A pH sensitive electrode for a wearable pH sensor for wound state diagnosis and application thereof

Abstract

The application provides a pH sensitive electrode of a wearable pH sensor for wound state diagnosis and application thereof. The electrode overcomes the problem of biological incompatibility defects of a conventional Ir-based electrode, realizes biological compatibility of the electrode sensitive material by introducing a biocompatible coating material to the surface of the Ir-based material, and solves the potential risk of application of the Ir-based pH sensor in a wearable application scenario.

Description

Technical Field

[0001] This invention belongs to the field of electrochemical detection and analysis and chemical sensor technology, and relates to a pH-sensitive electrode that constitutes a wearable pH sensor for wound condition diagnosis and its application. Background Technology

[0002] The treatment and management of trauma, especially chronic trauma, is a crucial task in the medical field. Currently, long-term care for chronic trauma primarily relies on three-dimensional dressings, but monitoring the physiological state of the wound during this period remains a technically challenging problem. The prevailing clinical practice involves continuous observation of the wound by experienced healthcare professionals to determine its physiological or pathological state. However, this method only provides a prognostic assessment of the wound's condition and cannot predict impending deterioration, resulting in insufficient accuracy. Furthermore, it requires frequent physical intervention, causing significant pain for the patient. pH is a vital physiological and biochemical indicator of wound health. A normal wound pH ranges from approximately 4.5 to 6.5. A suitable low pH level not only helps resist infection and inflammation but also significantly shortens the wound healing period. A rising natural pH indicates bacterial proliferation, and a high pH level signifies significant bacterial infection, indicating the wound is already in or about to enter a state of infection and inflammation. Therefore, timely and accurate measurement of the pH of the wound surface can accurately determine the physiological state of the wound and predict whether it faces the risk of infection.

[0003] Precise, stable, and rapid pH quantification is the core task in pH detection. While mature pH sensor products exist, their reliance on frequent calibration and difficulty in achieving planar design prevent them from meeting the demands of wound condition monitoring scenarios requiring both high accuracy and stability. Furthermore, most pH sensor sensitive materials suffer from low biocompatibility, particularly posing safety risks when applied directly to wound surfaces. These issues severely hinder the development of wearable pH sensors for wound care. Summary of the Invention

[0004] To overcome the problem that traditional electrochemical pH sensing technology relies on repeated calibration and cannot achieve long-term in-situ fully automatic monitoring, the present invention aims to provide a pH-sensitive electrode that constitutes a wearable pH sensor for wound condition diagnosis.

[0005] Another objective of this invention is to provide an application of the above-mentioned pH-sensitive electrode in an electrochemical calibration-free pH sensing method.

[0006] The sensor provided by this invention features a planar, flexible design, allowing it to be well-adhered to the wound surface; furthermore, this invention utilizes a polymer-Ir-IrO. x Nanoparticle composites overcome Ir-IrOx It overcomes its own bioincompatibility and enhances its electrochemical signal stability, and utilizes Ir-IrO x The electrochemical activity of the sensor indicates the pH of the wound surface. The method provided overcomes the inherent tissue incompatibility of the sensitive material and solves the safety issues when using the sensor in situ on the wound surface.

[0007] A pH-sensitive electrode comprising a wearable pH sensor for wound condition diagnosis is prepared by the following method:

[0008] (1) Use triethanolamine DMSO solution to adjust the pH of IrCl4 solution to 2.0-6.0, add DMSO solution of 3-methyl-1,2-cyclopentanedione, mix well and let stand in the dark;

[0009] (2) Heat the mixed solution obtained in step (1) to 75-100℃, add L-ascorbic acid sodium solution to it under stirring, react for 0.2-4.5h, collect the precipitate i.e. IrNP, and disperse it in water to obtain IrNP dispersion;

[0010] (3) Prepare an aqueous solution of 4,4'-diamino-5,5”-(dithiophen-2-yl)-2,2'-bipyridine, add the solution to the IrNP dispersion under stirring, and react at 45-70℃ for 12-24 h. After the reaction is completed, collect the precipitate, redisperse the precipitate with water, and prepare a dispersion system with a mass fraction of 0.05-0.5%.

