Cell electrochemical sensor and application thereof in evaluating anti-inflammatory activity of capsaicin

Abstract

The invention belongs to the technical field of biosensing, and particularly relates to a cell electrochemical sensor and application thereof in evaluating the anti-inflammatory activity of capsaicin. Sb2O4 / rGO is firstly prepared, a DDAB-HIMIMPF6 composite material with stable current response and low biotoxicity is synthesized in one step, then the DDAB-HIMIMPF6 composite material is combined with the Sb2O4 / rGO, an electrochemical sensor is used for loading, and the cell electrochemical sensor with high selectivity and high sensitivity is finally obtained and can be applied to evaluation of the anti-inflammatory activity of capsaicin. Moreover, the cell electrochemical sensor provided by the invention not only can realize the purpose of evaluating the anti-inflammatory effect of capsaicin, but also has the outstanding advantages of low cost, high detection speed, simplicity in operation and miniaturization, and has a wide application prospect.

Description

Technical Field

[0001] The invention belongs to the technical field of biosensors, and in particular relates to a cell electrochemical sensor and an application thereof in evaluating the anti-inflammatory effect of capsaicin. Background Art

[0002] Capsaicin (8-methyl-N-vanillyl-6-nonanamide) is a derivative of benzylamine and the main compound in the capsaicinoid family, accounting for nearly 70% of the total number of capsaicinoids in spices. The content of capsaicinoids is an important quality indicator for commercial peppers and other peppers. Many people consider capsaicinoids to be the king of medicinal materials because they believe that it can bring many health benefits. It is reported that capsaicinoids not only increase the flavor of food, but also have a variety of pharmacological and physiological activities, including: analgesia, anti-cancer, anti-inflammatory, antioxidant, improve the gastrointestinal system, and lose weight. Among them, the natural anti-inflammatory ability of capsaicinoids can relieve symptoms such as arthritis, heart disease and diabetes, and reduce their risk of onset. Therefore, testing the intracellular anti-inflammatory activity of capsaicinoids has important clinical and practical significance.

[0003] The anti-inflammatory activity of capsaicin is a newly developed method. The anti-inflammatory activity test of capsaicin has obvious advantages because it simulates the cell growth process, including cell absorption and metabolism. At present, A549 cells are widely used as an in vitro model of type II alveolar epithelial cells and are the preferred cells for establishing an in vitro cell model of acute lung injury (ALI). Endotoxin is an important factor leading to ALI, and its main component lipopolysaccharide (LPS) plays an important role in ALI. LPS-induced A549 cells can establish a more ideal in vitro inflammatory injury model of ALI. Nitric oxide (NO) was selected as a representative of inflammatory factors. Studies have shown that the amount of NO released from cells is proportional to the concentration of LPS, and capsaicin pretreatment can significantly reduce LPS-induced acute lung injury in mice. Therefore, the anti-inflammatory activity of capsaicin is evaluated by detecting the reduction in NO release.

[0004] The main methods for detecting nitric oxide include gas chromatography-mass spectrometry, fluorescence, spectral analysis and chemical analysis. Spectral analysis includes infrared absorption spectroscopy and ultraviolet absorption spectroscopy. Chemical analysis mainly uses the determination of the reaction products of nitric oxide and other compounds to indirectly quantify the NO concentration. Common methods include Griess reagent method and iron salt method. However, due to the cumbersome detection steps of traditional methods, expensive detection reagents and the short half-life of NO, it is imperative to select a fast, simple, low-cost and efficient method for detecting nitric oxide.

[0005] Compared with other detection methods, electrochemical sensors have small electrode size, non-destructive analysis mode, minimal or no reagent requirements, high sensitivity, simplicity and low cost. Therefore, electrochemical detection is a feasible method for measuring NO released by cells. In the existing technology, the patent "A fluorescent probe compound for detecting nitric oxide, preparation method and application thereof" (CN118126073A) publishes a highly stable and highly selective nitric oxide fluorescent probe, but the preparation process of the probe is complicated and the preparation time is long. The patent "A fluorescent probe capable of separately and simultaneously detecting nitric oxide and nitrosyl, preparation method and application thereof" (CN116655692A) publishes a highly selective and non-invasive visual fluorescent detection probe, but the fluorescence is easily quenched and the fluorescence intensity is weak, which also reduces the detection effect. Summary of the invention

[0006] In view of the above problems, the present invention aims to solve one of the problems; the present invention provides a cell electrochemical sensor and its use for evaluating the anti-inflammatory effect of capsaicin, which has outstanding advantages such as low cost, fast detection, simple operation, and strong miniaturization capability.

