A paper-based nitrite ion detection chip and a preparation method and application thereof

Abstract

The application relates to the field of water quality detection, in particular to a paper-based nitrite ion detection chip and a preparation method and application thereof. The paper-based nitrite ion detection chip comprises a bottom plate and a paper-based layer on the bottom plate; the paper-based layer comprises a color developing area and a sample inlet area which are in communication with each other; a detection reagent is prepositioned in the color developing area, and the detection reagent comprises mannitol, p-aminobenzenesulfonic acid, N-1-naphthyl ethylenediamine and ascorbic acid; wherein the detection reagent solution is dropped into the color developing area and then freeze-dried, so that the detection reagent is prepositioned in the color developing area. In the application, the filter paper is combined with the freeze-drying technology, the detection reagent is uniformly covered on the surface of the filter paper, a loose structure is formed, the uniformity of a color developing layer is effectively improved, and the reaction time is reduced.

Description

Technical Field

[0001] This invention relates to the field of water quality testing, specifically to a paper-based nitrite ion detection chip, its preparation method, and its application. Background Technology

[0002] Nitrite ions are nitrite anions (NO2) - Nitrite ions, widely present in the environment, are the most common nitrogen-containing compounds in nature. They have various applications in daily life and industry, such as in food processing, as additives in cosmetics and personal care products, water treatment, and chemical synthesis reactions. Large amounts of nitrite ions entering water bodies can cause eutrophication, leading to excessive algal growth, depletion of dissolved oxygen, algal blooms, food chain disruption, reduced biodiversity, and habitat degradation in rivers, lakes, and coastal waters. Excessive nitrates and nitrite ions also pose significant health risks. Statistics show that drinking water in areas with high rates of esophageal cancer generally has high nitrate levels. This is because nitrite ions react with human blood to oxidize methemoglobin, converting it to ferrous methemoglobin, thus impairing its oxygen-carrying capacity and causing methemoglobinemia. Therefore, the detection of nitrite ions in water bodies is of great importance for environmental protection and human health.

[0003] Among numerous methods for detecting nitrite ions, paper-based chip technology relies on the capillary action of the paper itself for liquid transport, avoiding the need for additional driving equipment and accelerating the portability of detection devices. Its low cost and environmental friendliness have also made it popular among researchers. Paper-based chip detection methods have been gradually applied to the research and development of environmental monitoring and pollutant analysis technologies, achieving a series of significant advancements. For nitrite ion detection using paper-based chips, the detection reagent needs to be pre-fabricated onto the paper chip. The method of reagent pre-fabrication has a crucial impact on the detection results. How to pre-fabricate the detection reagent onto the paper chip and how to design a suitable paper chip structure are problems that urgently need to be solved.

[0004] Currently, the detection of nitrite ions is attracting increasing attention. Patent application CN114798021B discloses a three-dimensional paper-based microfluidic chip for rapid detection of nitrite ions in water, its preparation method, and its application, relating to the field of microfluidic technology. The three-dimensional paper-based microfluidic chip has a four-layer structure: a cellophane cover, a quincunx-shaped paper chip, a circular colorimetric layer, and a cellophane pad layer; the cellophane cover also has a sample dispensing port located in the center of the cellophane cover. Patent application CN 115166173 A discloses a paper microfluidic system and detection method based on a smartphone app for multiple detection. This system uses a detection program installed on a smartphone and its camera to capture images of the paper microfluidic chip. Data analysis is then performed on the images of the paper microfluidic chip captured by the image acquisition module to obtain the HSV values ​​of each detection image. Finally, based on data stored in a database, the system detects the types and concentrations of ions in the solution to be tested, as well as the pH value of the solution. Patent application CN 116879285 A provides a kit, detection system, and related method for detecting trace amounts of nitrite ions. The kit includes a nitrite ion detection reagent comprising the following components in parts by weight: 0.5 to 50.0 parts of p-aminobenzenesulfonic acid, 0.1 to 20.0 parts of naphthylethylenediamine hydrochloride, 10.0 to 90.0% weighting agent, 1.0 to 60.0 parts of masking agent, and 0 to 10.0 parts of accelerator.

[0005] The aforementioned patent applications all involve directly soaking or adding the detection reagent to filter paper. However, the uniformity of color distribution after the reagent reacts with nitrite ions needs improvement, and the color development speed is relatively slow. Therefore, how to rapidly and uniformly detect nitrite ions is a problem worthy of further research. Summary of the Invention

[0006] The purpose of this invention is to overcome the problems of slow color development and uneven color distribution in existing nitrite ion detection methods, and to provide a paper-based nitrite ion detection chip, its preparation method, and its application.

