing the spurt structure on the sites. Through the electro-deposition technology, the flower-shaped nano gold particle structure with the same appearance, uniform size and uniform distribution and the spurs is deposited. Because the existence of the spurs enables the specific surface area of the gold nanoparticles to be larger, the structure can accelerate the electron transfer rate of the electrode surface, generate an oxidation peak and detect uric acid.
Optionally, the first polymer solution is a polyacrylic acid solution, and the second polymer solution is a polyethyleneimine solution;
preferably, the first polymer solution is a polyacrylic acid solution with a mass concentration of 15-30wt%, more preferably 20wt%; and/or
The second polymer solution is a polyethyleneimine solution with a mass concentration of 15-30wt%, and more preferably 20%.
The two polymer solutions are selected to react at normal temperature, and the polymer polyethyleneimine with positive charges and the polymer polyacrylic acid with negative charges can form physical crosslinking, so that an interface film formed on the surface of the electrode after the two polymer solutions are soaked layer by layer is tighter. Because the interfacial film is compact, the formed gold nucleation sites are also denser, and the polyethyleneimine and polyacrylic acid gel have homogeneous structures, and the homogeneous properties of the polyethyleneimine and polyacrylic acid gel can enable the nucleation sites to be more uniform and dispersed, so that the deposited gold nanoparticles have the effects of uniform size and uniform distribution.
The amino group can react with the carboxyl group to form an ionic bond, and the amino group can react with the carbonyl group to form a covalent bond. The polyethyleneimine can be combined with different substances due to the polar group (amino group) and hydrophobic group (vinyl) structures, meanwhile, the polyethyleneimine exists as a polymeric cation in water and can neutralize and adsorb anionic substances, and the polyacrylic acid solution exists as a polymeric anion in solution and can neutralize and adsorb cationic substances.
The mass concentration of the first polymer solution is 15-30wt%, and the mass concentration of the second polymer solution is 15-30wt%, and the arrangement can improve the uniformity of film formation in the reaction process of the first polymer and the second polymer, improve the adhesion capability of the nano gold particles, and effectively increase the specific surface area of the nano gold.
Optionally, the polymerization degree of the polymer in the first polymer solution is 80-120, preferably 100; the polymerization degree of the polymer in the second polymer solution is 80 to 120, preferably 100.
If the molecular weight of the polymer is too large, the reaction is difficult to occur; if the molecular weight is too small, the crosslinking density is small and the film-forming property of the electrode surface is poor. When the polymerization degree is about 100, the polyethyleneimine can be used for trapping metals, namely deposition and attachment of nanoparticles.
Optionally, dripping chloroauric acid solution with the concentration of 50-100 mM on the surface of the working electrode modified by the polymer, and performing electrodeposition to form flower-shaped nano gold particles;
preferably, the dropping amount of the chloroauric acid solution is 10-30 mu L.
Under the concentration, the chloroauric acid solution has the largest specific surface area of the deposition appearance, the most uniform size of the deposition particles and the most uniform distribution. The dropping amount of the chloroauric acid solution is within the range, the surface tension of the solution dropped in the working area of the electrode is the largest, and gathered liquid drops are easily formed, so that the electrodeposition action is more uniform in the working electrode area. The dropping amount is too large, and the liquid drops are easy to disperse, so that the deposition effect is influenced; the dropping amount is too small, the size of the liquid drop is small, and the deposited nano gold particles are few, or the three-electrode system cannot be communicated, so that the electrodes are broken and cannot work.
Optionally, the electrodeposition is performed by potentiostatic deposition;
preferably, the deposition voltage of the electrodeposition is-0.4 to-0.2V, and the deposition time of the electrodeposition is 300 to 900s.
Optionally, post-processing the working electrode modified by the flower-shaped nano gold particles by adopting a cyclic voltammetry method and a current time method to obtain the electrochemical test paper,
preferably, the working electrode modified by the flower-shaped nano gold particles is subjected to post-treatment by sequentially adopting a primary cyclic voltammetry method and a primary current time method.