[0007] In order to achieve the above objectives, the present invention adopts the following technical solutions:

[0008] A method for preparing a cell electrochemical sensor, the steps are as follows:

[0009] Step 1: Preparation of flower-like nanostructured Sb2O4 / rGO;

[0010] (1) dissolving antimony trichloride in ethanol to obtain an antimony trichloride ethanol solution; then adding a graphene oxide suspension to the antimony trichloride ethanol solution, mixing, and subjecting to ultrasonic treatment to obtain a mixed solution;

[0011] (2) sealing the mixed solution obtained in step (1) in a reactor for heating reaction, collecting the generated black precipitate after the heating reaction, and then soaking it in ultrapure water overnight, and finally collecting the precipitate again by filtration, and freeze-drying the precipitated product to obtain the final product, namely Sb2O4 / rGO;

[0012] Step 2: Preparation of DDAB-HIMIMPF6 composite material;

[0013] The didodecyl dimethyl ammonium bromide solution was mixed with 1-hexyl-3-methylimidazolium hexafluorophosphate, and the obtained mixed material was recorded as DDAB-HIMIMPF6 composite material (it will become a solid state when left at room temperature, so if it is in a solid state, it needs to be heated to become a liquid before use);

[0014] Step 3: preparing an electrochemical sensor;

[0015] (1) Weighing the Sb2O4 / rGO prepared in step 1, dissolving it in pure water and stirring it evenly to obtain a mixed solution; then dropping the mixed solution onto a screen printed electrode (SPCE), drying it with nitrogen gas, and obtaining a treated screen printed 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 dripped onto the screen-printed electrode treated in step 1 and dried at room temperature to obtain a cell electrochemical sensor.

[0017] Preferably, in step (1) of the present invention, the dosage of antimony trichloride, ethanol and graphene oxide suspension is in the ratio of 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, the heating reaction temperature in step (2) is 180° C. and the time is 21 h.

[0019] Preferably, in step 2, the volume ratio of the didodecyl dimethyl ammonium bromide solution to 1-hexyl-3-methyl imidazole hexafluorophosphate is 5:1, wherein the concentration of the didodecyl dimethyl ammonium bromide solution is 0.1M.

[0020] Preferably, in step three (1), the dosage 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, the water bath heating temperature in step 3 (2) is 20-40° C., and the amount added dropwise is 5 μL.

[0022] The use of cell-based electrochemical sensors to evaluate the anti-inflammatory effect 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, stir evenly to obtain a mixed solution; then dilute the mixed solution 3-5 times with phosphate buffer to terminate the reaction and obtain a reaction solution; dialyze the reaction solution obtained in step 1 to obtain a dialysate; then filter the dialysate, and pre-freeze the obtained filtrate to obtain a GelMA prepolymer; wherein the amount of gelatin, phosphate buffer, and methacrylate anhydride is 5g:50ml:4-6ml; the stirring temperature is 50°C, when diluting with phosphate buffer, the temperature of the phosphate buffer is 50°C, and the dilution is 1 times each time; the dialysis temperature is 50°C, and the time is 7-14d; the filtration is performed using a membrane with a pore size of 0.22μm; the pre-freezing temperature is -20°C or -80°C, and the pre-freezing time is 5-10d.

[0025] (2) GelMA prepolymer was weighed into a brown bottle, and then PBS containing photoinitiator Irgacure 2959 was added. After dissolving in a warm bath, the solution was filtered to obtain a GelMA solution. The ratio of GelMA prepolymer to PBS was 0.25-1 g: 5 ml. The photoinitiator was Irgacure 2959, and the concentration in PBS was 0.5% (w / v). The temperature of the warm bath was 40° C., and the filtration was performed using a membrane with a pore size of 0.22 μm.

