A Colorimetric Detection Method for Foodborne Pathogens Based on Fe3O4@MIL-100 (Fe) Magnetic Separation and Nanozyme Catalysis

Abstract

The application discloses a colorimetric detection method for foodborne pathogenic bacteria based on Fe3O4@MIL-100(Fe) magnetic separation and nanocatalysis, which mainly comprises the following steps: preparing Fe3O4@MIL-100(Fe) material through a solvothermal method and an in-situ growth method, and modifying the material with an aptamer to obtain Fe3O4@MIL-100(Fe)@aptamer, so as to realize the capture and magnetic separation of foodborne pathogenic bacteria. The nanocatalytic activity of the Fe3O4@MIL-100(Fe)@aptamer is utilized, TMB is used as a substrate to catalyze a product, and foodborne pathogenic bacteria can be rapidly detected. The method disclosed by the application has the advantages of high detection sensitivity, strong specificity, short time consumption and the like, and is suitable for rapid detection of foodborne pathogenic bacteria in food samples.

Description

Technical Field

[0001] This invention belongs to the field of food safety technology, and relates to nanotechnology and biotechnology. Specifically, it relates to a colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100 (Fe) magnetic separation and nanozyme catalysis, which utilizes the magnetic separation characteristics of magnetic MOFs and the principle of nanozyme activity to detect foodborne pathogens with high sensitivity. Background Technology

[0002] Food is the material basis for human survival, and food safety is a major concern for the international community. Among these issues, diseases caused by foodborne pathogens have become the most serious problem in the field of food safety, attracting widespread attention from researchers. Therefore, developing accurate, efficient, and rapid methods for detecting foodborne pathogens in food samples is crucial. However, due to the complexity of food matrices, effectively separating, enriching, and accurately detecting Listeria monocytogenes is quite challenging. In recent years, magnetic materials have developed rapidly and have been widely applied in fields such as biology and medicine. Among them, magnetic Fe3O4 particles possess excellent superparamagnetism and can be used for the separation and enrichment of target substances; metal-organic frameworks (MOFs) are porous coordination polymers composed of metal ion clusters and organic ligands. Compared with traditional porous materials, MOFs have high volumetric adsorption capacity, large specific surface area, effective loading capacity, and excellent catalytic properties, and have been studied and applied in adsorption, separation, catalysis, and drug delivery.

[0003] Immunomagnetic separation technology utilizes the magnetic responsiveness and biocompatibility of magnetic materials to achieve precise and rapid separation and enrichment of pathogenic bacteria, and has been extensively studied in the pretreatment of complex food samples. Meanwhile, biosensor technology, with its advantages of short detection time and high sensitivity, is widely used in the field of food safety testing. Therefore, this invention combines magnetic separation technology and biosensor technology to construct a rapid detection method for Listeria monocytogenes based on Fe3O4@MOFs magnetic separation, providing a new option for purifying food matrices and enriching trace targets, and offering a new direction for developing new technologies for high-sensitivity and high-efficiency food safety testing. In view of this, the present invention is proposed. Summary of the Invention

[0004] The purpose of this invention is to overcome the defects of the prior art and provide a colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100 (Fe) magnetic separation and nanozyme catalysis. This detection method has the characteristics of high detection sensitivity and strong specificity.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] A colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis includes the following steps:

[0007] a) Fe3O4@MIL-100(Fe) was washed with ultrapure water and resuspended in ultrapure water. After adding the aptamer, it was shaken at room temperature for a period of time. After washing with ultrapure water, the supernatant was removed by magnetic separation to obtain Fe3O4@MIL-100(Fe)@aptamer as a catalytic probe, which was stored at 4℃.

[0008] b) The catalytic probe Fe3O4@MIL-100(Fe)@aptamer prepared in step a) was added to bovine serum albumin solution, blocked at room temperature for a period of time, washed with ultrapure water, and magnetically separated to remove the supernatant to obtain Fe3O4@MIL-100(Fe)@aptamer@BSA as a capture probe, which was stored at 4℃.

