A porphyrin-based metal-organic framework nanozyme-mediated chemiluminescence optical fiber biosensor for ricin detection

Abstract

The present application belongs to the technical field of environmental pollution detection, and relates to a porphyrin-based metal-organic framework nanoenzyme-mediated chemiluminescence optical fiber biosensor for ricin detection. The biosensor of the present application realizes the detection of ricin based on nanoenzyme catalysis, indirect competition nucleic acid aptamer affinity and optical fiber biosensing principle. The porphyrin-based metal-organic framework nanoenzyme probe labeled with nucleic acid aptamer is used to catalyze the luminol / hydrogen peroxide system to generate a luminescence signal. The higher the concentration of ricin is, the more nanoenzyme probe binding sites are occupied, which leads to the reduction of nanoenzyme probes combined with the optical fiber probe of the modified complementary DNA chain, and the generated luminescence signal is also reduced. Based on the high specificity of nucleic acid aptamer, the biosensor has good selectivity and anti-interference for ricin detection. The sensor has simple structure, high sensitivity and good selectivity, and can realize the rapid detection of ricin in medical and environmental samples.

Description

Technical Field

[0001] This invention belongs to the field of environmental pollution detection technology and relates to a porphyrin-based metal-organic framework nanoenzyme-mediated chemiluminescent fiber optic biosensor for the detection of ricin. Background Technology

[0002] Ricin, isolated from castor beans, is a highly toxic plant protein composed of two polypeptide chains, A and B, linked by disulfide bonds. Ricin is extremely toxic, approximately 6000 times more toxic than cyanide, with a lethal oral dose for humans of about 5–10 μg / kg. In recent years, terrorist organizations have increasingly used it in their terrorist activities. Castor beans are widely available and highly heat-resistant; foods containing ricin remain toxic even after high-temperature processing, and ricin poisoning incidents are common. Currently, there are no antidotes, vaccines, or other specific effective treatments for ricin poisoning or prevention. Therefore, developing rapid and reliable ricin detection methods is crucial for preventing the harm caused by ricin and is an urgent need in fields such as medical diagnostics, food safety, environmental safety, and counter-terrorism.

[0003] Among various types of biosensors, chemiluminescent biosensors possess advantages such as high sensitivity, low background signal, and strong anti-interference ability, making them particularly effective for highly sensitive detection of biological toxins. Chemiluminescence is a photoluminescence phenomenon that accompanies a chemical reaction. Due to the limited quantum yield of luminescent reagents, most luminescent systems produce weak chemiluminescence intensity; therefore, enhancing luminescence intensity has always been a key focus and challenge in chemiluminescence detection technology research. Nanozymes, nanomaterials with enzyme-like catalytic activity, offer advantages such as high catalytic activity, good stability, and low cost. Based on nanozyme catalytic signal amplification strategies, nanozyme-mediated chemiluminescent biosensors with high sensitivity and detection accuracy can be designed and constructed.

[0004] Heme / G-quadruplex DNAases can be prepared by tightly binding heme to guanine-rich single-stranded DNA via intramolecular G-quadruplex binding. These DNAases can then be embedded in metal-organic frameworks (MOFs) with micro / nanoporous structures to construct porphyrin-based MOF nanozymes that catalyze persistent chemiluminescent signals. The combination of these two active components enhances the effective loading and increases the number of active sites. Simultaneously, the micro / nanoporous structure of the MOF material prolongs the interaction time between the catalytic active site and the luminescent substrate, resulting in a more stable and persistent chemiluminescent signal. Using ricin as the detection target, porphyrin-based MOF nanozymes labeled with nucleic acid aptamers serve as chemiluminescent reaction catalysts. By integrating nanozyme catalysis, indirect competitive nucleic acid aptamer affinity, and fiber optic biosensing principles, a porphyrin-based metal-organic framework nanozyme-mediated chemiluminescent fiber optic biosensor was constructed. Summary of the Invention

[0005] The purpose of this invention is to address the problems of complex operation and difficulty in rapid and sensitive detection of ricin using traditional detection techniques, and to provide a porphyrin-based metal-organic framework nanoenzyme-mediated chemiluminescent fiber optic biosensor for ricin detection.