Optionally, dripping uric acid buffer solution on the electrochemical test paper with the working electrode modified by the flower-shaped gold nanoparticles, setting the test voltage interval to be-0.2-0.6V and the number of scanning cycles to be 1 cycle, and performing cyclic voltammetry treatment to obtain an oxidation peak potential value; taking the oxidation peak potential value as a circuit voltage, and carrying out current-time method treatment to obtain post-treated electrochemical test paper;
preferably, 300-1000 μ M of uric acid is dissolved in 0.2M PBS buffer to obtain a uric acid buffer solution with pH of 7.35-7.45, and the dropping amount of the uric acid buffer solution is 15-25 μ L.
The post-treatment can change the sharp state of the surface of the nano gold particles to make the nano gold particles blunt. The tip is oxidized and ground flat and is irreversibly changed, so that the electrochemical test paper is stable, and the detection effect is improved.
Optionally, the electrochemical test strip further comprises a reference electrode;
preferably, the working electrode, the counter electrode and the reference electrode in the electrochemical test paper are screen-printed electrodes, wherein the working electrode is a carbon electrode, the counter electrode is a carbon electrode, and the reference electrode is a silver/silver chloride electrode.
According to another aspect of the present application, there is provided an electrochemical test strip obtained by the method of any one of the above-mentioned methods.
According to still another aspect of the present application, the present application provides a use of the electrochemical test paper obtained by any one of the above-mentioned preparation methods in the detection of uric acid.
The beneficial effects of the invention comprise at least one of the following:
1. the polymer provided by the application has low cost, simple film-forming preparation steps and high film-forming speed, meanwhile, the modification of the polymer film can enable the gold nanostructure to be more stable and not to fall off easily, and the interface film is formed by selecting the conductive polymer with the reduction property, so that the in-situ reduction of an interface can be realized, and the load capacity can be improved;
2. the nano-gold particle functionalized uric acid test paper is prepared by electrodeposition on the surface of a working electrode modified by a polymer, the polymer enables deposited gold particles not to fall off easily, and simultaneously can induce the crystal face orientation growth of gold again, effectively increase the specific surface area of nano-gold and accelerate the electron transfer rate of the nano-gold particles on the surface of the test paper, does not need an electron mediator, and has the capability of directly detecting uric acid in a micro system;
3. the preparation process is simple, the cost is low, and industrial production is easy to realize;
4. the sample to be detected in the application does not need pretreatment, is simple to operate, has short detection time, can directly detect the blood sample, has small using amount, and can meet the real-time detection requirement;
5. the application provides an electrochemical test paper of uric acid detection, has broken through the bottleneck that detects whole blood uric acid and can receive the pollution, and antipollution ability is good to can also be with blood volume low to 2 microlitres in, still keep good repeatability, detection accuracy is fine.
Drawings
The accompanying drawings, which are included to provide a further understanding of the application, are incorporated in and constitute a part of this application, illustrate embodiment(s) of the application and together with the description serve to explain the application and not to limit the application. In the drawings:
fig. 1 shows a scanning electron microscope image of electrodeposited flower-like nano-gold particles prepared from electrochemical test paper # 1 in exemplary example 1 of the present invention;
fig. 2 shows a scanning electron microscope image of flower-like nano-gold particles obtained in the process of preparing electrochemical test paper 4# in exemplary embodiment 1 of the present invention;
fig. 3 shows a scanning electron microscope image of flower-like gold nanoparticles obtained during the preparation of electrochemical test paper 10# in exemplary embodiment 1 of the present invention;
fig. 4 shows a scanning electron microscope image of flower-like gold nanoparticles obtained during the preparation of electrochemical test paper D3# in exemplary example 1 of the present invention;
FIG. 5 shows a cyclic voltammogram of the electrochemical test paper for uric acid detection # 4 test prepared in exemplary example 1 of the present invention;
fig. 6 shows a current time graph of the electrochemical test paper 4# for uric acid detection prepared in the exemplary embodiment 1 of the present invention.
