Molecularly imprinted magnetic covalent organic framework as well as preparation method and application thereof

Abstract

The invention discloses a molecularly imprinted magnetic covalent organic framework as well as a preparation method and application thereof, and relates to the technical field of functional materials. The preparation method comprises the following steps: performing prepolymerization reaction on template molecules and functional monomers in a pore-foaming agent, adding a magnetic metal covalent organic framework for mixing reaction, then adding a cross-linking agent and an initiator for polymerization reaction, separating out a product under the action of an external magnetic field after the reaction is finished, and eluting to remove the template molecules, thereby obtaining the magnetic metal covalent organic framework. The molecularly imprinted magnetic covalent organic framework is obtained; the template molecule is doxycycline. The molecularly imprinted magnetic covalent organic framework has the advantages of efficient magnetic separation and high selectivity, and can simply, stably and reliably detect DOX in a complex sample.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of functional materials, in particular to a molecularly imprinted magnetic covalent organic framework as well as a preparation method and application thereof. BACKGROUND

[0002] Doxycycline (DOX), also known as strong mycin or deoxytetracycline, as a typical representative of tetracycline antibiotics, has good antibacterial effect due to its chemical structure composed of a beta-diketone group. The chemical structural formula of DOX is as follows: .

[0003] Due to good gastrointestinal absorption, strong tissue penetration ability and long biological half-life, DOX is widely used in the field of livestock and poultry breeding for the prevention and treatment of bacterial infections in respiratory tract, digestive tract, urinary tract, etc. At present, the detection methods of DOX mainly include high performance liquid chromatography (HPLC), enzyme-linked immunoassay, thin layer chromatography, capillary electrophoresis, fluorescence probe analysis, electrochemistry and voltammetry, etc. However, the matrix of DOX in meat is complex, and these detection methods have certain shortcomings. Wang Minghui et al. used HPLC to detect chlortetracycline and DOX in shellfish at the same time, which had a relatively long analysis time and consumed a large amount of reagents (see the literature "Wang Minghui, Zheng Lit, Li Shuyi, et al. Simultaneous determination of chlortetracycline and doxycycline in shellfish products by high performance liquid chromatography [J]. Fujian Light Textile, 2023, (12): 7-11+18." ). Zhang Qing studied the elimination rules of DOX soluble powder in pigs and chickens, and found that enzyme-linked immunoassay was not suitable for simultaneous analysis of multiple components (see the literature "Zhang Qing. Residue elimination rules of doxycycline hydrochloride soluble powder in pigs and chickens [D]. Henan Agricultural University, 2022." ). Choma I et al. used a simple thin-layer chromatographic method to determine flumequine and DOX in milk, which had poor repeatability (see the literature "Choma I, Grenda D, Malinowska I, et al. Determination of flumequine and doxycycline in milk by a simple thin-layer chromatographic method [J]. Journal of Chromatography B, Biomedical Sciences and Applications, 1999, 734(1): 7-14" ). Yu Qin used low eutectic solvent pretreatment technology combined with capillary electrophoresis to detect veterinary drug residues, which had high requirements for sample pretreatment and low detection throughput (see the literature "Yu Qin. Sample pretreatment technology based on low eutectic solvent combined with capillary electrophoresis for detection of veterinary drug residues [D]. Suzhou University of Science and Technology, 2023" ). In the study of fluorescence probe detection of tetracycline by Yin Ziyu et al., environmental changes easily led to inaccurate detection results (see the literature "Yin Ziyu, Zhou Shuai, Yao Zhiyi. Research progress of rapid detection method of tetracycline based on nano-fluorescent probe [J]. Meat Research, 2023, 37(07): 60-70" ). Chen Linlin used electrochemical sensing to detect veterinary drug residues, which was easy to be contaminated and poisoned, increasing the detection cost and time (see the literature "Chen Linlin, Zhang Jiaxin, Fan Tianjiao, et al. Application of nano-based electrochemical sensors in detection of veterinary drug residues [J]. Journal of Food Safety and Quality Inspection, 2021, 12(18): 7112-7118" ).Therefore, given the limitations of the above detection methods, there is an urgent need to develop accurate and efficient methods for DOX detection. Summary of the Invention

[0004] The purpose of this invention is to provide a molecularly imprinted magnetic covalent organic framework, its preparation method, and its applications, to solve the problems existing in the prior art. This molecularly imprinted magnetic covalent organic framework has the advantages of efficient magnetic separation and high selectivity, enabling simple, stable, and reliable detection of DOX in complex samples.