[0007] To achieve the above objectives, the present invention provides a paper-based nitrite ion detection chip, the paper-based nitrite ion detection chip comprising a base plate and a paper base layer located on the base plate;

[0008] The paper substrate includes a color development area and a sample injection area that are interconnected;

[0009] The colorimetric region is pre-filled with a detection reagent, which includes mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid;

[0010] In this process, the test reagent solution is added dropwise to the color development area and then freeze-dried, so that the test reagent is pre-positioned in the color development area.

[0011] Preferably, the base plate is made of PP, PET or PDMS;

[0012] Preferably, the base plate is a rectangle with a length of 30-50mm and a width of 15-30mm.

[0013] Preferably, the color development area is a circle with a diameter of 3-10 mm, and the sample injection area 3 is a strip with a length of 5-15 mm and a width of 2-4 mm.

[0014] Preferably, the weight ratio of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid is 20-35:0.8-2.4:1.8-4.5:1.

[0015] A second aspect of the present invention provides a method for preparing a paper-based nitrite ion detection chip, the method comprising the following steps:

[0016] (1) The filter paper is cut using a laser engraving machine to obtain a paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected.

[0017] (2) The paper substrate is placed in liquid nitrogen for pre-freezing, and then the detection reagent solution is dropped onto the color development area, and then freeze-dried so that the detection reagent is pre-placed in the color development area;

[0018] (3) Provide a base plate and place the paper base obtained in step (2) on the base plate;

[0019] The detection reagents include mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine, and ascorbic acid.

[0020] Preferably, the color development area is a circle with a diameter of 3-10 mm, and the sample injection area is a strip with a length of 5-15 mm and a width of 2-4 mm.

[0021] Preferably, the weight ratio of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid is 20-35:0.8-2.4:1.8-4.5:1.

[0022] Preferably, the concentration of the test reagent in the test reagent solution is 0.5-1.5 wt%.

[0023] Preferably, the freeze-drying conditions include: a pressure of 5-20 Pa and a time of 10-25 h.

[0024] A third aspect of the present invention provides a paper-based nitrite ion detection chip prepared according to the method described above.

[0025] The fourth aspect of this invention provides the application of the paper-based nitrite ion detection chip described above in the detection of nitrite ion concentration.

[0026] The fifth aspect of this invention provides a method for detecting the concentration of nitrite ions in a solution, the method comprising the following steps:

[0027] The initial RGB value of the color development area of ​​the paper-based nitrite ion detection chip is tested. Then, the test solution is added to the paper-based nitrite ion detection chip. The measured RGB value of the color development area after the reaction is then tested. The chromaticity distance D is determined based on the initial RGB value and the measured RGB value. Then, the concentration of nitrite ions in the test solution is determined by the chromaticity distance D.

[0028] The paper-based nitrite ion detection chip is the same as the paper-based nitrite ion detection chip described above.

[0029] Preferably, the formula for calculating the chromaticity distance is:

[0030]

[0031] Where D is the chromaticity distance, R, G and B are the measured R, G and B values ​​respectively, and R0, G0 and B0 are the initial R, G and B values ​​respectively.

[0032] Compared with the prior art, the present invention has the following beneficial effects:

[0033] 1. The paper-based nitrite ion detection chip of this invention uses filter paper as a substrate. The filter paper can filter out large particulate matter that affects color development, ensuring that the test solution is not interfered with by particulate matter when it comes into contact with the colorimetric reagent. Furthermore, by combining color development with photographic extraction of colorimetric values, accurate test results can be obtained quickly.

[0034] 2. In this invention, by combining filter paper with freeze-drying technology, the detection reagent is uniformly covered on the surface of the filter paper to form a loose structure, which effectively improves the uniformity of the color development layer and reduces the reaction time.

[0035] 3. The paper-based nitrite ion detection chip of the present invention is simple to prepare, economical, and easy to promote and apply on a large scale. Attached Figure Description

[0036] Figure 1 This is a schematic diagram of the structure of the paper-based nitrite ion detection chip described in this invention;

[0037] Figure 2 These are images of the paper-based nitrite ion detection chip prepared in Example 1;

[0038] Figure 3This is the standard curve obtained by fitting in Test Example 3.

[0039] Explanation of reference numerals in the attached figures

[0040] 1. Base plate 2. Color display area

[0041] 3 Sample Injection Zone Detailed Implementation

[0042] The specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings. It should be understood that the specific embodiments described herein are for illustration and explanation only and are not intended to limit the present invention.

