CN115015354A - Current type nitrite sensor based on bionic enzyme ion selective membrane and preparation - Google Patents
Current type nitrite sensor based on bionic enzyme ion selective membrane and preparation Download PDFInfo
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Abstract
The invention relates to a current type nitrite sensor based on a bionic enzyme ion selective membrane, a preparation method and application. The sensor comprises a working electrode, a porous conductive skeleton, chemically modified conductive particles and a conductive electrode; the porous conductive skeleton and the bionic enzyme ion selective membrane are covalently combined through chemically modified conductive particles, and the material of the bionic enzyme ion selective membrane comprises vitamins, proteins, metal oxides, high molecular polymers and the like which have a chelation effect on nitrite. When the sensor is used for detecting the nitrite content in a sample, nitrite can be specifically identified and chelated, and the sensor is insensitive to other interfering substances, so that specific detection of nitrite is realized. Meanwhile, after the bionic enzyme ion selective membrane chelates nitrite, the generation of oxidation-reduction reaction can be rapidly catalyzed, electron transfer is promoted through the porous conductive framework, and the sensitive and rapid detection of the current type nitrite is realized.
Description
Technical Field
The invention relates to the technical field of sensors, in particular to a current type nitrite sensor based on a bionic enzyme ion selective membrane, a preparation method and application.
Background
Nitrate and nitrite are nitrogen-containing compounds that are commonly found in nature, and are widely present in water, air, soil, plants, and the like. Wherein, the action of nitrite on the body includes both beneficial regulation and harmful regulation. Continuous excess nitrite is easy to cause serious diseases such as upper digestive tract cancer, lactic acidosis, spontaneous abortion and the like; the nitrite with proper concentration can effectively prevent and treat fatty liver and regulate cardiovascular function. Therefore, the development of a detection device for nitrite content is of great significance.
For the detection of nitrite content, the traditional detection methods mainly include high performance liquid chromatography, chemiluminescence methods relying on azo compound color development, catalytic luminescence methods relying on fluorescent probes, and the like. Although the methods can sensitively detect the content of the nitrite, the methods usually need to use complex instruments and equipment, and the sample pretreatment is complex, the detection time is long, the methods can only be intensively detected in a hospital laboratory, and the methods are difficult to popularize in the aspects of home self-detection and the like. Subsequently, a detection method based on an electrochemical sensor is carried out, the detection method greatly simplifies the detection flow of the nitrite content, improves the detection efficiency of the nitrite content, and promotes the portable detection of the nitrite content. However, the traditional detection method based on the electrochemical sensor has difficulty in combining high sensitivity and specificity, which restricts the development of the sensor for electrochemically detecting nitrite to a certain extent.
Disclosure of Invention
Based on the above, there is a need for an amperometric nitrite sensor based on a biomimetic enzyme ion selective membrane, and a preparation method and application thereof. The sensor can effectively give consideration to both the sensitivity and the specificity of nitrite content detection.
In order to solve the technical problems, the technical scheme of the invention is as follows:
a current type nitrite sensor based on a bionic enzyme ion selective membrane comprises a working electrode, a porous conductive framework, chemically modified conductive particles and the bionic enzyme ion selective membrane; the porous conductive framework is positioned on the surface of the working electrode, and the porous conductive framework and the bionic enzyme ion selective membrane are covalently combined through the chemically modified conductive particles; the bionic enzyme ion selective membrane is made of at least one of vitamins, proteins, metal oxides and polymers which have a chelating effect on nitrite.
In one embodiment, the material of the biomimetic enzyme ion selective membrane comprises at least one of a vitamin containing cobalt ions, an oxide, a polymer and a nitrite transporter.
In one embodiment, the bionic enzyme ion selective membrane is made of cobalt ion-containing cobalamin and Co 3 O 4 At least one of cobalt (II) -disalicylaldehyde o-phenylenediamine, cobalt (II) -tetraaminophthalocyanine and nitrite oxidase.
In one embodiment, the material of the porous conductive skeleton comprises at least one of a porous conductive metal and a porous conductive non-metal.
In one embodiment, the porous conductive metal comprises at least one of gold foam, nickel foam, and titanium foam.