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

[0006] This invention provides a method for preparing a molecularly imprinted magnetic covalent organic framework for the separation of doxycycline, comprising the following steps:

[0007] After the template molecule and functional monomer undergo a prepolymerization reaction in a porogen, a magnetic metal covalent organic framework is added for a mixed reaction. Then, a crosslinking agent and an initiator are added for a polymerization reaction. After the reaction is completed, the product is separated under the action of an external magnetic field, and the template molecule is eluted to remove it, thus obtaining the molecularly imprinted magnetic covalent organic framework.

[0008] The template molecule is doxycycline.

[0009] Furthermore, the preparation method of the magnetic metal covalent organic framework includes the following steps:

[0010] Fe3O4 powder was dispersed in a solvent, and then p-phenylenediamine and 2,4,6-tricarboxymethyl phloroglucinol were added to carry out a mixed reaction. After the reaction was completed, the product was separated by an external magnetic field to obtain the magnetic metal covalent organic framework.

[0011] Furthermore, the solvent is dimethyl sulfoxide.

[0012] Furthermore, the functional monomer is 4-vinylpyridine.

[0013] Furthermore, the pore-forming agent is ethanol.

[0014] Further, the crosslinking agent is ethylene glycol dimethacrylate; and / or

[0015] The initiator is azobisisobutyronitrile.

[0016] Furthermore, the mass ratio of the template molecule, the functional monomer, and the crosslinking agent is 1:5:30.

[0017] The present invention also provides a molecularly imprinted magnetic covalent organic framework prepared according to the above preparation method.

[0018] The present invention also provides the application of the above-described molecularly imprinted magnetic covalent organic framework in the separation of doxycycline.

[0019] The present invention also provides a method for separating doxycycline, comprising the step of adsorbing and separating doxycycline in a sample to be tested using the above-described molecularly imprinted magnetic covalent organic framework.

[0020] The present invention discloses the following technical effects:

[0021] This invention utilizes surface molecular imprinting technology with molecularly modified carbon (MCOFs) as the supporting material to successfully prepare novel MCOFs@MIPs nanomaterials. These MCOFs@MIPs nanomaterials exhibit considerable saturation magnetization and excellent thermal stability. Static adsorption, kinetic adsorption, and selectivity experiments demonstrate that this material exhibits excellent adsorption capacity, rapid binding kinetics, and significant selectivity for the target analyte DOX. Furthermore, six batches of MCOFs@MIPs samples showed good reproducibility, with a relative standard deviation (RSD) of less than 6.5%. Leveraging the advantages of efficient magnetic separation and high selectivity of MCOFs@MIPs, DOX in complex samples can be detected simply, stably, and reliably. In summary, this invention provides a promising candidate material and a practical analytical strategy for the selective detection of trace DOX in food matrices. Attached Figure Description

[0022] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0023] Figure 1 A schematic diagram of the synthesis of MCOFs@MIPs;

[0024] Figure 2 SEM image of Fe3O4;

[0025] Figure 3 FT-IR images of different materials; where a: Fe3O4; b: MCOFs prepared by reaction in a reactor for 48 h; c: MCOFs prepared by reaction in a reactor for 24 h; d: MCOFs prepared by stirring in an oil bath for 30 min;

[0026] Figure 4 TEM images of MCOFs;

[0027] Figure 5 The image shows the UV spectrum of the prepolymerization system.

[0028] Figure 6TEM images of MCOFs@MIPs;

[0029] Figure 7 EDS plot of MCOFs@MIPs;

[0030] Figure 8 FT-IR plot of MCOFs@MIPs;

[0031] Figure 9 VSM diagram of MCOFs@MIPs;

[0032] Figure 10 XRD plot of MCOFs@MIPs;

[0033] Figure 11 Thermal stability analysis diagram of MCOFs@MIPs;

[0034] Figure 12 The graph shows the detection results of the static adsorption experiment.

[0035] Figure 13 The results of fitting and analyzing the experimental data using an isothermal adsorption model are shown in the figure.

[0036] Figure 14 The graph shows the detection results of the dynamic adsorption experiment.

[0037] Figure 15 The figure shows the results of fitting the experimental data using a kinetic model.

[0038] Figure 16 The structural formulas of five antibiotics;

[0039] Figure 17 A statistical chart showing the selective adsorption of five antibiotics by MCOFs@MIPs and MCOFs@NIPs;

[0040] Figure 18 Selective adsorption chromatograms of five antibiotics by MCOFs@MIPs and MCOFs@NIPs; where (a): initial concentration of the five antibiotics; (b): supernatant after adsorption by MCOFs@NIPs; (c): supernatant after adsorption by MCOFs@MIPs.

[0041] Figure 19 Chromatograms of pork samples analyzed by MCOFs@MIPs material; where (a): DOX standard solution; (b): extract of blank pork sample; (c): initial extract of DOX-spiked pork sample; (d): residual liquid of DOX-spiked pork sample after adsorption by MCOFs@MIPs; (e): MCOFs@MIPs eluent. Detailed Implementation

[0042] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0043] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0044] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0045] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0046] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0047] Example 1

[0048] 1. Experimental Materials

[0049] The main reagents used in this embodiment are shown in Table 1.

