Nano composite material based on zirconium-based metal organic framework, electrochemical sensor and preparation and application of nano composite material

Abstract

The invention provides a nano composite material based on a zirconium-based metal organic framework, PEI / MCNs is prepared through a step-by-step synthesis method, AuNPs is introduced into NH2-UIO-66 to form a functionalized nano composite material, and then the AuNPs / NH2-UIO-66 is directionally loaded on the surface of the PEI / MCNs under the coordination action. The invention also provides an electrochemical sensor for detecting the LPS, the AuNPs / NH2-UIO-66 / PEI / MCNs nano composite material is modified on a glassy carbon electrode, an electrochemical sensing platform based on a three-electrode system is constructed, and through the synergistic effect of the components in the links of enrichment-recognition-signal, the problem of specific recognition of the sensor is solved, and the sensitivity of the sensor is improved. The detection sensitivity, the response speed and the stability are remarkably improved, and a reliable material support is provided for efficient detection of endotoxin in a complex system.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biosensors, in particular to a zirconium-based metal organic framework-based nanocomposite, an electrochemical sensor and preparation and application thereof. BACKGROUND

[0002] Endotoxin, as an important biological toxin, has a wide range of biological activities, from mild inflammatory response to severe systemic inflammatory response syndrome and multiple organ dysfunction syndrome, and even can cause death. Endotoxin is the main component of the cell wall of gram-negative bacteria, mainly composed of lipopolysaccharide (LPS), and lipid A is the core of its toxicity. When bacteria die or lyse, endotoxin is released into the surrounding environment, which can enter the human body through food, water or air, and trigger a series of pathological reactions. Therefore, specific and sensitive detection of endotoxin is of great significance for food safety and clinical treatment.

[0003] The traditional detection method of endotoxin is limited by the lack of sensitivity and specificity of the limulus reagent method, complex operation and time-consuming, high requirements for samples, dependence on limulus reagent, sustainability and lack of accuracy, which limits its further development. In order to adapt to the modern detection needs of endotoxin detection method, endotoxin detection method will develop towards faster, higher sensitivity and lower cost, providing more reliable protection for food safety and biological product safety.

[0004] At present, the electrochemical aptamer biosensor for detecting LPS mainly includes biosensors prepared based on LPS binding peptides, proteins, cells, antibodies, aptamers and other ligands. These sensors use different biomolecules as recognition elements, which specifically bind to LPS to convert biological recognition events into electrical signals, thereby realizing the detection of LPS. For example, the electrochemical aptamer biosensor with antibody as recognition element utilizes the specific binding of antibody to LPS antigenic determinant to occur immune reaction on the electrode surface, causing changes in the electrochemical properties of the electrode surface, and then realizing the quantitative analysis of LPS by detecting the electrical signal; the sensor with aptamer as recognition element utilizes the high affinity and specific binding between aptamer and LPS to construct an electrochemical sensing system for detection.

[0005] However, endotoxin often exists in a complex solution system in an actual environment, rather than a single solute solution. In the detection process, the existing sensor is easily affected by other interference substances in the solution. These interference substances can be non-specifically combined with the recognition element of the sensor, or interfere with the biological recognition process, thereby seriously affecting the specificity of the biosensor, causing the detection result to deviate, and false positives and false negatives frequently occur. At the same time, the presence of interference substances also destroys the stability of the sensor, so that the detection performance of the sensor decreases, such as detection sensitivity and repeatability, during multiple detections or long-term use. Considering the stability, specificity and cost required by the biosensor, some sensors also have problems such as complex synthesis process, difficult raw material acquisition and high cost, which are difficult to meet the needs of large-scale promotion and use in actual applications. SUMMARY

[0006] The present application aims at the problems of slow speed and insufficient reliability of the existing LPS detection technology, and provides a zirconium-based metal organic framework-based nanocomposite, an electrochemical sensor and a preparation and application thereof.

[0007] According to a first aspect of the present application, a zirconium-based metal organic framework-based nanocomposite is provided, which takes polyethyleneimine modified mesoporous carbon nanospheres (PEI / MCNs) as a carrier, and AuNPs / NH2-UIO-66 is directionally loaded on the surface of the PEI / MCNs through coordination, wherein the AuNPs / NH2-UIO-66 refers to gold nanoparticles (AuNPs) modified on the surface of an amino-functionalized zirconium-based metal-organic framework material (NH2-UIO-66).

[0008] As an optional implementation, the particle size distribution range of the NH2-UIO-66 is 15 nm-20 nm, and the particle size of the AuNPs is 10 nm-20 nm.

[0009] According to a second aspect of the present application, a preparation method of a zirconium-based metal organic framework-based nanocomposite is provided, which comprises the following steps: Mixing and stirring mesoporous carbon nanospheres (MCNs) and polyethyleneimine (PEI) to obtain PEI / MCNs; Mixing, ultrasonicating and drying gold nanoparticles (AuNPs) and an amino-functionalized zirconium-based metal-organic framework material (NH2-UIO-66) to obtain AuNPs / NH2-UIO-66; Mixing and ultrasonicating PEI / MCNs and AuNPs / NH2-UIO-66 to obtain AuNPs / NH2-UIO-66 / PEI / MCNs nanocomposite.

[0010] As an optional embodiment, the specific preparation process of the PEI / MCNs is as follows: After the MCNs are dissolved in ultrapure water and uniformly dispersed by ultrasonic, the PEI is added dropwise and stirred to make the PEI and the MCNs complex, and then the PEI / MCNs are obtained by filtration, ultrapure water washing and drying; wherein the mass ratio of the PEI to the MCNs is (0.5-1.5):(1.5-4.5).

[0011] As an optional embodiment, the specific preparation process of the AuNPs / NH2-UIO-66 is as follows: The HAuCl4 is prepared into an aqueous solution, heated to boiling, and 1% trisodium citrate aqueous solution is accurately added under stirring, and the aqueous solution is continuously boiled until it is stably red, and then cooled to room temperature and restored to the original volume with distilled water to prepare a gold nanoparticle solution; The NH2-UIO-66 is dissolved in the gold nanoparticle solution, ultrasonically dispersed until the NH2-UIO-66 is completely dispersed, and then filtered and dried to prepare the AuNPs / NH2-UIO-66; wherein the mass ratio of the NH2-UIO-66 to the AuNPs is (600-800):1.

[0012] As an optional embodiment, the mass ratio of the PEI / MCNs to the AuNPs / NH2-UIO-66 is (0.5-0.8):(5-10).