Detailed Description
In order to more clearly explain the overall concept of the invention, the following detailed description is given by way of example in conjunction with the accompanying drawings.
In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention, however, the present invention may be practiced in other ways than those specifically described herein, and therefore the scope of the present invention is not limited by the specific embodiments disclosed below.
In addition, in the description of the present invention, it should be understood that the terms "top", "bottom", "inner", "outer", "axial", "radial", "circumferential", and the like refer to orientations or positional relationships based on those shown in the drawings, which are used only for convenience in describing the present invention and for simplicity in description, and do not indicate or imply that the device or element referred to must have a particular orientation, be constructed and operated in a particular orientation, and thus, are not to be construed as limiting the present invention.
In the present invention, unless otherwise expressly specified or limited, the terms "mounted," "connected," "secured," and the like are to be construed broadly and can, for example, be fixedly connected, detachably connected, or integrally formed; the connection can be mechanical connection, electrical connection or communication; either directly or indirectly through intervening media, either internally or in any other relationship. The specific meanings of the above terms in the present invention can be understood by those skilled in the art according to specific situations.
In the present invention, unless expressly stated or limited otherwise, the first feature "on" or "under" the second feature may be directly contacting the second feature or the first and second features may be indirectly contacting each other through intervening media. In the description of the specification, reference to the description of "one embodiment," "some embodiments," "an example," "a specific example," or "some examples" or the like means that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.
In an exemplary embodiment of the present invention, the electrochemical test strip for uric acid detection comprises a working electrode and a counter electrode, and the specific preparation method can be realized by the following steps.
(1) Preparation of Polymer films
And at least soaking the working electrode in a first polymer solution and a second polymer solution respectively to obtain the polymer modified working electrode, wherein one of the first polymer solution and the second polymer solution is a polymer solution containing primary amine and/or secondary amine, and the other is a polymer solution containing carboxyl. The first polymer solution may be any one of a polyacrylic acid solution and a polylactic acid-glycolic acid copolymer solution, however, the present invention is not limited thereto. The second polymer solution may be any one of a polyethyleneimine solution, a chitosan solution, and a sodium polyaspartate solution, however, the present invention is not limited thereto.
The polymerization degree of the polymer in the first polymer solution is 80 to 120, preferably 100. The polymerization degree of the polymer in the second polymer solution is 80 to 120, preferably 100.
(2) Preparation by electrodeposition method
Dripping chloroauric acid solution with the concentration of 50-100 mM on the surface of the working electrode modified by the polymer, and depositing the flower-shaped nano gold particles by adopting a constant potential deposition method for 300-900 s to obtain the electrochemical test paper.
(3) Working electrode post-treatment
Dropping uric acid buffer solution on the electrochemical test paper of the working electrode modified by the flower-shaped nano gold particles, setting the test voltage interval to be-0.2-0.6V and the number of scanning circles to be 1 circle, and carrying out cyclic voltammetry treatment to obtain the oxidation peak potential value. And (4) taking the oxidation peak potential value as a circuit voltage, and carrying out current time method treatment to obtain the post-treated electrochemical test paper.
Wherein, 300-1000 MuM of uric acid is prepared and dissolved in 0.2M of PBS buffer solution to obtain the uric acid buffer solution with the pH of 7.35-7.45, and the dripping amount of the uric acid buffer solution is 15-25 MuL.
Example 1
The electrochemical test paper 1# -10# and the comparative electrochemical test paper D1# -D3# are prepared by adopting the method, wherein the electrochemical test paper further comprises a reference electrode, a working electrode, a counter electrode and the reference electrode in the electrochemical test paper are screen-printed electrodes, the working electrode is a carbon electrode, the counter electrode is a carbon electrode, and the reference electrode is a silver/silver chloride electrode.