[0050] Table 1 Main experimental reagents

[0051] 2. Experimental Methods

[0052] 2.1 Preparation of Molecularly Imprinted Polymers MCOFs@MIPs

[0053] like Figure 1 As shown, the specific steps for synthesizing MCOFs@MIPs are as follows:

[0054] 2.1.1 Preparation of Fe3O4

[0055] 5.4 g of ferric chloride hexahydrate and 160 mL of ethylene glycol were added to a 250 mL flask and stirred until clear. 12.4 g of anhydrous sodium acetate and 1.6 g of sodium citrate were added and dissolved by sonication. The mixture was stirred in an oil bath at 150 °C for 30 min, then transferred to a reaction vessel and reacted in an oven at 200 °C for 6 h. After the reaction was complete, the product was separated under an external magnetic field, washed eight times alternately with anhydrous ethanol and water, and dried in a vacuum drying oven at 60 °C to obtain Fe3O4 powder.

[0056] 2.1.2 Preparation of Magnetic Metal-Covalent Organic Frameworks (MCOFs)

[0057] 150.0 mg of Fe3O4 powder was placed in a 250 mL flask, 40 mL of dimethyl sulfoxide was added and ultrasonically dispersed, 63.0 mg of TP and 48.6 mg of PA were added and ultrasonically dissolved, 2 mL of glacial acetic acid was slowly added dropwise, and the mixture was stirred in an oil bath at 100 °C for 30 min. After the reaction was completed, the product was separated under the action of an external magnetic field, washed 8 times alternately with anhydrous ethanol and tetrahydrofuran, and dried in a vacuum drying oven at 60 °C to obtain MCOFs.

[0058] 2.1.3 Preparation of Molecularly Imprinted Magnetic Covalent Organic Frameworks (MCOFs@MIPs)

[0059] 48.09 mg of DOX powder and 52.57 mg of 4-VP were dissolved in 50 mL of ethanol (porogen) and prepolymerized at room temperature for 2 h. 50 mg of MCOFs were added to a constant-temperature shaker (30 °C, 180 rpm) and reacted for 1 h. Then, 0.6 g of crosslinking agent EGDMA and 10.0 mg of initiator AIBN were added, and the reaction was carried out under N2 protection at 50 °C for 6 h and then at 60 °C for 18 h. After the reaction, the product was separated under an external magnetic field, washed five times alternately with ethanol and water, dried at 60 °C, and extracted with a methanol-glacial acetic acid (9 / 1, v / v) solution using a Soxhlet extract. The template molecules were eluted for 24 h, washed five times with methanol, and dried at 60 °C to obtain MCOFs@MIPs.

[0060] 2.1.4 Preparation of control MCOFs@NIPs

[0061] Without adding template molecules (i.e., DOX powder), the experimental steps in 2.1.3 were repeated to prepare control polymers MCOFs@NIPs.

[0062] 2.1.5 Optimization of preparation method

[0063] Based on section 2.1.3, adjustments were made to the functional monomers, porogens, and preparation ratios to prepare the MCOFs@MIPs shown in Table 2. The molar ratio of the bifunctional monomers was 1:1.

[0064] Table 2 Optimization of MCOFs@MIPs

[0065] 2.2 Test methods for adsorption performance

[0066] The prepared MCOFs@MIPs were added to a DOX standard solution prepared with an adsorption solvent, and the mixture was then shaken on a shaker at 200 rpm at room temperature for 4 h. After shaking, the supernatant was collected under an external magnetic field, and its peak area was determined by high-performance liquid chromatography (HPLC).

[0067] The adsorption capacity of DOX by MCOFs@MIPs at adsorption equilibrium was calculated using data obtained from high-performance liquid chromatography (HPLC). The calculation formula is as follows: ;

[0068] Where Q (μg / mg) is the adsorption capacity of the material at adsorption equilibrium, V is the volume (mL) of standard DOX solution added, m is the actual mass (mg) of MCOFs@MIPs added, and C0 and C2 are also present. e : Initial concentration (μg / mL) and adsorption equilibrium concentration (μg / mL) of DOX standard solution.

[0069] High performance liquid chromatography conditions:

[0070] The adsorption performance analysis of the samples was performed on an Agilent 1260 Infinity II DAD detector. The chromatographic conditions were as follows: TC-C18 (2) column (5 μm, 4.6 mm × 250 mm), column temperature: 35℃. Mobile phase A was 15 mM ammonium acetate-10 mM disodium ethylenediaminetetraacetate-triethylamine (AA-EDTA-TEA, 100 / 10 / 1, v / v / v), pH adjusted to 8.5 with glacial acetic acid. Mobile phase B was acetonitrile, A:B = 80:20, flow rate: 1.0 mL / min. Injection volume: 10 μL. Ultraviolet detector, detection wavelength: 280 nm.