[0011] (4) Add Ru-containing substances to the dispersion system obtained in step (3). 3+ The clean working surface of the substrate electrode is immersed in an aqueous solution and kept at a constant temperature of 40–60°C for 45–80 minutes. After rinsing, it is then immersed in a solution containing Ru. 3+ The pH-sensitive electrode, denoted as IR, is obtained by immersing the electrode in an aqueous solution of IR and then in a bipyridine solution for 60–180 min.

[0012] Furthermore, the concentration of the IrCl4 solution in step (1) is 5-15 mM, more preferably 10 mM; the IrCl4 solution is obtained by dissolving IrCl4 solid in HCl solution.

[0013] Furthermore, the concentration of triethanol in step (1) is 0.1-1M; the concentration of the DMSO solution of 3-methyl-1,2-cyclopentanedione is 0.1-0.25M.

[0014] Furthermore, the volume ratio of the IrCl4 solution to the DMSO solution of 3-methyl-1,2-cyclopentanedione in step (1) is 100:20-75.

[0015] Furthermore, the temperature for standing in step (1) is 20-35°C, and the standing time is 12-36 hours.

[0016] Furthermore, the volume ratio of the mixed solution obtained in step (1) to the L-ascorbic acid sodium solution is 12-18:5-25;

[0017] Furthermore, the mass fraction of the IrNP dispersion in step (2) is 0.01% to 2%, preferably 0.1% to 1%;

[0018] Furthermore, the reaction time in step (2) is 0.5 to 2.5 h.

[0019] Furthermore, the aqueous solution of 4,4'-diamino-5,5”-(dithiophen-2-yl)-2,2'-bipyridine in step (3) has a concentration of 10-20 mM, preferably 15 mM; and its volume ratio with the IrNP dispersion is 2:4-12.

[0020] Furthermore, the reaction temperature in step (3) is 50-60°C, more preferably 55°C, and the reaction time is 14-20h, more preferably 16h.

[0021] Furthermore, the mass fraction of the dispersion system described in step (3) is 0.1% to 0.5%.

[0022] Furthermore, step (4) first describes the Ru-containing... 3+ Ru in aqueous solution 3+ The concentration is 1–20 mM, preferably 5–15 mM, and more preferably 10 mM.

[0023] Furthermore, in step (4), the temperature of the first constant temperature treatment is 45-55°C, preferably 50°C, and the time is 50-70 min, preferably 60°C.

[0024] Furthermore, step (4) is the second Ru... 3+ The concentration of the aqueous solution is 1-20 mM, preferably 5-15 mM, and more preferably 10 mM; the second immersion time is 10-20 min.

[0025] Furthermore, the concentration of the bipyridine solution in step (4) is 2-10 mM, preferably 6 mM; the bipyridine solution is prepared using a PBS buffer solution with a pH of 6-8.

[0026] Furthermore, the immersion time in the bipyridine solution in step (4) is 100-140 min, preferably 120 min.

[0027] Furthermore, in step (4) Ru 3+An aqueous solution was obtained by dissolving RuCl3·6H2O in water.

[0028] Furthermore, the substrate electrode in step (4) is at least one of gold, silver, copper, nickel, iridium, platinum, chromium and their alloys, graphite, carbon black, carbon nanotubes, graphene and its derivatives, and indium tin oxide.

[0029] Furthermore, after obtaining IrNP in step (2), a modification step is also included, specifically dispersing IrNP in water containing NaOH, D- / L-cysteine ​​and glycerol, and refluxing at 65-100℃ for 16-48h to obtain IrNP-pS.

[0030] The concentrations of NaOH, D- / L-cysteine, and glycerol in water are 0.15-1.0M: 2-60mM: 0.02-0.2M, preferably 0.15-1.0M: 5-30mM: 0.02-0.2M.

[0031] The mass fraction of the IrNP in water is 0.05-1%, preferably 0.1-0.5%, and more preferably 0.2%.

[0032] By replacing the IrNP dispersion in step (2) with the IrNP-pS dispersion, the electrode IpR can be prepared.

[0033] Furthermore, a protective layer can be added to the IR or IpR surface. The protective layer material can be any one or more of the following: Nafion resin, hyaluronic acid, chitosan, agarose, cellulose, alginic acid, chondroitin sulfate, polyuronic acid, gelatin, collagen, polyurethane and its derivatives, silk fibroin, polyvinyl alcohol and its derivatives or copolymers. This can enhance the stability of the sensitive material on the electrode surface; improve the hydrophilicity / hydrophobicity of the electrode interface and its resistance to interference from specific ions; improve the biocompatibility of the electrode surface; and reduce the surface free energy of the electrode.