[0013] According to a third aspect of the object of the present application, an electrochemical sensor for detecting LPS is provided, comprising a substrate, a nanocomposite adsorbed on the surface of the substrate, and an aptamer grafted onto the nanocomposite; wherein the nanocomposite is the aforementioned nanocomposite based on a zirconium-based metal organic framework, and the substrate is a glassy carbon electrode.

[0014] According to a fourth aspect of the object of the present application, a preparation method of the aforementioned electrochemical sensor for detecting LPS is provided, comprising the following steps: The PEI / MCNs and the AuNPs / NH2-UIO-66 are mixed and ultrasonically dispersed, and then added into a Nafion solution to prepare a Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs suspension; wherein the mass concentration of the Nafion solution is 0.5%-2%, the concentration of the PEI / MCNs in the prepared suspension is 0.15 mg / mL-0.25 mg / mL, and the concentration of the AuNPs / NH2-UIO-66 in the prepared suspension is 5 mg / mL-8 mg / mL; The Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs suspension liquid is drop-coated on the surface of the pretreated glassy carbon electrode, and after drying, the aptamer solution is added dropwise and incubated, and then the BSA solution is added dropwise and incubated to block the non-specific binding sites, to obtain the electrochemical sensor.

[0015] According to a fifth aspect of the object of the present application, the electrochemical sensor for detecting LPS or the electrochemical sensor for detecting LPS prepared by the foregoing method is applied to the detection of endotoxins.

[0016] As an optional implementation, the detection adopts a three-electrode system, the three-electrode system includes a working electrode, a reference electrode and a counter electrode, and the working electrode is the electrochemical sensor; The detection includes: adding the to-be-detected liquid drop to the surface of the working electrode and incubating, then placing the three-electrode system in an electrolyte, and testing by cyclic voltammetry; wherein the electrolyte is a phosphate buffer solution containing [Fe(CN)6] 3- / 4- and KCl, and the electrochemical detection is implemented at a constant scanning rate of 100 mV / s in a potential scanning interval of-0.1 V to 0.6 V.

[0017] As can be seen from the technical solutions of the present application above, the nano-composite material based on zirconium-based metal organic framework proposed in the present application, PEI / MCNs is prepared by a step-by-step synthesis method, AuNPs is introduced into NH2-UIO-66 to form a functionalized nano-composite material, and then AuNPs / NH2-UIO-66 is directionally loaded on the surface of PEI / MCNs by coordination; in the present application, MCNs serve as a basic framework, providing an attachment platform for other components with high specific surface area and mesoporous structure, while allowing endotoxins and nucleic acid aptamers to quickly diffuse to the material surface; NH2-UIO-66 further adsorbs and enriches endotoxins in the solution through regular channels, improving the local concentration of endotoxins on the sensor surface and laying a foundation for subsequent recognition reactions; in the present application, The amino groups on the surface of NH2-UIO-66 are combined with nucleic acid aptamers through covalent bonds or electrostatic interactions, and PEI enhances the stability of this combination through cationic characteristics, ensuring that the nucleic acid aptamers are firmly fixed on the material surface and remain active; when endotoxins are present, the fixed nucleic acid aptamers are combined with endotoxins through specific recognition, completing the capture of target objects; The abundant reaction sites provided by the AuNPs further enhance the binding efficiency of the aptamer and the endotoxin, and the excellent electrical conductivity of the AuNPs accelerates the transmission of electrons in the material; the good electrical conductivity of the MCNs cooperates with the AuNPs, ensuring efficient transmission of electrons from the recognition site to the sensor electrode; the PEI optimizes the electron transmission path by adjusting the surface charge, reduces signal loss, and finally realizes significant amplification of the electrochemical signal; The bridging effect of the PEI enhances the binding force between the AuNPs, the NH2-UIO-66 and the MCNs, avoiding the shedding of components; the chemical stability and mechanical strength of the MCNs ensure that the material remains structurally intact in a complex detection environment, ensuring long-term stable operation of the sensor; In this way, through the synergistic effect of the components in the "enrichment-recognition-signal" link, both the specific recognition problem of the sensor and the detection sensitivity, response speed and stability are significantly improved, providing reliable material support for efficient detection of endotoxins in complex systems. BRIEF DESCRIPTION OF DRAWINGS

[0018] Figure 1 is a structural schematic diagram of the zirconium-based metal organic framework-based nanocomposite material of the present application.

[0019] Figure 2 is a detection principle diagram of the electrochemical sensor of the present application.

[0020] Figure 3 is a test diagram of the condition optimization of the electrochemical sensor in Example 1 of the present application; wherein, Figure 3 Part A of in is the MCNs:PEI ratio; Figure 3 Part B of in is the nanocomposite loading; Figure 3 Part C of in is the pH value of the electrolyte; Figure 3 Part D of in is the aptamer binding time.

[0021] Figure 4 is a TEM diagram of the material in Example 2 of the present application; wherein, Figure 4 Part A of in is NH2-UIO-66; Figure 4 Part B of in is AuNPs / NH2-UIO-66; Figure 4 Part C of in is MCNs; Figure 4 Part D of in is PEI / MCNs; Figure 4 Part E of in is a micrograph of AuNPs / NH2-UIO-66 / PEI / MCNs under a scale of 500 nm; Figure 4 Part F of in is a micrograph of AuNPs / NH2-UIO-66 / PEI / MCNs under 50 nm.

[0022] Figure 5is the XRD pattern of the material in Example 2 of the present application; wherein, Figure 5 Part A in is NH2-UIO-66; Figure 5 Part B in is MCNs, PEI / MCNs; Figure 5 Part C in is NH2-UIO-66, MCNs, PEI / MCNs and AuNPs / NH2-UIO-66 / PEI / MCNs samples.

[0023] Figure 6 is the electrochemical cyclic voltammogram of the working electrode under different modification steps in Example 2 of the present application; wherein, Figure 6 Part A in shows the cyclic voltammograms of NH2-UIO-66 / GCE (a), AuNPs / NH2-UIO-66 / GCE (b) and AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (c) in the potential range of -0.1 V to 0.6 V; Figure 6 Part B in shows the cyclic voltammograms of AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (a), Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (b), LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (c), BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (d), LPS / BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (e) in the potential range of 0.6 V to -0.1 V relative to Ag / AgCl.

[0024] Figure 7 is the standard curve of the electrochemical sensor of Example 2 of the present application.