[0009] c) Foodborne pathogens of different concentration gradients were reacted with the capture probe Fe3O4@MIL-100(Fe)@aptamer@BSA prepared in step b) at room temperature for 30-50 min. The supernatant was discarded by magnetic separation, and the mixture was washed with ultrapure water. The catalytic probe Fe3O4@MIL-100(Fe)@aptamer prepared in step a) was then added and coupled at room temperature for 30-50 min. The supernatant was discarded by magnetic separation and resuspended in ultrapure water. TMB and dilute hydrochloric acid were then added and mixed. The mixture was reacted at room temperature for 5-15 min. The UV absorption intensity of the supernatant was then detected by magnetic separation. A standard curve was plotted with UV absorption intensity as the ordinate and foodborne pathogen concentration as the abscissa.

[0010] d) The sample to be tested and the capture probe magnetic Fe3O4@MIL-100(Fe)@aptamer@BSA prepared in step b) are reacted at room temperature for 30-50 min. After magnetic separation, the supernatant is discarded, and the sample is washed with ultrapure water. Then, the catalytic probe Fe3O4@MIL-100(Fe)@aptamer prepared in step a) is added and coupled at room temperature for 30-50 min. After magnetic separation, the supernatant is discarded and resuspended in ultrapure water. TMB and dilute hydrochloric acid are added and mixed. The mixture is reacted at room temperature for 5-15 min. After magnetic separation, the UV absorption intensity of the supernatant is detected. The bacterial concentration in the sample is calculated by substituting the results into the standard curve.

[0011] Further, in step a), the preparation method of Fe3O4@MIL-100(Fe) is as follows: 1,3,5-benzenetricarboxylic acid is added to the aqueous dispersion of Fe3O4 and mixed evenly. The mixture is sonicated for 5-15 min, transferred to a reaction vessel, and reacted at 110-140℃ for 12-18 h. After cooling to room temperature, the mixture is washed and dried to obtain Fe3O4@MIL-100(Fe) powder.

[0012] Furthermore, 0.30-0.50 g of 1,3,5-benzenetricarboxylic acid was added to an aqueous dispersion containing 0.2 g of Fe3O4.

[0013] Furthermore, the Fe3O4 is obtained through the following treatment: ethylene glycol is mixed evenly with ferric chloride hexahydrate, anhydrous sodium acetate, and sodium citrate dihydrate, and then heated at 180-220℃ for 10-14 hours. After cooling to room temperature, it is washed and dried to obtain the final product. Specifically, 8-10g of ferric chloride hexahydrate, 14-15g of anhydrous sodium acetate, and 3-4g of sodium citrate dihydrate are added to 220-260mL of ethylene glycol. The Fe3O4 prepared by this method has carboxyl groups on its surface, which is more conducive to the growth of organic ligands on its surface.

[0014] Furthermore, in step a), 400 μL of a 2 μM aptamer is added to 0.2 mg Fe3O4@MIL-100 (Fe), and the mixture is shaken at room temperature for 50-70 min. The aptamer is a Listeria monocytogenes aptamer.

[0015] Further, in step b), 0.2 mg Fe3O4@MIL-100(Fe)@aptamer is added to 400 μL of 2% (W / V) bovine serum albumin solution and blocked at room temperature for 1.5-2.5 h.

[0016] Further, in step c), the foodborne pathogens are Listeria monocytogenes, Escherichia coli O157:H7, Salmonella typhimurium, Staphylococcus aureus, Vibrio parahaemolyticus, or Pseudomonas aeruginosa.

[0017] Furthermore, in step c), the volume of the foodborne pathogenic bacteria in different concentration gradients is 1 mL, and the dosage of Fe3O4@MIL-100(Fe)@aptamer@BSA and Fe3O4@MIL-100(Fe)@aptamer is 0.2 mg.

[0018] Furthermore, in step d), the sample to be tested is a pure culture of bacteria or a food sample containing bacteria. The volume of the sample to be tested is 1 mL, and the dosage of Fe3O4@MIL-100(Fe)@aptamer@BSA and Fe3O4@MIL-100(Fe)@aptamer is 0.2 mg.