[0043] The endpoints and any values ​​of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0044] The first aspect of this invention provides a paper-based nitrite ion detection chip, which is described in conjunction with the above. Figure 1 The paper-based nitrite ion detection chip includes a base plate 1 and a paper base layer located on the base plate;

[0045] The paper substrate includes a color development area 2 and a sample injection area 3 that are interconnected;

[0046] The color development area 2 is pre-filled with a detection reagent, which includes mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid;

[0047] In this process, the test reagent solution is added dropwise to the color development area 2 and then freeze-dried, so that the test reagent is pre-placed in the color development area 2.

[0048] In this invention, the paper-based nitrite ion detection chip uses filter paper as the base layer. The base layer is divided into interconnected colorimetric areas 2 and sample injection areas 3. Detection reagents are pre-placed in the colorimetric areas via freeze-drying, allowing the reagents to uniformly cover the filter paper surface, forming a loose structure that effectively improves the uniformity of the colorimetric layer and reduces reaction time. In use, the sample solution is added through the sample injection area 3, then permeates into the colorimetric area 2 to react with the detection reagents, resulting in color development. The concentration of nitrite ions in the sample is output by comparing the chromaticity distance value of the colorimetric area with a standard curve. Based on this paper-based nitrite ion detection chip, the concentration of nitrite ions in the solution can be obtained quickly and accurately. Preferably, the sample solution is added from the end of the sample injection area 3 furthest from the colorimetric area 2. During permeation, the filter paper effectively filters out large particles that may affect color development, ensuring that the sample solution is not interfered with by particulate matter when it comes into contact with the colorimetric reagents.

[0049] In this invention, the base plate is used to support the paper substrate. To reduce the influence of the base plate's color on color development, the base plate is preferably made of a transparent material. Preferably, the base plate is made of polypropylene (PP), polyethylene terephthalate (PET), or polydimethylsiloxane (PDMS). In a preferred embodiment, the base plate is rectangular, with a length of 30-50 mm and a width of 15-30 mm.

[0050] In this invention, the paper base layer is directly constructed from filter paper. Preferably, the paper base layer is made of Waterman 4# filter paper with a pore size of 25 μm.

[0051] In the preferred case, refer to Figure 1 The color development area 2 is a circle with a diameter (D) of 3-10 mm, and the sample injection area 3 is a long strip with a length (L) of 5-15 mm and a width (H) of 2-4 mm.

[0052] In this invention, to further improve the colorimetric effect and accuracy of the detection reagent, the amount of each component in the detection reagent can be reasonably controlled. Preferably, the weight ratio of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine, and ascorbic acid is 20-35:0.8-2.4:1.8-4.5:1.

[0053] A second aspect of the present invention provides a method for preparing a paper-based nitrite ion detection chip, the method comprising the following steps:

[0054] (1) The filter paper is cut using a laser engraving machine to obtain a paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected.

[0055] (2) The paper substrate is placed in liquid nitrogen for pre-freezing, and then the detection reagent solution is dropped onto the color development area, and then freeze-dried so that the detection reagent is pre-placed in the color development area;

[0056] (3) Provide a base plate and place the paper base obtained in step (2) on top of the base plate;

[0057] The detection reagents include mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine, and ascorbic acid.

[0058] In the conventional preparation of paper-based microarrays, filter paper is typically soaked in a solution or the solution is dropped onto the filter paper and then dried. These methods often result in uneven distribution of the reagents on the filter paper, leading to uneven color development and slow reaction times. In the method described in this invention, freeze-drying technology allows the detection reagents to be evenly distributed on the filter paper in a loose state, effectively improving color uniformity and reducing reaction time.

[0059] In a preferred embodiment, the filter paper used in step (1) is Waterman 4# filter paper with a pore size of 25 μm.

[0060] In a preferred embodiment, the colorimetric area is a circle with a diameter of 3-10 mm, and the sample injection area is a strip with a length of 5-15 mm and a width of 2-4 mm. A laser engraving machine can be used to cut the paper substrate to obtain the above-described shape. Setting the colorimetric area as a circle, as described above, facilitates the adhesion of the detection reagent solution to the colorimetric area during preparation and allows for lyophilization, resulting in a uniform distribution of the detection reagent.

[0061] In a preferred embodiment, to further improve the colorimetric effect and accuracy of the detection reagent, the amount of each component in the detection reagent can be reasonably controlled. Specifically, the weight ratio of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine, and ascorbic acid can be 20-35:0.8-2.4:1.8-4.5:1; more specifically, the weight ratio of mannitol to ascorbic acid can be 20:1, 23:1, 25:1, 2... The weight ratio of p-aminobenzenesulfonic acid to ascorbic acid can be 0.8:1, 1:1, 1.2:1, 1.4:1, 1.5:1, 1.6:1, 1.8:1, 2:1, 2.2:1, or 2.4:1, and the weight ratio of N-1-naphthylethylenediamine to ascorbic acid can be 1.8:1, 2:1, 2.5:1, 3:1, 3.5:1, 4:1, or 4.5:1.