[0025] Figure 8 is the selectivity statistical chart of the electrochemical sensor of Example 2 of the present application, and the interferents are magnesium sulfate, sodium nitrite, sodium chloride, citric acid and glucose, respectively.

[0026] Figure 9 is the stability performance statistical chart of the electrochemical sensor of Example 2 of the present application placed continuously for 0, 5 and 15 days.

[0027] Figure 10 is the reproducibility statistical chart of the electrochemical sensor of Example 2 of the present application of different batches. DETAILED DESCRIPTION

[0028] In order to better understand the technical content of the present application, specific embodiments are described below with the aid of the accompanying drawings.

[0029] Aspects of the present application are described in the disclosure by reference to the drawings, in which are shown by way of illustration various embodiments. The embodiments of the present disclosure are not necessarily intended to include all aspects of the present application. It should be understood that a variety of conceptual and embodiments introduced above, and those described in more detail below, can be implemented in any of a number of ways. Nanocomposite based on zirconium-based metal organic framework

[0030] In combination Figure 1 As shown, in a preferred embodiment of the present application, a zirconium-based metal-organic framework-based nanocomposite is provided, which takes polyethyleneimine modified mesoporous carbon nanospheres (PEI / MCNs) as a carrier, and AuNPs / NH2-UIO-66 is directionally loaded on the surface of PEI / MCNs through coordination, wherein AuNPs / NH2-UIO-66 refers to gold nanoparticles (AuNPs) modified on the surface of amino-functionalized zirconium-based metal-organic framework material (NH2-UIO-66); wherein the particle size distribution of NH2-UIO-66 is in the range of 15 nm to 20 nm, and the particle size of AuNPs is preferably in the range of 10 nm to 20 nm.

[0031] After modification by polyethyleneimine (PEI), a continuous coating layer is formed on the surface of MCNs due to the cross-linking effect of PEI molecular chains, resulting in the disappearance of the boundary between particles and the formation of a dense whole, which effectively enhances the chemical properties and electrical stability of mesoporous carbon nanospheres. Meanwhile, after the particle size of 10-20 nm gold nanoparticles (AuNPs) is modified on NH2-UIO-66, the gold nanoparticles can form a conductive path in the porous structure of NH2-UIO-66, which can significantly enhance the conductive effect and improve the signal stability; when AuNPs / NH2-UIO-66 and PEI / MCNs are further combined, the two can form a composite system with synergistic structure and performance through multiple interactions, and the comprehensive characteristics are significantly optimized. Process for the preparation of a nanocomposite based on zirconium-based metal organic framework

[0032] In another preferred embodiment of the present application, a preparation method of a zirconium-based metal-organic framework-based nanocomposite is provided, comprising the following steps: Mixing and stirring mesoporous carbon nanospheres (MCNs) with polyethyleneimine (PEI) to obtain PEI / MCNs; Mixing, ultrasonicating and drying gold nanoparticles (AuNPs) and amino-functionalized zirconium-based metal-organic framework material (NH2-UIO-66) to obtain AuNPs / NH2-UIO-66; The PEI / MCNs and the AuNPs / NH2-UIO-66 are mixed under ultrasonic to obtain the AuNPs / NH2-UIO-66 / PEI / MCNs nanocomposite.

[0033] In an optional example, the specific preparation process of the PEI / MCNs is as follows: The MCNs are dissolved in ultrapure water and uniformly dispersed under ultrasonic, then the PEI is added dropwise and stirred to make the PEI and the MCNs composite, and then the PEI / MCNs is obtained through filtration, ultrapure water cleaning and drying; wherein the mass ratio of the PEI to the MCNs is (0.5~1.5):(1.5~4.5).

[0034] In an optional example, the specific preparation process of the AuNPs / NH2-UIO-66 is as follows: The HAuC14 is prepared into an aqueous solution, heated to boiling, and 1% trisodium citrate aqueous solution is accurately added under stirring, and the heating and boiling are continued until the aqueous solution is stably red, then cooled to room temperature, and distilled water is used to restore to the original volume to prepare the gold nanoparticle solution; The NH2-UIO-66 is dissolved in the prepared gold nanoparticle solution, and ultrasonic is applied until the NH2-UIO-66 is completely dispersed, and then filtered and dried to prepare the AuNPs / NH2-UIO-66; wherein the mass ratio of the NH2-UIO-66 to the AuNPs is (600~800):1.

[0035] In an optional example, the mass ratio of the PEI / MCNs to the AuNPs / NH2-UIO-66 is (0.5~0.8):(5~10), and is particularly preferably 1:10. Electrochemical sensor for detecting LPS

[0036] In another preferred embodiment of the present application, an electrochemical sensor for detecting LPS prepared by using the aforementioned nanocomposite based on zirconium-based metal organic framework is provided, which comprises a substrate, a nanocomposite adsorbed on the surface of the substrate, and an aptamer grafted to the nanocomposite; wherein the nanocomposite is the aforementioned nanocomposite based on zirconium-based metal organic framework, and the substrate is a glassy carbon electrode. Process for the preparation of an electrochemical sensor for detecting LPS

[0037] In one preferred embodiment, the preparation method of the aforementioned electrochemical sensor for detecting LPS comprises the following steps: The Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs suspension is prepared by mixing PEI / MCNs and AuNPs / NH2-UIO-66 under ultrasonic and then adding a Nafion solution; wherein the mass concentration of the Nafion solution is 0.5% to 2%, the concentration of PEI / MCNs in the prepared suspension is 0.15 mg / mL to 0.25 mg / mL, and the concentration of AuNPs / NH2-UIO-66 in the prepared suspension is 5 mg / mL to 8 mg / mL.

[0038] The Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs suspension is drop-coated on the surface of a pretreated glassy carbon electrode, dried, and then the aptamer solution is added dropwise and incubated, followed by adding the BSA solution dropwise for incubation to block the non-specific binding sites, to obtain the electrochemical sensor. Use in the detection of endotoxins

[0039] In other exemplary embodiments, the application also provides a use of the aforementioned electrochemical sensor for detecting LPS in detecting endotoxins.

[0040] In an optional example, the detection adopts a three-electrode system, which includes a working electrode, a reference electrode and a counter electrode, and the working electrode is the electrochemical sensor; The detection includes: adding the to-be-detected liquid dropwise on the surface of the working electrode for incubation, then placing the three-electrode system in an electrolyte, and testing by cyclic voltammetry; wherein the electrolyte is a phosphate buffer solution containing [Fe(CN)6] 3- / 4- and KCl, and the electrochemical detection is implemented at a constant scanning rate of 100 mV / s in a potential scanning interval of -0.1 V to 0.6 V.