[0026] (3) A549 cells were resuspended in GelMA solution to obtain a cell-GelMA solution, wherein the cell density was 10 6 / ml; then drop the cell-GelMA solution into each well of the 96-well plate and perform photocuring under ultraviolet light to form a cell-GelMA hydrogel, which is recorded as A549 / GelMA; the amount of cell-GelMA solution is 100μL, and the ultraviolet light conditions are: 405nm, 50mW / cm 2 , the light curing time is 30s;

[0027] Step 2: Establishment of LPS-induced A549 cell inflammation model

[0028] First, LPS solution was prepared with DMEM culture medium, with concentrations of 0 to 20 μg / ml; then, an equal volume of LPS solution was added to A549 / GelMA for induction treatment; after the induction treatment, the supernatant was taken as the test solution, and the cell electrochemical sensor was used for testing, and the DPV method was used to measure the change of the current signal, and the corresponding current value was recorded as ILPS; the induction treatment time was 24 hours;

[0029] Step 3: Capsaicin anti-inflammatory activity assay

[0030] (1) First, a capsaicin solution was prepared with DMEM medium at a concentration of 0 to 180 μM; then, 100 μL of the capsaicin solution was added to the A549 / GelMA prepared in step 1 for incubation, wherein the incubation temperature and gas environment were 37° C. and 5% CO 2 , respectively; after incubation, 100 μL of LPS solution (at a concentration of 1 μg / ml) was added for induction for 24 h;

[0031] After the induction, the supernatant was taken as the test solution and tested using a cell electrochemical sensor. The DPV method was used to detect the peak current value of NO released by the cells, and the current value was recorded as ICAP.

[0032] The relative anti-inflammatory ability of capsaicin was evaluated by formula (I):

[0033]

[0034] Where: ILPS is the peak current value of DPV induced by LPS;

[0035] ICAP is the DPV current value of A549 cells incubated with capsaicin and then induced by LPS;

[0036] IA is the blank control, i.e., the DPV peak current without capsaicin and LPS treatment;

[0037] △NO: The reduction in NO release.

[0038] By comparing the NO current value released by A549 cells stimulated by LPS and the NO current value released by A549 cells first intervened with capsaicin and then stimulated with LPS; then the reduction in NO release can be calculated according to formula (I);

[0039] The anti-inflammatory ability of capsaicin was evaluated by the reduction in NO release (△NO). The greater the reduction, the stronger the anti-inflammatory ability of capsaicin.

[0040] Beneficial effects of the present invention:

[0041] On the one hand, due to the unique highly networked flower-like nanostructure of Sb2O4 / rGO, NO can easily enter the reaction center and be further captured and adsorbed, and then on Sb 3+ and Sb 4+ Rapid electron transfer occurs between the two electrodes and charge transport is enhanced by reducing graphene oxide sheets.

[0042] On the other hand, a novel biocompatible composite membrane of water-insoluble surfactant didodecyldimethylammonium bromide (DDAB) and hydrophobic room temperature ionic liquid (RTIL) 1-hexyl-3-methylimidazolium hexafluorophosphate (HIMIMPF6) as electrode coating was able to eliminate NO.2- , ascorbic acid and uric acid, thus improving the detection effect of the biosensor on NO gas.

[0043] In summary, the present invention provides a cell electrochemical sensor that can not only be used to evaluate the anti-inflammatory effect of capsaicin, but also has the outstanding advantages of low cost, fast detection, simple operation and miniaturization. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 This is the SEM image of GelMA prepolymer.

[0045] Figure 2 This is the H NMR spectrum of GelMA.

[0046] Figure 3 It is the standard curve of sodium nitrite solution with different concentrations at 540nm. DETAILED DESCRIPTION

[0047] The present invention will be described in more detail with the help of the following embodiments; the following embodiments are merely illustrative and the present invention is not limited to these embodiments.

[0048] Embodiment 1:

[0049] Step 1: Synthesis of Sb2O4 / rGO

[0050] 0.5 mol of antimony trichloride was dissolved in 15 ml of ethanol to provide antimony element, and liquid ① was obtained; 15 ml of 1 mg / ml graphene oxide suspension was added to liquid ①, and mixed, and ultrasonic treatment was performed for 30 minutes to obtain liquid ②.