Electrochemical test paper 1#
(1) Cleaning a working electrode of the electrochemical test paper by using ethanol, drying the working electrode by using nitrogen, and alternately soaking the working electrode in a polyacrylic acid solution with the mass concentration of 20% and a polyethyleneimine solution for 2 times respectively, wherein the polymerization degrees of polyacrylic acid and polyethyleneimine are both 100, and after each soaking, washing the working electrode by using ultrapure water and drying the working electrode by using nitrogen;
(2) 20 mu L of chloroauric acid solution with the concentration of 50mM is dripped on the surface of the working area of the electrochemical test paper, and the electrochemical workstation is utilized to carry out electrodeposition, wherein the solution needs to cover all areas of the working electrode, the counter electrode and the reference electrode. And (3) depositing for 900s under the condition of-0.3V of deposition voltage, washing with ultrapure water after deposition is finished, and drying with nitrogen to obtain the working electrode modified by the nano-gold particles, namely the electrochemical test paper No. 1, and storing for later use in a dry environment.
(1) Cleaning a working electrode of the electrochemical test paper with ethanol, drying the working electrode with nitrogen, alternately soaking the working electrode in a polyacrylic acid solution with the mass concentration of 15% and a polyethyleneimine solution with the mass concentration of 30% for 2 times respectively, wherein the polymerization degree of polyacrylic acid is 120, the polymerization degree of polyethyleneimine is 80, and the working electrode is washed clean with ultrapure water and dried with nitrogen after being soaked each time;
(2) Dripping 10 mu L of chloroauric acid solution with the concentration of 100mM on the surface of the working area of the electrochemical test paper, and carrying out electrodeposition by using an electrochemical workstation, wherein the solution needs to cover all areas of the working electrode, the counter electrode and the reference electrode. And (3) depositing for 900s under the condition of-0.25V of deposition voltage, washing with ultrapure water after deposition is finished, and drying with nitrogen to obtain the working electrode modified by the nano-gold particles, namely the electrochemical test paper No. 2, and storing for later use in a dry environment.
Electrochemical test paper 3#
(1) Cleaning a working electrode of the electrochemical test paper with ethanol, drying the working electrode with nitrogen, alternately soaking the working electrode in a polyacrylic acid solution with the mass concentration of 30% and a polyethyleneimine solution with the mass concentration of 15% for 2 times respectively, wherein the polymerization degree of polyacrylic acid is 80, the polymerization degree of polyethyleneimine is 120, and the working electrode is washed clean with ultrapure water and dried with nitrogen after being soaked each time;
(2) 30 mu L of chloroauric acid solution with the concentration of 50mM is dripped on the surface of the working area of the electrochemical test paper, and the electrochemical workstation is utilized to carry out electrodeposition, wherein the solution needs to cover all areas of the working electrode, the counter electrode and the reference electrode. And (3) depositing for 300s under the condition of-0.35V deposition voltage, washing with ultrapure water after deposition is finished, and drying with nitrogen to obtain the working electrode modified by the nano-gold particles, namely the electrochemical test paper 3#, and storing for later use in a dry environment.
Electrochemical test paper 4#
The electrochemical test sample No. 4 is obtained by performing post-treatment on the basis of the electrochemical test paper No. 1, and specifically comprises the following steps:
dripping 20 mu L of uric acid buffer solution on the electrochemical test paper 1#, wherein the uric acid buffer solution is obtained by dissolving 400 mu M of uric acid in 0.2M of PBS buffer solution, the test voltage interval is set to be-0.2-0.6V, the number of scanning circles is 1 circle, and performing cyclic voltammetry treatment to obtain an oxidation peak potential value; and taking the oxidation peak potential value as a circuit voltage, carrying out current time method treatment to obtain electrochemical test paper No. 4, and storing in a dry environment for later use.