[0041] The present application relies on aptamer as a recognition probe, which is a single-stranded DNA or RNA fragment screened by SELEX (Systematic Evolution of Ligands by Exponential Enrichment) technology, and has high specificity and affinity. Aptamer can specifically bind to endotoxin through base pairing, conformational changes and other ways, and this specific binding has high selectivity, which can effectively distinguish endotoxin from other interferents in the solution. Even in a complex solution system, aptamer can accurately identify endotoxin and reduce non-specific binding with interferents, thereby significantly improving the specificity of the sensor. For example, a specific aptamer can specifically bind to a specific domain on the surface of endotoxin, while not producing obvious interaction with other biological molecules or chemical substances, ensuring that the detection signal only comes from the binding event between endotoxin and aptamer, greatly reducing the influence of interferents on the detection result. In the electrochemical sensor of the present application, Gold nanoparticles have good biocompatibility and excellent electrical properties. Their large specific surface area can provide abundant reaction sites for the binding of aptamer and endotoxin, increase the binding efficiency, and thus improve the sensitivity of the sensor. At the same time, gold nanoparticles can accelerate electron transfer and enhance electrochemical signals, making the detection signal more obvious and easier to detect and analyze.

[0042] NH2-UIO-66 has a regular pore structure and a large specific surface area, which can effectively adsorb and enrich endotoxin, increase the concentration of endotoxin on the surface of the sensor, and further enhance the detection signal. Its amino functionalization feature makes its surface have abundant amino groups, which can stably bind with aptamer through covalent bond or electrostatic interaction, improve the fixing efficiency and stability of aptamer on the surface of the sensor, and ensure that aptamer can continuously play a specific recognition role.

[0043] PEI is a cationic polymer with good water solubility and high positive charge density. It can interact with negatively charged aptamer, endotoxin and the groups on the surface of NH2-UIO-66 through electrostatic interaction, play the role of bridge and connection, enhance the binding force between the components of the nanocomposite, and make the whole system more stable. At the same time, the presence of PEI can also adjust the charge properties of the sensor surface, optimize the electron transfer process, and improve the electrochemical performance of the sensor.

[0044] Mesoporous carbon nanoparticles have high specific surface area, large pore volume and good electrical conductivity. Their mesoporous structure is conducive to the diffusion and transmission of endotoxin and aptamer, and promotes the rapid progress of biological recognition reaction. Good electrical conductivity can ensure efficient electron transfer in the sensor, further improving the detection sensitivity and response speed of the sensor. In addition, MCNs also have certain chemical stability and mechanical strength, which can enhance the overall stability of the nanocomposite and maintain good performance in complex detection environment.

[0045] Through the synergistic effect of the components, not only the specificity of the sensor is solved, but also the stability and detection performance of the sensor are significantly improved. The specific recognition of aptamer combined with the efficient signal amplification and stable support of the nanocomposite enables the sensor to accurately and stably detect endotoxin in complex solution system. At the same time, the nanocomposite has the characteristics of simple synthesis process, easy access to raw materials, environmental friendliness and low cost, which meets the demand of practical application from the cost point of view, and has broad application prospect.

[0046] Next, combined with the following Figure 2 In the exemplary embodiments of the present application, the preparation and detection process of the electrochemical sensor for detecting LPS is as follows:

[0047] (1) Preparation of AuNPs / NH2-UIO-66 nanocomposites First, HAuCl4 was prepared into a 0.01% aqueous solution, 100 mL of which was heated to boiling, and a certain amount of 1% aqueous sodium citrate solution was accurately added under stirring, and boiling was continued for 5 min. At this time, it was observed that the light yellow aqueous solution of chloroauric acid quickly turned gray after the addition of sodium citrate, and then turned black, and then gradually stabilized into red. The whole process took about 2-3 min. After cooling to room temperature, the original volume was restored with distilled water, and a gold nanoparticle solution was prepared. The molar ratio of HAuCl4 to sodium citrate was 1:(8-9).

[0048] NH2-UIO-66 was dissolved in the prepared gold nanoparticle solution (mass concentration about 1%), and ultrasonic was applied until NH2-UIO-66 was completely dispersed. After filtration and drying, AuNPs / NH2-UIO-66 nanocomposites were prepared.

[0049] (2) Preparation of PEI / MCNs nanocomposites The mesoporous carbon nanospheres were dissolved in ultrapure water and ultrasonicated to disperse uniformly. Then, PEI was added dropwise and stirred for 12 h. After filtration, the product was washed with ultrapure water three times and dried to obtain PEI functionalized mesoporous carbon nanospheres (PEI / MCNs nanocomposites).

[0050] (3) Preparation of Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs AuNPs / NH2-UIO-66 and PEI / MCNs were mixed according to a mass ratio of (5-10):(0.5-0.8) and ultrasonicated for 30 min. Finally, 100 μL of Nafion solution was added to modify the electrode surface and improve the stability of the electrode. Finally, Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs was prepared.

[0051] (4) Preparation of BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE The glassy carbon electrode was polished with 0.3 μm and 0.05 μm alumina slurry in the shape of a "8" on a piece of suede, and then ultrasonically cleaned in ultrapure water or anhydrous ethanol (10 s / time, repeated 3-5 times) until a mirror-like luster was formed on the surface of the electrode.

[0052] Afterwards, the Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs was ultrasonically treated for 30 min, 20 μL of the suspension was dropped on the surface of the pretreated glassy carbon electrode, and a uniform film was formed after drying at room temperature, thus obtaining the Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs modified glassy carbon electrode (Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE).

[0053] After the electrode material was dried on the surface of the glassy carbon electrode, 2 μL of the aptamer (LBA) solution was added, and incubated at 4 ℃ for 60-120 min, and the free nucleic acid aptamer was washed with ultrapure water (the nucleotide sequence of the aptamer is 5'-SH2-(CH2)6-CTTCTGCCCGCCTCCTTCCTAGCCGGATCGCGCTGGCCAGATGATATAAAGGGTCAGCCCCCCAGGAGACGAGATAGGCGGACACT-3').

[0054] 10 μL of a 1%-5% bovine serum albumin (BSA) solution was continuously added to block the unbound active sites, and incubated at 4 ℃ for 15-60 min, and the unbound bovine serum albumin was washed with water to obtain the BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE.