In one embodiment, the porous conductive non-metal comprises at least one of carbon foam, conductive polymer, conductive media doped insulating material.
In one embodiment, the conductive medium includes at least one of graphene, carbon nanotubes, and nano-metal particles.
In one embodiment, the insulating material comprises at least one of a protein, an insulating polymer, and a ceramic.
In one embodiment, the chemically modified conductive particles include at least one of amino-modified conductive particles, carboxyl-modified conductive particles, and thiol-modified conductive particles.
In one embodiment, the conductive particles include at least one of carbon nanotubes, nano-metal particles, and conductive polymers.
A preparation method of the amperometric nitrite sensor based on the bionic enzyme ion selection membrane in any embodiment comprises the following steps;
dispersing the material of the porous conductive framework and the chemically modified conductive particles in a solvent to prepare a dispersion liquid;
mixing the material of the bionic enzyme ion selective membrane with the dispersion liquid to prepare electrode modification liquid;
transferring the electrode modification liquid to the surface of the working electrode to prepare a sensor preform;
and carrying out heat treatment on the sensor preform.
In one embodiment, the solvent is water.
In one embodiment, the mass ratio of the material of the porous conductive skeleton, the material of the bionic enzyme ion selective membrane and the chemically modified conductive particles is (0.6-8): (0.3-3): 0.1-1).
The amperometric nitrite sensor based on the bionic enzyme ion selection membrane, which is described in any embodiment, or the amperometric nitrite sensor based on the bionic enzyme ion selection membrane, which is prepared by the preparation method described in any embodiment, is applied to nitrite content detection.
In one embodiment, the application comprises the steps of:
dropwise adding a solution to be detected on the surface of the bionic enzyme ion selective membrane, detecting a steady-state current generated by the solution to be detected, and further obtaining the nitrite content of the solution to be detected according to the steady-state current.
The current type nitrite sensor based on the bionic enzyme ion selection membrane comprises a working electrode, a porous conductive framework, chemically modified conductive particles and the bionic enzyme ion selection membrane; the porous conductive skeleton and the bionic enzyme ion selective membrane are covalently combined through chemically modified conductive particles, and the bionic enzyme ion selective membrane is made of vitamins, proteins, metal oxides, high molecular polymers and the like which have a chelating effect on nitrite. When the sensor is used for detecting the nitrite content in a sample, the sensor can specifically identify and chelate nitrite, is insensitive to other interfering substances, can intercept other interfering substances in the sample, and realizes the specific detection of nitrite. Meanwhile, after the bionic enzyme ion selective membrane chelates nitrite, the generation of oxidation-reduction reaction can be quickly catalyzed, and electron transfer is promoted through the porous conductive framework, so that sensitive and quick detection of nitrite is realized. In addition, the covalent binding effect provided by the chemically modified conductive particles effectively increases the adsorption density of the bionic enzyme material on the surface of the porous conductive framework, optimizes the binding stability of the bionic enzyme ion selection membrane on the surface of the porous conductive framework, improves the specific binding capacity of the sensor on nitrite, and better considers the sensitivity and specificity of the sensor on nitrite detection.
Further, the nitrite sensor based on current detection is obtained by designing the nitrite sensor, and the content of nitrite in the solution to be detected can be obtained through the current generated by the solution to be detected. Compared with the traditional mode of detecting the nitrite content based on electric potential, the invention provides a novel nitrite sensor.
Furthermore, the current type nitrite sensor based on the bionic enzyme ion selective membrane with small volume is obtained by designing the sensor, and the sensor can be conveniently integrated with other common equipment to be made into nitrite detection equipment which is convenient to carry and has wide application range.
In the preparation method of the current type nitrite sensor based on the bionic enzyme ion selective membrane, the electrode modification solution is prepared by dispersion and mixing, the electrode modification solution is transferred to the surface of the working electrode to prepare the preformed sensor product, and then the preformed sensor product is subjected to heating treatment.
