Cell electrochemical sensor and use thereof in evaluation of Anti-inflammatory activity of capsaicin
By preparing a cellular electrochemical sensor based on a flower-like nanostructure of Sb2O4/rGO and DDAB-HIMIMPF6 composite material, and combining it with a GelMA hydrogel model, the cumbersome and costly problems of capsaicin anti-inflammatory activity detection were solved, achieving rapid, low-cost, and convenient detection results and improving detection sensitivity.
Patent Information
- Authority / Receiving Office
- WO · WO
- Patent Type
- Applications
- Current Assignee / Owner
- JIANGSU UNIV
- Filing Date
- 2025-03-31
- Publication Date
- 2026-05-07
AI Technical Summary
Existing methods for detecting the anti-inflammatory activity of capsaicin are cumbersome, costly, and have low sensitivity, and cannot effectively simulate the intracellular growth process.
A cell electrochemical sensor was prepared using a flower-like nanostructured Sb2O4/rGO and DDAB-HIMIMPF6 composite material. Combined with a GelMA hydrogel model, the effect of capsaicin on NO release was detected by the DPV method to evaluate its anti-inflammatory ability.
This method enables rapid, low-cost, and convenient detection of capsaicin's anti-inflammatory activity, effectively mimicking intracellular growth processes and improving detection sensitivity and accuracy.
Smart Images
Figure CN2025086117_07052026_PF_FP_ABST
Abstract
Description
A cell electrochemical sensor and its use in assessing the anti-inflammatory activity of capsaicin. Technical Field
[0001] This invention belongs to the field of biosensing technology, specifically relating to a cell electrochemical sensor and its use in evaluating the anti-inflammatory effects of capsaicin. Background Technology
[0002] Capsaicin (8-methyl-N-vanillyl-6-nonanoamide), a derivative of benzylamine, is a major compound in the capsaicin family, accounting for nearly 70% of all capsaicin compounds in spices. Capsaicin content is an important quality indicator for commercial chilies and other peppers. Many people consider capsaicin the king of medicinal herbs because they believe it offers numerous health benefits. It has been reported that capsaicin not only enhances the flavor of food but also possesses various pharmacological and physiological activities, including analgesia, anticancer, anti-inflammatory, antioxidant, gastrointestinal improvement, and weight loss. Among these, capsaicin's natural anti-inflammatory properties can alleviate symptoms of arthritis, heart disease, and diabetes, and reduce their risk. Therefore, testing the intracellular anti-inflammatory activity of capsaicin has significant clinical and practical implications.
[0003] The anti-inflammatory activity of capsaicin is being investigated using a newly developed method. Capsaicin anti-inflammatory assays offer significant advantages because they mimic cell growth processes, including uptake and metabolism. Currently, A549 cells are widely used as an in vitro model of type II alveolar epithelial cells and are the preferred cell type for establishing in vitro cell models of acute lung injury (ALI). Endotoxin is a crucial factor contributing to ALI, and its main component, lipopolysaccharide (LPS), plays a vital role in ALI. LPS-induced A549 cells can establish a relatively ideal in vitro ALI inflammatory injury model. Nitric oxide (NO) was chosen as a representative inflammatory factor. Studies have shown that the amount of NO released from cells is directly proportional to LPS concentration, and capsaicin pretreatment significantly alleviates LPS-induced acute lung injury in mice. Therefore, the anti-inflammatory activity of capsaicin was evaluated by detecting the reduction in NO release.
[0004] The main methods for detecting nitric oxide (NO) include gas chromatography-mass spectrometry, fluorescence spectrometry, spectroscopic analysis, and chemical analysis. Spectroscopic analysis includes infrared absorption spectroscopy and ultraviolet absorption spectroscopy. Chemical analysis primarily utilizes the determination of reaction products between nitric oxide and other compounds to indirectly quantify NO concentration; common methods include the Griess reagent method and the ferric salt method. However, due to the cumbersome detection steps, expensive reagents, and short half-life of NO in traditional methods, it is imperative to select a rapid, simple, inexpensive, and efficient method for detecting nitric oxide.