[0055] (5) The nucleic acid aptamer biosensor interface was introduced into the test liquid, and the sensing system was placed in a low-temperature environment at 4 ℃, and incubated for 1-2 hours to ensure that the specific binding between LPS and the aptamer was fully carried out; after the binding reaction was terminated, the sensing interface was washed with a suitable buffer solution (for example, PBS buffer solution, TF buffer solution) for multiple times to effectively remove the free LPS molecules that did not occur binding.

[0056] Subsequently, the cleaned biosensor was completely immersed in the electrolyte system of the phosphate buffer solution containing [Fe(CN)6] 3- / 4- and KCl prepared in advance, and electrochemical detection was carried out at a constant scanning rate of 100 mV / s in the potential scanning interval of-0.1 V to 0.6 V, and the corresponding response signal was obtained.

[0057] The application constructs an electrochemical sensing platform based on AuNPs / NH2-UIO-66 / PEI / MCNs nanocomposite material, and successfully applies it to efficient detection of endotoxin in food. In view of the characteristics of food matrix complexity and many interference substances, the sensor improves the anti-interference ability through multi-stage cooperative interface design: polyethyleneimine modified mesoporous carbon nanospheres (PEI / MCNs) are used as a substrate, the signal stability is significantly improved by combining the directional loading characteristics of NH2-UIO-66 and the conductive enhancement effect of gold nanoparticles (AuNPs). The sensor exhibits excellent detection performance, with a detection limit as low as 40 fg / mL, and can realize trace identification of lipopolysaccharide (LPS). At the same time, it shows good selectivity to common interference substances in food, which helps to quickly and accurately detect LPS.

[0058] In order to better understand, the application will be further described below in conjunction with several specific examples, but the preparation process is not limited thereto, and the content of the application is not limited thereto.

[0059] Unless otherwise specified, the materials in the examples are prepared according to existing methods or directly purchased from the market.

[0060] The glassy carbon electrodes used in the following examples are treated as follows: The glassy carbon electrode is successively polished with 0.3 μm and 0.05 μm alumina slurries in a "8" shape, and then immersed in ultrapure water or anhydrous ethanol for ultrasonic cleaning (10 s / time, repeated 3-5 times) until a mirror-like luster is formed on the surface of the electrode. Example 1

[0061] [Preparation of AuNPs / NH2-UIO-66 nanocomposite material]

[0062] HAuCl4 is first prepared into a 0.01 % aqueous solution, 100 mL of which is heated to boiling, and 2 mL of 1 % aqueous trisodium citrate solution is accurately added under stirring, and boiling is continued for 5 min. At this time, it can be observed that the light yellow aqueous solution of chloroauric acid quickly turns gray after the addition of sodium citrate, and then turns black, and then gradually stabilizes into red. The whole process takes about 2-3 min. After cooling to room temperature, the original volume is restored with distilled water to prepare a gold nanoparticle solution.

[0063] 60 mg of NH2-UIO-66 is dissolved in the prepared gold nanoparticle solution, and ultrasonic treatment is performed until the NH2-UIO-66 is completely dispersed. After filtration and drying, an AuNPs / NH2-UIO-66 nanocomposite material is prepared. Example 2

[0064] [Preparation of PEI / MCNs nanocomposite material]

[0065] The mesoporous carbon nanospheres MCNs were dissolved in ultrapure water and ultrasonically dispersed, then PEI was added dropwise and stirred for 12 h, and then filtered, washed with ultrapure water three times, and dried to obtain PEI functionalized mesoporous carbon nanospheres (PEI / MCNs nanocomposites) (the mass ratio of MCNs to PEI was variable 1).

[0066] The specific values of variable 1 are shown in Table 1.

[0067] Table 1

[0068] Sample 1-5 10 mg was dispersed in 20 mL of ultrapure water, and then drop-coated on the surface of a pretreated glassy carbon electrode to obtain the corresponding electrode. The corresponding electrode was tested by cyclic voltammetry (5 mM [Fe(CN)6] 3- / 4- and 0.1 M KCl phosphate buffer solution (PBS) pH = 7.4, and the electrochemical detection was carried out at a constant scan rate of 100 mV / s in a potential scan range of -0.1 V to 0.6 V, and then the corresponding response signal was obtained), and the results are shown in Part A of Figure 3

[0069] When the ratio of MCNs was increased to 3:1, the electrochemical activity of the composite material was significantly enhanced, and the oxidation peak current value increased significantly, with an increase of 7%. The ratio of MCNs:PEI = 3:1 was finally selected as the optimal composite ratio for subsequent research. Example 3

[0070] [Preparation of Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs nanocomposite]

[0071] AuNPs / NH2-UIO-66 (60 mg, prepared by the method of Example 1) and PEI / MCNs (6 mg, prepared by the method of Sample 5) were mixed and ultrasonicated for 30 min, and finally 100 μL of Nafion solution (mass concentration 0.5%) was added to prepare the Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs composite material.

[0072] Preparation of BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE ​The Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs nanocomposite material is ultrasonically treated for 30 min, and a suspension (volume as a variable 2) is taken and dropped on the surface of the pretreated glassy carbon electrode, and after drying at room temperature to form a uniform film, a Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE is prepared.

[0073] The specific value of the variable 2 is shown in Table 2.

[0074] Table 2

[0075] Samples 6-10 are tested by cyclic voltammetry (5 mM [Fe(CN)6] 3- / 4- and 0.1 M KCl in a phosphate buffer solution (PBS) with pH = 6, and electrochemical detection is performed at a constant scanning rate of 100 mV / s in a potential scanning interval of -0.1 V to 0.6 V, and the corresponding response signal is obtained, and the results are shown in Part C of Table 3. Figure 3

[0076] Under the condition of the electrolyte with pH = 6, when the volume of the material solution increases from 4 μL to 20 μL, the electrochemical signal intensity shows a gradient enhancement trend, which fully verifies the excellent conductive characteristics of the nanocomposite material of the application, and at the same time, it shows that within the experimental test range, the electrode performance is positively correlated with the material loading amount; and 20 μL is comprehensively selected as the optimal loading amount.