[0062] In the method described in this invention, if the concentration of the detection reagent in the detection reagent solution is too low, the amount of reagent will be insufficient, and the colorimetric reaction will not proceed completely; if the concentration of the detection reagent is too high, the amount of reagent in the colorimetric area after lyophilization will be excessive, and some reagent will fall off the filter paper, resulting in uneven distribution of the detection reagent and thus affecting the uniformity of color development. Preferably, the concentration of the detection reagent in the solution is 0.5-1.5 wt%. The solvent used in the detection reagent solution can be water.

[0063] In the method described in this invention, if the volume of the solution added to the paper substrate is too large, the solution cannot adhere to the paper substrate, and the droplet will fall off the paper substrate; if the volume is too small, the amount of reagent in the colorimetric area will be insufficient, and the colorimetric reaction cannot proceed completely. Preferably, the ratio of the volume of the detection reagent solution to the area of ​​the colorimetric area is 0.3-1 μL: 1 mm. 2 .

[0064] In a preferred embodiment, the freeze-drying conditions include a pressure of 5-20 Pa and a time of 10-25 h.

[0065] In the method described in this invention, conventional methods in the art, such as adhesives, can be used to fix the paper base to the substrate. Transparent adhesives are preferred to minimize the impact on color development.

[0066] A third aspect of this invention provides a paper-based nitrite ion detection chip prepared according to the method described above. This paper-based nitrite ion detection chip can rapidly and accurately detect the concentration of nitrite ions in a solution, and its preparation process is simple, its size is small, and it is easy to promote large-scale application.

[0067] The fourth aspect of this invention provides the application of the paper-based nitrite ion detection chip described above in the detection of nitrite ion concentration.

[0068] The fifth aspect of this invention provides a method for detecting the concentration of nitrite ions in a solution, the method comprising the following steps:

[0069] The initial RGB value of the color development area of ​​the paper-based nitrite ion detection chip is tested. Then, the test solution is added to the paper-based nitrite ion detection chip. The measured RGB value of the color development area after the reaction is then tested. The chromaticity distance D is determined based on the initial RGB value and the measured RGB value. Then, the concentration of nitrite ions in the test solution is determined by the chromaticity distance D.

[0070] The paper-based nitrite ion detection chip is the same as the paper-based nitrite ion detection chip described above.

[0071] In this invention, the detection reagent pre-placed in the color development area of ​​the paper-based nitrite ion detection chip only needs 2 minutes to complete the reaction with nitrite ions in the solution; it has the advantages of fast detection speed and accurate detection results. In a preferred embodiment, the test solution is added dropwise to the sample introduction area of ​​the paper-based nitrite ion detection chip. After the test solution reaches the color development area, wait 2-4 minutes, and then test the measured RGB value of the color development area after the reaction.

[0072] In this invention, the RGB values ​​include R, G, and B values, which refer to the brightness values ​​of the three basic colors: red (R), green (G), and blue (B), respectively. The initial and measured RGB values ​​of the color display area can be obtained by taking a picture using a camera (e.g., a mobile phone), and then processing the picture using software (Photoshop or ImageJ) to obtain the RGB values ​​of the color display area.

[0073] In the method described in this invention, the formula for calculating the chromaticity distance D is:

[0074]

[0075] Where D is the chromaticity distance, R, G and B are the measured R, G and B values ​​respectively, and R0, G0 and B0 are the initial R, G and B values ​​respectively.

[0076] In a preferred embodiment, the concentration of nitrite ions in the solution is quantitatively detected using the external standard method. At least five nitrite ion standard solutions of known concentrations are prepared. The chromaticity distance D of each standard solution is measured using a paper-based nitrite ion detection chip, and the relationship between concentration and chromaticity distance D is calculated. Then, the chromaticity distance D of the test solution is measured using the same method. The concentration of nitrite ions in the test solution is calculated using the relationship between chromaticity distance D, concentration, and chromaticity distance D. The paper-based nitrite ion detection chips used for the standard solutions and the water sample are from the same batch (i.e., prepared under the same conditions), and the injection volumes of the standard solution and the water sample are the same during detection. The standard solutions can be prepared by mixing nitrite and water.

[0077] According to some specific embodiments of the present invention, the method for detecting the concentration of nitrite ions in a solution includes the following steps:

[0078] S1: Test the initial RGB value of the color development area of ​​the paper-based nitrite ion detection chip, and then add the standard solution to the sample injection area of ​​the paper-based nitrite ion detection chip. After the standard solution reaches the color development area, wait for 2-4 minutes and test the measured RGB value of the color development area after the reaction. Determine the chromaticity distance D of the standard solution based on the initial RGB value and the measured RGB value.

