Double-sided ink-jet paper-based sweat four-index detection system and method

Abstract

The invention relates to the technical field of sweat detection, in particular to a double-sided ink-jet paper-based sweat four-index detection system and method.The detection system comprises a double-sided ink-jet printing paper-based micro-fluidic chip, an acquisition unit, a transmission unit and a detection unit, the acquisition unit, the transmission unit and the detection unit are sequentially stacked to form a three-dimensional stacked structure; the four-index chromogenic reaction system is preloaded on the detection unit and comprises chromogenic substrates of a glucose detection area, a lactic acid detection area, a chloride ion detection area and a pH detection area; the standardized imaging and quantitative analysis device comprises a closed color acquisition terminal and an image processing module, wherein the closed color acquisition terminal consists of a short-focus camera and an annular light source. The channel precision is improved through a double-sided ink-jet configuration and three-dimensional folding, skin-attaching efficient collection and diversion can be achieved, four indexes of glucose, lactic acid, chloride ions and pH in sweat can be synchronously detected, and RGB quantitative analysis can be achieved.

Description

Technical Field

[0001] This invention relates to the field of sweat detection technology, specifically a double-sided inkjet paper-based sweat four-index detection system and method. Background Technology

[0002] Sweat, as a continuously secreted and non-invasive biological sample, contains key physiological indicators such as chloride ions, lactic acid, glucose, and pH value. Changes in its concentration can reflect exercise intensity, metabolic state, and disease progression. Existing analytical methods (such as chromatography and electrochemical methods) rely on large instruments and specialized operations, which are costly and cumbersome, making it difficult to achieve simultaneous detection of multiple indicators.

[0003] Paper-based microfluidics, using cellulose paper as a carrier, drives the transport of trace amounts of sweat through capillary action, combined with colorimetric reactions, providing a new approach for low-cost, portable sweat analysis. However, this technology still faces significant bottlenecks in practical applications: (1) Unstable sweat collection and micro-transport: Common wearable sweat patches often use simple absorbent materials or two-dimensional channel structures, which are prone to uneven sweat dispersion, low aggregation efficiency, and large differences in sample volume in different detection areas, resulting in poor quantification.

[0004] (2) Difficulty in simultaneous detection of multiple indicators: Integrating multiple colorimetric systems or enzyme-catalyzed systems on the same paper-based platform can easily lead to channel crosstalk, cross-contamination, and mutual influence of reagents. At the same time, different reactions require different optimal reaction conditions, resulting in insufficient consistency and stability of multiple indicators.

[0005] (3) Insufficient molding precision and durability of paper-based hydrophobic channels: Traditional wax printing, screen printing and other methods have problems such as limited resolution, channel size drift caused by heating diffusion, and poor batch consistency; when constructing hydrophobic boundaries on one side, liquid may penetrate into the paper thickness direction, causing lateral leakage and crossflow.

[0006] (4) Color reading is greatly affected by ambient light: Color comparison by the naked eye or direct mobile phone photography is usually significantly affected by lighting conditions, shooting distance, angle and automatic exposure of the camera, and differences in mobile phone models, resulting in unstable relationship between RGB color intensity and concentration, making it difficult to achieve repeatable quantitative analysis. Summary of the Invention

[0007] The purpose of this invention is to overcome the shortcomings of the prior art and propose an integrated double-sided inkjet paper-based sweat four-index detection system and method that integrates "collection-transmission-detection".