[0034] An electrochemical calibration-free pH sensing method includes the following steps:

[0035] The pH of a pH standard buffer solution was detected using an electrode system containing the aforementioned pH-sensitive and pH-insensitive electrodes.

[0036] The pH indicator parameter au was defined by voltammetric analysis. Where E f ΔI1 and ΔI2 are the peak potentials, HWHM is the half-peak half-width potential, ΔI1 and ΔI2 are the peak current intensities corresponding to the pH-sensitive electrode and the pH-insensitive electrode, respectively, and a is the electrode constant.

[0037] A regression curve was generated based on the relationship between au value and pH value of standard buffer solution;

[0038] An electrode system consisting of the same pH-sensitive electrode and pH-insensitive electrode was used to detect wound exudate at the wound site, and the pH value was determined based on the regression curve.

[0039] The current and potential signals generated by the pH-sensitive electrode will respond to changes in the pH being measured.

[0040] The current and potential signals generated by the pH-insensitive electrode do not respond to changes in pH.

[0041] Furthermore, the current and potential signals generated by the pH-sensitive electrode will respond to changes in the pH being measured and environmental interference factors; the current and potential signals generated by the pH-insensitive electrode will not change with pH but will only respond to changes in environmental interference factors.

[0042] Furthermore, the interfering factors mainly include ion types, ion concentrations, solution viscosity, strong acids and bases, extreme temperatures, fluorides, sulfides, adhering substances, biological pollutants, etc. These are all environmental factors that seriously interfere with the accuracy of traditional methods. This solution has good anti-interference performance against these known interfering factors.

[0043] The pH-insensitive electrode is at least one of gold, silver, copper, nickel, iridium, platinum, chromium and their alloys, graphite, carbon black, carbon nanotubes, graphene and its derivatives, or indium tin oxide, or is prepared by modifying the above electrode materials with functionalized materials or introducing functionalized materials in situ.

[0044] Furthermore, the functional materials for the pH-insensitive electrode include, but are not limited to, at least one of the following: pure metals, alloys, carbon materials, metal chelates and their derivatives, benzene compounds and their derivatives, heterocyclic aromatic hydrocarbons and their derivatives, and electrochemically active biomolecules and their derivatives. Even further, the functional materials for the pH-insensitive electrode include, but are not limited to, at least one of the following: silver, gold, gold amalgam, tin-lead alloy, boron-doped diamond, carbon black, glassy carbon, ruthenium bipyridine, phenanthroline iron, (poly)phenylthiol, and poly(4-vinylpyridine).

[0045] Furthermore, the modification method of the functionalized material on the substrate electrode is at least one of chemical vapor deposition, vacuum sputtering, evaporation, electroplating, electrochemical deposition, chemical in-situ synthesis, electrochemical in-situ synthesis, and film formation.

[0046] Furthermore, the electrode system is one of a two-electrode, a three-electrode, or a four-electrode system.

[0047] Furthermore, the electrode can be a screen-printed electrode, a planar sputtered thin film electrode, or a planar MEMS microelectrode.

[0048] Furthermore, the electrode substrate material is at least one of polyester, polycarbonate, polyurethane, polyimide, polyethylene, polyvinylidene fluoride, polytetrafluoroethylene, polyvinyl chloride, paper, silk, nylon, polyethersulfone, polydimethylsiloxane, or ethylene-vinyl acetate copolymer.

[0049] Furthermore, the current-voltage analysis method includes, but is not limited to, at least one of linear current-voltage analysis, cyclic current-voltage analysis, pulse current-voltage analysis, differential pulse current-voltage analysis, and square wave pulse current-voltage analysis.

[0050] Furthermore, for linear voltammetry and cyclic voltammetry, the scanning potential window is -2.5 to +2.5 V; the scanning rate is 0.01 mV / s to 10 V / s; for pulse voltammetry, the potential increment is 0.01 to 1000 mV / s, the potential amplitude is 1 to 500 mV, and the frequency is 1 to 100000 Hz.