The electrochemical test 5# is obtained by performing post-treatment on the basis of electrochemical test paper 1# and specifically comprises the following steps:
dripping 15 mu L of uric acid buffer solution on the electrochemical test paper 1#, wherein the uric acid buffer solution is obtained by dissolving 600 mu M of uric acid in 0.2M of PBS buffer solution, the test voltage interval is set to be-0.2-0.6V, the number of scanning circles is 1 circle, and performing cyclic voltammetry treatment to obtain an oxidation peak potential value; and taking the oxidation peak potential value as a circuit voltage, carrying out current time method treatment to obtain the electrochemical test paper No. 5, and storing for later use in a dry environment.
The electrochemical test paper 6# is different from the electrochemical test paper 1 #: and (2) alternately soaking the polylactic acid-glycolic acid copolymer solution with the mass concentration of 20% and the chitosan solution twice in the step (1), and obtaining the electrochemical test paper No. 6 by the same steps as the electrochemical test paper No. 1, and storing the electrochemical test paper No. 6 in a dry environment for later use.
Electrochemical test paper 7#
The difference between the electrochemical test paper 7# and the electrochemical test paper 1# is that: the polymerization degrees of polyacrylic acid and polyethyleneimine are both 150, and the rest steps are the same as those of the electrochemical test paper No. 1, so that the electrochemical test paper No. 7 is obtained and stored in a dry environment for later use.
The difference between the electrochemical test paper 8# and the electrochemical test paper 1# is that: in the step (2), the concentration of the chloroauric acid solution is 150mM, and the rest steps are the same as the electrochemical test paper No. 1, so that the electrochemical test paper No. 8 is obtained and stored in a dry environment for later use.
Electrochemical test paper 9#
The difference between the electrochemical test paper 9# and the electrochemical test paper 1# is as follows: in the step (2), the dripping amount of the chloroauric acid solution is 50 muL, and the rest steps are the same as the electrochemical test paper 1#, so that the electrochemical test paper 9# is obtained and stored in a dry environment for later use.
The electrochemical test paper 10# is different from the electrochemical test paper 1 #: in the step (2), the chloroauric acid solution is deposited for 1200s under the condition that the deposition voltage is minus 0.5V, and the rest steps are the same as the electrochemical test paper No. 1, so that the electrochemical test paper No. 10 is obtained and is stored for later use in a dry environment.
Comparative electrochemical test paper D1#
The difference between the electrochemical test paper D1# and the electrochemical test paper 1# is as follows: the first polymer solution is a polyethylene glycol solution, the second polymer solution is a polyvinylpyrrolidone solution, and the rest steps are the same as the steps of the electrochemical test paper 1#, so that the electrochemical test paper D1# is obtained and stored in a dry environment for later use.
Comparative electrochemical test paper D2#
The difference between the comparative electrochemical test paper D2# and the electrochemical test paper 1# is as follows: and repeatedly soaking the test paper in a polyacrylic acid solution with the mass concentration of 20% for 2 times, and obtaining the electrochemical test paper D2# by the same steps as the electrochemical test paper 1# and storing the test paper for later use in a dry environment.
Comparative electrochemical test paper D3#
The difference between the electrochemical test paper D3# and the electrochemical test paper 1# is as follows: soaking the test paper in a polyethyleneimine solution with the mass concentration of 20% once, soaking the test paper in a polyacrylic acid solution with the mass concentration of 20% twice, waiting for the electrodeposition of the nanogold, soaking the test paper in a polyacrylic acid solution with the mass concentration of 20% once, and obtaining the electrochemical test paper D3# by the same steps as the electrochemical test paper 1# and storing the test paper in a dry environment for later use.
Test example 1 uric acid detection
The detection instrument of the test example is carried out by using Chenghua CHI1000C electrochemical workstation, but the electrochemical test paper detection of the invention is not limited to the instrument and is also suitable for other electrochemical workstations.