[0008] To achieve the above objectives, the technical solution specifically adopted by the present invention is as follows: A double-sided inkjet paper-based sweat four-index detection system includes: The double-sided inkjet-printed paper-based microfluidic chip includes a collection unit, a transmission unit, and a detection unit, which form hydrophilic microchannels by inkjet printing hydrophobic patterns on both sides of a filter paper substrate. The collection unit, transmission unit, and detection unit are stacked sequentially to form a three-dimensional stacked structure, so that sweat is collected, diverted, and undergoes a color reaction in the sequence of "collection unit → transmission unit → detection unit". The four-index colorimetric reaction system is preloaded onto the detection unit, including colorimetric substrates for glucose detection, lactic acid detection, chloride ion detection, and pH detection. When sweat reaches the detection area, the reaction is automatically triggered and a color change occurs. The standardized imaging and quantitative analysis device includes a short-focal-length camera, a closed color acquisition terminal consisting of a ring light source, and an image processing module. The closed color acquisition terminal, consisting of the short-focal-length camera and the ring light source, is used to acquire images of the detection area under fixed geometric and lighting conditions. The image processing module is used to extract the RGB color intensity of each detection area and match it with a pre-established standard curve to output the concentration of the target indicator.

[0009] Furthermore, the filter paper substrate is made of Whatman No.1 filter paper, with a nominal thickness of approximately 0.18 mm and a typical particle rejection rate of approximately 11 μm. This allows for self-driven liquid transport through capillary action generated by the internal pore structure of the filter paper. Simultaneously, this substrate exhibits excellent patterning compatibility with hydrophobic inks, enabling the formation of hydrophobic barriers and hydrophilic channels on the filter paper surface via inkjet printing. This results in microfluidic structures with clear boundaries, stable patterns, and excellent fabrication repeatability.

[0010] In this invention, the collection unit is disposed on the skin-contact side of the chip and includes a sweat inlet area and a snowflake-shaped hydrophilic collection structure. The structure consists of multiple radially distributed hydrophilic branch bands, and the outer edge is defined by a hydrophobic fence to enhance the ability to capture, divert, and converge sweat.

[0011] The transmission unit includes four symmetrical hydrophilic branch channels. The length, width, curvature, and cross-sectional resistance of each channel are designed to achieve approximately equal volume distribution of sweat to the detection unit.

[0012] The detection unit contains four isolated hydrophilic detection zones, each connected to the transmission unit via an independent branch channel. The detection zones are circular reaction zones.

[0013] Furthermore, the hydrophobic pattern is formed by double-sided inkjet printing. Double-sided printing allows the hydrophobic material to penetrate into the thickness direction of the paper and form a through-hydrophobic barrier between the fibers, thereby significantly reducing lateral leakage, cross-flow, and cross-contamination caused by liquid penetration along the paper thickness direction. The hydrophobic ink is based on a sol-gel system, including tetraethyl orthosilicate (TEOS), 1H,1H,2H,2H-perfluorooctyltriethoxysilane (PFOTS), anhydrous ethanol, water, and hydrochloric acid catalyst. The preparation steps include: adding hydrochloric acid to anhydrous ethanol and deionized water, stirring, adding TEOS for hydrolysis and condensation, and adding PFOTS for co-condensation to obtain the hydrophobic ink. The prepared ink is stored at low temperature (4°C) to improve the stability of the system.

[0014] In this invention, inkjet printing is performed using an Epson R330 inkjet printer, and the steps include: (1) Print the hydrophobic area on the front of the filter paper according to the design pattern; (2) After the surface is initially dry, flip it over and print the corresponding hydrophobic pattern on the back of the filter paper; (3) Based on the hydrophobic effect and the channel forming accuracy, repeat the printing 2-3 times to improve the hydrophobicity.

[0015] (4) After printing, place the filter paper in a forced-air drying oven for baking and curing at 60 ℃ for 1 h. Baking is used to promote further condensation and fixation of the hydrophobic layer of the sol, forming a stable hydrophobic channel. If the baking time is too short, the hydrophobic layer may not be fully cured, while if it is too long, the ink may spread on the paper surface, affecting the channel resolution.

[0016] In this invention, the loading method of the four-index colorimetric reaction system includes: Glucose detection zone: immobilized chitosan, glucose oxidase (GOD), horseradish peroxidase (HRP), trehalose, potassium iodide (KI), the reaction is based on the enzyme-iodine colorimetric principle; Lactic acid detection area: Immobilized with ABTS, lactate oxidase (L-LOx), and HRP; the reaction is based on the LOx-HRP-ABTS colorimetric principle. Chloride ion detection zone: immobilized with TPTZ, HgSO4, and FeSO4; reaction based on Hg 2+ -Fe 2+ -TPTZ competitive color development principle; pH detection area: Immobilized methyl red and bromothymol blue mixed indicator in a 3:2 ratio; The sample volume for each detection area was 4.5 μL, and the drying conditions were room temperature drying in the dark for 30 min.