[0051] Furthermore, after the data is measured by the volt-ampere analysis method, the data undergoes primary processing, including but not limited to linear filtering and linear parameter extraction; further still, the linear filtering includes at least one of differential filtering, high-pass filtering, and low-pass filtering; the linear parameter extraction includes at least one of derivative function analysis, nonlinear fitting, machine learning fitting, and interpolation.

[0052] Compared with the prior art, the present invention has the following technical features:

[0053] (1) This invention overcomes the problem of biocompatibility defects in conventional Ir-based electrodes. It achieves biocompatibility of electrode sensitive materials by introducing biocompatible coating materials onto the surface of Ir-based materials, thus solving the potential risks of using Ir-based pH sensors in wearable applications.

[0054] (2) The solution provided by this invention adopts a novel, non-traditional principle, achieving accurate quantification of pH without calibration. The finished product is ready to use upon leaving the factory and does not require any calibration or standardization during its service life, thus fully meeting the requirements for in-situ long-term online pH sensing of wound surfaces. At the same time, the electrode achieves a miniaturized and planar design, suitable for contact use on wound surfaces. Attached Figure Description

[0055] Figure 1 The following are the characterization results of Ir-based sensitive materials, where (a) is a TEM image of IrNP in Example 1; (b) is a TEM image of IrNP-pS in Example 2; (c) is a fine XPS C1s spectrum of IpR in Example 1; and (d) is a fine XPS Ir4f spectrum of IpR in Example 1.

[0056] Figure 2The square pulse voltammogram (a) of the IpR screen-printed electrode in Britton-Robinson buffer solution at pH 2-12 in Example 4, and the au-pH regression curve (b) plotted after converting the au value from the result.

[0057] Figure 3 The images show (a) and (b) of the simulated long-term wear test of the IpR flexible electrode in wound contact in Example 5.

[0058] Figure 4 The results of quantitative analysis (a) and simultaneous empirical judgment (b) of the wound health status in animal experiments using IpR screen-printed electrodes, as shown in Comparative Example 1.

[0059] Figure 5 The left and right graphs show the changes of two cytokines during the experimental period in Comparative Example 2. The left graph shows IL-13 and the right graph shows IL-4. Detailed Implementation

[0060] Example 1

[0061] (1) Fabrication of IpR screen-printed electrodes

[0062] IrCl4 was dissolved in 0.5M HCl to prepare 100 mL of an aqueous solution containing 10 mM IrCl4. The pH of the solution was adjusted to 4.0 with DMSO solution containing 0.5M triethanolamine. 20 mL of 0.2M DMSO solution of 3-methyl-1,2-cyclopentanedione was added. The mixture was thoroughly mixed and allowed to stand at room temperature in the dark for 24 h before use. The mixture was heated to 85°C, and 100 mL of 0.1M sodium L-ascorbate solution was added dropwise with stirring. After the reaction was completed for 2 h, the reaction solution was cooled to room temperature and centrifuged at 12000 rpm for 15 min. The precipitate was collected to obtain IrNP. An aqueous solution containing 15 mM 4,4'-diamino-5,5”-(dithiophene-2-yl)-2,2'-bipyridine was prepared using 0.1 M PBS buffer at pH 7.4. 20 mL of this solution was added dropwise to 80 mL of IrNP dispersion (0.5% by mass) with stirring, and the reaction was carried out at 55 °C for 16 h. After the reaction was complete, the reaction mixture was cooled to room temperature and centrifuged at 10,000 rpm for 30 min, and the precipitate was collected.

[0063] The sediment was redispersed with pure water to prepare a dispersion system with a mass fraction of 0.25%. A solution containing 10 mM RuCl3·6H2O was prepared by pre-dissolving RuCl3·6H2O in 0.1 M HCl solution. 3+The clean stone-ground rod working electrode was immersed in the solution and kept at 50°C for 60 min before being removed and washed. A 10 mM RuCl3·6H2O solution was prepared using 0.1 M HCl solution. The working surface of the electrode was immersed in the solution and kept at room temperature for 15 min before being removed and washed. The working surface of the electrode was partially immersed in a 6 mM bipyridine solution prepared with 0.1 M PBS buffer at pH 7.0 and kept at room temperature for 120 min before being removed and washed to prepare the IR electrode. The electrode constant a was determined to be 0.176.