Drawings
FIG. 1 is a schematic structural diagram of an amperometric nitrite sensor based on a biomimetic enzyme ion selective membrane according to an embodiment of the present invention;
FIG. 2 is a schematic view of an electrode interface of the sensor corresponding to FIG. 1;
FIG. 3 is a physical representation of a microscopic topology of the electrode interface of the corresponding sensor of FIG. 1;
FIG. 4 is a schematic diagram showing the sensitivity of the biomimetic enzyme ion selective membrane-based amperometric nitrite sensor in nitrite calibration solution obtained in embodiment 1 of the present invention;
FIG. 5 is a schematic diagram of the selectivity test of the amperometric nitrite sensor based on the biomimetic enzyme ion selective membrane obtained in example 1 of the present invention;
fig. 6 is a schematic diagram of the detection performance of the amperometric nitrite sensor based on the bionic enzyme ion selective membrane in the artificial saliva simulation sample obtained in embodiment 1 of the present invention.
The symbols in the figure illustrate:
100. an amperometric nitrite sensor based on a bionic enzyme ion selective membrane; 101. a working electrode; 102. a porous conductive skeleton; 103. chemically modified conductive particles; 104. a bionic enzyme ion selection membrane; 105. a counter electrode; 106. a reference electrode; 200. an electrode substrate.
Detailed Description
In order to make the aforementioned objects, features and advantages of the present invention comprehensible, embodiments accompanied with figures are described in detail below. In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. As used herein, the term "and/or" includes any and all combinations of one or more of the associated listed items.
Referring to fig. 1 and 2, an embodiment of the invention provides an amperometric nitrite sensor 100 based on a biomimetic enzyme ion selective membrane. The current type nitrite sensor 100 based on the bionic enzyme ion selective membrane comprises a working electrode 101, a porous conductive framework 102, chemically modified conductive particles 103 and a bionic enzyme ion selective membrane 104; the porous conductive framework 102 is positioned on the surface of the working electrode 101, and the porous conductive framework 102 and the bionic enzyme ion selective membrane 104 are covalently combined through chemically modified conductive particles 103; the bionic enzyme ion selective membrane 104 is made of at least one of vitamins, proteins, metal oxides and polymers having a chelating effect on nitrite.
It will be appreciated that the biomimetic enzyme ion selective membrane 104 is more porous than the conductive backbone 102, further away from the surface of the working electrode 101.
When the nitrite sensor 100 in the embodiment is used for detecting the nitrite content in a sample, nitrite can be specifically identified and chelated, and the nitrite sensor is insensitive to other interfering substances, so that other interfering substances in the sample can be effectively intercepted, and the specific detection of nitrite is realized. Meanwhile, after the bionic enzyme ion selective membrane 104 chelates nitrite, the occurrence of oxidation-reduction reaction can be rapidly catalyzed, and electron transfer is promoted through the porous conductive framework 102, so that sensitive and rapid detection of nitrite is realized. In addition, the covalent binding effect provided by the chemically modified conductive particles 103 effectively optimizes the binding stability of the bionic enzyme ion selective membrane 104 on the surface of the porous conductive framework 102. The nitrite sensor 100 in this embodiment improves the specific binding ability of the sensor to nitrite, and better considers the sensitivity and specificity of the sensor to nitrite detection.
Further, in the present embodiment, by designing the sensor, the nitrite sensor 100 based on current detection is obtained, and the content of nitrite in the solution to be detected can be obtained through the current generated by the solution to be detected. Compared with the traditional mode of detecting the nitrite content based on electric potential, the embodiment provides a novel nitrite sensor.
In one specific example, the material of working electrode 101 includes, but is not limited to, conductive carbon paste, silver paste, gold paste, conductive polymer, and the like. Further, the conductive paste may be modified using prussian blue or prussian blue analogs.
In a specific example, the nitrite sensor 100 selects the bionic enzyme ion selective membrane 104 having a chelating effect on nitrite, so that specific recognition and chelation can be performed on nitrite, and then the redox reaction of nitrite is catalyzed under a given voltage, thereby improving the specificity and selectivity of the sensor. Optionally, the given voltage range is 0.4V to 1.2V, and the working voltage can be selected according to the actual use condition and the specific modification material of the working electrode. The electrons lost by the nitrite in the oxidation reaction are then transferred to the electrode interface through the porous conducting skeleton 102, at which time the corresponding current signal can be acquired using an electrochemical workstation. Meanwhile, the porous conductive framework 102 improves the transfer efficiency of electrons, and can effectively improve the sensitivity of the sensor.