[0017] This invention also provides a method for detecting four indicators of sweat on double-sided inkjet paper, comprising the following steps: (1) Stack and assemble the paper chips and package them into a patch form so that the collection unit comes into contact with the sweat on the skin; (2) Sweat enters the snowflake-shaped collection area of ​​the collection unit, where it is collected under capillary action and distributed along the branch channels of the transmission unit; (3) Sweat enters the four detection zones and reacts with the pre-loaded substrate to produce color changes; (4) After the reaction is 30 min, the image of the detection area is acquired and the color is analyzed. Specifically, ImageJ software is used to select a circular area of ​​the detection area with a fixed area, extract the average RGB value, and match the pre-established standard curve to output the concentration. The color acquisition terminal is connected to a smartphone or computer via wireless network.

[0018] Furthermore, it also includes optional extensions: the reagents in the detection area can be replaced with colorimetric / fluorescent / electrochemical reagent systems of uric acid, creatinine, Na⁺ / K⁺ or cortisol; the readout method can use a mobile phone camera with a standard light source box or a portable reflectance photometer; the encapsulation material uses medical tape, elastic polymer film or breathable and waterproof membrane.

[0019] This invention has the following characteristics and beneficial effects: (1) Low cost and easy manufacturing: Using filter paper as the substrate and inkjet printing as the patterning method, it does not require expensive equipment such as photolithography and plasma processing, making it suitable for large-scale manufacturing and single-use.

[0020] (2) Double-sided hydrophobic molding reduces interference: Hydrophobic ink is printed and cured on both sides, forming a hydrophobic barrier, which significantly reduces leakage and cross-contamination, and improves the reliability of simultaneous detection in multiple detection areas.

[0021] (3) Integrated collection-transmission-detection: The snowflake-shaped collection structure improves the sweat capture efficiency and the symmetrical branch channels achieve stable diversion, integrating sample collection and analysis into the same paper chip, which can work automatically when applied to the skin.

[0022] (4) Simultaneous detection of four indicators: The colorimetric system of four typical sweat indicators, namely glucose, lactic acid, chloride ion and pH, is integrated on the same chip, which can simultaneously reflect the metabolic and electrolyte and acid-base balance, and is suitable for scenarios such as exercise monitoring and chronic disease management.

[0023] (5) Stable quantitative reading: By introducing a short-focal-length camera and a closed ring light source for standardized imaging conditions, combined with RGB extraction and standard curves, relatively stable quantitative analysis is achieved, breaking through the limitations of traditional semi-quantitative analysis by the naked eye and strong environmental dependence.

[0024] (6) High scalability: The reagents in the detection area can be replaced to expand to other sweat or body fluid (such as saliva, tears) indicators; the readout end can also be expanded to various forms such as mobile phones and portable photometers, which facilitates product iteration. Attached Figure Description

[0025] Other features, objects, and advantages of the present invention will become more apparent from the following detailed description of non-limiting embodiments with reference to the accompanying drawings: Figure 1 This is a schematic diagram of a paper chip.

[0026] Figure 2 The specific dimensions of the chip; In the figure: (a) is the front of the collection, transmission and detection unit; (b) is the back of the collection, transmission and detection unit.

[0027] Figure 3 A schematic diagram of a black box for 3D image acquisition.

[0028] Figure 4 This is a schematic diagram showing the paper chip stacking method and the distribution of the detection area. In the diagram: 1-Chloride ion detection area; 2-pH detection area; 3-Glucose detection area; 4-Lactic acid detection area. Detailed Implementation

[0029] The present invention will now be described in detail with reference to specific embodiments. These embodiments will help those skilled in the art to further understand the present invention, but do not limit the invention in any way. It should be noted that, unless otherwise specified, the embodiments and features described in the present invention can be combined with each other.