[0064] Example 2

[0065] The IrNP obtained in Example 1 was dispersed in ultrapure water containing 0.5M NaOH, 10mM L-cysteine, and 0.1M glycerol (the mass fraction of IrNP was 0.2%), refluxed at 100°C for 16-48h, cooled to room temperature, centrifuged at 12000rpm for 15min, and the precipitate was collected to obtain IrNP-pS.

[0066] Example 3

[0067] An aqueous solution containing 15 mM 4,4'-diamino-5,5”-(dithiophene-2-yl)-2,2'-bipyridine was prepared using 0.1 M PBS buffer at pH 7.4. 20 mL of this solution was added dropwise to 80 mL of an aqueous dispersion (0.5% by mass) of IrNP-pS prepared in Example 2, with stirring. The reaction was carried out at 55 °C for 16 h. After the reaction was complete, the reaction mixture was cooled to room temperature and centrifuged at 10,000 rpm for 30 min, and the precipitate was collected.

[0068] A 10 mM RuCl3·6H2O solution was prepared using 0.1 M HCl solution. The clean stone-ground rod working electrode was immersed in this solution and kept at 50 °C for 60 min, then removed and washed. A 10 mM RuCl3·6H2O solution was also prepared using 0.1 M HCl solution. The working surface of the electrode was immersed in this solution and kept at room temperature for 15 min, then removed and washed. Finally, the working surface of the electrode was partially immersed in a 6 mM bipyridine solution prepared using 0.1 M PBS buffer (pH 7.0) and kept at room temperature for 120 min, then removed and washed to prepare the electrode IpR. The electrode constant a was determined to be 0.178.

[0069] Example 4

[0070] The electrode IpR prepared in Example 3 was used as the working electrode (2×2 mm), the stone-ground rod counter electrode (2×2 mm), and the Ag reference electrode (2×1 mm) to form a CC-Ag three-electrode screen-printed electrode system. A 40 mM Britton-Robinson standard buffer solution was prepared, and the pH was adjusted to 2-12 using 0.5 M standard hydrochloric acid and potassium hydroxide. Differential pulse voltammetry was used to measure the changes in current and potential of the polymer voltammetric pH electrode with pH in the buffer sample (scanning potential from 800 to -500 mV, pulse potential increment of 5 mV, pulse amplitude of 50 mV, pulse frequency of 20 Hz). The pH response curve was recorded. The differential pulse voltammetry plots in Britton-Robinson standard buffer solutions containing 1 wt% bovine serum albumin (BSA) at pH 2-12 are shown below. Figure 2 As shown in (a). The microcontroller software automatically calculates E based on the curve results data. f , HWHM, ΔI1, ΔI2, a and au The values ​​of au and pH of the standard buffer were recorded to generate a regression curve. The results are as follows: Figure 2 As shown in (b).

[0071] (2) Wound exudate testing using IpR screen-printed electrodes

[0072] Ten samples of chronic trauma aspirated using a vacuum aspiration (VSD) device were collected. Five samples came from patients diagnosed with healthy wounds, while the other five came from patients diagnosed with infected wounds. The pH of the samples was independently and quantitatively analyzed using IpR screen-printed electrodes and a commercially available imported precision glass bulb pH meter. The results are shown in Table 1.

[0073] Table 1 Comparison of quantitative pH test results for wound exudate (N=5)

[0074]

[0075] The comparative test results show that the IpR screen-printed electrode, used to measure the pH value of wound exudate, exhibits extremely high consistency with commercially available imported precision glass bulb pH meters, accurately reflecting the pH level of wound exudate and differentiating the pH status of wounds from different sources and in different states. Furthermore, the analysis revealed that saline-assisted treatment slightly decreases the pH of wound exudate; common bacterial infections lead to a significant increase in the pH of wound exudate. These observations are consistent with medical practice and basic research experience, indicating that the IpR screen-printed electrode can accurately reflect the pH status of wound exudate.