[0079] S2: Fit the equations with chromaticity distance D as the x-axis and nitrite ion concentration as the y-axis to obtain the relationship between chromaticity distance D and nitrite ion concentration.

[0080] S3: Test the initial RGB value of the color development area of ​​the paper-based nitrite ion detection chip, and then drop the test solution into the sample injection area of ​​the paper-based nitrite ion detection chip. After the test solution reaches the color development area, wait for 2-4 minutes, and then test the measured RGB value of the color development area after the reaction. Determine the chromaticity distance D of the test solution based on the initial RGB value and the measured RGB value. Substitute the chromaticity distance D of the test solution into the relationship obtained in step S2 for calculation to obtain the concentration of nitrite ions in the test solution.

[0081] The formula for calculating the chromaticity distance D is as follows:

[0082]

[0083] Where D is the chromaticity distance, R, G and B are the measured R, G and B values ​​respectively, and R0, G0 and B0 are the initial R, G and B values ​​respectively.

[0084] In the method described in this invention, in a preferred embodiment, when detecting the concentration of nitrite ions in a solution, the paper-based nitrite ion detection chips used for the standard solution and the test solution are from the same batch (i.e., the preparation conditions are the same), and the volume of the standard solution and the test solution used is the same.

[0085] The method for detecting the concentration of nitrite ions in a solution described in this invention utilizes the aforementioned paper-based nitrite ion detection chip, which can quickly and accurately detect the concentration of nitrite ions in the solution. Since the detection chip described in this invention can be used with a mobile phone, it has advantages such as ease of operation and fast detection speed, providing a convenient and effective new approach for non-professionals to conduct real-time on-site detection of nitrite ion concentration in water.

[0086] The present invention will be described in detail below through embodiments, but the scope of protection of the present invention is not limited thereto.

[0087] Example 1

[0088] Preparation of paper-based nitrite ion detection chip S1:

[0089] (1) Use a laser engraving machine to cut the Waterman 4# filter paper to obtain the paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected; wherein, the sample injection area is a long strip with a length of 8mm and a width of 2.6mm, and the color development area is a circle with a diameter of 6mm;

[0090] (2) The test reagent is mixed with water to prepare a test reagent solution with a concentration of 0.8 wt%. The test reagent is composed of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid in a weight ratio of 25:2:2.5:1.

[0091] The paper substrate obtained in step (1) was placed in liquid nitrogen for pre-freezing, and then 16 μL of the detection reagent solution was dropped onto the colorimetric area. Then, freeze-drying was performed to pre-place the detection reagent in the colorimetric area. The freeze-drying conditions included a pressure of 10 Pa and a time of 18 h.

[0092] (3) Use a laser engraving machine to cut the PP material to obtain a base plate with a length of 40mm and a width of 20mm. Place the paper base layer obtained in step (2) on top of the base plate and fix it with adhesive. The resulting paper-based nitrite ion detection chip is shown in the image below. Figure 2 As shown.

[0093] Example 2

[0094] Preparation of paper-based nitrite ion detection chip S2:

[0095] (1) Use a laser engraving machine to cut the Waterman 4# filter paper to obtain the paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected; wherein, the sample injection area is a long strip with a length of 8mm and a width of 2.6mm, and the color development area is a circle with a diameter of 6mm;

[0096] (2) The test reagent is mixed with water to prepare a test reagent solution with a concentration of 0.8 wt%. The test reagent is composed of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid in a weight ratio of 25:1.6:2.5:1.

[0097] The paper substrate obtained in step (1) was placed in liquid nitrogen for pre-freezing, and then 16 μL of the detection reagent solution was dropped onto the colorimetric area. Then, freeze-drying was performed to pre-place the detection reagent in the colorimetric area. The freeze-drying conditions included a pressure of 10 Pa and a time of 18 h.

[0098] (3) Use a laser engraving machine to cut the PP material to obtain a base plate with a length of 40mm and a width of 20mm. Place the paper base layer obtained in step (2) on top of the base plate and fix it with adhesive.

[0099] Example 3

[0100] Preparation of paper-based nitrite ion detection chip S3:

[0101] (1) Use a laser engraving machine to cut the Waterman 4# filter paper to obtain the paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected; wherein, the sample injection area is a long strip with a length of 8mm and a width of 2.6mm, and the color development area is a circle with a diameter of 6mm;

[0102] (2) The test reagent is mixed with water to prepare a test reagent solution with a concentration of 0.8 wt%. The test reagent is composed of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid in a weight ratio of 25:2.2:2.4:1.