[0071] 3. Results and Discussion

[0072] 3.1 Preparation of MCOFs

[0073] 3.1.1 Scanning electron microscopy (SEM) characterization of Fe3O4

[0074] The scanning electron microscopy results of Fe3O4 are as follows: Figure 2 As shown. In Figure 2The Fe3O4 particles are clearly spherical and densely distributed, present in large quantities within the field of view. The particle size is relatively uniform, with no obvious abnormal distribution of large or small particles.

[0075] 3.1.2 Preparation of MCOFs

[0076] To obtain optimal MCOFs, this invention investigated the preparation of MCOFs using three different methods. The other two methods are similar to the MCOF preparation process described in 2.1.2, except that the oil bath stirring for 30 min in 2.1.2 is replaced with a reaction in a reactor at 120°C for 24 h or 48 h. Characterization was performed using infrared spectroscopy, such as... Figure 3 As shown.

[0077] In the infrared spectrum, Fe3O4 can be observed at 596 cm⁻¹. -1 There is an absorption peak at ( Figure 3 (a) shows the stretching vibration peak of Fe-O, confirming the successful synthesis of Fe3O4. When COFs are coated on the surface of Fe3O4, from... Figure 3 Images b, c, and d clearly show that the MCOFs prepared by the three methods have a lower viscosity at 596 cm⁻¹. -1 The Fe-O absorption peak is still present, further indicating that MCOFs have a magnetic core. The C=O peak (1609 cm⁻¹) is also present. -1 ) and C=C (1583 cm) -1 The stretching vibration bands merged, resulting in peak broadening and the appearance of a shoulder peak. Additionally, at 1400 cm⁻¹... -1 The C=C stretching vibration peak and 1261 cm -1 The CN stretching vibration is attributed to the keto-enol tautomerism of MCOFs. These absorption peaks collectively indicate successful MCOF coating. Comparative analysis revealed that oil bath stirring resulted in a characteristic peak at 1400 cm⁻¹. -1 The C=C absorption peak response is stronger at this point, and higher production yields can be achieved. Furthermore, the preparation time with oil bath stirring is shorter, helping to save time and costs. Based on these advantages, this invention decided to use an oil bath stirring reaction to prepare MCOFs in subsequent experiments.

[0078] Transmission electron microscopy (TEM) results of MCOFs prepared by oil bath stirring reaction are as follows: Figure 4 As shown. In Figure 4 It can be clearly observed that the MCOFs have a core-shell structure, with a relatively uniform thickness distribution, complete morphology, and complete coating, indicating that the COFs layer has been successfully coated on the Fe3O4 surface.

[0079] 3.2 Preparation of MCOFs@MIPs

[0080] 3.2.1 Selection of Functional Units

[0081] The specific recognition ability of molecularly imprinted polymers is largely related to the magnitude of the interaction forces during the pre-assembly of template molecules and functional monomers. Therefore, the selection of functional monomers is crucial. This invention selected five functional monomers—MAA, AM, 4-VP, [COOHpvim]Br, and [VAFMIM]Cl—to evaluate the adsorption performance of MCOFs@MIPs for DOX. Table 3 shows that, under the same preparation conditions, MCOFs@MIPs prepared with MAA and 4-VP as functional monomers exhibited the highest adsorption capacities, at 7.74 and 7.53 μg / mg, respectively. Therefore, the adsorption capacities of polymers prepared with MAA and 4-VP as functional monomers will be further investigated.

[0082] Table 3 Selection of Functional Monomers

[0083]

[0084] Note: The template is DOX, the porogen is ethanol, the crosslinking agent is EGDMA, the initiator is AIBN, and the molar ratio of template:functional monomer:crosslinking agent is 1:5:25.

[0085] To further investigate the functional monomers and attempt to improve the adsorption capacity of the prepared MCOFs@MIPs, bifunctional monomers were used (Table 4). The adsorption capacity of MCOFs@MIPs prepared using bifunctional monomers decreased significantly, and was lower than that of their monomers.

[0086] Table 4 Selection of Bifunctional Monomers

[0087]

[0088] Note: The template is DOX, the porogen is ethanol, the crosslinking agent is EGDMA, the initiator is AIBN, the molar ratio of template:functional monomer:crosslinking agent is 1:5:25, and the molar ratio of bifunctional monomer is 1:1.