[0019] This invention first synthesizes a catalytic probe Fe3O4@MIL-100(Fe)@aptamer and a capture probe Fe3O4@MIL-100(Fe)@aptamer@BSA. The capture probe is used for the isolation and enrichment of target bacteria. Then, utilizing the peroxidase-like activity of the catalytic probe and its catalytic color-changing mechanism for TMB, a highly sensitive method for detecting foodborne pathogens is provided. The main steps include: firstly, synthesizing Fe3O4 via a solvothermal method, then growing MIL-100(Fe) in situ on its surface to obtain Fe3O4@MIL-100(Fe), and then attaching an aptamer to its surface via adsorption to obtain Fe3O4@MIL-100(Fe)@aptamer, which is used for specific recognition of target bacteria and catalysis of TMB color change; then blocking with bovine serum albumin solution to specifically capture target bacteria in food samples; this method utilizes the dual recognition of target bacteria by Fe3O4@MIL-100(Fe)@aptamer and blocked Fe3O4@MIL-100(Fe)@aptamer, and utilizes the presence of target bacteria to cover and block the active sites of Fe3O4@MIL-100(Fe)@aptamer, thereby inhibiting the catalytic effect of Fe3O4@MIL-100(Fe)@aptamer on TMB, resulting in a decrease in ultraviolet absorption intensity to achieve detection.

[0020] The detection method of this invention involves adding target bacteria to cover and block the active sites of the Fe3O4@MIL-100(Fe)@aptamer, thereby inhibiting its catalytic effect on TMB and resulting in a decrease in UV absorption intensity. As the concentration of target bacteria increases, more active sites are covered, and the UV absorption intensity decreases accordingly. This allows the detection results to exhibit a good linear relationship within a certain range, providing a basis for quantitative analysis. This detection method has high sensitivity, with a detection range of 1.4 × 10⁻⁶ for pure cultures. 2 CFU / mL -1.4×10 7 There is a good linear relationship between CFU / mL; it also has the characteristics of high detection sensitivity and strong specificity.

[0021] Compared with other existing traditional detection methods, the present invention has the following significant features and beneficial effects:

[0022] 1) The materials and reagents used in the test are stable, simple, and easy and quick to operate;

[0023] 2) This invention achieves good specificity of detection results through the specific recognition effect of aptamers on target bacteria;

[0024] 3) This invention uses the ultraviolet absorption intensity reduction effect for detection, and the ultraviolet absorption intensity has a good linear relationship, which enables quantitative detection;

[0025] 4) This invention offers rapid and highly sensitive detection, with a detection time within 90 minutes. The detection limits for pure culture bacteria and bacteria-containing food samples (chicken breast and milk) are 14 CFU / mL and 1.7 × 10⁻⁶ CFU / mL, respectively. 2 CFU / mL, 2.1×10 2 CFU / mL. Attached Figure Description

[0026] Figure 1 FE-TEM images of Fe3O4(a) and Fe3O4@MIL-100(Fe)(b) prepared for this invention at different magnifications;

[0027] Figure 2 XRD patterns of Fe3O4 and Fe3O4@MIL-100(Fe) prepared in this invention;

[0028] Figure 3 FT-IR spectra of Fe3O4 and Fe3O4@MIL-100(Fe) prepared in this invention;

[0029] Figure 4 VSM hysteresis curves of Fe3O4 and Fe3O4@MIL-100(Fe) prepared for this invention;

[0030] Figure 5 TG curves of Fe3O4 and Fe3O4@MIL-100(Fe) prepared in this invention;

[0031] Figure 6 Zeta potential diagrams of Fe3O4, Fe3O4@MIL-100(Fe) and Fe3O4@MIL-100(Fe)@aptamer prepared in this invention;

[0032] Figure 7 The images show the ultraviolet absorption spectra (a) of Listeria monocytogenes at different concentrations of pure culture, the standard curve (b) of ultraviolet absorption intensity versus Listeria monocytogenes concentration, and the visualization results (c) of Listeria monocytogenes at different concentrations of pure culture.

[0033] Figure 8 The images show the visualization results (a) and corresponding UV absorption spectra (b) of chicken breast samples containing different concentrations of Listeria monocytogenes, and the visualization results (c) and UV absorption spectra (d) of milk samples containing different concentrations of Listeria monocytogenes.

[0034] Figure 9 This diagram illustrates the detection specificity of Listeria monocytogenes in this invention. Detailed Implementation

[0035] The present invention will be further described below with reference to specific embodiments. It should be understood that the following embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Those skilled in the art can make some non-essential improvements and adjustments based on the above-described invention.