[0030] In a first aspect, this invention provides a double-sided inkjet paper-based sweat four-index detection system to achieve efficient collection and stable diversion of sweat under skin-contact conditions; simultaneous colorimetric detection of four indicators—glucose, lactic acid, chloride ions, and pH—on the same chip; improved channel forming accuracy and reduced interference through double-sided inkjet hydrophobic patterning and a three-dimensional folded structure; and obtained repeatable quantitative results by combining standardized imaging and RGB analysis under a short-focal-length camera and enclosed light source conditions. In one embodiment, the sweat four-index detection system includes: The double-sided inkjet-printed paper-based microfluidic chip includes a collection unit, a transmission unit, and a detection unit, which form hydrophilic microchannels by inkjet printing hydrophobic patterns on both sides of a filter paper substrate. The collection unit, transmission unit, and detection unit are stacked sequentially to form a three-dimensional stacked structure, so that sweat is collected, diverted, and undergoes a color reaction in the sequence of "collection unit → transmission unit → detection unit". The four-index colorimetric reaction system is preloaded onto the detection unit, including the colorimetric substrates in glucose detection zone 3, lactic acid detection zone 4, chloride ion detection zone 1, and pH detection zone 2. When sweat reaches the detection zone, the reaction is automatically triggered and a color change occurs. The standardized imaging and quantitative analysis device includes a short-focal-length camera, a closed color acquisition terminal consisting of a ring light source, and an image processing module. The closed color acquisition terminal, consisting of the short-focal-length camera and the ring light source, is used to acquire images of the detection area under fixed geometric and lighting conditions. The image processing module is used to extract the RGB color intensity of each detection area and match it with a pre-established standard curve to output the concentration of the target indicator.

[0031] In this embodiment, the filter paper substrate is Whatman No.1 filter paper with a nominal thickness of approximately 0.18 mm and a typical particle rejection level of approximately 11 μm. It can achieve self-driven liquid transport through capillary action generated by the internal pore structure of the filter paper. Simultaneously, this substrate has good patterning compatibility with hydrophobic inks, allowing the formation of hydrophobic barriers and hydrophilic channels on the filter paper surface via inkjet printing. This results in a microfluidic structure with clear boundaries, stable patterns, and good reproducibility.

[0032] In this embodiment, the filter paper substrate is formed into a three-dimensional structure in the following way: Acquisition unit, transmission unit, and detection unit areas are planned on the same filter paper substrate. After double-sided inkjet printing and drying / curing, the substrate is cut along the hydrophobic edge line. The cut surfaces are then stacked sequentially to form a "acquisition-transmission-detection" three-dimensional structure, wherein: The collection unit is located on the skin-contact side of the chip and includes a sweat inlet area and a snowflake-shaped hydrophilic collection structure. This structure consists of multiple radially distributed hydrophilic branch bands, and the outer edge is defined by a hydrophobic fence to enhance the ability to capture, divert, and converge sweat.

[0033] The transmission unit includes four symmetrical hydrophilic branch channels. The length, width, curvature, and cross-sectional resistance of each channel are designed to achieve approximately equal volume distribution of sweat to the detection unit.

[0034] The detection unit contains four isolated hydrophilic detection zones, each connected to the transmission unit via an independent branch channel. The detection zones are circular reaction zones.

[0035] In this embodiment, the hydrophobic pattern is formed by double-sided inkjet printing. Double-sided printing allows the hydrophobic material to penetrate into the thickness direction of the paper and form a through-hole hydrophobic barrier between the fibers, thereby significantly reducing lateral leakage, cross-flow, and cross-contamination caused by liquid penetration along the paper thickness direction. The hydrophobic ink is based on a sol-gel system, and its key components include: Silicon source precursor: Tetraethyl orthosilicate (TEOS); Fluorosilane modifier: 1H,1H,2H,2H-perfluorooctyltriethoxysilane (abbreviated as PFOTS, or similar perfluoroalkyl silanes); Solvent: Anhydrous ethanol; Water: Used for hydrolysis and condensation; Acid catalyst: hydrochloric acid (HCl), used to promote the hydrolysis and condensation reaction of TEOS.