The working electrode of the electrochemical test paper 1# -10# prepared in the example 1 is connected to a Chenhua working circuit, 2 μ L of uric acid standard solution samples with the concentrations of 300 μ M, 500 μ M and 1000 μ M are respectively dropped into the working area of the electrode, each group measures 8 times, a Chenhua CV program is run, the measuring potential range is set to be-0.2V to 0.6V, the number of scanning cycles is one circle, the sensitivity range is proper, and the other sets are defaults and the program is run.
And (3) continuously running an it program by using the sample, setting the potential to be 0.3V, setting the scanning time to be 20s, and after the program is run, washing with pure water, drying by blowing, placing and storing.
The test data of each group was measured 8 times, and the average value, standard deviation and relative standard deviation were calculated from the current values at 9s to obtain the test data statistical table shown in table 1.
Table 1 statistical table of test data
As shown in Table 1, the relative standard deviation of the electrochemical test paper 4# and 5# for testing three uric acid samples with the concentrations is less than 4%, the electrochemical test paper 1# to 3# is not subjected to post-treatment, and the relative standard deviation of the uric acid samples with the concentrations of 300 mu M is larger than 18%, which indicates that the electrochemical test paper is stable and the detection effect is improved by the post-treatment. The types of polymers used in the electrochemical test paper 6# are different, and the relative standard deviation of uric acid samples with three concentrations is greater than that of the electrochemical test paper 4# and 5#, so that the polymer polyethyleneimine solution and the polyacrylic acid solution have better film forming uniformity than other polymers containing amino and carboxyl, the adhesion capability of the gold nanoparticles can be improved, and the stability is improved. The polymerization degrees of the polymers used in the electrochemical test paper 7# are different, and the relative standard deviation of uric acid samples with three concentrations is greater than that of the electrochemical test paper 4# and 5#, so that the polyethyleneimine can be used for trapping metal, namely deposition and attachment of nanoparticles when the polymerization degree is about 100, and if the molecular weight of the polymers is too large, the stability of the electrochemical test paper is not facilitated. The concentrations of chloroauric acid solutions used in the electrochemical test paper 8# are different, the relative standard deviations of uric acid samples with three concentrations are larger than those of the electrochemical test paper 4# and 5#, the specific surface area of the deposition morphology is largest, the size of deposition particles is the most uniform, and the deposition particles are distributed most uniformly when the chloroauric acid solutions are at the concentration of 50-100 mM, and the stability of the electrochemical test paper is not facilitated when the chloroauric acid solutions exceed the concentrations. The electrochemical test paper 9# chloroauric acid solution has different dropping amounts, and the relative standard deviation of uric acid samples with three tested concentrations is greater than that of electrochemical test paper 4# and 5# because the solution dropped in the working area of the electrode has the largest surface tension in the range of 10-30 muL, and gathered liquid drops are easily formed, so that the electrodeposition action is more uniform in the working electrode area, the dropping amount is large, the liquid drops are easily dispersed, and the deposition effect is influenced. The deposition time of the electrochemical test paper 10# is different, and the relative standard deviation of the uric acid samples with the three concentrations is greater than that of the electrochemical test paper 4# and 5#, so that the long electrodeposition time is not beneficial to the stability of the electrochemical test paper.
The polymer used for the comparative electrochemical test paper D1# has no amino group or carboxyl group, the comparative electrochemical test paper D2# is only soaked in polyacrylic acid solution and not soaked in polyethyleneimine solution, the comparative electrochemical test paper D3# is soaked in two polymers in different sequences, and the final test result is far greater than that of the electrochemical test paper 4# and 5#, because polyacrylic acid solution and polyethyleneimine solution can react at normal temperature, and polyethyleneimine with positive charge and polyacrylic acid with negative charge can form physical crosslinking, so that an interface film formed on the surface of an electrode after being soaked layer by layer is tighter. Meanwhile, polyethyleneimine exists in water as a polymeric cation and can neutralize and adsorb anionic substances, and polyacrylic acid solution exists in solution as a polymeric anion and can neutralize and adsorb cationic substances, so that the electrochemical test paper has good repeatability and high stability.