Further, the nitrite sensor 100 of the present embodiment relies on strong field interactions between ions and redox reactions, without involving biological enzymes or other labile substances. Therefore, the nitrite sensor 100 is less affected by conditions such as external temperature and pH during storage and use, and has high stability and long service life under daily use conditions.
Furthermore, the nitrite sensor 100 in this embodiment has a small volume, compared with the conventional nitrite sensor, the nitrite sensor has a small and portable volume, the detection process is simple and quick, complex sample pretreatment steps are not needed, the dependence on large-scale instruments and professional operators in the conventional nitrite content detection process is eliminated, the difficulty in nitrite content detection is effectively reduced, and great convenience is brought to the nitrite content detection by consumers. In addition, the nitrite sensor 100 in the present embodiment can be integrated with a detection device such as a POCT (point-of-care testing) device or a microfluidic chip, so as to construct a medical apparatus for non-disease diagnosis, health management, and remote medical treatment, and has good versatility. In addition, the nitrite sensor 100 in this embodiment has good biocompatibility, and can be integrated with devices such as a flexible wearable device and an implanted microneedle, so as to realize real-time dynamic monitoring of nitrite content in an organism.
In one particular example, nitrite sensor 100 further comprises counter electrode 105 and reference electrode 106, with counter electrode 105, reference electrode 106, and working electrode 101 forming a three-electrode system. In the example, the nitrite sensor based on the electrochemical three-electrode system is constructed, which is beneficial to further improving the accuracy of the sensor in detecting the nitrite content. It will be appreciated that in the design and manufacture of nitrite sensor 100, the size, lead location, length, number, etc. of the electrodes may be designed according to actual requirements.
It will be appreciated that the nitrite sensor 100 further comprises an electrode substrate 200, the electrode substrate 200 being adapted to carry the working electrode 101. Alternatively, the electrode substrate 200 includes a hard substrate or a flexible substrate. As an alternative example of the material of the hard substrate, the material of the hard substrate includes at least one of silicon, glass, cyclic olefin polymer (COC), Polytetrafluoroethylene (PTFE), and polymethyl methacrylate (PMMA). As an alternative example of the material of the flexible substrate, the material of the flexible substrate includes at least one of Polydimethylsiloxane (PDMS), Polyamide (PI), Polyurethane (PU), and polyethylene terephthalate (PET). It will also be appreciated that during the preparation of the nitrite sensor 100, a suitable electrode substrate 200 may be selected depending on the actual integration scenario and usage scenario of the sensor, etc.
In a specific example, the nitrite sensor 100 further comprises a signal processor (not shown). A signal processor is connected to the working electrode 101 for collecting and processing the electrical signal transmitted from the working electrode. Specifically, the signal processor is connected to the working electrode 101, the counter electrode 105 and the reference electrode 106 respectively for collecting and processing the electric signals transmitted from the electrodes. Alternatively, the connection between the signal processor and the electrode can be made by means of a lead wire led out of the electrode.
As the material selection of the biomimetic enzyme ion selective membrane 104, the material of the biomimetic enzyme ion selective membrane 104 includes at least one of vitamins having a chelating effect on nitrite, proteins having a chelating effect on nitrite, metal oxides having a chelating effect on nitrite, and polymers having a chelating effect on nitrite. Specifically, the material of the bionic enzyme ion selective membrane 104 comprises a vitamin containing cobalt ions,Nitrite transport protein, oxide containing cobalt, iron, nickel and other metals. Optionally, the bionic enzyme ion selective membrane 104 comprises cobalamin and Co 3 O 4 At least one of cobalt (II) -disalicylaldehyde o-phenylenediamine, cobalt (II) -tetraaminophthalocyanine and Nitrite Oxidase (NOR). Specifically, the material of the biomimetic enzyme ion selective membrane 104 includes cyanocobalamin (VB) 12 )。
As a material choice for the porous conductive skeleton 102, the material of the porous conductive skeleton 102 includes at least one of a porous conductive metal and a porous conductive non-metal. Optionally, the porous conductive metal comprises at least one of gold foam, nickel foam, and titanium foam. The porous conductive non-metal comprises at least one of carbon foam, conductive polymer, conductive media doped insulating material. Specifically alternatively, the conductive polymer may be polyethylenedioxythiophene.