[0103] The paper substrate obtained in step (1) was placed in liquid nitrogen for pre-freezing, and then 16 μL of the detection reagent solution was dropped onto the colorimetric area. Then, freeze-drying was performed to pre-place the detection reagent in the colorimetric area. The freeze-drying conditions included a pressure of 10 Pa and a time of 18 h.

[0104] (3) Use a laser engraving machine to cut the PP material to obtain a base plate with a length of 40mm and a width of 20mm. Place the paper base layer obtained in step (2) on top of the base plate and fix it with adhesive.

[0105] Example 4

[0106] Preparation of paper-based nitrite ion detection chip S4:

[0107] (1) Use a laser engraving machine to cut the Waterman 4# filter paper to obtain the paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected; wherein, the sample injection area is a long strip with a length of 8mm and a width of 2.6mm, and the color development area is a circle with a diameter of 6mm;

[0108] (2) The test reagent is mixed with water to prepare a test reagent solution with a concentration of 0.8 wt%. The test reagent is composed of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid in a weight ratio of 25:2:2.5:1.

[0109] The paper substrate obtained in step (1) was pre-frozen in liquid nitrogen, and then 14 μL of the detection reagent solution was dropped onto the colorimetric area. Then, freeze-drying was performed to pre-place the detection reagent in the colorimetric area. The freeze-drying conditions included a pressure of 10 Pa and a time of 18 h.

[0110] (3) Use a laser engraving machine to cut the PP material to obtain a base plate with a length of 40mm and a width of 20mm. Place the paper base layer obtained in step (2) on top of the base plate and fix it with adhesive.

[0111] Example 5

[0112] Preparation of paper-based nitrite ion detection chip S5:

[0113] (1) Use a laser engraving machine to cut the Waterman 4# filter paper to obtain the paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected; wherein, the sample injection area is a long strip with a length of 8mm and a width of 2.6mm, and the color development area is a circle with a diameter of 6mm;

[0114] (2) The test reagent is mixed with water to prepare a test reagent solution with a concentration of 0.8 wt%. The test reagent is composed of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid in a weight ratio of 25:2:2.5:1.

[0115] The paper substrate obtained in step (1) was pre-frozen in liquid nitrogen, and then 19 μL of the detection reagent solution was dropped onto the colorimetric area. Then, it was freeze-dried so that the detection reagent was pre-placed in the colorimetric area. The freeze-drying conditions included: pressure of 10 Pa and time of 18 h.

[0116] (3) Use a laser engraving machine to cut the PP material to obtain a base plate with a length of 40mm and a width of 20mm. Place the paper base layer obtained in step (2) on top of the base plate and fix it with adhesive.

[0117] Example 6

[0118] Preparation of paper-based nitrite ion detection chip S6:

[0119] (1) Use a laser engraving machine to cut the Waterman 4# filter paper to obtain the paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected; wherein, the sample injection area is a long strip with a length of 8mm and a width of 2.6mm, and the color development area is a circle with a diameter of 6mm;

[0120] (2) The test reagent is mixed with water to prepare a test reagent solution with a concentration of 0.9 wt%. The test reagent is composed of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid in a weight ratio of 25:2.2:2.5:1.

[0121] The paper substrate obtained in step (1) was pre-frozen in liquid nitrogen, and then 21 μL of the detection reagent solution was dropped onto the colorimetric area. Then, freeze-drying was performed to pre-place the detection reagent in the colorimetric area. The freeze-drying conditions included a pressure of 18 Pa and a time of 22 h.

[0122] (3) Use a laser engraving machine to cut the PP material to obtain a base plate with a length of 40mm and a width of 20mm. Place the paper base layer obtained in step (2) on top of the base plate and fix it with adhesive.

[0123] Example 7

[0124] Preparation of paper-based nitrite ion detection chip S7:

[0125] (1) Use a laser engraving machine to cut the Waterman 4# filter paper to obtain the paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected; wherein, the sample injection area is a strip with a length of 9mm and a width of 3mm, and the color development area is a circle with a diameter of 6.5mm;

[0126] (2) The test reagent is mixed with water to prepare a test reagent solution with a concentration of 0.8 wt%. The test reagent is composed of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid in a weight ratio of 25:2:2.5:1.

[0127] The paper substrate obtained in step (1) was pre-frozen in liquid nitrogen, and then 20 μL of the detection reagent solution was dropped onto the colorimetric area. Then, it was freeze-dried so that the detection reagent was pre-placed in the colorimetric area. The freeze-drying conditions included: pressure of 6 Pa and time of 15 h.

[0128] (3) Use a laser engraving machine to cut the PP material to obtain a base plate with a length of 40mm and a width of 20mm. Place the paper base layer obtained in step (2) on top of the base plate and fix it with adhesive.