[0036] Example 1

[0037] A method for preparing Fe3O4@MIL-100(Fe)@aptamer specifically includes the following steps:

[0038] 1) Dissolve 9.73 g of ferric chloride hexahydrate, 14.40 g of anhydrous sodium acetate, and 3.53 g of sodium citrate separately in 80 mL of ethylene glycol and stir magnetically for 15 min. Then, mix the three solutions together and sonicate for 30 min. Transfer the resulting solution to a 300 mL Teflon-lined reactor and heat at 200 °C for 12 h. After cooling to room temperature, wash the black Fe3O4 product four times with methanol and dry at room temperature for 24 h to obtain Fe3O4 powder.

[0039] 2) Take 0.2g of the Fe3O4 powder obtained in step 1) above and 0.4g of H3BTC, dissolve them separately in 30mL of ultrapure water, and sonicate for 10min; then, mix the two solutions together and sonicate for 10min. Transfer the resulting solution to a 300mL Teflon-lined reactor and heat at 125℃ for 15h. After cooling to room temperature, wash the brown product several times with anhydrous ethanol and ultrapure water, and dry at 60℃ for 24h to obtain Fe3O4@MIL-100(Fe) powder;

[0040] 3) Take 0.2 mg of the Fe3O4@MIL-100(Fe) powder prepared in step 2) above, wash twice with ultrapure water, and magnetically separate to remove the supernatant. Add 400 μL of 2 μM Listeria monocytogenes aptamer (sequence 5'-TACTATCGCGGAGACAGCGCGGGAGGCACCGGGGA-3'). Add 290 μL of ultrapure water to 100 OD of aptamer powder to obtain a 100 μM aptamer solution, and then dilute with ultrapure water to a concentration of 2 μM. Gently shake at room temperature for 1 h, magnetically separate and discard the supernatant to obtain Fe3O4@MIL-100(Fe)@aptamer, and store at 4℃ for later use.

[0041] 4) Add 400 μL of 2% (W / V) bovine serum albumin solution to 0.2 mg of Fe3O4@MIL-100(Fe)@aptamer prepared in step 3) and block at room temperature for 2 h. Then wash twice with ultrapure water and remove the supernatant by magnetic separation to obtain Fe3O4@MIL-100(Fe)@aptamer@BSA. Store at 4℃ for later use.

[0042] Figure 1 The images show FE-TEM images of Fe3O4 (Fig. a) and Fe3O4@MIL-100(Fe) (Fig. b) prepared in Example 1 at different magnifications. The FE-TEM images show that the Fe3O4@MIL-100(Fe) prepared in this invention has a uniform distribution and morphology, and is a core-shell structure with Fe3O4 as the core and MIL-100(Fe) wrapped around it.

[0043] Figure 2 The XRD patterns of Fe3O4 and Fe3O4@MIL-100(Fe) prepared in Example 1 are shown. The XRD patterns show the characteristic peaks of Fe3O4@MIL-100(Fe), and the corresponding lattice planes are (428)(088)(4814)(6618)(9321) of MIL-100(Fe) and (220)(311)(400)(511)(440) of Fe3O4. The XRD patterns show that the Fe3O4@MIL-100(Fe) prepared in this invention is reliable.

[0044] Figure 3 The FT-IR spectra of Fe3O4 and Fe3O4@MIL-100(Fe) prepared in Example 1 are shown below. The functional groups of Fe3O4 mainly consist of the following two parts: 580 cm⁻¹ -1 The corresponding Fe-O stretching and 1640-1390cm -1 The corresponding carboxyl group. Compared to Fe3O4, Fe3O4@MIL-100(Fe) at 1571 cm⁻¹ -1 and 1447cm -1 The characteristic peaks belong to the stretching vibrations of aromatic rings, indicating the growth of H3BTC on the Fe3O4 surface. Based on the above analysis, the Fe3O4@MIL-100(Fe) prepared in this invention is reliable.

[0045] Figure 4 The VSM hysteresis curves of Fe3O4 and Fe3O4@MIL-100(Fe) prepared in Example 1 are shown. The VSM hysteresis curves show that the saturation magnetization of Fe3O4 and Fe3O4@MIL-100(Fe) is 59.7 emu. -1 and 16.8emu.g -1 Although the magnetization of Fe3O4@MIL-100(Fe) is weaker than that of Fe3O4, it still has strong magnetism and can meet the requirements for separation.