In a particular example, the conductive medium includes at least one of graphene, carbon nanotubes, and nano-metal particles. The insulating material includes at least one of a protein, an insulating polymer, and a ceramic. It is understood that the protein may be a porous protein, the polymer may be a porous polymer, and the ceramic may be a porous ceramic.
Preferably, the porous conductive framework 102 adopts a porous protein framework with good biocompatibility as a substrate, and a conductive medium is used for coating and modifying to construct a protein framework structure with conductive performance, that is, a porous protein framework doped with a conductive medium is constructed as the porous conductive framework 102. The porous conductive framework 102 provides a larger specific surface area for doping of a conductive medium, greatly improves the fixation density of the bionic enzyme ion selective membrane 104 on the surface of the working electrode 101, provides more attachment sites for a bionic enzyme material, and simultaneously realizes the improvement of detection sensitivity and selectivity. Alternatively, the protein may be Bovine Serum Albumin (BSA), ovalbumin, casein, hemoglobin, myoglobin, cytochrome C, and the like.
Further, referring to fig. 3, the porous conductive skeleton 102 is a conductive protein skeleton structure with uniform voids. The bionic enzyme ion selective membrane 104 is covalently modified on the surface of the porous conductive framework 102 by the chemically modified conductive particles 103 to form a nitrite sensor electrode interface together. At this time, the nitrite sensor 100 sufficiently chelates nitrite in the sample to be detected, thereby realizing rapid electron transfer, effectively intercepting interfering substances in the sample to be detected, and ensuring the specificity and accuracy of detection.
In a specific example, the chemically modified conductive particles 103 include at least one of amino-modified conductive particles, carboxyl-modified conductive particles, and thiol-modified conductive particles. The amino-modified conductive particles, the carboxyl-modified conductive particles and the mercapto-modified conductive particles are more beneficial to improving the stability of the electrode interface of the nitrite sensor. Optionally, the conductive particles comprise at least one of carbon nanotubes, nano-metal particles, and conductive polymers. Alternatively, the conductive particles may be multi-walled carbon nanotubes, gold nanoparticles, silver nanoparticles, or the like.
The invention also provides a preparation method of the current type nitrite sensor based on the bionic enzyme ion selective membrane. The preparation method comprises the following steps; dispersing the material of the porous conductive framework and the chemically modified conductive particles in a solvent to prepare a dispersion liquid; mixing the material of the bionic enzyme ion selection membrane with the dispersion liquid to prepare an electrode modification liquid; transferring the electrode modification liquid to the surface of the working electrode to prepare a sensor preform; and carrying out heating treatment on the sensor preform. In the preparation method, electrode modification liquid is prepared through dispersion and mixing, the electrode modification liquid is transferred to the surface of a working electrode, a sensor preformed product is prepared, and then the sensor preformed product is subjected to heating treatment. The preparation method is simple and easy to implement, does not need to depend on severe conditions and equipment, and is suitable for large-scale popularization.
In one specific example, the transferring of the electrode modification solution to the surface of the working electrode is dropping the electrode modification solution to the surface of the working electrode.
In a specific example, the material of the biomimetic enzyme ion selective membrane is mixed with the dispersion by dropping the material of the biomimetic enzyme ion selective membrane into the dispersion.
In one particular example, the solvent is water. It is understood that ultrapure water can be used as the solvent to improve the preparation accuracy of the sensor.
In the preparation method of the nitrite sensor, the mass ratio of the material of the porous conductive framework, the material of the bionic enzyme ion selective membrane and the chemically modified conductive particles is (0.6-8): (0.3-3): 0.1-1). Specifically, the mass percent of the porous conductive skeleton material is 0.6-8%, the mass percent of the bionic enzyme ion selective membrane material is 0.3-3%, and the mass percent of the chemically modified conductive particles is 0.1-1%. Alternatively, the mass percentage of the material of the porous conductive skeleton may be 0.6%, 0.8%, 1%, 1.5%, 1.8%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, or 8% in terms of mass percentage of the solvent. Alternatively, the mass percentage of the material of the biomimetic enzyme ion selective membrane may be 0.3%, 0.5%, 0.8%, 1%, 1.5%, 1.8%, 2%, 2.5% or 3% by mass of the solvent. Alternatively, the chemically modified conductive particles may be 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, or 1% by mass in terms of a mass percentage of the solvent.