[0129] Comparative Example 1

[0130] The method described in Example 1 is implemented, except that in step (2), 16 μL of the detection reagent solution is dropped onto the color development area and then placed on a hot plate for heating so that the detection reagent is pre-placed in the color development area; wherein, the heating temperature is 60°C and the heating time is 5 min.

[0131] Test Example 1

[0132] The color development effect of the detection chips prepared in the examples and comparative examples was tested, and the test results are shown in Table 1.

[0133] The detection method is as follows: take a picture of the color display area of ​​the detection chip with a smartphone, and then process the picture with Photoshop software to obtain the initial RGB value of the color display area;

[0134] Take 15 μL of nitrite solution (nitrite ion concentration of 0.6 mg / L) with a pipette and inject it into the detection chip from the end of the injection area away from the color development area. After the solution reaches the color development layer, wait for 2 minutes. Take a picture of the color development area of ​​the detection chip with a smartphone and then process the picture with Photoshop software to obtain the measured RGB value of the color development area.

[0135] The chromaticity distance D is calculated based on the initial RGB values ​​and the measured RGB values. The calculation formula is as follows:

[0136]

[0137] Where D is the chromaticity distance, R, G and B are the measured R, G and B values ​​respectively, and R0, G0 and B0 are the initial R, G and B values ​​respectively.

[0138] Table 1

[0139]

[0140]

[0141] As shown in Table 1, the detection chip prepared in the examples has the characteristics of fast color development rate and good color development effect, which can shorten the detection time of nitrite ions. However, the color distance of the detection chip prepared in Comparative Example 1 is significantly smaller than that of the examples. This is because the detection reagent in Comparative Example 1 is directly dropped onto the filter paper during the preparation process, which makes the detection reagent particles compact and without a loose structure, resulting in a slower reaction rate.

[0142] Test Example 2

[0143] The color uniformity of the detection chips prepared in Example 1 and Comparative Example 1 was tested.

[0144] Detection method: Take a picture of the color display area of ​​the detection chip with a smartphone to obtain the picture. Randomly select 8 areas of the same size from the color display area, and then process the picture with Photoshop software to obtain the initial RGB values ​​of the above 8 areas.

[0145] Take 15 μL of nitrite solution (nitrite ion concentration of 0.6 mg / L) with a pipette and inject it into the detection chip from the end of the sample injection area away from the color development area. After the solution reaches the color development layer, wait for a certain period of time, take a picture of the color development area of ​​the detection chip with a smartphone, and then process the picture with Photoshop software to obtain the measured RGB values ​​of the above 8 areas.

[0146] The chromaticity distance D is calculated based on the initial RGB values ​​and the measured RGB values. The calculation formula is the same as that in Test Example 1.

[0147] When using the detection chip prepared in Example 1, after the solution reached the chromogenic layer, the process was allowed to proceed for 2 minutes before imaging. The chromatic distances of the eight detected regions were 102, 96, 106, 101, 97, 105, 105, and 96, with a relative standard deviation of 4.17%.

[0148] In the case where the detection chip prepared in Comparative Example 1 was used, after the solution reached the chromogenic layer, the process was allowed to proceed for 5 minutes before imaging. The chromatic distances of the eight regions obtained were 101, 112, 90, 96, 106, 102, 94, and 105, with a relative standard deviation of 7.09%.

[0149] The above test results show that the paper-based nitrite ion detection chip described in this invention can quickly and uniformly react with nitrite ions to produce a colorimetric reaction. In contrast, the detection chip prepared in Comparative Example 1, even with sufficient reaction between the detection reagent and the solution, exhibits significantly lower color uniformity compared to Example 1.

[0150] Test Example 3

[0151] Six paper-based nitrite detection chips prepared in Example 1 were prepared with nitrite ion standard solutions at concentrations of 0.1 mg / L, 0.2 mg / L, 0.4 mg / L, 0.6 mg / L, 0.8 mg / L, and 1 mg / L. The chips were then tested according to the method described in Test Example 1, yielding chromatic distances of 42, 55, 71, 102, 123, and 142, respectively. A standard curve was constructed with nitrite ion concentration as the ordinate and chromatic distance as the abscissa. The results are shown below. Figure 3 As shown, the nitrite ion concentration and the chromaticity distance exhibit a good functional relationship, y = 0.00879x - 0.267, where x is the chromaticity distance, y is the nitrite ion concentration, and R... 2 =0.994. Based on this mathematical relationship, after measuring the color distance, the concentration of nitrite ions in the solution can be calculated, thus achieving quantitative detection of nitrite ions.