[0006] This invention provides a real-time and rapid detection technology for ricin in the environment, and also offers theoretical and technical support for the development of on-site detection instruments for biotoxins. The invention achieves ricin detection based on nanozyme catalysis, indirect competitive nucleic acid aptamer affinity, and fiber optic biosensing principles. The porphyrin-based metal-organic framework nanozyme probe labeled with nucleic acid aptamers catalyzes the generation of a luminescent signal in a luminol / hydrogen peroxide system. Higher ricin concentrations occupy more nanozyme probe binding sites, leading to a reduction in the number of nanozyme probes binding to the fiber optic probe modified with complementary DNA strands, and consequently, a decrease in the generated luminescent signal.

[0007] This biosensor utilizes a porphyrin-based metal-organic framework nanozyme probe labeled with nucleic acid aptamers and a luminol / hydrogen peroxide substrate, employing a chemiluminescent fiber optic biosensing platform. It offers a powerful detection method for ricin in both medical and environmental fields. Based on the high specificity of nucleic acid aptamers, this biosensor exhibits excellent selectivity and interference resistance for ricin detection.

[0008] The technical solution adopted in this invention is as follows:

[0009] In a first aspect, the present invention provides a method for preparing porphyrin-based metal-organic framework nanozyme probes, specifically comprising the following steps:

[0010] S1: 8.5–34 parts by mass of zirconium chloride and 12–48 parts by mass of 5,10,15,20-tetra(4-carboxyphenyl)porphyrin ferric chloride are dissolved and dispersed uniformly in N,N-dimethylformamide; formic acid of 4–32% by volume of N,N-dimethylformamide is added to the above mixture to form a homogeneous solution; the mixture is transferred and refluxed in an oil bath at 110–130°C; after the reflux reaction is completed, the mixture is centrifuged, washed, dried, and cooled to obtain the product PCN;

[0011] S2: Disperse PCN evenly in deionized water to form a homogeneous PCN solution with a concentration of 1-4 mg / mL; add chloroplatinic acid solution with a concentration of 1-2 wt.% to the PCN solution, with a volume of 0.3-2% of the PCN solution, and stir evenly at room temperature; then add sodium borohydride solution with a concentration of 2-6 mg / mL dropwise, with a volume of 0.6-4% of the PCN solution, and continue stirring evenly at room temperature until Pt 4+ The product was completely reduced; after the reaction was complete, the product was centrifuged, washed, dried, and cooled to obtain Pt-PCN.

[0012] S3: Pt-PCN is uniformly dispersed in Tris-HCl to form a homogeneous Pt-PCN solution with a concentration of 1–4 mg / mL; 0.2–0.3% (by volume of the Pt-PCN solution) of a guanine-rich DNA solution with a concentration of 0.3–3.6 μM, wherein the guanine-rich DNA sequence is 5′-CCCATAGGGAAGTGGGGGA-3′, and 0.2–0.3% (by volume of the Pt-PCN solution) of a nucleic acid aptamer solution with a concentration of 0.3–3.6 μM, wherein the nucleic acid aptamer sequence is 5′-ACACCCACCGCAGGCAGACGCAACGCCTCGGAGACTAGCC-3′;

[0013] After the reaction is complete at 35–40℃, add 0.1–0.3% (by volume) of Pt-PCN solution in a 1.3–2.6 mol / L NaCl solution to the reaction system for aging treatment; after aging, centrifuge and dissolve the centrifuged product in a 25–50 mmol / L solution containing K. + The solution was then added to a Tris-HCl buffer solution, followed by the addition of 20–40% of the Tris-HCl buffer volume of heme solution (0.5–2 mg / mL). The reaction was continued at 35–40°C until complete, followed by centrifugation. The centrifuged product was resuspended in a solution containing K+. + In Tris-HCl buffer, porphyrin-based metal-organic framework nanozyme probes coupled with nucleic acid aptamers were obtained, which are referred to as nanozyme probes for short.

[0014] Preferably, in step S2, the specific operations of centrifugation, washing, drying and cooling are as follows: centrifuge at 10000 rpm for 10 min, wash three times alternately with anhydrous ethanol and deionized water, dry and cool at 60°C to obtain the product Pt-PCN.