In one particular example, the material of the porous framework comprises a conductive dielectric doped insulating material. At this time, the preparation method of the nitrite sensor comprises the following steps; dispersing a conductive medium, an insulating material and chemically modified conductive particles in a solvent to prepare a dispersion liquid; mixing the material of the bionic enzyme ion selection membrane with the dispersion liquid to prepare an electrode modification liquid; transferring the electrode modification liquid to the surface of the working electrode to prepare a sensor preform; the sensor preform is subjected to a heat treatment.
In one particular example, the material of the porous framework comprises a conductive dielectric doped insulating material. In this example, the mass ratio of the conductive medium to the insulating material is (0.1-3): 0.5-5. Specifically, the mass percent of the conductive medium is 0.1-3%, and the mass percent of the insulating material is 0.5-5%, based on the mass percent of the solvent. Alternatively, the mass percentage of the conductive medium may be 0.1%, 0.3%, 0.5%, 0.8%, 1%, 1.5%, 1.8%, 2%, 2.5%, or 3% in terms of mass percentage of the solvent. Alternatively, the mass percentage of the insulating material may be 0.5%, 0.6%, 0.8%, 1%, 1.5%, 1.8%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5% in terms of mass percentage of the solvent.
Further, the insulating material is protein. In this case, the mass ratio of the conductive medium to the protein is (0.1-3) to (0.5-5). Specifically, the mass percent of the conductive medium is 0.1-3%, and the mass percent of the protein is 0.5-5%, based on the mass percent of the solvent. Alternatively, the mass percentage of the conductive medium may be 0.1%, 0.3%, 0.5%, 0.8%, 1%, 1.5%, 1.8%, 2%, 2.5%, or 3% in mass percentage of the solvent. Alternatively, the mass percentage of the protein may be 0.5%, 0.6%, 0.8%, 1%, 1.5%, 1.8%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5% in terms of mass percentage of the solvent.
In a specific example, the temperature of the heating treatment is 37 ℃ to 55 ℃, and the time of the heating treatment is 30min to 120 min. Optionally, the temperature of the heat treatment is 37 ℃, 40 ℃, 42 ℃, 45 ℃, 48 ℃, 50 ℃, 52 ℃ or 55 ℃. The heating time is 30min, 40min, 50min, 60min, 70min, 80min, 90min, 100min, 110min or 120 min.
It is understood that when the insulating material is protein, the protein undergoes secondary structure restructuring to form a porous protein skeleton structure upon heat treatment.
The invention also provides an embodiment of the current-mode nitrite sensor based on the bionic enzyme ion selective membrane, or the application of the current-mode nitrite sensor based on the bionic enzyme ion selective membrane prepared by the preparation method in nitrite content detection.
Specifically, another embodiment of the present invention provides an amperometric nitrite sensor based on a biomimetic enzyme ion selective membrane, or an application of the amperometric nitrite sensor based on the biomimetic enzyme ion selective membrane prepared by the above preparation method in nitrite content detection for non-disease diagnosis purposes.
In one specific example, the application includes the steps of: dropwise adding a solution to be detected on the surface of the bionic enzyme ion selective membrane, detecting a steady-state current generated by the solution to be detected, and further obtaining the nitrite content of the solution to be detected according to the steady-state current.
It is understood that for working electrodes modified with different materials and proportions, the following steps are also included: carrying out nitrite content-steady state current calibration on the current type nitrite sensor based on the bionic enzyme ion selection film.
In one embodiment, the nitrite content-steady state current calibration comprises the steps of: dripping calibration solutions with different known nitrite contents on the surface of a bionic enzyme ion selection membrane of the sensor, and detecting calibration steady-state currents generated by the different calibration solutions; and obtaining the relation of the change of the steady-state calibration current along with the nitrite content of the calibration solution.