[0046] Figure 5The TG thermogravimetric diagrams for Fe3O4 and Fe3O4@MIL-100(Fe) prepared in Example 1 are shown. Under high temperature conditions, the solvent molecules and sodium citrate on the Fe3O4 surface are evaporated and decomposed successively, resulting in a weight loss of 14.32%. From 30 to 800 °C, the total mass loss of Fe3O4@MIL-100(Fe) is approximately 59.25%, which is mainly attributed to the decomposition of the MIL-100(Fe) framework and sodium citrate.

[0047] Figure 6 The Zeta potential diagrams of Fe3O4 and Fe3O4@MIL-100(Fe) and Fe3O4@MIL-100(Fe)@aptor prepared for this invention are shown. The Zeta potential diagrams show that the potentials of Fe3O4 and Fe3O4@MIL-100(Fe) and Fe3O4@MIL-100(Fe)@aptor are -41.6mV, -14.5mV and -25.8mV, respectively. The potential changes indicate that the aptor has been successfully bonded to the Fe3O4@MIL-100(Fe) surface.

[0048] Example 2

[0049] A method for detecting foodborne pathogens using Fe3O4@MIL-100(Fe)@aptamer specifically includes the following steps:

[0050] (1) Listeria monocytogenes (ATCC 15313) in the logarithmic growth phase was serially diluted to 1.4 × 10⁻⁶ under aseptic conditions. 1 CFU / mL, 1.4×10 2 CFU / mL, 1.4×10 3 CFU / mL, 1.4×10 4 CFU / mL, 1.4×10 5 CFU / mL, 1.4×10 6 CFU / mL and 1.4×10 7 CFU / mL, with sterile PBS buffer as a control. 1 mL of each diluted bacterial suspension was added to 0.2 mg of Fe3O4@MIL-100(Fe)@aptamer@BSA prepared in step 4) of Example 1, and the mixture was reacted at room temperature for 40 min for capture. Three replicates were performed for each concentration. The supernatant was discarded after magnetic separation, and the mixture was washed twice with ultrapure water. 0.2 mg of Fe3O4@MIL-100(Fe)@aptamer prepared in step 3) of Example 1 was added, and the mixture was reacted at room temperature for 40 min. The supernatant was then discarded after magnetic separation, and the mixture was resuspended in 200 μL of ultrapure water. 100 μL of TMB solution (0.01 mol / L) was then added. -1 ), 200 μL dilute hydrochloric acid solution (0.2 mol L -1The reaction was carried out at room temperature in the dark for 10 min, followed by magnetic separation and detection of the ultraviolet absorption intensity (350-800 nm) of the supernatant. A standard curve was plotted with ultraviolet absorption intensity as the ordinate and bacterial concentration as the abscissa (see...). Figure 7 a);

[0051] (2) 1 mL of Listeria monocytogenes bacterial suspension was used as the test sample and mixed with Fe3O4@MIL-100(Fe)@aptamer@BSA (0.2 mg) prepared in step 4) of Example 1. The mixture was reacted at room temperature for 40 min, the supernatant was discarded by magnetic separation, and the mixture was washed twice with ultrapure water. Then, Fe3O4@MIL-100(Fe)@aptamer (0.2 mg) prepared in step 3) of Example 1 was added, and the mixture was reacted at room temperature for 40 min. The supernatant was discarded by magnetic separation, and the mixture was resuspended in 200 μL of ultrapure water. Then, 100 μL of TMB solution (0.01 mol / L) was added. -1 ), 200 μL dilute hydrochloric acid solution (0.2 mol L -1 The sample was reacted at room temperature in the dark for 10 minutes, and the supernatant was magnetically separated and the ultraviolet absorption intensity (350-800nm) was detected. The sample was then substituted into the standard curve to calculate the bacterial concentration in the sample.