[0036] The preparation steps include: (1) Add anhydrous ethanol and deionized water to a round-bottom flask and stir thoroughly; (2) Add hydrochloric acid as a catalyst and continue stirring to form an acidic hydrolysis system; (3) Add TEOS under stirring conditions to hydrolyze and condense it to form a sol; (4) PFOTS is then added dropwise to co-condense with the sol network to obtain a hydrophobic ink that can be used for inkjet printing; (5) Store the prepared ink at a low temperature (4°C) to improve the stability of the system.

[0037] In this embodiment, computer-aided design software (CAD) is used to draw the hydrophobic pattern of the paper chip. The pattern includes at least: the hydrophobic / hydrophilic boundary of the snowflake-shaped collection area of ​​the acquisition unit; the branch channel network of the transmission unit; and the detection area and isolation fence of the detection unit.

[0038] In this embodiment, inkjet printing uses an Epson R330 inkjet printer, and the steps include: (1) Print the hydrophobic area on the front of the filter paper according to the design pattern; (2) After the surface is initially dry, flip it over and print the corresponding hydrophobic pattern on the back of the filter paper; (3) Based on the hydrophobic effect and the channel forming accuracy, repeat the printing 2-3 times to improve the hydrophobicity.

[0039] (4) After printing, place the filter paper in a forced-air drying oven for baking and curing at 60 ℃ for 1 h. Baking is used to promote further condensation and fixation of the hydrophobic layer of the sol, forming a stable hydrophobic channel. If the baking time is too short, the hydrophobic layer may not be fully cured, while if it is too long, the ink may spread on the paper surface, affecting the channel resolution.

[0040] In this embodiment, the loading method of the four-index colorimetric reaction system includes: Glucose detection area (enzyme-iodine colorimetric reaction) Principle: Glucose is converted into hydrogen peroxide under the catalysis of glucose oxidase (GOD); under the catalysis of horseradish peroxidase (HRP), iodide ions (I⁻) are oxidized to iodine (I₂) / triiodide ions, which are brown in color, and the color intensity is related to the glucose concentration.

[0041] The loading solution consists of chitosan, GOD, HRP, trehalose, and potassium iodide (KI).

[0042] Immobilization steps: After preparing the substrate solution, add chitosan solution, glucose oxidase solution, potassium iodide solution and horseradish peroxidase solution dropwise to the glucose detection area, and let it air dry for later use.

[0043] Lactic acid detection area (LOx-HRP-ABTS colorimetric assay) Principle: Lactic acid is converted into pyruvate and hydrogen peroxide under the catalysis of lactate oxidase (L-LOx); HRP catalyzes the oxidation of ABTS to produce a green product, and the color intensity is related to the lactic acid concentration.

[0044] Solid carrier fluid composition: ABTS, L-LOx and HRP.

[0045] Immobilization steps: Lactate oxidase, horseradish peroxidase and ABTS solution are added dropwise to the lactate detection area in sequence, and then allowed to air dry for later use.

[0046] Chloride ion detection zone (Hg²⁺-Fe²⁺-TPTZ competitive colorimetric development) Principle: In Hg 2+ Fe 2+ In a mixed solution of TPTZ, TPTZ preferentially reacts with Hg. 2+ Complexation, producing colorless Hg [TPTZ] 2. When chloride ions are present, Hg 2+ With Cl - HgCl2 precipitate is formed, and the released TPTZ reacts with Fe. 2+ Complexation, forming blue Fe [TPTZ] 2, thereby achieving a colorimetric response to Cl⁻.

[0047] Composition of the carrier liquid: TPTZ is soluble in methanol, HgSO4 solution and FeSO4 solution.