In one specific example, the application comprises the steps of: dripping calibration solutions with different known nitrite contents on the surface of a bionic enzyme ion selection membrane of the sensor, and detecting calibration steady-state currents generated by the different calibration solutions; obtaining the change relation of the steady-state calibration current along with the nitrite content of the calibration solution; and dripping the solution to be detected on the surface of the bionic enzyme ion selection membrane, detecting the steady-state current generated by the solution to be detected, and further obtaining the nitrite content of the solution to be detected according to the relation between the steady-state current and the change. In the example, the sensor is calibrated to obtain the variation relation of the current with the nitrite content of the solution, and then the nitrite content of the solution to be detected is obtained according to the variation relation and the current of the solution to be detected. The nitrite content and the current obtained based on the sensor in the embodiment have high correlation, and the nitrite content in the sample to be detected can be accurately determined according to the current information.
In another embodiment, the invention provides a device for detecting nitrite content. The device comprises the current type nitrite sensor based on the bionic enzyme ion selective membrane.
Specifically, the device for detecting the nitrite content comprises a POCT device, a microfluidic chip and portable electronic equipment.
The following are specific examples.
Example 1
The preparation method of the current-mode nitrite sensor based on the bionic enzyme ion selective membrane comprises the following steps:
s101: in terms of mass percent of ultrapure water, 0.5% of multi-walled carbon nanotubes (MWCNTs), 2% of BSA and 1% of carboxyl modified gold nanoparticles (more than 99%, purchased from Macklin company) in mass percent are added into ultrapure water to be uniformly dispersed, so as to obtain a dispersion liquid.
S102: 3 percent of cyanocobalamin (VB) by mass percentage 12 ) And adding the mixture into the dispersion liquid to obtain the electrode modification liquid.
S103: and uniformly dropwise adding 8 mu L of electrode modification liquid on the surface of a working electrode with the diameter of 4mm to obtain a sensor preform.
S104: and heating the sensor preform at 55 ℃ for 60min to obtain the sensor. In the heating process, the sensor preform is incubated, so that the BSA protein is subjected to secondary structure reconstruction to form a porous protein skeleton structure, and meanwhile, the carboxyl modified carbon nano tube modifies the protein skeleton to obtain a biocompatible protein skeleton structure with good electronic conduction performance and large specific surface area. One end of the nano gold particle modified by carboxyl as an immobilization layer is combined with the amino of the biomimetic enzyme cyanocobalamin, the other end is combined with the amino residue in BSA, and the cyanocobalamin biomimetic enzyme ion selective membrane is immobilized on the surface of the electron transfer layer.
And (3) calibrating the sensor by adopting nitrite solutions with different concentration gradients, wherein the working voltage is 1V. The results are shown in FIG. 4. Wherein the upper left panel of figure 4 is a plot of nitrite sensing electrode calibration at low concentrations. As can be seen from fig. 4, in the sensor of the present embodiment, the nitrite content and the current have a high correlation, and the nitrite content in the sample to be measured can be accurately determined according to the current information. The sensor of this embodiment exhibits a significantly increased detection range and a reduced detection limit compared to conventional nitrite sensing electrodes. The detection range of the sensor in the embodiment spans 0.025 mM-45 mM, and the lowest detection limit can reach 1 nM. The detection range of the traditional nitrite sensing electrode is only 0.1 mM-0.7 mM, and the lowest detection limit is higher than 170 nM.
Fig. 5 shows a schematic diagram of the selective testing of the sensor in the present embodiment. As can be seen from fig. 5, the sensor does not generate a current response to the interferent in the solution to be measured, and is shown to be capable of well trapping the interferent in the solution to be measured.
Fig. 6 illustrates the detection performance of a sensor in an artificial saliva simulation sample in an embodiment of the invention. As can be seen from fig. 6, the detection accuracy in the simulated sample is as high as 98%, and the response current exhibits good linearity.
From the above experimental results, the sensor in this embodiment has great advantages in sensitivity, specificity, selectivity, and detection time.
All possible combinations of the technical features of the above embodiments may not be described for the sake of brevity, but should be considered as within the scope of the present disclosure as long as there is no contradiction between the combinations of the technical features.