[0015] Preferably, in step S3, the aged solution is centrifuged at 13000 rpm for 10 min; after adding heme solution and reacting, it is centrifuged at 7500 rpm for 10 min.

[0016] Preferably, the homogenized PCN solution in step S2 is 20-30 mL.

[0017] Furthermore, in step S3, the centrifuged product after aging is dissolved in 500 μL of solution containing K. + Tris-HCl buffer.

[0018] In a second aspect, the present invention provides a porphyrin-based metal-organic framework nanozyme-mediated chemiluminescent fiber optic biosensor for ricin detection, comprising a porphyrin-based metal-organic framework nanozyme probe 1, a fiber optic probe modified with complementary DNA strands 2, a microchannel sample cell module 3, a photomultiplier tube module 4, a control circuit and signal processing module 5, and a tablet computer 6.

[0019] The porphyrin-based metal-organic framework nanozyme probe 1 is coupled to the fiber optic probe 2 modified with complementary DNA strands. One end of the fiber optic probe 2 modified with complementary DNA strands is connected to the microchannel sample cell module 3, and the other end is connected to the photomultiplier tube module 4. The photomultiplier tube module 4, the control circuit and the signal processing module 5, and the computer 6 are connected in sequence.

[0020] The nanozyme probe 1 is coupled to the fiber optic probe 2 modified with complementary DNA strands; the sequence of the complementary DNA strand modified on the fiber optic probe 2 is as follows:

[0021] 5′-GGCTAGTCTCCGGAGGCGTTGCGTCTGCCTGCGGTGGGTGT-3′.

[0022] Preferably, the photomultiplier tube module 4 has a detection wavelength of 420nm.

[0023] Thirdly, the present invention provides a method for detecting ricin using the biosensor described in the second aspect, the specific steps of which are as follows:

[0024] Sample preparation:

[0025] Sample a: Luminol / hydrogen peroxide substrate

[0026] A mixed solution of luminol and hydrogen peroxide is prepared. This mixed solution can generate blue light with a wavelength of 420 nm under the catalysis of nanozyme probe 1. The light is finally collected by fiber optic probe 3 and detected by photomultiplier tube 5.

[0027] Sample b: Nanozyme probe solution

[0028] Prepare a solution of nanozyme probe 1 for biological samples that do not contain ricin;

[0029] Sample c: A mixed solution of nanozyme probe and ricin sample to be detected.

[0030] Prepare sample c, which is a mixed solution of nanozyme probe and ricin sample to be detected; ricin in the sample can specifically bind to nanozyme probe and occupy its active site, resulting in a reduction of nanozyme probe binding to complementary DNA strands immobilized on optical fiber. As the concentration of ricin increases, the luminescence signal catalyzed by sample a decreases significantly.

[0031] Insert fiber optic probe 2, and then pass in Tris-HCl buffer solution;

[0032] Samples b and c were then introduced into the microchannel sample cell module 3, and then Tris-HCl buffer solution was introduced.

[0033] Sample a is then passed into the microchannel sample cell module 3; the instrument detects the emission signals of the samples respectively.

[0034] The concentration of ricin in the test sample c was determined by the relationship between the concentration of ricin in the standard sample and the quantity of luminescence signal.

[0035] Preferably, the regeneration method for the tested fiber optic probe 2 is as follows: the fiber optic probe 2 is regenerated by introducing SDS solution, and then cleaned by introducing Tris-HCl buffer solution.

[0036] Preferably, the concentration of luminol solution in sample a is 1.5–4 μmol, and the volume is 40–80 μL; the concentration of hydrogen peroxide is 1.5–4 mmol, and the volume is 40–80 μL; the amount of nanozyme probe 1 solution used in sample b is 20–100 μL, and the concentration is 25–75 μg / mL; the amount of nanozyme probe 1 solution used in sample c is 20–100 μL, and the concentration is 25–75 μg / mL, and the concentration of ricin after mixing does not exceed 200 ng / mL.

[0037] Preferably, the detection range for ricin in human serum samples is 4.71–195.04 ng / mL (y = 0.401 + 0.617 / [1 + (x / 28.459) 0.908], R 2 =0.999), and the detection limit is 1.71 ng / mL.