[0005] Compared to other detection methods, electrochemical sensors offer advantages such as small electrode size, non-destructive analysis, minimal or no reagent requirements, high sensitivity, simplicity, and low cost. Therefore, electrochemical detection is a feasible method for measuring NO released by cells. Among existing technologies, the patent "A fluorescent probe compound for detecting nitric oxide and its preparation method and application" (CN118126073A) discloses a highly stable and selective fluorescent probe for nitric oxide, but its preparation process is complex and time-consuming. The patent "A fluorescent probe capable of separately and simultaneously detecting nitric oxide and nitrosyl and its preparation method and application" (CN116655692A) discloses a highly selective, non-invasive, visualized fluorescent detection probe; however, the fluorescence is easily quenched and the fluorescence intensity is weak, which also reduces the detection effect. Summary of the Invention
[0006] To address the aforementioned problems, this invention aims to solve one of them; this invention provides a cell electrochemical sensor and its application in evaluating the anti-inflammatory effects of capsaicin, which has outstanding advantages such as low cost, fast detection, simple operation, and strong miniaturization capability.
[0007] To achieve the above objectives, the present invention adopts the following technical solution;
[0008] A method for preparing a cell electrochemical sensor, comprising the following steps:
[0009] Step 1: Preparation of flower-like nanostructured Sb₂O₄ / rGO;
[0010] (1) Dissolve antimony trichloride in ethanol to obtain an antimony trichloride ethanol solution; then add graphene oxide suspension to the antimony trichloride ethanol solution and mix, and then treat with ultrasound to obtain a mixed solution;
[0011] (2) The mixed solution obtained in step (1) is sealed in a reaction vessel and heated to react. After the heating reaction, the generated black precipitate is collected and then soaked in ultrapure water overnight. Finally, the precipitate is collected again after filtration. The precipitate product is freeze-dried to obtain the final product, which is Sb2O4 / rGO.
[0012] Step 2: Preparation of DDAB-HIMIMPF6 composite material;
[0013] The mixture of bis(dodecyl dimethyl ammonium bromide) solution and 1-hexyl-3-methylimidazolium hexafluorophosphate is called DDAB-HIMIMPF6 composite material (it will become solid at room temperature, so if it is solid before use, it needs to be heated to become liquid).
[0014] Step 3: Fabrication of the electrochemical sensor;
[0015] (1) Weigh the Sb2O4 / rGO prepared in step one and dissolve it in pure water and stir evenly to obtain a mixed solution; then take the mixed solution and drop it onto the screen printing electrode (SPCE), blow it dry with nitrogen gas to obtain the treated screen printing electrode.
[0016] (2) The DDAB-HIMIMPF6 composite material prepared in step 2 is heated in a water bath to form a liquid state, and then dropped onto the screen-printed electrode after step 1. After drying at room temperature, the cell electrochemical sensor is obtained.
[0017] Preferably, in step (1), the ratio of antimony trichloride, ethanol and graphene oxide suspension is 0.5 mol: 15 ml: 15 ml, wherein the concentration of graphene oxide suspension is 1 mg / ml; and the ultrasonic treatment time is 30 min.
[0018] Preferably, in step (2), the heating reaction temperature is 180°C and the time is 21 hours.
[0019] Preferably, in step two, the volume ratio of the dodecyl dimethyl ammonium bromide solution to 1-hexyl-3-methylimidazolium hexafluorophosphate is 5:1, wherein the concentration of the dodecyl dimethyl ammonium bromide solution is 0.1M.
[0020] Preferably, in step three (1), the ratio of Sb2O4 / rGO to pure water is 1 mg: 1 ml, and the amount of the mixed solution added is 5 μL.
[0021] Preferably, in step three (2), the water bath heating temperature is 20-40℃, and the amount added is 5μL.
[0022] The application of cell electrochemical sensors to evaluate the anti-inflammatory effects of capsaicin is as follows:
[0023] Step 1: Preparation of cell-methacrylated gelatin hydrogel (GelMA);
[0024] (1) Dissolve gelatin in phosphate-buffered saline (PBS), stir, and then add methacrylate anhydride dropwise, stirring until homogeneous to obtain a mixed solution; then dilute the mixed solution 3-5 times with phosphate-buffered saline to terminate the reaction, obtaining a reaction solution; dialyze the reaction solution obtained in step 1 to obtain a dialysate; then filter the dialysate, and pre-freeze the filtrate to obtain GelMA prepolymer; wherein the ratio of gelatin, phosphate-buffered saline, and methacrylate anhydride is 5g:50ml:4-6ml; the stirring temperature is 50℃, and the temperature of the phosphate-buffered saline is 50℃ when diluting, and the dilution is 1-fold each time; the dialyzing temperature is 50℃, and the time is 7-14 days; the filtration is performed using a membrane with a pore size of 0.22μm; the pre-freezing temperature is -20℃ or -80℃, and the pre-freezing time is 5-10 days.