[0089] At the same concentration, the UV detection images of different functional monomers prepolymerized with template DOX for 2 h are shown below. Figure 5 As shown, it can be observed that when the functional monomer is 4-VP, its interaction with DOX shows the most significant change, with a significant increase in absorbance. This indicates that the new complex formed by 4-VP and DOX has the strongest stability, proving that 4-VP is more suitable as the functional monomer of this polymer.

[0090] 3.2.2 Investigation of Adsorbed Solvents

[0091] To obtain the optimal adsorption capacity and chromatogram, this invention investigated the enrichment of DOX using methanol and water / acetonitrile (8 / 2, v / v) as adsorption solvents (Table 5). It can be clearly seen that, under the same preparation conditions, the adsorption capacity generally increases when the adsorption solvent is water / acetonitrile (8 / 2, v / v), and the adsorption capacity reaches 10.01 μg / mg when the functional monomer is 4-VP, making it the optimal functional monomer. In contrast, the adsorption capacity of MCOFs@MIPs prepared with MAA decreases, indicating that the prepared material is unstable, further proving that 4-VP is the optimal functional monomer.

[0092] Table 5. Investigation of Adsorption Solvents

[0093]

[0094] Note: The template is DOX, the porogen is ethanol, the crosslinking agent is EGDMA, the initiator is AIBN, and the molar ratio of template:functional monomer:crosslinking agent is 1:5:25.

[0095] 3.2.3 Selection of porogen

[0096] The porogen serves as the site for the prepolymerization of the template and functional monomer. Different porogens have varying effects on the pore volume, specific surface area, and average pore size of the prepolymer; therefore, selecting a suitable porogen is crucial. This invention investigated the synthesis of MCOFs@MIPs using 4-VP as the functional monomer and different porogens (Table 6). Under the same preparation conditions, ethanol was used as the porogen, resulting in the highest adsorption capacity of 10.01 μg / mg. Therefore, ethanol was selected as the porogen for subsequent investigations.

[0097] Table 6 Investigation of pore-forming agents

[0098]

[0099] Note: The template is DOX, the functional monomer is 4-VP, the crosslinking agent is EGDMA, the initiator is AIBN, and the molar ratio of template:functional monomer:crosslinking agent is 1:5:25.

[0100] 3.2.4 Investigation of ionic liquids as functional monomers and porogens

[0101] MCOFs@MIPs were prepared using [COOHpvim]Br and [VAFMIM]Cl as functional monomers and ethanol + [BmIm]PF6 as a porogen. As shown in Table 7, the adsorption capacities were 3.50 and 3.51 μg / mg, respectively, both lower than those prepared with 4-VP. Therefore, using 4-VP as the functional monomer and ethanol as the porogen is the optimal method for preparing MCOFs@MIPs.

[0102] Table 7. Investigation of ionic liquids as functional monomers and porogens

[0103]

[0104] Note: The template is DOX, the crosslinking agent is EGDMA, the initiator is AIBN, and the molar ratio of template:functional monomer:crosslinking agent is 1:5:25.

[0105] 3.2.5 Investigation into the ratio of template to functional monomer and crosslinking agent

[0106] Through the above experiments, this invention determined that 4-VP is the optimal functional monomer. Based on this, this invention further investigated the effect of different ratios of template, functional monomer, and crosslinking agent on DOX adsorption capacity. The results are shown in Table 8. When the ratio was set to 1:5:30, the adsorption effect was optimal, reaching 19.44 μg / mg. Therefore, this invention will use this ratio for further in-depth research in the subsequent preparation of MCOFs@MIPs.

[0107] Table 8. Investigation of the ratio of template to functional monomer and crosslinking agent

[0108]

[0109] Note: The porogen is ethanol, and the initiator is AIBN.

[0110] 3.2.6 Characterization of MCOFs@MIPs

[0111] To further evaluate MCOFs@MIPs, TEM morphology analysis was performed. TEM images of some of the prepared MCOFs@MIPs are shown in section 2.1.3. Figure 6 .from Figure 6 As can be observed, a relatively light-colored coating layer exists on the surface of the microspheres, indicating that a thin imprint layer has been successfully coated on the surface of MCOFs@MIPs. Furthermore, energy dispersive spectroscopy (EDS) analysis based on TEM images shows that the Fe element content in the material decreases, while the O, C, and N element contents increase (…). Figure 7 These results further confirm the successful preparation of the imprinted layer.

[0112] FT-IR characterization revealed that, in addition to the characteristic absorption peaks of MCOFs mentioned above, the MCOFs@MIPs polymer also exhibited a peak at 1722 cm⁻¹. - An absorption peak appears at position ¹, which is attributed to the C=O bond in EGDMA ( Figure 8 The results show that MCOFs@MIPs were successfully prepared.