[0038] The principle behind this method is:

[0039] A method for detecting ricin based on nanozyme catalysis, indirect competitive nucleic acid aptamer affinity, and fiber optic biosensing principles:

[0040] When ricin is absent from the detection system, the nanozyme probe binds to a fiber optic probe modified with complementary DNA strands via DNA double-strand hybridization, and then catalyzes the luminol / hydrogen peroxide system to generate a strong luminescent signal, which is detected by a photomultiplier tube.

[0041] When ricin is present in the detection system, the higher the concentration of ricin, the more nanozyme probe binding sites it occupies. This leads to a reduction in the number of nanozyme probes that bind to the fiber optic probe modified with complementary DNA strands, resulting in a decrease in the luminescent signal. The luminescent signal is then detected by a photomultiplier tube.

[0042] In one detection cycle, ricin is first mixed with a certain concentration of nanozyme probes and then the mixture is passed into a microchannel sample cell. When there is no ricin in the sample, more nanozyme probes bind to the fiber optic probe, generating a strong luminescence signal. When ricin is present in the sample, the binding sites of the nanozyme probes are occupied, reducing the number of nanozyme probes binding to the fiber optic probe and weakening the luminescence signal. The higher the ricin concentration, the lower the chemiluminescence signal value. Based on the relationship between the chemiluminescence signal intensity and the ricin concentration, ricin is detected.

[0043] The beneficial effects of this invention are: the sensor has a simple structure, high sensitivity, and good selectivity, enabling rapid detection of ricin in medical and environmental samples. This invention can be used for detection in serum samples, and the detection results are highly consistent with those obtained by liquid chromatography-mass spectrometry. Attached Figure Description

[0044] Figure 1 Schematic diagram of the structure of a porphyrin-based metal-organic framework nanozyme-mediated chemiluminescent fiber optic biosensor

[0045] In the figure: 1. Porphyrin-based metal-organic framework nanozyme probe, 2. Fiber optic probe modified with complementary DNA strands, 3. Microchannel sample cell module, 4. Photomultiplier tube module, 5. Control circuit and signal processing module, 6. Tablet PC.

[0046] Figure 2 Schematic diagram of luminescence signal generation in luminol-hydrogen peroxide system catalyzed by porphyrin-based metal-organic framework nanozymes.

[0047] In the figure: Ⅰ, 0.1 μg / mL porphyrin-based metal-organic framework nanozyme; Ⅱ, 0.5 μg / mL porphyrin-based metal-organic framework nanozyme HGP-PCN; Ⅲ, 1 μg / mL porphyrin-based metal-organic framework nanozyme.

[0048] Figure 3 Flowchart of the detection method of porphyrin-based metal-organic framework nanozyme-mediated chemiluminescent fiber optic biosensor and luminescence signal curves for detecting different concentrations of ricin.

[0049] In the diagram: A, Tris-HCl buffer is introduced; B, sample b or sample c is introduced; C, Tris-HCl buffer is introduced; D, sample a is introduced; E, SDS solution is introduced; F, Tris-HCl buffer is introduced; G, 0 ng / mL ricin; H, 1 ng / mL ricin; I, 10 ng / mL ricin; J, 60 ng / mL ricin; K, 125 ng / mL ricin; L, 500 ng / mL ricin. Detailed Implementation

[0050] Example 1

[0051] A porphyrin-based metal-organic framework nanozyme-mediated chemiluminescent fiber optic biosensor for ricin detection includes a porphyrin-based metal-organic framework nanozyme probe (1), a fiber optic probe modified with complementary DNA strands (2), a microchannel sample cell module (3), a photomultiplier tube module (4), a control circuit and signal processing module (5), and a tablet computer (6).

[0052] The porphyrin-based metal-organic framework nanozyme probe (1) is coupled to the fiber optic probe (2) modified with complementary DNA strands. One end of the fiber optic probe (2) modified with complementary DNA strands is connected to the microchannel sample cell module (3), and the other end is connected to the photomultiplier tube module (4). The photomultiplier tube module (4), the control circuit and signal processing module (5), and the computer (6) are connected in sequence.