Test example 2 uric acid assay
The electrochemical test paper prepared in example 1 was stored in a dry environment for 6 months, and then 2. Mu.L of uric acid standard solution samples having concentrations of 300. Mu.M, 500. Mu.M, and 1000. Mu.M were dropped onto the electrode working area, respectively, according to the test method in test example 1, and the measurement data of each group was measured 8 times, and the average value, the standard deviation, and the relative standard deviation were calculated from the current value at 9 seconds, to obtain the statistical table of the test data shown in Table 2.
Table 2 statistical table of test data
As shown in table 2, the test results of the electrochemical test paper prepared in example 1 after being stored in a dry environment for 6 months have the same trend as the data in table 2, and particularly, the relative standard deviation of uric acid samples with three concentrations tested after 6 months in the electrochemical test paper 4# and 5# is still less than 4%, which indicates that the electrochemical test paper has good stability, long shelf life and good precision.
As shown in fig. 1, which is a scanning electron microscope image of the flower-like nano-gold particles obtained by electrodeposition during the preparation of the electrochemical test paper 1# in example 1, it can be seen from fig. 1 that the nano-gold particles obtained by electrodeposition form gold popcorn structures with many spurs, because the polymer polyethyleneimine with positive charge and the polymer polyacrylic acid with negative charge can form physical cross-linking, a film formed on the surface of an electrode after being soaked layer by layer is more compact. Because the interfacial film is compact, the formed gold nucleation sites are more dense, and the polyethyleneimine and polyacrylic acid gel have homogeneous structures, and the homogeneous properties of the polyethyleneimine and polyacrylic acid gel can enable the nucleation sites to be more uniform and dispersed, so that the deposited gold nanoparticles have the effects of uniform size and uniform distribution. The conductive polymer polyethyleneimine with the reduction property can help to realize in-situ reduction of an interface and improve the load capacity. The conductive polymer enables the deposited gold particles not to fall off easily, and can induce the crystal face orientation growth of gold again, and the polyacrylic acid and the polyethyleneimine polymer used in the application can induce the deposited gold particles to form a gold popcorn-shaped structure with a plurality of spurs, so that the specific surface area of gold is increased, the electron transfer rate is accelerated, and the detection of uric acid is facilitated.
As shown in fig. 2, which is a scanning electron microscope image of the flower-like gold nanoparticles obtained by electrodeposition during the preparation of electrochemical test paper 4#, it can be seen from fig. 2 that the shapes of the gold nanoparticles obtained by electrodeposition are blunt-ended but have many protrusions on the particle surface due to post-treatment.
As shown in FIG. 3, which is a scanning electron microscope image of flower-like gold nanoparticles obtained by electrodeposition during the preparation of the electrochemical test paper No. 10#, it can be seen from FIG. 3 that the gold nanoparticles obtained by electrodeposition have snowflake dendritic structures, are highly non-uniform, and are non-uniform in size.
As shown in fig. 4, which is a scanning electron microscope image comparing flower-like gold nanoparticles obtained by electrodeposition during the preparation of electrochemical test paper D3#, it can be seen from fig. 4 that the gold nanoparticles obtained by electrodeposition form large clusters and small clusters which are alternately distributed, most of the flower-like clusters have rounded ends, and the flower petals are rare.
Fig. 5 is a graph obtained by detecting electrochemical test paper 4# by cyclic voltammetry, and it can be seen from fig. 5 that the highest current of uric acid standard solution samples with concentrations of 300, 500, and 1000 μ M is obtained when the oxidation peak potential is about 0.3V, and it can be determined that the sample to be detected contains uric acid.