[0077] The loading amount of the nanocomposite material on the glassy carbon electrode is 20 μL, and 6 samples are prepared according to the preparation process of Sample 10, and are detected in 5 mM [Fe(CN)6] 3- / 4- and 0.1 M KCl in a phosphate buffer solution (PBS) with pH = 6, 6.5, 7, 7.5, 8.0 and 8.5, and electrochemical detection is performed at a constant scanning rate of 100 mV / s in a potential scanning interval of -0.1 V to 0.6 V, and the corresponding response signal is obtained, and the results are shown in Part C of Table 3. Figure 3

[0078] The pH of the electrolyte is from 6 to 8.5, the peak current intensity first increases and then decreases, and reaches the maximum at pH = 6, and therefore, pH = 6 is selected as the optimal electrolyte pH. Example 4

[0079] ​​After the Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs electrode material on the surface of the glassy carbon electrode is dried (Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE, prepared according to the process of preparing sample 10), 2 μL of an aptamer solution (Shanghai Biotech, nucleotide sequence 5'-SH2-(CH2)6-CTTCTGCCCGCCTCCTTCCTAGCCGGATCGCGCTGGCCAGATGATATAAAGGGTCAGCCCCCCAGGAGACGAGATAGGCGGACACT-3') is added dropwise, and incubated at 4 ℃ (the time is a variable 3), the free nucleic acid aptamer is washed with ultrapure water, and after drying at room temperature, a modified electrode LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE is obtained.

[0080] 10 μL of a 1% bovine serum protein solution is added dropwise to block the unbound active sites, and incubated at 4 ℃ for 15 min, and the unbound bovine serum protein is washed with water to obtain a BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE.

[0081] The specific values of variable 3 are shown in Table 3.

[0082] Table 3

[0083] Samples 11-15 are tested by cyclic voltammetry (5 mM [Fe(CN)6] 3- / 4- and 0.1 M KCl phosphate buffer solution (PBS) pH=6, and electrochemical detection is performed at a constant scan rate of 100 mV / s in a potential scan interval of -0.1 V to 0.6 V, and the corresponding response signal is obtained), and the results are shown in Part D of Table 4. Figure 3

[0084] As the fixation time is extended to 60 min, the decay rate of the peak current is significantly reduced and tends to be stable, so 60 min is selected as the optimal fixation time. Example 5

[0085] The same preparation method as in Examples 1-4 is used, where (variable 1 is 3:1, variable 2 is 20 μL, and variable 3 is 60 min), and the obtained BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE is sample 16.

[0086] ​The samples used in the following tests are sample 16 and the corresponding intermediate sample prepared by the method in Example 5. Example 6

[0087] The morphology of the sample was tested, and the results are as follows: Figure 4 As shown.

[0088] Figure 4 Part A shows that the NH2-UIO-66 crystals exhibit a well-dispersed spherical nanocrystalline structure with a particle size distribution range of 15-20 nm. The crystals are clear and there is no significant agglomeration, indicating that the amino functionalization modification did not destroy the crystallinity and morphological integrity of the UIO-66 framework.

[0089] Figure 4 In part B, gold nanoparticles (AuNPs) with a particle size of 10-20 nm were observed to be uniformly anchored on the surface of NH2-UIO-66, and the interface between the two was in close contact, confirming that the abundant amino functional groups on the surface of NH2-UIO-66 can effectively stabilize metal nanoparticles through coordination.

[0090] Figure 4 Part C in the figure shows that unmodified mesoporous carbon nanospheres (MCNs) are formed by physical stacking of primary carbon nanounits with a diameter of less than 5 nm to form a three-dimensional porous network structure, but there are obvious boundaries between particles.

[0091] Figure 4 Part D shows that after modification with polyethyleneimine (PEI), a continuous coating layer is formed on the surface of MCNs due to the cross-linking of PEI molecular chains, resulting in the disappearance of interparticle boundaries and the formation of a dense whole. This indicates that PEI can significantly enhance the structural integration of MCNs through chemical bonding.

[0092] Further morphological analysis of the composite material was conducted. Figure 4 (Conducting studies on the E and F portions of the composite material), it was found that AuNPs and NH2-UIO-66 were preferentially loaded on the surface of PEI / MCNs, and the three components formed a stable heterostructure through amino-carboxyl electrostatic attraction and metal-ligand interactions. The amorphous carbon matrix of AuNPs, NH2-UIO-66, and PEI / MCNs was clearly distinguishable in high-resolution TEM images, and the components were uniformly distributed without phase separation, confirming the successful construction of the AuNPs / NH2-UIO-66 / PEI / MCNs multi-component composite material. Example 7

[0093] The samples were characterized using XRD technology, and the results are as follows: Figure 5 As shown.

[0094] Figure 5Part A of the graph clearly shows a strong peak at 2θ = 7.290°, indicating the presence of a nanodefect structure that can improve conductivity to some extent. Meanwhile, the characteristic peaks of NH2-UIO-66 are consistent with the XRD patterns reported in previous studies, indicating that NH2-UIO-66 was successfully synthesized.

[0095] from Figure 5 In the B part of the MCNs, two distinct characteristic peaks appear at 25° and 43°, corresponding to the (002) and (101) crystal planes of the carbon structure, which illustrates the amorphous nature of the mesoporous carbon nanospheres. When the MCNs are modified with PEI, PEI may reduce the amorphous region by filling the micropores or defects on the surface of the MCNs, thereby enhancing the crystallinity and resulting in a decrease in the half-width at half-maximum. The PEI layer may also change the scattering path of X-rays on the material surface, leading to an apparent increase in peak intensity, as can be clearly seen from the significant increase in the peak values ​​at 25° and 43°.

[0096] from Figure 5 Part C shows that the diffraction peak positions of the composite are basically consistent with those of pure NH2-UIO-66, indicating that the loading process did not destroy the crystal structure of the MOF. Meanwhile, in Figure 5 The C AuNPs / NH2-UIO-66 / PEI / MCNs composite material shows a weak peak at 38-40°. The low peak intensity may be related to the small size (10-15nm) and high dispersion of Au nanoparticles. Some particles may be embedded in the pores of MOF, resulting in signal dilution.

[0097] XRD analysis showed that the AuNPs / NH2-UIO-66 / PEI / MCNs composite successfully loaded highly dispersed Au nanoparticles while retaining the crystal structure of NH2-UIO-66, and the introduction of PEI / MCNs did not destroy its crystal form.