[0025] (2) Weigh the GelMA prepolymer into a brown bottle, then add PBS containing the photoinitiator Irgacure 2959, dissolve it in a warm bath, and then filter it to obtain a GelMA solution; wherein the ratio of GelMA prepolymer to PBS is 0.25-1g:5ml; the photoinitiator is Irgacure 2959, and the concentration in PBS is 0.5% (w / v); the temperature of the warm bath is 40℃, and the filtration is carried out using a membrane with a pore size of 0.22μm;
[0026] (3) A549 cells were resuspended in GelMA solution to obtain cell-GelMA solution, in which the cell density was 10-1. 6 Cells / ml; then, cell-GelMA solution was added to each well of a 96-well plate and photocured under UV light to form a cell-GelMA hydrogel, denoted as A549 / GelMA; the volume of cell-GelMA solution used was 100 μL, and the UV light conditions were: 405 nm, 50 mW / cm². 2 The photocuring time is 30 seconds;
[0027] Step 2: Establishment of an LPS-induced A549 cell inflammation model
[0028] First, LPS solutions were prepared in DMEM medium at concentrations of 0–20 μg / ml. Then, an equal volume of LPS solution was added to A549 / GelMA for induction treatment. After induction treatment, the supernatant was used as the test solution, and the cell electrochemical sensor was used for testing. The change in current signal was measured using the DPV method, and the corresponding current value was recorded as ILPS. The induction treatment time was 24 h.
[0029] Step 3: Capsaicin anti-inflammatory activity assay
[0030] (1) First, a capsaicin solution was prepared using DMEM medium with a concentration of 0–180 μM; then, 100 μL of capsaicin solution was added to the A549 / GelMA prepared in step 1 for incubation, with an incubation temperature of 37 °C and a gas environment of 5% CO2; after incubation, 100 μL of LPS solution (concentration of 1 μg / ml) was added for induction for 24 h.
[0031] After induction, the supernatant was used as the test solution, and the cell electrochemical sensor was used for testing. The peak current value of NO released by the cells was detected by the DPV method, and the current value was recorded as ICAP.
[0032] The relative anti-inflammatory capacity of capsaicin was evaluated using formula (I):
[0033] Where: ILPS is the peak current value of DPV induced by LPS treatment;
[0034] ICAP is the DPV current value of A549 cells after incubation with capsaicin followed by LPS induction.
[0035] IA is the blank control, i.e., the peak current of DPV without capsaicin and LPS treatment;
[0036] △NO: The amount of NO release reduced.
[0037] By comparing the NO current value released by A549 cells stimulated by LPS with the NO current value released by A549 cells stimulated by LPS after intervention with capsaicin, the reduction in NO release can be calculated according to formula (I).
[0038] The anti-inflammatory ability of capsaicin is assessed by the reduction in NO release (ΔNO). The greater the reduction, the stronger the anti-inflammatory ability of capsaicin.
[0039] The beneficial effects of this invention are:
[0040] On the one hand, the unique, highly networked flower-like nanostructure of Sb₂O₄ / rGO, due to its unique three-dimensional flower-like structure, allows NO to easily enter the reaction center for further capture and adsorption, subsequently forming a crystalline structure within the Sb₂O₄ / rGO nanostructure. 3+ and Sb 4+ Rapid electron transfer occurs between them, and charge transport is enhanced by reducing graphene oxide sheets.
[0041] On the other hand, a novel biocompatible composite membrane consisting of the water-insoluble surfactant bis(dodecyl dimethyl ammonium bromide) (DDAB) and the hydrophobic room-temperature ionic liquid (RTIL) 1-hexyl-3-methylimidazolium hexafluorophosphate (HIMIMPF6) can be used as an electrode coating to eliminate NO. 2-The interference from ascorbic acid and uric acid improves the detection efficiency of this biosensor for NO gas.
[0042] In summary, the present invention provides a cell electrochemical sensor that can not only be used to evaluate the anti-inflammatory effects of capsaicin, but also has the outstanding advantages of low cost, fast detection, simple operation, and miniaturization. Attached Figure Description
[0043] Figure 1 shows the SEM image of the GelMA prepolymer.
[0044] Figure 2 shows the hydrogen NMR spectrum of GelMA.
[0045] Figure 3 shows the standard curves of sodium nitrite solutions of different concentrations at 540 nm. Detailed Implementation
[0046] The invention will be described in more detail by means of the following embodiments; the following embodiments are merely illustrative and the invention is not limited to these embodiments.