[0048] Immobilization steps: Prepare a detection reagent by mixing TPTZ, HgSO4 and FeSO4 solutions in a ratio of 1:2:2, then add it dropwise to the chloride ion detection area and allow it to air dry for later use.

[0049] pH detection area (indicator colorimetric method) Principle: A pH indicator is used to show different colors at different pH levels, thus enabling colorimetric determination of sweat pH.

[0050] Composition of the carrier liquid: methyl red solution and bromothymol blue solution.

[0051] Immobilization steps: Prepare a mixed indicator by mixing methyl red and bromothymol blue in a 3:2 ratio, then add it dropwise to the pH detection area and let it air dry for later use.

[0052] Among them, the general process parameters for reagent immobilization a) Spotting volume: 4.5 μL per detection zone; b) Drying conditions: Dry at room temperature in the dark for 30 minutes; c) Stabilizer: Trehalose is added to the enzyme system to improve its stability during drying and storage; d) Storage: After drying, place in a sealed bag and store at 4 ℃ away from light.

[0053] In a second aspect, the present invention provides a method for detecting four indicators of sweat on double-sided inkjet paper based on the detection system described in the first aspect, comprising the following steps: (1) Stack and assemble the paper chips and package them into a patch form so that the collection unit comes into contact with the sweat on the skin; (2) Sweat enters the snowflake-shaped collection area of ​​the collection unit, where it is collected under capillary action and distributed along the branch channels of the transmission unit; (3) Sweat enters the four detection zones and reacts with the pre-loaded substrate to produce color changes; (4) After a reaction of 30 minutes, images of the detection area are acquired and color analysis is performed. Specifically, to reduce ambient light interference, this invention uses a short-throw camera and a ring light source to construct a closed color capture black box. The black box contains a short-throw camera and a fixed light source, and is equipped with a wireless network. When a smartphone or computer connects to the black box's unique wireless network, the image captured by the black box can be viewed. After image acquisition is completed, (1) Image region selection: Using ImageJ software, each detection region is selected using the circular selection tool, and the area of ​​the region is fixed to ensure that the area of ​​the selected region is consistent in all subsequent images; (2) RGB extraction: Record the average R, G, and B values ​​of the selected area; (3) Establishment of standard curve: Use standard solutions of known concentration to perform detection and obtain the linear fitting relationship between color intensity and concentration, which will serve as the basis for subsequent quantification; (4) Unknown sample determination: Substitute the color intensity obtained from the detection of unknown sweat sample into the standard curve to obtain the concentration of glucose, lactic acid, chloride ion and pH value.

[0054] It is worth noting that the reagents in the detection area can be replaced with colorimetric / fluorescent / electrochemical reagent systems of uric acid, creatinine, Na⁺ / K⁺ or cortisol; the readout method can use a mobile phone camera with a standard light source box or a portable reflectance photometer; the packaging material uses medical tape, elastic polymer film or breathable and waterproof membrane.

[0055] Example 1: Preparation of Fluorosilane Modified Sol-Based Hydrophobic Ink (1) Solvent and hydrolysis system: Add 24 mL of anhydrous ethanol and 2 mL of deionized water to a round-bottom flask and mix well; add 3 μL of 36% hydrochloric acid and stir for 10 min; (2) Add silicon source and fluorosilane: Add 2 mL TEOS and 1.5 mL PFOTS under continuous stirring, and continue stirring for 1 h to allow it to be fully hydrolyzed; (3) Aging and storage: Let the sol stand for aging, filter it with a 0.22 μm filter membrane and put it into the ink cartridge; when not in use, store it at 4 ℃ away from light.