[0113] The hysteresis properties of the samples were characterized using a vibrating sample magnetometer (VSM). Figure 9As can be seen, the saturation magnetizations of Fe3O4, MCOFs, and MCOFs@MIPs are 60.5, 34.3, and 32.9 emu / g, respectively. This difference can be attributed to the magnetic shielding effect of the sequentially deposited COFs and MIPs layers on the Fe3O4 surface. Despite the reduced saturation magnetization, MCOFs@MIPs still maintain excellent magnetic response performance. Figure 9 As shown in the inset, under the influence of an external magnetic field, MCOFs@MIPs can achieve magnetic separation from a completely dispersed solution within 30 s, indicating their potential for rapid separation and recycling in future applications.

[0114] The crystal structure of the polymer was characterized using X-ray diffraction (XRD), and the results are as follows: Figure 10 As shown, distinct diffraction peaks were clearly observed at 2θ = 30.2° (220), 35.4° (311), 43.1° (400), 54.2° (442), 57.4° (511), and 62.6° (440), which are consistent with the standard crystal data of Fe3O4. Furthermore, both MCOFs and MCOFs@MIPs retained all the characteristic diffraction peaks of the original Fe3O4, and no new crystal phases were detected. This result indicates that the subsequent grafting of MIPs layers did not alter its crystal structure.

[0115] The thermal stability of the polymer was characterized using TGA. The results showed that the Fe3O4 nanoparticles experienced a mass loss of 11.83%, likely due to the evaporation of bound water within the material. For the MCOFs curve, an additional mass loss of approximately 34.22% was observed, primarily attributed to the pyrolysis of the COFs layer. Furthermore, the degradation percentage of MCOFs@MIPs increased by 1.60% compared to MCOFs, indicating that the MIPs were successfully coated onto the MCOFs surface. Additionally, as... Figure 11 As shown, MCOFs@MIPs maintain excellent stability at temperatures below 200°C, ensuring reliable application at high temperatures.

[0116] Example 2

[0117] Taking the MCOFs@MIPs prepared in section 2.1.3 of Example 1 as an example (with the MCOFs@NIPs prepared in section 2.1.4 as a control), the adsorption performance was investigated.

[0118] 1. Experimental Methods

[0119] 1.1 Static Adsorption

[0120] The prepared MCOFs@MIPs and MCOFs@NIPs were added to a series of DOX standard solutions, and the mixtures were then shaken on a shaker at 200 rpm at room temperature for 4 h. After shaking, the supernatant was collected under an external magnetic field, and its peak area was determined by HPLC. The adsorption capacity calculation formula is the same as in section 2.2 of Example 1.

[0121] Next, the static adsorption processes of MCOFs@MIPs and MCOFs@NIPs were mathematically fitted using the Langmuir and Freundlich isotherm adsorption equations. The calculation formulas are as follows:

[0122] Langmuir isothermal adsorption equation: ;

[0123] Freundlich isothermal adsorption equation: ;

[0124] In the formula, Q max (mg / g) is the maximum adsorption capacity calculated after fitting, K L (mL / mg) and K F (mg / g) are the Langmuir and Freundlich adsorption equilibrium constants, respectively, where m represents the Freundlich exponent of heterogeneity.

[0125] 1.2 Dynamic Adsorption

[0126] MCOFs@MIPs and MCOFs@NIPs were added to 3 mL of 100 μg / mL DOX standard solution, respectively, and the mixture was shaken at 200 rpm for 10, 20, 40, 60, 80, 100, 120, 150, 180, 210, and 240 min. After the reaction was completed, the solution and adsorbent were quickly separated from the adsorbent using a magnet, and the supernatant was collected. Finally, the peak area was determined by HPLC, and the adsorption capacity was calculated.

[0127] Subsequently, the dynamic adsorption process of MCOFs@MIPs and MCOFs@NIPs was mathematically fitted using pseudo-first-order and pseudo-second-order kinetic equations, and the calculation formulas are as follows:

[0128] Pseudo-first-order dynamic equation: ;

[0129] Pseudo-second-order dynamic equation: ;

[0130] In the formula, Q t (mg / g) and Q e(mg / g) represents the adsorption amount after equilibrium and at time t (min), respectively. v0 is the initial adsorption rate (mg / g / min), and K1 ( / min) and K2 (g / mg / min) are the equilibrium rate constants of the pseudo-first-order and pseudo-second-order kinetic equations, respectively.