FIG. 6 shows the test curve of electrochemical test paper 4# by current time method, and FIG. 6 shows that the uric acid standard solution sample current value of 1000 μ M uric acid standard solution sample current value > 500 μ M uric acid standard solution sample current value > 300 μ M uric acid standard solution sample current value, therefore, the uric acid concentration and the current value are in positive correlation, and the higher the uric acid concentration, the larger the current.
The above description is only an example of the present invention and is not intended to limit the present invention. Various modifications and alterations to this invention will become apparent to those skilled in the art. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention should be included in the scope of the claims of the present invention.
Claims (10)
1. The preparation method of the electrochemical test paper for uric acid detection comprises a working electrode and a counter electrode, and is characterized by comprising the following steps of:
at least respectively soaking the working electrode in a first polymer solution and a second polymer solution to obtain a polymer modified working electrode, wherein one of the first polymer solution and the second polymer solution is a polymer solution containing primary amine and/or secondary amine, and the other is a polymer solution containing carboxyl;
and depositing flower-shaped nano gold particles on the surface of the working electrode modified by the polymer to obtain the electrochemical test paper.
2. The method according to claim 1, wherein the first polymer solution is a polyacrylic acid solution, and the second polymer solution is a polyethyleneimine solution;
preferably, the first polymer solution is a polyacrylic acid solution with the mass concentration of 15-30wt%, and more preferably 20wt%; and/or
The second polymer solution is a polyethyleneimine solution with a mass concentration of 15-30wt%, and more preferably 20%.
3. The method according to claim 1, wherein the degree of polymerization of the polymer in the first polymer solution is 80 to 120, preferably 100;
the polymerization degree of the polymer in the second polymer solution is 80 to 120, preferably 100.
4. The preparation method according to any one of claims 1 to 3, characterized in that a chloroauric acid solution with a concentration of 50-100 mM is dripped on the surface of the polymer modified working electrode, and flower-shaped nano gold particles are formed by electrodeposition;
preferably, the dropping amount of the chloroauric acid solution is 10-30 mu L.
5. The production method according to claim 4, wherein the electrodeposition employs a potentiostatic deposition method;
preferably, the deposition voltage of the electrodeposition is-0.4 to-0.2V, and the deposition time of the electrodeposition is 300 to 900s.
6. The preparation method according to any one of claims 1 to 3, characterized in that the electrochemical test paper is obtained by post-treating the working electrode modified by the flower-shaped nano-gold particles by cyclic voltammetry and amperometry,
preferably, the working electrode modified by the flower-shaped nano gold particles is subjected to post-treatment by sequentially adopting a primary cyclic voltammetry method and a primary current time method.
7. The preparation method according to claim 6, characterized in that uric acid buffer solution is dripped on the electrochemical test paper of the working electrode modified by the flower-shaped nano gold particles, the test voltage interval is set to be-0.2-0.6V, the number of scanning cycles is 1 cycle, and cyclic voltammetry treatment is carried out to obtain the oxidation peak potential value; taking the oxidation peak potential value as a circuit voltage, and carrying out current-time method treatment to obtain post-treated electrochemical test paper;
preferably, 300-1000 μ M of uric acid is dissolved in 0.2M PBS buffer to obtain a uric acid buffer solution with pH of 7.35-7.45, and the dropping amount of the uric acid buffer solution is 15-25 μ L.
8. The method of any one of claims 1 to 3, wherein the electrochemical test strip further comprises a reference electrode;
preferably, the working electrode, the counter electrode and the reference electrode in the electrochemical test paper are screen-printed electrodes, wherein the working electrode is a carbon electrode, the counter electrode is a carbon electrode, and the reference electrode is a silver/silver chloride electrode.
9. An electrochemical test strip obtained by the method of any one of claims 1 to 8.
10. Use of the electrochemical test strip obtained by the preparation method according to any one of claims 1 to 8 for detecting uric acid.
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