[0051] 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; the black precipitate generated after the heating reaction was collected and then soaked in ultrapure water overnight to remove impurities, the precipitate was collected after filtration, and the product obtained after freeze-drying was the final product Sb2O4 / rGO.

[0052] Step 2: Synthesis of DDAB-HIMIMPF6

[0053] 1 ml of didodecyl dimethyl ammonium bromide solution (0.1 M) was mixed with 200 μL of 1-hexyl-3-methylimidazolium hexafluorophosphate, and the resulting mixed material was recorded as DDAB-HIMIMPF6 composite material (it would become solid when left at room temperature, so if it was in a solid state, it was necessary to heat the composite material to become liquid before use);

[0054] Step 3: Preparation of modified nitric oxide electrochemical sensor

[0055] Weigh 1 mg of Sb2O4 / rGO and dissolve it in 1 ml of pure water and stir evenly to obtain a mixed solution; take 5 μL of the mixed solution and drop it on the screen-printed electrode, blow dry with nitrogen gas, and obtain the treated screen-printed electrode;

[0056] The DDAB-HIMIMPF6 composite material was then heated in a 40°C water bath to melt into a liquid state, and then 5 μL was aspirated and dropped onto the screen-printed electrode treated in step 1 and dried at room temperature to obtain a cell electrochemical sensor.

[0057] Embodiment 2:

[0058] The use of cell-based electrochemical sensors to evaluate the anti-inflammatory effect of capsaicin is as follows:

[0059] Step 1: Preparation of cell-methacrylated gelatin hydrogel (GelMA)

[0060] (1) Dissolve 5 g of gelatin in 50 ml of phosphate buffered saline and stir gently. Then add 4 ml of methacrylate anhydride dropwise to the gelatin solution and stir at 50 °C for 2 h. Dilute the solution with 50 °C phosphate buffered saline, dilute 1-fold each time, and dilute 5 times to terminate the reaction to obtain a reaction solution; dialyze the reaction solution obtained in step 1 at 50 °C for 7 days to remove low molecular weight impurities (molecular weight cutoff: 0–22 kDa) to obtain a dialysate; then pass the dialysate through a 0.22 μm membrane filter, pre-freeze at -80 °C, and then freeze for 5 days to obtain a GelMA prepolymer.

[0061] Figure 1 This is a field emission scanning electron microscope image of GelMA. Its structure is loose and porous, which is conducive to wrapping cells and transmitting NO produced by cells. Figure 2 : is the hydrogen nuclear magnetic resonance spectrum of GelMA. It can be seen from the attached figure that GelMA is synthesized from gelatin. After calculation, it is found that the synthesized GelMA precursor has a substitution degree of up to 76%.

[0062] (2) Take 0.25 g of GelMA prepolymer and put it in a brown bottle. Add 5 ml of 0.5% (w / v) photoinitiator Irgacure 2959 in PBS, dissolve it in a warm bath at 40°C, and filter it with a 0.22 μm membrane filter while it is hot to sterilize it.

[0063] (3) A549 cells were resuspended in GelMA solution to obtain a cell-GelMA solution, wherein the cell density was 10 6 Then, 100 μL of cell-GelMA solution was dripped into each well of a 96-well plate and illuminated under ultraviolet light (405 nm, 50 m W / cm 2 ) for 30 s to form cell-GelMA hydrogel, denoted as A549 / GelMA;

[0064] Step 2: Establishment of LPS-induced A549 cell inflammation model

[0065] To the A549 / GelMA obtained in step 1, 100 μL 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 LPS solutions prepared with DMEM medium were added and induced for 24 h.

[0066] After the induction treatment, the supernatant was taken as the test solution and tested using a cell electrochemical sensor. The DPV method was used to measure the change in current signal, and the corresponding current value was recorded as ILPS.

[0067] Step 3: Verification (NO kit assay)

[0068] (1) The NO production level was measured using a NO assay kit based on the Griess reaction.

[0069] RAW264.7 cells (100 μL, 10 5 / ml) were induced with 0, 0.4μg / ml, 0.8μg / ml, 1μg / ml, 2μg / ml, 4μg / ml, 5μg / ml, 10μg / ml, and 20μg / ml LPS solution in 96-well plates for 24h.