[0056] Example 2: Preparation of double-sided inkjet printing paper chips (1) Pattern design: Use CAD to draw the three-unit structure pattern (snowflake-shaped collection area of ​​the acquisition unit, four-branch channel of the transmission unit, and four detection areas of the detection unit); (2) Printing: Load the hydrophobic ink into the inkjet printer and select the "high quality photo" mode; print the hydrophobic pattern on the front of the Whatman filter paper, let it dry, flip it over and print the corresponding hydrophobic pattern on the back; repeat the printing 3 times to improve the permeability of the hydrophobic barrier; (3) Curing: Place the printed filter paper in a 60 ℃ drying oven and bake for 60 min to cure the hydrophobic layer. (4) Stacking: Arrange the acquisition unit on the top layer, the transmission unit in the middle, and the detection unit on the bottom layer in sequence to ensure that the channel entrance and the detection area are aligned; double-sided tape can also be used to fix the frame.

[0057] Example 3: Reagent loading in the four-index detection area (1) Glucose detection solution: Prepare 1 mg / mL GOD and 0.2 mg / mL HRP (both with 5% (w / v) trehalose to protect enzyme activity) enzyme solution with sodium citrate buffer solution, prepare 1 mg / mL chitosan solution (0.25% (v / v) acetic acid) and 1 MKI, and add 4.5 μL chitosan solution, 4.5 μL GOD, 4.5 μL KI and 4.5 μL HRP dropwise to glucose detection area and dry at room temperature in the dark; (2) Lactic acid detection solution: Add 4.5 μL of 180 U / mL LOx, 0.2 mg / mL HRP and 1 mg / mL ABTS to the lactic acid detection area in sequence, and dry at room temperature in the dark; (3) Chloride ion detection solution: Dissolve 2,4,6-tris(2-pyridyl)-s-triazine (TPTZ) in methanol to a concentration of 50 x 10⁻⁶. -3 M. Prepare solutions of 20 mM HgSO4 and 20 mM FeSO4, and make a chloride ion detection solution by volume ratio TPTZ:HgSO4:FeSO4=1:2:2. Add 4.5 uL to the chloride ion detection area and dry at room temperature in the dark. (4) pH indicator: Add 4.5 μL of a mixed indicator of methyl red and bromothymol blue = 3:2 to the pH detection area and dry at room temperature in the dark; (5) After drying, the paper chips are placed in a sealed drying bag and stored at 4 ℃ away from light for later use.

[0058] Example 4: Detection and Quantitative Analysis of Sweat Samples (1) Standard curve: Prepare standard solutions of glucose, lactic acid, Cl⁻ and different pH buffers at different concentrations; add a fixed volume to the inlet of the acquisition unit, wait for it to be diverted to each detection area and develop color, and then take a picture; extract the average RGB value of each detection area and fit it to obtain the standard curve; (2) Sweat detection: The paper chip is packaged into a patch and attached to the sweating area of ​​the skin (or a sweat sample is directly dripped into the inlet). After reacting for 30 minutes, a short-focus camera terminal is used to take a picture. (3) Image processing: Select each detection area using ImageJ and record the average RGB value; substitute it into the standard curve to obtain the concentrations of glucose, lactic acid, Cl⁻ and pH results; (4) Result output: The calculation model can be written into the mobile phone program to achieve automatic recognition and result display.

[0059] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely preferred examples and are not intended to limit the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed. The scope of protection of the present invention is defined by the appended claims and their equivalents.

Claims

1. A four-index detection system for sweat on double-sided inkjet paper, characterized in that: include: A double-sided inkjet-printed paper-based microfluidic chip includes a collection unit, a transmission unit, and a detection unit, which are formed by inkjet printing hydrophobic patterns on both sides of a filter paper substrate to create hydrophilic microchannels. The collection unit, transmission unit, and detection unit are stacked sequentially to form a three-dimensional stacked structure. A four-index colorimetric reaction system, preloaded onto the detection unit, includes colorimetric substrates for glucose detection, lactate detection, chloride ion detection, and pH detection zones; The standardized imaging and quantitative analysis device includes a short-focal-length camera, a closed color acquisition terminal consisting of a ring light source, and an image processing module.

2. The double-sided inkjet paper-based sweat four-index detection system as described in claim 1, characterized in that: The filter paper substrate uses Whatman No.1 filter paper with a nominal thickness of 0.18 mm and a particle rejection level of 11 μm, and achieves self-driven liquid transport through capillary action.