[0076] Example 5

[0077] (1) Fabrication of IpR flexible electrode

[0078] IrCl4 was dissolved in 0.5M HCl to prepare 100 mL of an aqueous solution containing 10 mM IrCl4. The pH of the solution was adjusted to 4.0 with DMSO solution containing 0.5 M triethanolamine. 20 mL of 0.2 M 3-methyl-1,2-cyclopentanedione solution was added. The mixture was thoroughly mixed and allowed to stand at room temperature in the dark for 24 h before use. The mixture was heated to 95 °C, and 100 mL of 0.06 M L-ascorbic acid sodium solution was added dropwise with stirring. After the reaction was completed, the reaction solution was cooled to room temperature and centrifuged at 10,000 rpm for 20 min. The precipitate was collected to obtain IrNP. IrNP was dispersed in ultrapure water containing 1 M triethanolamine and 15 mM L-cysteine ​​(IrNP mass fraction 0.2%), refluxed at 100 °C for 6 h, cooled to room temperature, and centrifuged at 10,000 rpm for 30 min. The precipitate was collected to obtain IrNP-pS. An aqueous solution containing 15 mM 4,4'-diamino-5,5”-(dithiophene-2-yl)-2,2'-bipyridine was prepared using 0.1 M PBS buffer at pH 7.4. 20 mL of this solution was added dropwise to 80 mL of LIrNP dispersion (0.5% by mass), and the reaction was carried out at 55 °C for 16 h. After the reaction was complete, the solution was cooled to room temperature and centrifuged at 10,000 rpm for 30 min, and the precipitate was collected.

[0079] A 10 mM RuCl3·6H2O solution was prepared using 0.1 M HCl solution. The clean working electrode portion of the stone-ground rod was immersed in this solution and kept at 80 °C for 10 min before being removed and washed. A 10 mM RuCl3·6H2O solution was also prepared using 0.1 M HCl solution. The working surface of the electrode was immersed in this solution and kept at room temperature for 15 min before being removed and washed. The working surface of the electrode was then immersed in a 6 mM bipyridine solution prepared using 0.1 M PBS buffer (pH 7.0) and kept at room temperature for 120 min before being removed and washed to prepare the IR electrode. Using a similar method, the IrNP dispersion was replaced with IrNP-pS to prepare the IpR electrode. The electrode constant a was determined to be 0.181.

[0080] The prepared electrode IpR was used as the working electrode (2×2 mm), the stone-ground rod as the counter electrode (2×2 mm), and the Ag reference electrode (2×1 mm) to form a CC-Ag three-electrode screen-printed electrode system. A 40 mM Britton-Robinson standard buffer solution was prepared, and the pH was adjusted to 2-12 using 0.5 M standard hydrochloric acid and potassium hydroxide. Differential pulse voltammetry (scanning potential from 800 to -500 mV, pulse potential increment of 5 mV, pulse amplitude of 50 mV, pulse frequency of 20 Hz) was used to test the changes in current and potential of the polymer voltammetric pH electrode with pH in the buffer sample. The pH response curve was recorded, and the microcontroller software automatically calculated E based on the curve results. f , HWHM, ΔI1, ΔI2, a and au The values ​​of au and pH were recorded to generate a regression curve. The measured values ​​of au = -16.02·pH + 5.521, R0 2 =0.998, and this regression equation was used for subsequent testing.

[0081] (2) Simulation test of contact wound surface pH sensing using IpR flexible electrode

[0082] Ballistic gelatin was prepared to simulate the surface of an open wound: gelatin particles were completely dissolved in deionized water to obtain a 30% by weight gelatin solution, which was then poured into a homemade mold and allowed to cool naturally to room temperature, forming 30 mm cubes. Subsequently, the gelatin cubes were transferred to a homemade electrochemical cell. Figure 3 a) The newly prepared IpR flexible electrode sheet was tightly adhered to the gelatin and sealed with a cap. The battery was then connected to a peristaltic pump to form a circulating, closed system, with artificial wound fluid (AWF) prepared according to the following composition: 2% human serum albumin, 0.36% sodium chloride, 0.05% sodium bicarbonate, 0.02% sodium citrate, 0.1% sodium lactate, 0.1% glucose, 0.01% calcium chloride dihydrate, 0.02% magnesium chloride, and 0.01% urea) as the fluid phase. The system was sterilized by exposing it to ultraviolet light overnight. The system was continuously tested for 60 days in SWV mode, and the pH value of the simulated environment was calculated from the results. In the experimental group, in addition to pumping AWF containing 0.04% NaHCO3 daily from day 20 to simulate the wound infection process, and pumping AWF containing 0.25% L-lactic acid from day 40 to 60 to simulate the wound healing process, a parallel system without any treatment was also tested for 60 days of continuous pH change.