[0070] Then, 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. The plate was incubated for 15 min and the absorbance at 540 nm was measured on an ELISA reader.

[0071] (2) Using the same method to measure the NaNO2 standard solution, the generated standard curve is as follows: Figure 3 , based on the standard curve, the detection of nitric oxide can be achieved and compared with the electrochemical results of this embodiment, thereby determining the reliability of this method.

[0072] Step 4: Capsaicin anti-inflammatory activity assay

[0073] First, prepare a capsaicin solution with a concentration of 0 to 180 μM using DMEM medium; then add 100 μL of the capsaicin solution to the A549 / GelMA prepared in step 1 for incubation, wherein the incubation temperature and gas environment are 37°C and 5% CO2, respectively; after incubation, add 100 μL of LPS solution (with a concentration of 1 μg / ml) for induction for 24 hours;

[0074] After the induction, the supernatant was taken as the test solution and tested using a cell electrochemical sensor. The DPV method was used to detect the peak current value of NO released by the cells, and the current value was recorded as ICAP.

[0075] The relative anti-inflammatory ability of capsaicin was evaluated by formula (I):

[0076]

[0077] Where: ILPS is the peak current value of DPV induced by LPS;

[0078] ICAP is the DPV current value of A549 cells incubated with capsaicin and then induced by LPS;

[0079] IA is the DPV peak current of the blank control group without capsaicin and LPS treatment;

[0080] △NO: The reduction in NO release.

[0081] By comparing the NO current value released by A549 cells stimulated by LPS and the NO current value released by A549 cells first intervened with capsaicin and then stimulated with LPS; then the reduction in NO release can be calculated according to formula (I);

[0082] The anti-inflammatory ability of capsaicin is evaluated by the reduction in its NO release. The greater the reduction, the stronger the anti-inflammatory ability of capsaicin.

[0083] 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 this specification has described the present invention in detail with reference to the above embodiments, a person skilled in the art should understand that the present invention can still be modified or replaced by equivalents; and all technical solutions and improvements thereof that do not depart from the spirit and scope of the present invention should be included in the scope of the claims of the present invention.

Claims

1. A method for preparing a cell electrochemical sensor, characterized in that: The following steps are involved: Step 1: Preparation of flower-like nanostructured Sb2O4 / rGO; (1) dissolving antimony trichloride in ethanol to obtain an antimony trichloride ethanol solution; then adding a graphene oxide suspension to the antimony trichloride ethanol solution, mixing, and subjecting to ultrasonic treatment to obtain a mixed solution; (2) sealing the mixed solution obtained in step (1) in a reactor for heating reaction, collecting the generated black precipitate after the heating reaction, and then soaking it in ultrapure water overnight, and finally collecting the precipitate again by filtration, and freeze-drying the precipitated product to obtain the final product, namely Sb2O4 / rGO; Step 2: Preparation of DDAB-HIMIMPF6 composite material; The didodecyl dimethyl ammonium bromide solution was mixed with 1-hexyl-3-methyl imidazolium hexafluorophosphate, and the obtained mixed material was recorded as DDAB-HIMIMPF6 composite material; Step 3: preparing an electrochemical sensor; (1) Weighing the Sb2O4 / rGO prepared in step 1, dissolving it in pure water and stirring it evenly to obtain a mixed solution; then dropping the mixed solution onto the screen-printed electrode, blowing it dry with nitrogen gas, and obtaining a treated screen-printed electrode; (2) The DDAB-HIMIMPF6 composite material prepared in step 2 is heated in a water bath to form a liquid state, and then dripped onto the screen-printed electrode treated in step 1 and dried at room temperature to obtain a cell electrochemical sensor.

2. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that: In step (1) of the present invention, the dosage of antimony trichloride, ethanol and graphene oxide suspension is in the ratio of 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.

3. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that: The heating reaction temperature in step 2 (2) is 180° C. and the reaction time is 21 h.