[0052] Analysis of detection results: A standard curve for detecting Listeria monocytogenes was obtained by fitting the relationship between the UV absorption intensity at 452 nm at various concentrations and the bacterial suspensions of different concentrations (see [link to curve]). Figure 7 In section b), the formula is Y = 2.49674 - 0.16671X, R 2 =0.94512, with a detection limit as low as 14 CFU / mL, and the relative standard deviation of the detection results for each concentration gradient does not exceed 6%, indicating good detection specificity and that the relative standard deviation of the detection results for various types of pathogenic bacteria does not exceed 5%, demonstrating that the detection method of this invention has good repeatability and reliability. Compared with this method, the traditional plate count method is time-consuming, requiring 1-2 days, and cannot detect bacteria in an unculturable state.

[0053] Figure 7 The images show the UV absorption spectra (a) and the standard curve (b) of UV absorption intensity versus Listeria monocytogenes concentration for detecting different concentrations of pure cultured Listeria monocytogenes in this invention. The standard curve for detecting Listeria monocytogenes was obtained by fitting the relationship between the UV absorption intensity at 452 nm and the concentration of different bacterial solutions. Figure 7 The standard curve equation is shown to be Y = 2.49674 - 0.16671X, R. 2 =0.94512, with a detection limit as low as 14 CFU / mL, and the relative standard deviation of the detection results for each concentration gradient does not exceed 6%.

[0054] Figure 8 The images show the visualization results (a) and corresponding UV absorption spectra (b) of chicken breast samples containing different concentrations of Listeria monocytogenes, and the visualization results (c) and UV absorption spectra (d) of milk samples containing different concentrations of Listeria monocytogenes. The processing of the chicken breast samples followed GB 4789.30-2016, specifically: different concentrations of Listeria monocytogenes were uniformly inoculated onto the surface of 25g of chicken breast. The treated chicken breast was then placed in 225mL of PBS, and a bacterial suspension was obtained by tapping and shaking to obtain a final concentration of 1.7 × 10⁻⁶. 1 CFU / mL, 1.7×10 2 CFU / mL, 1.7×10 3 CFU / mL, 1.7×10 4 CFU / mL, 1.7×10 5 CFU / mL, 1.7×10 6 CFU / mL, 1.7×10 7 CFU / mL and 1.7×10 8 Listeria monocytogenes sample solution from chicken breast was prepared at a final concentration of 2.1 × 10⁻⁶ CFU / mL. Milk was treated similarly to chicken breast, yielding a solution containing [unclear - likely a specific concentration]. 1 CFU / mL, 2.1×10 2 CFU / mL, 2.1×10 3 CFU / mL, 2.1×10 4 CFU / mL, 2.1×10 5 CFU / mL, 2.1×10 6 CFU / mL, 2.1×10 7 CFU / mL and 2.1×10 8 Listeria monocytogenes milk sample solution at CFU / mL.

[0055] Specificity detection: Take a concentration of 10 7 CFU / mL of Listeria monocytogenes, Escherichia coli O157:H7, Salmonella typhimurium, Staphylococcus aureus, Pseudomonas aeruginosa, and Vibrio parahaemolyticus were detected according to step 2) above. The UV absorption intensity of each sample was compared to evaluate the detection specificity of the method of the present invention.

[0056] Figure 9 This is a diagram illustrating the detection specificity of the pure bacterial culture solution according to the present invention. Figure 9The results showed that, compared with the blank control Blank, the UV absorbance of Listeria monocytogenes was significantly lower than that of the blank control and the other five pathogenic bacteria. This indicates that the method of the present invention has good detection specificity.

[0057] The foregoing has shown and described the basic principles and main features of the present invention, as well as its advantages. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely illustrative of the principles of the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed. The scope of protection of this invention is defined by the appended claims and their equivalents.