[0098] As shown by the above tests, the present invention has successfully synthesized AuNPs / NH2-UIO-66 / PEI / MCNs nanocomposite materials. Example 8

[0099] Cyclic voltammetry experiments were conducted on the electrode materials and electrodes at different stages of the modification process to understand the conductivity of the materials at each stage. A sample containing 5 mM [Fe(CN)6] was used. 3- / 4- A phosphate-buffered saline (PBS) solution of 0.1 M KCl was used as the electrolyte to detect the electrochemical properties of various electrodes. The results are as follows: Figure 6 As shown.

[0100] Figure 6Part A in FIG. 1 shows the cyclic voltammograms of NH2-UIO-66 / GCE (a), AuNPs / NH2-UIO-66 / GCE (b) and AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (c) in the potential range of -0.1 V to 0.6 V. Figure 6 Part B in FIG. 1 shows the cyclic voltammograms of AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (a), Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (b), LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (c), BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (d), LPS / BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE (e) in the potential range of 0.6 V to -0.1 V vs. Ag / AgCl.

[0101] wherein, the preparation of NH2-UIO-66 / GCE is by dispersing NH2-UIO-66 in ultrapure water (concentration 6 mg / mL) and dropping onto the surface of the pretreated glassy carbon electrode as needed; the preparation of AuNPs / NH2-UIO-66 / GCE is by dispersing AuNPs / NH2-UIO-66 in ultrapure water (concentration 0.5 mg / mL) and dropping onto the surface of the pretreated glassy carbon electrode as needed; the preparation of AuNPs / NH2-UIO-66 / PEI / MCNs / GCE is by dispersing AuNPs / NH2-UIO-66 (concentration 6 mg / mL) and PEI / MCNs (concentration 0.2 mg / mL) in ultrapure water and dropping onto the surface of the pretreated glassy carbon electrode as needed; Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE, LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE and BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE are obtained according to the method of Example 2; LPS / BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE is obtained by adding 10 μL of 100 ng / mL LPS on the surface of BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE.

[0102] As can be seen from the figure, when the AuNPs / NH2-UIO-66 / PEI / MCNs is modified on the GCE, the peak current is maximum; when the Nafion is introduced into the AuNPs / NH2-UIO-66 / PEI / MCNs composite system, the double characteristics of its electron insulation and proton selective conduction cause the cyclic voltammetry peak current to decrease.

[0103] In the hierarchical composite sensing interface constructed based on Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs, after the APT aptamer is directionally immobilized through Au-S covalent bond, due to the non-conductivity of LBA as a biological macromolecule and the electrostatic repulsion between the phosphate skeleton of the aptamer and the redox probe [Fe(CN)6] 3- / 4- , the diffusion mass transfer is hindered, and the electrochemical cyclic voltammetry response current further decreases.

[0104] After the LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE is incubated with BSA, due to the dense protein layer formed by the BSA molecules, the physical hindering effect increases the electron transfer resistance of the electrode / solution interface, and the three-dimensional structure of the BSA molecules limits the diffusion process of the electroactive substance to the electrode surface, so that the current signal further decreases.

[0105] When the LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE composite electrode specifically combines with lipopolysaccharide (LPS), the spatial conformation matching between the target substance and the aptamer induces the molecular self-assembly process, the three-dimensional conformation change caused by the combination of the aptamer and the LPS significantly increases the electron transport barrier of the solid-liquid interface, and the diffusion mass transfer process of the electroactive substance [Fe(CN)6] 3- / 4- is inhibited through the steric hindering effect, so that the peak current further decreases.

[0106] From the above, it can be seen that each step of modification of the sensor of the application successfully achieves the corresponding modification purpose, and it is proved that the sensor prepared by the application can be applied to the actual detection of endotoxin. Example 9

[0107] [Linear range of the electrochemical biosensor for LPS detection]

[0108] Based on the electrochemical biosensors constructed in the above steps, their surfaces were modified with LPS solutions at concentrations of 0.0001 ng / mL, 0.001 ng / mL, 0.01 ng / mL, 0.1 ng / mL, 1 ng / mL, 10 ng / mL, and 100 ng / mL, respectively. After incubation at low temperature for 60 min, the electrochemical biosensor interfaces that have completed recognition and detection were obtained. Each concentration of LPS solution corresponds to one electrochemical biosensor, and there is a one-to-one correspondence between the concentration and the electrochemical biosensor.

[0109] A three-electrode system (GCE working electrode, Pt counter electrode, Ag / AgCl reference electrode) was used on a CHI 760E electrochemical workstation, selecting the cyclic voltammetry (CV) detection step. The electrolyte system contained 5 mM [Fe(CN)6]. 3- / 4- The detection conditions were cyclic voltammetry, using a PBS solution containing 0.1 M KCl, and scanning potentials from -0.1 to 0.6 V at a scan rate of 100 mV.

[0110] Calculate the LPS concentration in the standard solution, where x is lg CLPS and y is I. 空白 -I LPS The linear regression equation was calculated based on x and y, and the equation formula is I (μA) = 0.864lgC (ng / mL) + 3.83 (R² = 0.996), which was used as the linear equation in the actual detection.

[0111] pass Figure 7 It can be seen that the proposed sensing strategy has a linear range of 0.0001 ng / mL to 100 ng / mL for LPS detection, spanning 7 orders of magnitude, indicating that the sensor of the present invention can be successfully applied to practical detection. Example 10

[0112] [Selectivity Analysis of Electrochemical Sensors]

[0113] In food processing systems, various functional additives are often added to maintain product freshness and regulate textural properties. However, these compounds may cause matrix interference effects on the detection performance of aptamer biosensors. To systematically evaluate the selectivity characteristics of the sensor, this test constructed a mixed interference system containing magnesium sulfate, sodium nitrite, sodium chloride, citric acid, and glucose, with the concentrations of each component consistent with the target analyte detection conditions.

[0114] like Figure 8As shown, the test results show that the oxidation peak current values of each group of solutions have no significant difference with the blank control, and when the detection system contains 100 ng / mL of LPS, the oxidation peak current value appears a significant decrease, which is in sharp contrast with the blank solution. The comparison experiment shows that the sensor of the present application has a high specific recognition ability for LPS detection, effectively excluding the interference of common components in the food matrix. Example 11

[0115] [Stability analysis of electrochemical sensor]

[0116] After the preparation of BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE, it is stored at 4 °C. The influence of time on the performance of the sensor is evaluated by regularly monitoring the endotoxin level at a constant concentration and observing the fluctuation of the peak current.