The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the patent of the present invention should be subject to the appended claims, and the description and the drawings can be used for explaining the contents of the claims.
Claims (10)
1. A current type nitrite sensor based on a bionic enzyme ion selective membrane is characterized by comprising a working electrode, a porous conductive framework, chemically modified conductive particles and the bionic enzyme ion selective membrane; the porous conductive framework is positioned on the surface of the working electrode, and the porous conductive framework and the bionic enzyme ion selective membrane are covalently combined through the chemically modified conductive particles; the bionic enzyme ion selective membrane is made of at least one of vitamins, proteins, metal oxides and polymers which have a chelating effect on nitrite.
2. The amperometric nitrite sensor based on the biomimetic enzyme ion selective membrane of claim 1, wherein the material of the biomimetic enzyme ion selective membrane comprises cobalamin containing cobalt ions, Co 3 O 4 At least one of cobalt (II) -disalicylaldehyde o-phenylenediamine, cobalt (II) -tetraaminophthalocyanine and nitrite oxidase; and/or the presence of a gas in the gas,
the material of the porous conductive skeleton comprises at least one of a porous conductive metal and a porous conductive nonmetal.
3. The biomimetic enzyme ion selective membrane based amperometric nitrite sensor of claim 2, wherein the porous conductive metal comprises at least one of gold foam, nickel foam, and titanium foam; and/or the presence of a gas in the gas,
the porous conductive non-metal comprises at least one of carbon foam, conductive polymer and conductive medium doped insulating material.
4. The amperometric nitrite sensor based on the biomimetic enzyme ion-selective membrane of claim 3, wherein the conductive medium comprises at least one of graphene, carbon nanotubes, and nano-metal particles; and/or the presence of a gas in the gas,
the insulating material includes at least one of a protein, an insulating polymer, and a ceramic.
5. The amperometric nitrite sensor based on the biomimetic enzyme ion-selective membrane of claim 1, wherein the chemically modified conductive particles comprise at least one of amino-modified conductive particles, carboxyl-modified conductive particles, and thiol-modified conductive particles.
6. The biomimetic enzyme ion selective membrane based amperometric nitrite sensor of claim 5, wherein the conductive particles comprise at least one of carbon nanotubes, nano-metal particles, and conductive polymers.
7. The preparation method of the bionic enzyme ion selective membrane-based amperometric nitrite sensor as defined in any one of claims 1 to 6, which comprises the following steps;
dispersing the material of the porous conductive framework and the chemically modified conductive particles in a solvent to prepare a dispersion liquid;
mixing the material of the bionic enzyme ion selective membrane with the dispersion liquid to prepare electrode modification liquid;
transferring the electrode modification liquid to the surface of the working electrode to prepare a sensor preform;
and carrying out heat treatment on the sensor preform.
8. The production method according to claim 7, characterized in that the solvent is water; and/or the presence of a gas in the atmosphere,
the mass ratio of the material of the porous conductive framework, the material of the bionic enzyme ion selective membrane and the chemically modified conductive particles is (0.6-8): (0.3-3): 0.1-1).
9. The use of the biomimetic enzyme ion selective membrane based amperometric nitrite sensor of any one of claims 1-6 or the biomimetic enzyme ion selective membrane based amperometric nitrite sensor prepared by the preparation method of any one of claims 7-8 in nitrite content detection.
10. Use according to claim 9, characterized in that it comprises the following steps:
dropwise adding a solution to be detected on the surface of the bionic enzyme ion selective membrane, detecting a steady-state current generated by the solution to be detected, and further obtaining the nitrite content of the solution to be detected according to the steady-state current.
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| CN112748169A (en) * | 2020-12-24 | 2021-05-04 | 西南大学 | Preparation method of nano-particle bionic enzyme sensitive element, product and application thereof |
| CN113447544A (en) * | 2020-03-27 | 2021-09-28 | 清华大学 | Degradable transient NO sensor and preparation method thereof |
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| CN104316580A (en) * | 2014-10-20 | 2015-01-28 | 中南林业科技大学 | Method for detecting nitrites in water by using nanogold enzyme sensor |
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