Claims

1. A colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100 (Fe) magnetic separation and nanozyme catalysis, characterized in that, Includes the following steps: a) Fe3O4@MIL-100(Fe) was washed with ultrapure water and resuspended in ultrapure water. After adding the aptamer, it was shaken at room temperature for a period of time. After washing with ultrapure water, the supernatant was removed by magnetic separation to obtain Fe3O4@MIL-100(Fe)@aptamer as a catalytic probe, which was stored at 4℃. b) The catalytic probe Fe3O4@MIL-100(Fe)@aptamer obtained in step a) was added to bovine serum albumin solution, blocked at room temperature for a period of time, washed with ultrapure water, and magnetically separated to remove the supernatant to obtain Fe3O4@MIL-100(Fe)@aptamer@BSA as a capture probe, which was stored at 4℃. c) Foodborne pathogens of different concentration gradients were reacted with the capture probe Fe3O4@MIL-100(Fe)@aptamer@BSA prepared in step b) at room temperature for 30-50 min. The supernatant was discarded by magnetic separation, and the mixture was washed with ultrapure water. The catalytic probe Fe3O4@MIL-100(Fe)@aptamer prepared in step a) was then added and coupled at room temperature for 30-50 min. The supernatant was then discarded by magnetic separation and resuspended in ultrapure water. TMB and dilute hydrochloric acid were then added and mixed. The mixture was reacted at room temperature for 5-15 min. The UV absorption intensity of the supernatant was then detected by magnetic separation. A standard curve was plotted with UV absorption intensity as the ordinate and foodborne pathogen concentration as the abscissa. d) The sample to be tested and the capture probe magnetic Fe3O4@MIL-100(Fe)@aptamer@BSA prepared in step b) were reacted at room temperature for 30-50 min. After magnetic separation, the supernatant was discarded, and the sample was washed with ultrapure water. Then, the catalytic probe Fe3O4@MIL-100(Fe)@aptamer prepared in step a) was added and coupled at room temperature for 30-50 min. After magnetic separation, the supernatant was discarded and resuspended in ultrapure water. TMB and dilute hydrochloric acid were added and mixed. The mixture was reacted at room temperature for 5-15 min. After magnetic separation, the UV absorption intensity of the supernatant was detected. The bacterial concentration in the sample was calculated by substituting the results into the standard curve. The aptamer in step a) is the Listeria monocytogenes aptamer, with the sequence 5'-TACTATCGCGGAGACAGCGCGGGAGGCACCGGGGA-3'. In step c), the foodborne pathogen is Listeria monocytogenes.

2. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 1, characterized in that, In step a), the preparation method of Fe3O4@MIL-100(Fe) is as follows: 1,3,5-benzenetricarboxylic acid is added to the aqueous dispersion of Fe3O4 and mixed evenly. The mixture is sonicated for 5-15 min, transferred to a reaction vessel, and reacted at 110-140℃ for 12-18 h. After cooling to room temperature, the mixture is washed and dried to obtain Fe3O4@MIL-100(Fe) powder.

3. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 2, characterized in that, Add 0.30-0.50 g of 1,3,5-benzenetricarboxylic acid to an aqueous dispersion containing 0.2 g of Fe3O4.

4. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 2 or 3, characterized in that, The Fe3O4 was obtained by the following treatment: ethylene glycol was mixed with ferric chloride hexahydrate, anhydrous sodium acetate and sodium citrate dihydrate, and then heated at 180-220℃ for 10-14 h. After cooling to room temperature, it was washed and dried to obtain the Fe3O4.

5. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 4, characterized in that, Add 8-10g of ferric chloride hexahydrate, 14-15g of anhydrous sodium acetate, and 3-4g of sodium citrate dihydrate to 220-260mL of ethylene glycol.

6. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 1, characterized in that, In step a), 400 μL of a 2 μM aptamer is added to 0.2 mg Fe3O4@MIL-100(Fe), and the mixture is shaken at room temperature for 50-70 min. The aptamer is a Listeria monocytogenes aptamer.

7. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 1, characterized in that, In step b), 0.2 mg Fe3O4@MIL-100(Fe)@aptamer is added to 400 μL of 2% (W / V) bovine serum albumin solution and blocked at room temperature for 1.5-2.5 h.

8. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 1, characterized in that, In step c), the bacterial culture volume of foodborne pathogens at different concentration gradients is 1 mL, and the dosage of Fe3O4@MIL-100(Fe)@aptamer@BSA and Fe3O4@MIL-100(Fe)@aptamer is 0.2 mg.

9. The colorimetric detection method for foodborne pathogens based on Fe3O4@MIL-100(Fe) magnetic separation and nanozyme catalysis according to claim 1, characterized in that, In step d), the sample to be tested is a pure culture of bacteria or a food sample containing bacteria. The volume of the sample to be tested is 1 mL, and the amount of Fe3O4@MIL-100(Fe)@aptamer@BSA and Fe3O4@MIL-100(Fe)@aptamer is 0.2 mg.

Images