[0131] 1.3 Selective Adsorption

[0132] To evaluate the selective adsorption performance of MCOFs@MIPs and MCOFs@NIPs, tigecycline (TGC) and oxytetracycline (OTC) were selected as structural analogs, and norfloxacin (NOR) and chloramphenicol (CHL) were selected as reference analogs. 10 mg of the polymer was added to 3 mL of a mixed solution containing DOX and the four analogs (each substance concentration was 100 µg / mL). The mixture was incubated at 30 °C with shaking for 210 min. The supernatant was separated by an external magnetic field and analyzed by HPLC. The adsorption capacity of the polymer for the target analyte was determined according to section 2.2 of Example 1, and the imprinting factor (IF) was calculated using the following formula:

[0133] Imprint factor (IF) = Q MIPs / Q NIPs ;

[0134] In the formula, Q MIPs and Q NIPs The adsorption amounts of the target analytes by MCOFs@MIPs and MCOFs@NIPs are respectively.

[0135] 1.4 Reproducibility Experiment

[0136] To assess the reproducibility of the polymer, six batches of 10.0 mg imprinted polymer were weighed, and after adsorption of DOX, the supernatant was obtained under an external magnetic field. Finally, the peak area was determined by HPLC, and the adsorption capacity was calculated.

[0137] 2. Experimental Results

[0138] 2.1 Static Adsorption

[0139] Static adsorption experiments were conducted on MCOFs@MIPs and MCOFs@NIPs by varying the adsorption concentration. Figure 12 As shown, the adsorption capacity of DOX by MCOFs@MIPs increases with increasing concentration, reaching adsorption equilibrium at 100 µg / mL; while the adsorption capacity of MCOFs@NIPs is lower and the variation is minimal, reaching equilibrium at 80 µg / mL. These results indicate that the surface of MCOFs@MIPs possesses abundant specific recognition sites, enabling it to exhibit excellent specific recognition ability for target analytes.

[0140] To further elucidate the adsorption mechanisms of MCOFs@MIPs and MCOFs@NIPs, the Langmuir and Freundlich isothermal adsorption models were used to fit and analyze the experimental data. Figure 13 As shown, the correlation coefficient (R0) of the Langmuir isotherm model between MCOFs@MIPs and MCOFs@NIPs is shown. 2 The correlation coefficients were 0.9918 and 0.9973, respectively, both higher than the correlation coefficients of the Freundlich isotherm model, indicating that the adsorption process of both polymers follows the monolayer homogeneous adsorption mechanism.

[0141] 2.2 Dynamic Adsorption

[0142] like Figure 14 The adsorption capacities of MCOFs@MIPs and MCOFs@NIPs for DOX were demonstrated. The results showed that the adsorption capacity of MCOFs@MIPs increased more slowly, reaching adsorption equilibrium at 210 min with a maximum adsorption capacity of 20.0 mg / g; while MCOFs@NIPs reached adsorption equilibrium at 180 min, and its maximum adsorption capacity was significantly lower than that of MCOFs@MIPs. These results indicate that MCOFs@MIPs have a longer equilibrium time and a higher adsorption capacity, a phenomenon that can be attributed to the abundant specific recognition sites on its surface.

[0143] To further investigate the adsorption kinetics of MCOFs@MIPs and MCOFs@NIPs, pseudo-first-order and pseudo-second-order kinetic models were used to fit and analyze the experimental data. Details of the fitting curves and corresponding parameters for the pseudo-first-order kinetic model are shown below. Figure 15 As shown in the figure. The comparative analysis results show that the pseudo-first-order kinetic model can most accurately describe the adsorption behavior of MCOFs@MIPs (R²≥0.9936), indicating that the adsorption process is mainly dominated by the physical adsorption mechanism.

[0144] 2.3 Selective adsorption:

[0145] To evaluate the specific adsorption performance of MCOFs@MIPs for DOX, a competitive adsorption experiment was conducted using four antibiotics: tigecycline (TGC), oxytetracycline (OTC), norfloxacin (NOR), doxycycline (DOX), and chloramphenicol (CHL). The structural formulas of these antibiotics are shown below. Figure 16 As shown in the figure. The adsorption capacity of MCOFs@MIPs for structural analogs and the chromatograms of the samples after adsorption are shown in the figure. Figure 17 and Figure 18As shown in the figure. The results indicate that MCOFs@MIPs exhibit a significantly higher adsorption capacity for DOX compared to other competitors. This phenomenon is attributed to the spatial structural matching between the imprinted layer and DOX. MCOFs@NIPs, on the other hand, show lower selectivity for DOX, and their adsorption capacity for various structural analogs does not exhibit a clear regularity. Further analysis using the imprinting factor (IF) and partition coefficient (K) further... d The selectivity of MCOFs@MIPs was evaluated using the selectivity coefficient (K) and relative selectivity coefficient (K′) (Table 9). The IF values ​​of MCOFs@MIPs for TGC, OTC, NOR, DOX, and CHL were 0.78, 8.73, 0.89, 4.35, and 0.44, respectively, confirming its excellent selectivity for DOX. The relatively high IF value for OTC is due to the negligible adsorption capacity of MCOFs@MIPs for this compound. In addition, the partition coefficient K′ for DOX... d The K′ value (0.5) is greater than that of other analytes, indicating that MCOFs@MIPs have better separation performance for DOX. The relative selectivity coefficient K′ results show that, except for OTC, the K′ values ​​for all other analytes are greater than 1. This finding indicates that, compared with MCOFs@NIPs, MCOFs@MIPs have higher binding performance and selectivity for the target analyte DOX, and compared with other structural analogs, it has better specific recognition of DOX. The abnormal K′ value of OTC is consistent with the aforementioned abnormal IF value. In summary, these data confirm that the imprinting effect of MCOFs@MIPs enables it to selectively separate template molecules from complex real-world samples.