[0083] The experimental results are recorded as follows: Figure 3b: In the experimental group, the pH changes recorded by the IpR flexible electrode matched the two characteristic changes in wound pH, indicating that the sensor could accurately record the wound pH value. In the control group, the pH recorded by the sensor did not change significantly and remained at the initial value level. This demonstrates that the sensor can maintain high stability and signal reproducibility under long-term in-situ testing conditions, and can be used for continuous and accurate measurement of wound pH.

[0084] Comparative Example 1

[0085] IpR screen-printed electrodes are used in VSD-assisted treatment to improve the efficiency of chronic trauma prediagnosis.

[0086] Electrode IpR screen-printed electrodes were prepared using the sensor fabrication method described in Example 1. The electrodes were installed inside the reservoir of a VSD-specific 3D dressing, with the electrode tip positioned outside the reservoir and connected to the sensing device. Sixteen healthy rats with a weight difference ≤10g and similar age and physical condition were selected and divided into two groups (50% male and 50% female) respectively. A wound was created on the back of each rat. The control group had the wound wrapped with a three-dimensional dressing containing IpR screen-printed electrodes; the experimental group had 2 mL of bacterial cells at a density of 100 CFU / mL inoculated into the wound before wrapping. -1 S. aureus (ATCC 25923) culture medium was used. During the experiment, animals were provided with complete nutrition, and the rearing environment was maintained at 24±1℃ and 55±10% humidity. pH readings were continuously recorded using an IpR screen-printed electrode. During the experiment, 100 μL of wound exudate was periodically aspirated from the storage bottle, and the IL-6 level was measured using ELISA. A blinded experimental method was used, requiring participating medical personnel to visually and empirically determine the timeline for wound deterioration in the experimental animals. Results are as follows... Figure 4 As shown:

[0087] Figure 4 The results showed that from the second day after inoculation with the infected strain, the expression level of the wound inflammation indicator IL-6 began to increase significantly, reaching its maximum rate of change on the fourth day, and approaching the highest level observed in experiments by the sixth day. Simultaneous pH measurements of the wound recorded by IpR screen-printed electrodes showed a trend almost identical to the changes in IL-6 levels, and the pH level exceeded neutral on days 3-4, indicating the onset of significant wound infection. Figure 4The results (b) indicate that, based on the experience of healthcare professionals, 65% of them judged the wound infection to have occurred on day 6, 15% on day 7, and only 20% on day 5. This demonstrates that relying on experience to correctly determine the timing of wound infection can be 2-3 days later than precise quantitative analysis, potentially leading to missed opportunities for early intervention in clinical practice. The results demonstrate that IpR screen-printed electrodes can effectively record real-time dynamic changes in the pH of chronic wounds, and these changes can indicate the onset of wound infection and inflammation. Their proper use can significantly shorten the time required for healthcare professionals to detect wound infection, providing valuable time for early intervention.

[0088] Comparative Example 2

[0089] IpR flexible electrodes are used for safe real-time pH monitoring of VSD contact wound surfaces.

[0090] The IR flexible electrode described in Example 1 and the IpR flexible electrode described in Example 5 were respectively installed within the wound surface contact protective layer of a VSD-specific three-dimensional dressing, with the electrode ends extending outside the dressing and connected to a sensing device. Ten healthy rats with a weight difference ≤10g and the same age and physical condition were selected and divided into two groups (50% male and 50% female) respectively. A wound was created on the back of each rat. The control group had their wounds wrapped with a three-dimensional dressing containing an IR flexible electrode; the experimental group had their wounds wrapped with a three-dimensional dressing containing an IpR flexible electrode. During the experiment, the animals were provided with complete nutrition, and the rearing environment was maintained at 24±1℃ and 55±10% humidity. pH readings from the IR and / or IpR flexible electrodes were continuously recorded.