[0047] Example 1:
[0048] Step 1: Synthesis method of Sb2O4 / rGO
[0049] 0.5 mol of antimony trichloride was dissolved in 15 ml of ethanol to provide antimony element, resulting in solution ①; 15 ml of graphene oxide suspension at 1 mg / ml was added to solution ① and mixed, and the mixture was sonicated for 30 min to obtain solution ②.
[0050] Liquid ② was sealed in a polytetrafluoroethylene-lined reactor and heated at 180°C for 21 hours to reduce graphene oxide and generate antimony tetroxide. After the heating reaction, the generated black precipitate was collected and then soaked in ultrapure water overnight to remove impurities. After filtration, the precipitate was collected and freeze-dried to obtain the final product Sb2O4 / rGO.
[0051] Step 2: Synthesis method of DDAB-HIMIMPF6
[0052] Mix 1 ml of bis(dodecyl)dimethylammonium bromide solution (0.1 M) with 200 μ L of 1-hexyl-3-methylimidazolium hexafluorophosphate to obtain a mixed material denoted as DDAB-HIMIMPF6 composite material (it will become solid at room temperature, therefore, if it is in a solid state before use, it needs to be heated to become liquid).
[0053] Step 3: Preparation of the modified nitric oxide electrochemical sensor
[0054] Weigh 1 mg of Sb2O4 / rGO and dissolve it in 1 ml of pure water. Stir well to obtain a mixed solution. Take 5 μL of the mixed solution and drop it onto the screen-printed electrode. Blow dry with nitrogen gas to obtain the treated screen-printed electrode.
[0055] The DDAB-HIMIMPF6 composite material was then heated in a 40°C water bath until it melted into a liquid state. 5 μL of the liquid was then added to the screen-printed electrode treated in step 1 and allowed to air dry at room temperature to obtain the cell electrochemical sensor.
[0056] Example 2:
[0057] The application of cell electrochemical sensors to evaluate the anti-inflammatory effects of capsaicin is as follows:
[0058] Step 1: Preparation of Cell-Methacrylated Gelatin Hydrogel (GelMA)
[0059] (1) Dissolve 5g of gelatin in 50ml of phosphate-buffered saline solution and stir gently. Then add 4ml of methacrylate anhydride dropwise to the gelatin solution and stir at 50℃ for 2h. Dilute the solution with 50℃ phosphate-buffered saline solution, diluting by 1 time each time, for 5 times to terminate the reaction, to obtain the reaction solution; dialyze the reaction solution obtained in step 1 at 50℃ for 7d to remove low molecular weight impurities (molecular weight cutoff: 0–22kDa), to obtain the dialysate; then pass the dialysate through a 0.22μm membrane filter, pre-freeze at -80℃, and then freeze for 5d to obtain the GelMA prepolymer.
[0060] Figure 1 shows a field emission scanning electron microscope image of GelMA. Its loose and porous structure is conducive to encapsulating cells and transporting NO produced by cells. Figure 2 shows the proton nuclear magnetic resonance spectrum of GelMA. As can be seen from the attached figure, GelMA was synthesized from gelatin, and calculations showed that the synthesized GelMA precursor had a substitution degree as high as 76%.
[0061] (2) Take 0.25g of GelMA prepolymer into a brown bottle, add 5ml of 0.5% (w / v) photoinitiator Irgacure 2959 in PBS, dissolve in a 40°C bath, and filter with a 0.22μm membrane filter while hot to sterilize.
[0062] (3) A549 cells were resuspended in GelMA solution to obtain cell-GelMA solution, in which the cell density was 10-1. 6 Cells / ml; then 100 μL of cell-GelMA solution was added to each well of a 96-well plate and incubated under UV light (405 nm, 50 mW / cm²). 2 After curing under light for 30 seconds, a cell-GelMA hydrogel was formed, denoted as A549 / GelMA.
[0063] Step 2: Establishment of an LPS-induced A549 cell inflammation model
[0064] Add 100 μL of LPS solutions prepared in DMEM medium at concentrations of 0, 0.4 μg / ml, 0.8 μg / ml, 1 μg / ml, 2 μg / ml, 4 μg / ml, 5 μg / ml, 10 μg / ml, and 20 μg / ml to the A549 / GelMA obtained in step 1 for induction treatment for 24 h.
[0065] After the induction treatment, the supernatant was used as the test solution and tested using a cell electrochemical sensor. The change in current signal was measured by the DPV method and the corresponding current value was recorded as ILPS.