[0146] Table 9 Selectivity Evaluation Results of MCOFs@MIPs

[0147]

[0148] 2.4 Reproducibility Experiment

[0149] The reproducibility of MCOFs@MIPs was investigated by preparing six independent batches of polymer, with each batch subjected to three parallel adsorption experiments for DOX. The results showed that the average adsorption capacity of MCOFs@MIPs for DOX was 19.91 mg / g, with a relative standard deviation (RSD) of less than 6.5%. These results indicate that MCOFs@MIPs exhibits good reproducibility.

[0150] Example 3

[0151] Pork was weighed and DOX standard solution was added, and the mixture was allowed to stand at 4°C for 12 h. Subsequently, 1% acetic acid-acetonitrile solution (v / v) was added to the spiked sample for ultrasonic extraction, and the supernatant was collected by centrifugation. This extraction process was repeated three times, and all supernatants were combined and concentrated using a rotary evaporator at 45°C. The concentrated residue was transferred with acetonitrile, dried under nitrogen purging, and then dissolved in an acetonitrile-water mixture (v / v 2:8). The solution was extracted using MCOFs@MIPs-17 from Example 1. The adsorbent was then separated under an external magnetic field and eluted with a methanol-acetic acid mixture (v / v 9:1) using ultrasound. After the eluent was dried under nitrogen purging, the residue was reconstituted with the solvent and analyzed by HPLC.

[0152] To verify the practicality of the novel MCOFs@MIPs material in actual sample analysis, it was applied to the determination of DOX in pork samples. The chromatogram is shown below. Figure 19 As shown. According to Figure 19 As can be seen, no endogenous DOX was detected in the blank pork sample. After treatment with MCOFs@MIPs, the peak area of ​​DOX in the residual liquid of the spiked sample was significantly reduced compared with the initial spiked extract. In addition, the peak area of ​​DOX in the MCOFs@MIPs eluent was significantly enhanced, with an enrichment factor of 30. These results confirm that the MCOFs@MIPs nanomaterials prepared in this invention can efficiently and conveniently enrich and separate DOX from complex pork samples.

[0153] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A method for preparing a molecularly imprinted magnetic covalent organic framework for the separation of doxycycline, characterized in that, Includes the following steps: After the template molecule and functional monomer undergo a prepolymerization reaction in a porogen, a magnetic metal covalent organic framework is added for a mixed reaction. Then, a crosslinking agent and an initiator are added for a polymerization reaction. After the reaction is completed, the product is separated under the action of an external magnetic field, and the template molecule is eluted to remove it, thus obtaining the molecularly imprinted magnetic covalent organic framework. The template molecule is doxycycline.

2. The preparation method according to claim 1, characterized in that, The method for preparing the magnetic metal covalent organic framework includes the following steps: Fe3O4 powder was dispersed in a solvent, and then p-phenylenediamine and 2,4,6-tricarboxymethyl phloroglucinol were added to carry out a mixed reaction. After the reaction was completed, the product was separated by an external magnetic field to obtain the magnetic metal covalent organic framework.

3. The preparation method according to claim 2, characterized in that, The solvent is dimethyl sulfoxide.

4. The preparation method according to claim 1, characterized in that, The functional monomer is 4-vinylpyridine.

5. The preparation method according to claim 1, characterized in that, The porogen is ethanol.

6. The preparation method according to claim 1, characterized in that, The crosslinking agent is ethylene glycol dimethacrylate; and / or The initiator is azobisisobutyronitrile.

7. The preparation method according to claim 1, characterized in that, The mass ratio of the template molecule, the functional monomer, and the crosslinking agent is 1:5:

30.

8. A molecularly imprinted magnetic covalent organic framework prepared by the preparation method according to any one of claims 1-7.

9. The application of a molecularly imprinted magnetic covalent organic framework as described in claim 8 in the separation of doxycycline.

10. A method for separating doxycycline, characterized in that, The method includes the step of adsorbing and separating doxycycline from the sample to be tested using the molecularly imprinted magnetic covalent organic framework as described in claim 8.

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