[0117] As shown in Figure 9 Compared with day 0, the peak current of 5 days and 15 days increased by about 0.1 % and decreased by 0.2 %, respectively. Based on these data, it can be concluded that the sensor of the present application exhibits excellent stability and is suitable for long-term storage. Example 12

[0118] [Repeatability analysis of electrochemical sensor]

[0119] In this study, five batches of BSA / LBA / Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs / GCE composite electrodes were successfully prepared under standardized production conditions (prepared according to the method of Example 2).

[0120] As shown in Figure 10 The five groups of independently constructed sensors exhibit excellent signal consistency in the same detection system, with an inter-group relative standard deviation (RSD) of the oxidation peak current of only 0.118 %, which confirms that the sensor preparation process of the present application has excellent batch-to-batch stability.

[0121] As can be seen from the above, the nanocomposite of the present application, through the synergistic effect of each component, not only solves the problem of specificity of the sensor, but also significantly improves the stability and detection performance of the sensor; the specific recognition of the aptamer combined with the efficient signal amplification and stable support of the nanocomposite enables the sensor of the present application to accurately and stably detect endotoxin in a complex solution system; at the same time, the nanocomposite has the characteristics of simple synthesis process, easy access to raw materials, environmental friendliness and low cost, etc., which meets the demand of practical application from the cost point of view, and has a broad application prospect.

[0122] While the application has been described by way of example with reference to preferred embodiments, it is to be understood that this application is not limited to the embodiments disclosed, but is intended to cover modifications and variations within the spirit and scope of the application. Therefore, the scope of the application is defined not by the detailed description of the application but by the following claims, wherein reference to an alternative embodiment includes reference to all features describing that embodiment.

Claims

1. A nanocomposite based on a zirconium-based metal-organic framework, characterized in that, The nanocomposite takes polyethyleneimine modified mesoporous carbon nanospheres (PEI / MCNs) as a carrier, and AuNPs / NH2-UIO-66 is directionally loaded on the surface of the PEI / MCNs through coordination, wherein the AuNPs / NH2-UIO-66 refers to gold nanoparticles (AuNPs) modified on the surface of an amino-functionalized zirconium-based metal-organic framework (NH2-UIO-66).

2. The zirconium-based metal-organic framework based nanocomposite of claim 1, wherein, The particle size distribution of the NH2-UIO-66 ranges from 15 nm to 20 nm, and the particle size of the AuNPs ranges from 10 nm to 20 nm.

3. A method for preparing a nanocomposite based on a zirconium-based metal organic framework, characterized in that, The method comprises the following steps: MCNs are mixed with polyethyleneimine (PEI) to obtain PEI / MCNs; AuNPs are mixed with the amino-functionalized zirconium-based metal-organic framework (NH2-UIO-66), ultrasonicated, and dried to obtain AuNPs / NH2-UIO-66; PEI / MCNs are mixed with AuNPs / NH2-UIO-66 and ultrasonicated to obtain the AuNPs / NH2-UIO-66 / PEI / MCNs nanocomposite.

4. The method of claim 3, wherein the zirconium-based metal organic framework-based nanocomposite is prepared by a method comprising: (a) preparing a mixture of a zirconium-based metal organic framework and a polymer; and (b) heating the mixture to form the zirconium-based metal organic framework-based nanocomposite. The specific preparation process of the PEI / MCNs is as follows: After MCNs are dissolved in ultrapure water and ultrasonicated to be uniformly dispersed, PEI is added dropwise and stirred to allow the PEI to be compounded with the MCNs, and then the PEI / MCNs are obtained through filtration, ultrapure water washing, and drying; wherein the mass ratio of the PEI to the MCNs is (0.5-1.5):(1.5-4.5).

5. The method for preparing the zirconium-based metal-organic framework nanocomposite material according to claim 3, characterized in that, The specific preparation process of the AuNPs / NH2-UIO-66 is as follows: HAuC14 is prepared into an aqueous solution, heated to boiling, and 1% trisodium citrate aqueous solution is accurately added under stirring, and the aqueous solution is continuously boiled until it is stably red, and then cooled to room temperature and restored to the original volume with distilled water to prepare a gold nanoparticle solution; NH2-UIO-66 is dissolved in the gold nanoparticle solution, ultrasonicated until the NH2-UIO-66 is completely dispersed, and then filtered and dried to obtain AuNPs / NH2-UIO-66; wherein the mass ratio of the NH2-UIO-66 to the AuNPs is (600-800):

1.

6. The method of claim 3, wherein the zirconium-based metal-organic framework-based nanocomposite is prepared by a method comprising: (a) preparing a mixture of a zirconium-based metal-organic framework and a polymer; and (b) heating the mixture to form the zirconium-based metal-organic framework-based nanocomposite. The mass ratio of the PEI / MCNs to the AuNPs / NH2-UIO-66 is (0.5-0.8):(5-10).

7. An electrochemical sensor for detecting LPS, characterized in that, The method comprises the following steps:

8. A method of preparing an electrochemical sensor for detecting LPS as claimed in claim 7, wherein, The method comprises the following steps: The method comprises the following steps: The Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs suspension is prepared by mixing PEI / MCNs and AuNPs / NH2-UIO-66 under ultrasonic and then adding a Nafion solution; wherein the mass concentration of the Nafion solution is 0.5% to 2%, the concentration of PEI / MCNs in the prepared suspension is 0.15 mg / mL to 0.25 mg / mL, and the concentration of AuNPs / NH2-UIO-66 in the prepared suspension is 5 mg / mL to 8 mg / mL; The Nafion / AuNPs / NH2-UIO-66 / PEI / MCNs suspension is drop-coated on the surface of a pretreated glassy carbon electrode, dried, and then the aptamer solution is added dropwise and incubated, followed by adding the BSA solution dropwise for incubation to block the non-specific binding sites, to obtain the electrochemical sensor.

9. Use of the electrochemical sensor for detecting LPS according to claim 7 or the electrochemical sensor for detecting LPS prepared by the method according to claim 8 in detecting endotoxins.

10. Use according to claim 9, characterized in that, The detection uses a three-electrode system, which comprises a working electrode, a reference electrode and a counter electrode, and the working electrode is the electrochemical sensor. The detection comprises: incubating the to-be-tested liquid droplet on the surface of the working electrode, then placing the three-electrode system in an electrolyte, and testing by cyclic voltammetry; wherein the electrolyte is a phosphate buffer solution containing [Fe(CN)6] 3- / 4- and KCl, and the electrochemical detection is performed at a constant scanning rate of 100 mV / s in a potential scanning interval of -0.1 V to 0.6 V.

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