[0091] During the experiment, 100 μL of wound exudate was periodically aspirated from the storage bottle, and the levels of IL-4 and IL-13 were measured using ELISA. The changes in these two cytokines throughout the experimental period are shown below. Figure 5As shown, during a 12-day monitoring period, the IR electrode induced a significant increase in IL-4 levels at the wound site after 6 days of wear, and a significant increase in IL-13 levels after 12 days of wear. IL-13 and IL-4 are important Th2 cytokines that play a crucial role in allergic reactions. They can stimulate B cells to differentiate into plasma cells and promote the production of IgE antibodies by these cells, which is an important step in allergic reactions. In addition, IL-4 can also promote the activation and proliferation of mast cells and basophils, thereby enhancing allergic reactions. Changes in their levels can directly reflect the intensity of allergic reactions. This indicates that conventional iridium-based electrodes, as wearable electrode materials for wounds, can induce certain allergic reactions and pose a potential risk. In contrast, no allergic reactions were observed in the wear test of the IpR electrode, indicating that it has ideal biocompatibility and is suitable as a sensing material for long-term wearable devices for wounds, directly applied in pH monitoring scenarios that require direct contact with the wound surface.

Claims

1. A pH-sensitive electrode comprising a wearable pH sensor for wound condition diagnosis, characterized in that... It is prepared by the following method: (1) Use triethanolamine DMSO solution to adjust the pH of IrCl4 solution to 2.0-6.0, add DMSO solution of 3-methyl-1,2-cyclopentanedione, mix well and let stand in the dark; (2) Heat the mixed solution obtained in step (1) to 75-100 °C, add L-ascorbic acid sodium solution under stirring, react for 0.2-4.5 h, collect the precipitate i.e. IrNP, and disperse it in water to obtain IrNP dispersion; (3) Prepare an aqueous solution of 4,4'-diamino-5,5''-(dithiophen-2-yl)-2,2'-bipyridine, add the solution to the IrNP dispersion under stirring, and react at 45~70℃ for 12~24 h. After the reaction is completed, collect the precipitate, redisperse the precipitate with water, and prepare a dispersion system with a mass fraction of 0.05~0.5%; (4) Add Ru-containing substances to the dispersion system obtained in step (3). 3+ The cleaned substrate electrode is immersed in an aqueous solution and kept at a constant temperature of 40-60 °C for 45-80 min. After rinsing, it is then immersed in a solution containing Ru. 3+ The pH-sensitive electrode, denoted as IR, is obtained by immersing the electrode in an aqueous solution of IR and then in a bipyridine solution for 60-180 min. The concentration of the IrCl4 solution in step (1) is 5~15 mM; the concentration of the DMSO solution of 3-methyl-1,2-cyclopentanedione is 0.1-0.25 M; the volume ratio of the IrCl4 solution to the DMSO solution of 3-methyl-1,2-cyclopentanedione is 100:20~75. The volume ratio of the mixed solution obtained in step (1) to the L-ascorbic acid sodium solution is 12~18:5~25; Step (2) after obtaining IrNP includes a modification step, specifically dispersing IrNP in water containing NaOH, D- / L-cysteine ​​and glycerol, and refluxing at 65-100 °C for 16-48 h to obtain IrNP-pS; The electrode IpR can be prepared by replacing the IrNP dispersion in step (3) with the IrNP-pS dispersion. The concentrations of NaOH, D- / L-cysteine, and glycerol in water are 0.15-1.0 M: 2-60 mM: 0.02-0.2 M; The mass fraction of the IrNP in water is 0.05~1%.

2. The pH-sensitive electrode according to claim 1, characterized in that: The mass fraction of the IrNP dispersion in step (2) is 0.01%~2%; In step (3), the aqueous solution of 4,4'-diamino-5,5''-(dithiophen-2-yl)-2,2'-bipyridine has a concentration of 10~20 mM and its volume ratio with the IrNP dispersion is 2:4~12. The mass fraction of the dispersion system in step (3) is 0.1~0.5%.

3. The pH-sensitive electrode according to claim 1, characterized in that: Step (4) The first time the Ru-containing 3+ Ru in aqueous solution 3+ Concentration 1~20 mM; In step (4), the temperature for the first isothermal treatment is 45~55℃; Step (4) The second Ru 3+ The concentration of the aqueous solution is 1~20 mM; The concentration of the bipyridine solution in step (4) is 2~10 mM.

4. The pH-sensitive electrode according to claim 1, characterized in that: The concentration of triethanol in step (1) is 0.1~1 M; The temperature for standing in step (1) is 20~35℃ and the standing time is 12~36h.

5. The pH-sensitive electrode according to claim 1, characterized in that: The substrate electrode in step (4) is at least one of gold, silver, copper, nickel, iridium, platinum, chromium and their alloys, graphite, carbon black, carbon nanotubes, graphene and its derivatives, and indium tin oxide.

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