[0152] Solutions with nitrite ion concentrations of 0.3 mg / L, 0.5 mg / L, and 0.7 mg / L were prepared as test solutions. Three paper-based nitrite ion detection chips prepared according to the method described in Example 1 were prepared. The colorimetric distance was measured according to the method described in Test Example 1, and the concentration was calculated by substituting the values ​​into the above formula. The measured results were compared with the theoretical values ​​of the test solutions, and the results are shown in Table 1.

[0153] Table 1

[0154] Theoretical value Chromatic distance Detection value Error 0.3 mg / L 65.6 0.31 mg / L 3.3％ 0.5 mg / L 86.1 0.49 mg / L 2.0％ 0.7 mg / L 112.3 0.72 mg / L 2.9％

[0155] The results above show that the nitrite ion concentration and the chromaticity distance exhibit a good linear relationship, indicating that the paper-based nitrite ion detection chip described in this invention can accurately detect the nitrite ion concentration in the solution.

[0156] The preferred embodiments of the present invention have been described in detail above; however, the present invention is not limited thereto. Within the scope of the inventive concept, various simple modifications can be made to the technical solutions of the present invention, including combinations of various technical features in any other suitable manner. These simple modifications and combinations should also be considered as the content disclosed in the present invention and are all within the protection scope of the present invention.

Claims

1. A paper-based nitrite ion detection chip, characterized in that, The paper-based nitrite ion detection chip includes a base plate (1) and a paper base layer located on the base plate; The paper substrate includes an interconnected color development area (2) and a sample injection area (3); The color development area (2) is pre-filled with a detection reagent, which includes mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine and ascorbic acid; In this process, the test reagent solution is added dropwise to the color development area (2) and then freeze-dried, so that the test reagent is pre-placed in the color development area (2).

2. The paper-based nitrite ion detection chip according to claim 1, characterized in that, The base plate is made of PP, PET or PDMS; Preferably, the base plate is a rectangle with a length of 30-50mm and a width of 15-30mm.

3. The paper-based nitrite ion detection chip according to claim 1 or 2, characterized in that, The color development area (2) is a circle with a diameter of 3-10 mm, and the sample injection area (3) is a strip with a length of 5-15 mm and a width of 2-4 mm.

4. The paper-based nitrite ion detection chip according to any one of claims 1-3, characterized in that, The weight ratio of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine, and ascorbic acid is 20-35. 0.8-2.4:1.8-4.5:1。 5. A method for preparing a paper-based nitrite ion detection chip, characterized in that, The method includes the following steps: (1) The filter paper is cut using a laser engraving machine to obtain a paper base layer; the paper base layer includes a color development area and a sample injection area that are interconnected. (2) The paper substrate is placed in liquid nitrogen for pre-freezing, and then the detection reagent solution is dropped onto the color development area, and then freeze-dried so that the detection reagent is pre-placed in the color development area; (3) Provide a base plate and place the paper base obtained in step (2) on the base plate; The detection reagents include mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine, and ascorbic acid.

6. The method according to claim 5, characterized in that, The color development area is a circle with a diameter of 3-10 mm, and the sample injection area is a strip with a length of 5-15 mm and a width of 2-4 mm.

7. The method according to claim 5, characterized in that, The weight ratio of mannitol, p-aminobenzenesulfonic acid, N-1-naphthylethylenediamine, and ascorbic acid is 20-35. 0.8-2.4:1.8-4.5:1。 8. The method according to any one of claims 5-7, characterized in that, The concentration of the test reagent in the test reagent solution is 0.5-1.5 wt%.

9. The method according to claim 5, characterized in that, The conditions for freeze drying include: a pressure of 5-20 Pa and a time of 10-25 h.

10. The paper-based nitrite ion detection chip prepared by the method according to any one of claims 5-9.

11. The application of the paper-based nitrite ion detection chip according to any one of claims 1-4 and 10 in the detection of nitrite ion concentration.

12. A method for detecting the concentration of nitrite ions in a solution, characterized in that, The method includes the following steps: The initial RGB value of the color development area of ​​the paper-based nitrite ion detection chip is tested. Then, the test solution is added to the paper-based nitrite ion detection chip. The measured RGB value of the color development area after the reaction is then tested. The chromaticity distance D is determined based on the initial RGB value and the measured RGB value. Then, the concentration of nitrite ions in the test solution is determined by the chromaticity distance D. The paper-based nitrite ion detection chip is the paper-based nitrite ion detection chip according to any one of claims 1-4 and 10.

13. The method according to claim 12, characterized in that, The formula for calculating chromaticity distance is: Where D is the chromaticity distance, R, G and B are the measured R, G and B values ​​respectively, and R0, G0 and B0 are the initial R, G and B values ​​respectively.

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