[0066] Step 3: Validation (NO assay kit)
[0067] (1) NO generation level was determined using a NO assay kit based on Griess reaction.
[0068] RAW264.7 cells (100 μL, 10 5 (particles / ml) were induced in 96-well plates with 0, 0.4 μg / ml, 0.8 μg / ml, 1 μg / ml, 2 μg / ml, 4 μg / ml, 5 μg / ml, 10 μg / ml, and 20 μg / ml LPS solutions for 24 h.
[0069] Subsequently, 50 μL of culture supernatant was collected and mixed with 50 μL of Griess reagent I and 50 μL of Griess reagent II in an ELISA plate. After incubation for 15 min, the absorbance at 540 nm was measured using an ELISA reader.
[0070] (2) The standard solution of NaNO2 was measured using the same method. The resulting standard curve is shown in Figure 3. Nitric oxide can be detected based on the standard curve. The results were compared with the electrochemical results of this embodiment to determine the reliability of the method.
[0071] Step 4: Capsaicin anti-inflammatory activity assay
[0072] First, a capsaicin solution with a concentration of 0–180 μM was prepared using DMEM medium. Then, 100 μL of the capsaicin solution was added to the A549 / GelMA prepared in step 1 for incubation at a temperature of 37 °C and a gas environment of 5% CO2. After incubation, 100 μL of LPS solution (concentration of 1 μg / ml) was added for induction for 24 h.
[0073] After induction, the supernatant was used as the test solution, and the cell electrochemical sensor was used for testing. The peak current value of NO released by the cells was detected by the DPV method, and the current value was recorded as ICAP.
[0074] The relative anti-inflammatory capacity of capsaicin was evaluated using formula (I):
[0075] Where: ILPS is the peak current value of DPV induced by LPS treatment;
[0076] ICAP is the DPV current value of A549 cells after incubation with capsaicin followed by LPS induction.
[0077] IA represents the peak DPV current of the blank control group that was not treated with capsaicin and LPS.
[0078] △NO: The amount of NO release reduced.
[0079] By comparing the NO current value released by A549 cells stimulated by LPS with the NO current value released by A549 cells stimulated by LPS after intervention with capsaicin, the reduction in NO release can be calculated according to formula (I).
[0080] The anti-inflammatory ability of capsaicin is assessed by the amount of reduction in its NO release; the greater the reduction, the stronger the anti-inflammatory ability of capsaicin.
[0081] Note: The above embodiments are only used to illustrate the present invention and are not intended to limit the technical solutions described in the present invention. Therefore, although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the present invention. All technical solutions and improvements that do not depart from the spirit and scope of the present invention should be covered within the scope of the claims of the present invention.
Claims
1. A method for preparing a cell electrochemical sensor, characterized in that, Includes the following steps: Step 1: Preparation of flower-like nanostructured Sb₂O₄ / rGO; (1) Dissolve antimony trichloride in ethanol to obtain an antimony trichloride ethanol solution; then add graphene oxide suspension to the antimony trichloride ethanol solution and mix, and then treat with ultrasound to obtain a mixed solution; (2) The mixed solution obtained in step (1) is sealed in a reaction vessel and heated to react. After the heating reaction, the generated black precipitate is collected and then soaked in ultrapure water overnight. Finally, the precipitate is collected again after filtration. The precipitate product is freeze-dried to obtain the final product, which is Sb2O4 / rGO. Step 2: Preparation of DDAB-HIMIMPF6 composite material; The mixture of bis(dodecyl dimethyl)ammonium bromide solution and 1-hexyl-3-methylimidazolium hexafluorophosphate was denoted as DDAB-HIMIMPF6 composite material. Step 3: Fabrication of the electrochemical sensor; (1) Weigh the Sb2O4 / rGO prepared in step one, dissolve it in pure water and stir until uniform to obtain a mixed solution; then take the mixed solution and drop it onto the screen printing electrode, blow it dry with nitrogen gas to obtain the treated screen printing electrode. (2) The DDAB-HIMIMPF6 composite material prepared in step 2 is heated in a water bath to form a liquid state, and then dropped onto the screen-printed electrode after step 1. After drying at room temperature, the cell electrochemical sensor is obtained.
2. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that, In step one (1), the ratio of antimony trichloride, ethanol and graphene oxide suspension is 0.5 mol: 15 ml: 15 ml, where the concentration of graphene oxide suspension is 1 mg / ml; the ultrasonic treatment time is 30 min.
3. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that, In step two (2), the heating reaction is carried out at a temperature of 180°C for 21 hours.
4. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that, In step two, the volume ratio of the dodecyl dimethyl ammonium bromide solution to 1-hexyl-3-methylimidazolium hexafluorophosphate is 5:1, and the concentration of the dodecyl dimethyl ammonium bromide solution is 0.1M.
5. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that, In step three (1), the ratio of Sb2O4 / rGO to pure water is 1 mg: 1 ml, and the amount of the mixed solution added is 5 μL.
6. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that, In step three (2), the water bath heating temperature is 20-40℃, and the amount added is 5μL.
7. The use of the cell electrochemical sensor prepared by any one of claims 1-6 for evaluating the anti-inflammatory effects of capsaicin.
8. The use according to claim 7, characterized in that, The steps are as follows: Step 1: Preparation of cell-methacrylated gelatin hydrogel; (1) Dissolve gelatin in phosphate buffer, stir, and then add methacrylate anhydride dropwise, stirring until homogeneous to obtain a mixed solution; then dilute the mixed solution 3-5 times with phosphate buffer to terminate the reaction, obtaining a reaction solution; dialyze the reaction solution obtained in step 1 to obtain a dialysate; then filter the dialysate, and pre-freeze the filtrate to obtain GelMA prepolymer; wherein the ratio of gelatin, phosphate buffer, and methacrylate anhydride is 5g:50ml:4-6ml; the stirring temperature is 50℃, and the temperature of the phosphate buffer is 50℃ when diluting with phosphate buffer, and the dilution is 1-fold each time; the dialyzing temperature is 50℃, and the time is 7-14 days; the filtration is performed using a membrane with a pore size of 0.22μm; the pre-freezing temperature is -20℃ or -80℃, and the pre-freezing time is 5-10 days. (2) Weigh the GelMA prepolymer into a brown bottle, then add PBS containing the photoinitiator Irgacure 2959, dissolve it in a warm bath, and then filter it to obtain a GelMA solution; wherein the ratio of GelMA prepolymer to PBS is 0.25-1g:5ml; the concentration of the photoinitiator Irgacure 2959 in PBS is 0.5% (w / v); the temperature of the warm bath is 40℃, and the filtration is performed using a membrane with a pore size of 0.22μm; (3) A549 cells were resuspended in GelMA solution to obtain cell-GelMA solution, in which the cell density was 10-1. 6 Cells / ml; then, cell-GelMA solution was added to each well of a 96-well plate and photocured under UV light to form a cell-GelMA hydrogel, denoted as A549 / GelMA; the volume of cell-GelMA solution used was 100 μL, and the UV light conditions were: 405 nm, 50 mW / cm². 2 The photocuring time is 30 seconds; Step 2: Establishment of an LPS-induced A549 cell inflammation model First, LPS solutions were prepared in DMEM medium at concentrations of 0–20 μg / ml. Then, an equal volume of LPS solution was added to A549 / GelMA for induction treatment. After induction treatment, the supernatant was used as the test solution, and the cell electrochemical sensor was used for testing. The change in current signal was measured using the DPV method, and the corresponding current value was recorded as ILPS. The induction treatment time was 24 h. Step 3: Capsaicin anti-inflammatory activity assay (1) First, a capsaicin solution was prepared using DMEM medium with a concentration of 0–180 μM; then, 100 μL of capsaicin solution was added to the A549 / GelMA prepared in step 1 for incubation, with an incubation temperature of 37 °C and a gas environment of 5% CO2; after incubation, 100 μL of LPS solution (concentration of 1 μg / ml) was added for induction for 24 h. After induction, the supernatant was used as the test solution, and the cell electrochemical sensor was used for testing. The peak current value of NO released by the cells was detected by the DPV method, and the current value was recorded as ICAP. The relative anti-inflammatory capacity of capsaicin was evaluated using formula (I): Where: ILPS is the peak current value of DPV induced by LPS treatment; ICAP is the DPV current value of A549 cells after incubation with capsaicin followed by LPS induction. IA is the blank control, i.e., the peak current of DPV without capsaicin and LPS treatment; △NO: The amount of NO release reduced; By comparing the NO current value released by A549 cells stimulated by LPS with the NO current value released by A549 cells stimulated by LPS after intervention with capsaicin, the reduction in NO release can be calculated according to formula (I). The anti-inflammatory ability of capsaicin is assessed using the ΔNO value; the larger the ΔNO value, the stronger the anti-inflammatory ability of capsaicin.