[0053] The nanozyme probe (1) is coupled to a fiber optic probe (2) modified with complementary DNA strands; the sequence of the complementary DNA strand modified on the fiber optic probe (2) is as follows:

[0054] 5′-GGCTAGTCTCCGGAGGCGTTGCGTCTGCCTGCGGTGGGTGT-3′.

[0055] The photomultiplier tube module (4) has a detection wavelength of 420nm.

[0056] The preparation method of the porphyrin-based metal-organic framework nanozyme probe 1 is as follows:

[0057] 34 mg of zirconium chloride and 48 mg of 5,10,15,20-tetra(4-carboxyphenyl)porphyrin ferric chloride were dissolved in 30 mL of N,N-dimethylformamide and mixed thoroughly. 4.8 mL of formic acid was added to the mixture, followed by sonication until a homogeneous solution was formed. The mixture was transferred to a flask and refluxed at 120 °C in an oil bath for 24 h. After the reaction was complete, the mixture was centrifuged at 10,000 rpm for 10 min, washed three times alternately with anhydrous ethanol and N,N-dimethylformamide, dried at 60 °C, and cooled to obtain the product, abbreviated as PCN.

[0058] PCN was uniformly dispersed in deionized water to form a homogeneous solution of 4 mg / mL. Then, a 2 wt.% solution of chloroplatinic acid (100–400 μL) was added to this solution, and the mixture was stirred at room temperature for 1 h. Subsequently, a 2–6 mg / mL sodium borohydride solution (200–800 μL) was added dropwise to the mixture, and stirring was continued at room temperature for 3 h to ensure the PCN concentration was within acceptable limits. 4+The product was completely reduced. After the reaction was complete, the product was centrifuged at 10,000 rpm for 10 min, washed three times alternately with anhydrous ethanol and deionized water, dried at 60°C and cooled to obtain the product, which is referred to as Pt-PCN.

[0059] Pt-PCN was uniformly dispersed in Tris-HCl to form a homogeneous solution of 4 mg / mL. 60 μL of guanine-rich DNA solution (sequence 5′-CCCATAGGGAAGTGGGGGA-3′, concentration 0.3–3.6 μM) and 60 μL of nucleic acid aptamer solution (sequence...) were then added to this solution.

[0060] The reaction mixture (5′-ACACCCACCGCAGGCAGACGCAACGCCTCGGAGACTAGCC-3′) was reacted at 37℃ for 16 h. Then, NaCl solution (concentration 1.3–2.6 mol / L, volume 30–60 μL) was added to the reaction system, and the mixture was aged for 3 h. After centrifugation at 13000 rpm, the product was dissolved in 500 μL of a solution containing K. + The solution was added to Tris-HCl buffer (50 mmol / L), followed by the addition of heme solution (2 mg / mL, 200 μL). The reaction was continued at 37°C for 2 h, followed by centrifugation at 7500 rpm for 10 min. The product was resuspended in a solution containing K+. + In Tris-HCl buffer, porphyrin-based metal-organic framework nanozyme probes coupled with nucleic acid aptamers are obtained, which are simply referred to as nanozyme probes.

[0061] Example 2

[0062] The apparatus described in Example 1 was used to detect ricin in human serum sample c.

[0063] The specific steps are as follows:

[0064] Configure samples a, b, and c:

[0065] Sample a: Luminol / hydrogen peroxide substrate

[0066] A mixed solution of luminol and hydrogen peroxide was prepared. This mixed solution, catalyzed by nanozyme probe 1, produced blue light at a wavelength of 420 nm, which was ultimately collected by fiber optic probe 3 and detected by photomultiplier tube 5. The concentration of luminol solution was 4 μmol and the volume was 80 μL; the concentration of hydrogen peroxide was 4 mmol and the volume was 80 μL.

[0067] Sample b: Nanozyme probe solution

[0068] Prepare a 100 μL solution of nanozyme probe 1 for biological samples that do not contain ricin, at a concentration of 75 μg / mL.

[0069] Sample c: Human serum sample prepared as ricin sample

[0070] Ricin samples were prepared using human serum samples at concentrations of 0, 2, 20, 120, 250, and 1000 ng / mL. Biological samples c containing ricin were prepared, namely a mixture of nanozyme probes and ricin.