3. The double-sided inkjet paper-based sweat four-index detection system as described in claim 1, characterized in that: The collection unit is located on the skin-contact side of the chip and includes a sweat inlet area and a snowflake-shaped hydrophilic collection structure. This structure consists of multiple radially distributed hydrophilic branch bands, and the outer edge is defined by a hydrophobic fence to enhance the ability to capture, divert, and converge sweat.

4. The double-sided inkjet paper-based sweat four-index detection system as described in claim 1, characterized in that: The transmission unit includes four symmetrical hydrophilic branch channels. The length, width, curvature, and cross-sectional resistance of each channel are designed to achieve approximately equal volume distribution of sweat to the detection unit.

5. The double-sided inkjet paper-based sweat four-index detection system as described in claim 1, characterized in that: The detection unit contains four isolated hydrophilic detection zones, each connected to the transmission unit via an independent branch channel. The detection zones are circular reaction zones.

6. The double-sided inkjet paper-based sweat four-index detection system as described in claim 1, characterized in that: The hydrophobic pattern is formed by double-sided inkjet printing. The hydrophobic ink is based on a sol-gel system, including tetraethyl orthosilicate, 1H,1H,2H,2H-perfluorooctyltriethoxysilane, anhydrous ethanol, water, and hydrochloric acid catalyst. The preparation steps include: adding hydrochloric acid to anhydrous ethanol and deionized water, stirring, adding TEOS for hydrolysis and condensation, and adding PFOTS for co-condensation to obtain the hydrophobic ink.

7. The double-sided inkjet paper-based sweat four-index detection system as described in claim 1, characterized in that: The inkjet printing steps include: printing a hydrophobic pattern on the front side of the filter paper, flipping it over to print a hydrophobic pattern on the back side, and repeating the printing 2–3 times; baking and curing at 60°C for 1 hour.

8. The four-index detection system for sweat on double-sided inkjet paper as described in claim 1, characterized in that: The loading methods for the four-index colorimetric reaction system include: Glucose detection zone: immobilized chitosan, glucose oxidase, horseradish peroxidase, trehalose, and potassium iodide; the reaction is based on the enzyme-iodine colorimetric principle. Lactic acid detection area: ABTS, lactate oxidase, and HRP are immobilized, and the reaction is based on the LOx-HRP-ABTS colorimetric principle; Chloride detection zone: immobilized TPTZ, HgSO4, FeSO4, reaction based on Hg 2+ -Fe 2+ -TPTZ competitive color development principle; pH detection area: Immobilized methyl red and bromothymol blue mixed indicator in a 3:2 ratio; The sample volume for each detection area was 4.5 μL, and the drying conditions were room temperature drying in the dark for 30 min.

9. A method for detecting four indicators of sweat on double-sided inkjet paper as described in any one of claims 1-8, characterized in that: Includes the following steps: (1) Stack and assemble the paper chips and package them into a patch form so that the collection unit comes into contact with the sweat on the skin; (2) Sweat enters the snowflake-shaped collection area of ​​the collection unit, where it is collected under capillary action and distributed along the branch channels of the transmission unit; (3) Sweat enters the four detection zones and reacts with the pre-loaded substrate to produce color changes; (4) After the reaction is 30 min, the image of the detection area is acquired and color analysis is performed; specifically, ImageJ software is used to select a circular area of ​​fixed area of ​​the detection area, extract the average RGB value, and match the pre-established standard curve to output the concentration; the color acquisition terminal is connected to a smartphone or computer via wireless network.

10. The double-sided inkjet paper-based sweat four-index detection system as described in claim 1, characterized in that: Optional extensions are also included: the reagent in the detection zone can be replaced with a colorimetric / fluorescent / electrochemical reagent system of uric acid, creatinine, Na⁺ / K⁺ or cortisol; The readout method can use a mobile phone camera with a standard light source box or a portable reflectometer; the encapsulation material is medical tape, elastic polymer film or breathable and waterproof membrane.

Images