4. The method for preparing a cell electrochemical sensor according to claim 1, characterized in that: In the step 2, the volume ratio of the didodecyl dimethyl ammonium bromide solution to 1-hexyl-3-methyl imidazolium hexafluorophosphate is 5:1, wherein the concentration of the didodecyl 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 3 (1), the dosage of Sb2O4 / rGO and 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 3 (2), the water bath heating temperature is 20-40° C., and the amount added dropwise is 5 μL.

7. Use of the cell electrochemical sensor prepared according to any one of claims 1 to 6 for evaluating the anti-inflammatory effect of capsaicin.

8. The use according to claim 7, characterized in that Here are the steps: Step 1: Preparation of cell-methacrylated gelatin hydrogel; (1) Dissolve gelatin in phosphate buffer, stir and then add methacrylate anhydride dropwise, stir evenly to obtain a mixed solution; then dilute the mixed solution 3-5 times with phosphate buffer to terminate the reaction and obtain a reaction solution; dialyze the reaction solution obtained in step 1 to obtain a dialysate; then filter the dialysate, and pre-freeze the obtained filtrate to obtain a GelMA prepolymer; wherein the amount of gelatin, phosphate buffer, and methacrylate anhydride is 5g:50ml:4-6ml; the stirring temperature is 50°C, when diluting with phosphate buffer, the temperature of the phosphate buffer is 50°C, and the dilution is 1 times each time; the dialysis temperature is 50°C, and the time is 7-14d; the filtration is performed using a membrane with a pore size of 0.22μm; the pre-freezing temperature is -20°C or -80°C, and the pre-freezing time is 5-10d. (2) Weighing GelMA prepolymer into a brown bottle, then adding PBS containing photoinitiator Irgacure 2959, dissolving in a warm bath, and then filtering to obtain a GelMA solution; wherein the amount ratio of GelMA prepolymer to PBS is 0.25-1g:5ml; the concentration of photoinitiator Irgacure 2959 in PBS is 0.5% (w / v); the temperature of the warm bath is 40°C, and filtration is performed using a membrane with a pore size of 0.22μm; (3) A549 cells were resuspended in GelMA solution to obtain a cell-GelMA solution, wherein the cell density was 10 6 / ml; then drop the cell-GelMA solution into each well of the 96-well plate and perform photocuring under ultraviolet light to form a cell-GelMA hydrogel, which is recorded as A549 / GelMA; the amount of cell-GelMA solution is 100 μL, and the ultraviolet light conditions are: 405 nm, 50 m W / cm 2 , the light curing time is 30s; Step 2: Establishment of LPS-induced A549 cell inflammation model First, LPS solution was prepared with DMEM culture medium, with concentrations of 0 to 20 μg / ml; then, an equal volume of LPS solution was added to A549 / GelMA for induction treatment; after the induction treatment, the supernatant was taken as the test solution, and the cell electrochemical sensor was used for testing, and the DPV method was used to measure the change of the current signal, and the corresponding current value was recorded as ILPS; the induction treatment time was 24 hours; Step 3: Capsaicin anti-inflammatory activity detection (1) First, a capsaicin solution was prepared with DMEM medium at a concentration of 0 to 180 μM; then, 100 μL of the capsaicin solution was added to the A549 / GelMA prepared in step 1 for incubation, wherein the incubation temperature and gas environment were 37° C. and 5% CO 2 , respectively; after incubation, 100 μL of LPS solution (at a concentration of 1 μg / ml) was added for induction for 24 h; After the induction, the supernatant was taken as the test solution and tested using a cell electrochemical sensor. The DPV method was used to detect the peak current value of NO released by the cells, and the current value was recorded as ICAP. The relative anti-inflammatory ability of capsaicin was evaluated by formula (I): Where: ILPS is the peak current value of DPV induced by LPS; ICAP is the DPV current value of A549 cells incubated with capsaicin and then induced by LPS; IA is the blank control, i.e., the DPV peak current without capsaicin and LPS treatment; △NO: reduction in NO release; By comparing the NO current value released by A549 cells stimulated by LPS and the NO current value released by A549 cells first intervened with capsaicin and then stimulated with LPS; then the reduction in NO release can be calculated according to formula (I); The △NO value is used to evaluate the anti-inflammatory ability of capsaicin. The larger the △NO value, the stronger the anti-inflammatory ability of capsaicin.

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