[0071] Then, fiber optic probe 2 was inserted into microchannel sample cell module 3. Tris-HCl buffer was circulated into microchannel sample cell 3 at 300 μL / min to clean fiber optic probe 2 for 40 s, while obtaining a detection baseline. Next, samples b and c were circulated into microchannel sample cell module 3, where nanozyme probe 1 bound to fiber optic probe 2 with modified complementary DNA strands via DNA double-strand hybridization. Tris-HCl buffer was then circulated to clean fiber optic probe 2. Subsequently, sample a was circulated into microchannel sample cell 3, where a chemiluminescent signal was generated under the catalysis of nanozyme probe 1. The instrument automatically detected the signal, then SDS was circulated to regenerate fiber optic probe 2, and Tris-HCl buffer was circulated again to clean fiber optic probe 2 for the next detection. Data was read from tablet computer 6. Sample a showed a high luminescence signal of 2.5–2.8 × 10⁻⁶. 6 RLU and sample b showed low luminescence signal (1.2–2.4 × 10⁻⁶). 6 RLU.

[0072] The sensor's detection range for ricin in human serum samples is 4.71–195.04 ng / mL (y = 0.401 + 0.617 / [1 + (x / 28.459) 0.908], R 2 =0.999), the detection limit was 1.71 ng / mL, and the detection results were highly consistent with those of high performance liquid chromatography-mass spectrometry.

Claims

1. A method for preparing a porphyrin-based metal-organic framework nanozyme probe, characterized in that, Specifically, the steps include the following: S1: 8.5–34 parts by mass of zirconium chloride and 12–48 parts by mass of 5,10,15,20-tetra(4-carboxyphenyl)porphyrin ferric chloride are dissolved and dispersed uniformly in N,N-dimethylformamide; formic acid of 4–32% by volume of N,N-dimethylformamide is added to the above mixture to form a homogeneous solution; the mixture is transferred and refluxed in an oil bath at 110–130°C; after the reflux reaction is completed, the mixture is centrifuged, washed, dried, and cooled to obtain the product PCN; S2: Disperse PCN evenly in deionized water to form a homogeneous PCN solution with a concentration of 1-4 mg / mL; add chloroplatinic acid solution with a concentration of 1-2 wt.% to the PCN solution, with a volume of 0.3-2% of the PCN solution, and stir evenly at room temperature; then add sodium borohydride solution with a concentration of 2-6 mg / mL dropwise, with a volume of 0.6-4% of the PCN solution, and continue stirring evenly at room temperature until Pt 4+ The product was completely reduced; after the reaction was complete, the product was centrifuged, washed, dried, and cooled to obtain Pt-PCN. S3: Pt-PCN is uniformly dispersed in Tris-HCl to form a homogeneous Pt-PCN solution with a concentration of 1–4 mg / mL; 0.2–0.3% (by volume of the Pt-PCN solution) of a 0.3–3.6 μM guanine-rich DNA solution, wherein the guanine-rich DNA sequence is 5′-CCCATAGGGAAGTGGGGGA-3′, and 0.2–0.3% (by volume of the Pt-PCN solution) of a 0.3–3.6 μM guanine-rich DNA solution, are added to this Pt-PCN solution. A 3.6 μM nucleic acid aptamer solution, wherein the nucleic acid aptamer sequence is 5′-ACACCCACCGCAGGCAGACGCAACGCCTCGGAGACTAGCC-3′, is prepared. After sufficient reaction at 35–40 °C, 0.1–0.3% (by volume) of Pt-PCN solution in a 1.3–2.6 mol / L NaCl solution is added to the reaction system for aging treatment. After aging, the solution is centrifuged, and the centrifuged product is dissolved in a 25–50 mmol / L solution containing potassium. + The solution was then added to a Tris-HCl buffer solution, followed by the addition of 20–40% of the Tris-HCl buffer volume of heme solution (0.5–2 mg / mL). The reaction was continued at 35–40°C until complete, followed by centrifugation. The centrifuged product was resuspended in a solution containing K+. + In Tris-HCl buffer, porphyrin-based metal-organic framework nanozyme probes with coupled nucleic acid aptamers were obtained.

2. The method for preparing the porphyrin-based metal-organic framework nanozyme probe according to claim 1, characterized in that, In step S2, the specific operations of centrifugation, washing, drying and cooling are as follows: centrifuge at 10,000 rpm for 10 min, wash three times alternately with anhydrous ethanol and deionized water, dry and cool at 60°C to obtain the product Pt-PCN.

3. The method for preparing the porphyrin-based metal-organic framework nanozyme probe according to claim 1, characterized in that, In step S3, the aged solution is centrifuged at 13000 rpm for 10 min; after adding heme solution and reacting, it is centrifuged at 7500 rpm for 10 min.

4. The method for preparing the porphyrin-based metal-organic framework nanozyme probe according to claim 1, characterized in that, In step S2, the homogenized PCN solution is 20-30 mL.

5. The method for preparing the porphyrin-based metal-organic framework nanozyme probe according to claim 4, characterized in that, In step S3, the centrifuged product after aging is dissolved in 500 μL of solution containing K. + Tris-HCl buffer.

6. A porphyrin-based metal-organic framework nanozyme-mediated chemiluminescent fiber optic biosensor for ricin detection, characterized in that, The invention includes a porphyrin-based metal-organic framework nanozyme probe (1), a fiber optic probe modified with complementary DNA strands (2), a microchannel sample cell module (3), a photomultiplier tube module (4), a control circuit and signal processing module (5), and a tablet computer (6) prepared by the method described in any one of claims 1 to 5. The porphyrin-based metal-organic framework nanozyme probe (1) is coupled to the fiber optic probe (2) modified with complementary DNA strands. One end of the fiber optic probe (2) modified with complementary DNA strands is connected to the microchannel sample cell module (3), and the other end is connected to the photomultiplier tube module (4). The photomultiplier tube module (4), the control circuit and signal processing module (5), and the computer (6) are connected in sequence. The nanozyme probe (1) is coupled to the fiber optic probe (2) modified with complementary DNA strand; the sequence of the complementary DNA strand modified on the fiber optic probe (2) is 5′-GGCTAGTCTCCGGAGGCGTTGCGTCTGCCTGCGGTGGGTGT-3′.

7. The porphyrin-based metal-organic framework nanozyme-mediated chemiluminescent fiber optic biosensor for ricin detection according to claim 6, characterized in that, The photomultiplier tube module (4) has a detection wavelength of 420nm.

8. A method for detecting ricin using a biosensor as described in claim 6 or 7, characterized in that, The specific steps are as follows: Sample preparation: Sample a: Luminol / hydrogen peroxide substrate Prepare a mixed solution of luminol and hydrogen peroxide; Sample b: Nanozyme probe solution Prepare a solution of biological sample b nanozyme probe (1) without ricin; Sample c: A mixed solution of nanozyme probe and ricin sample to be detected. Prepare sample c, which is a mixed solution of nanozyme probe and ricin sample to be detected; Insert the fiber optic probe (2), and then pass in the Tris-HCl buffer solution; Samples b and c were then introduced into the microchannel sample cell module (3), and then Tris-HCl buffer solution was introduced. Then, sample a is passed into the microchannel sample cell module (3); the instrument detects the luminescence signals of the above samples respectively; The concentration of ricin in the test sample c was determined by the relationship between the concentration of ricin in the standard sample and the quantity of luminescence signal.

9. The method for detecting ricin using a biosensor according to claim 8, characterized in that, The regeneration method for the tested fiber optic probe (2) is as follows: introduce SDS solution to regenerate the fiber optic probe (2), and then introduce Tris-HCl buffer solution for cleaning.

10. The method for detecting ricin using a biosensor according to claim 8, characterized in that, The concentration of luminol solution in sample a is 1.5–4 μmol, and the volume is 40–80 μL; the concentration of hydrogen peroxide is 1.5–4 mmol, and the volume is 40–80 μL; the amount of nanozyme probe (1) solution used in sample b is 20–100 μL, and the concentration is 25–75 μg / mL; the amount of nanozyme probe (1) solution used in sample c is 20–100 μL, and the concentration is 25–75 μg / mL, and the concentration of ricin after mixing does not exceed 200 ng / mL.

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