Magnetic composites and their use in rapid detection of mycotoxins in grains

Abstract

This invention discloses a magnetic composite material and its application in the rapid detection of mycotoxins in grains, belonging to the field of mycotoxin detection technology. The magnetic composite material is prepared by the following method: FeCl3·6H2O and FeSO4·7H2O are dissolved in water, concentrated hydrochloric acid is added to prevent hydrolysis, and the solution is added dropwise to a NaOH solution preheated to 80°C. After stirring and reacting, sodium citrate is added for modification, followed by magnetic separation and washing, and drying to obtain Fe3O4 nanoparticles. A metal-organic framework material (UiO-66) is then prepared. The Fe3O4 nanoparticles, ionic liquid, UiO-66, and chitosan are dissolved in acetic acid, and then added dropwise to an alkaline solution using a syringe to form a hydrogel. This invention provides a magnetic composite material with excellent adsorption properties and a high specific surface area, achieving simultaneous detection and enrichment of various mycotoxins with high selectivity and high affinity. It exhibits high detection sensitivity, is suitable for trace analysis, has a wide range of applicable matrices, and demonstrates strong method robustness.

Description

Technical Field

[0001] This invention relates to the field of mycotoxin detection technology, and in particular to a magnetic composite material and its application in the rapid detection of mycotoxins in grains. Background Technology

[0002] Mycotoxins are common harmful contaminants in cereal foods, such as aflatoxin (AFTB1, AFTB2, AFTG1, AFTG2), deoxynivalenol (DON), zearalenone (ZEN), and ochratoxin A (OTA). These toxins possess strong carcinogenic, teratogenic, and mutagenic properties, seriously threatening human health and food security. Currently, the detection of mycotoxins mainly relies on thin-layer chromatography (TLC), enzyme-linked immunosorbent assay (ELISA), high-performance liquid chromatography (HPLC), and their coupling with mass spectrometry (such as HPLC-MS / MS). However, traditional methods have the following problems: (1) Cumbersome sample pretreatment: Traditional solid phase extraction (SPE) requires column packing, activation, and equilibration, which is complicated and time-consuming. (2) High material cost: Immunoaffinity columns rely on imports, are expensive, and are difficult to use on a large scale; (3) Limitations of single toxin detection: Most methods can only detect one or a few toxins, which is difficult to meet the needs of simultaneous screening of multiple toxins; (4) High consumption of organic solvents: A large amount of organic solvents are used in the pretreatment process, which does not meet the requirements of green analysis.

[0003] In recent years, magnetic solid-phase extraction (MSPE) technology has shown great promise in sample pretreatment due to its advantages such as simple operation, easy separation of adsorbents, and ability to be functionalized. Ionic liquids (ILs) and metal-organic frameworks (MOFs) are often used to construct high-performance adsorbent materials due to their tunable structure, large specific surface area, and rich functional groups. Chitosan (CS), as a natural polymer, has advantages such as good biocompatibility, biodegradability, and rich amino and hydroxyl groups, making it an ideal matrix for constructing green adsorbent materials.

[0004] However, existing magnetic composite materials still suffer from problems such as small specific surface area, limited adsorption sites, and insufficient ability to simultaneously recognize multiple toxins. Therefore, there is an urgent need to develop a magnetic composite material and its supporting detection method that combines high adsorption capacity, simultaneous recognition of multiple toxins, simple operation, and low cost. Summary of the Invention

[0005] This invention provides a magnetic composite material and its application in the rapid detection of mycotoxins in grains, in order to solve the problems of small specific surface area, limited adsorption sites, and insufficient ability to simultaneously identify multiple toxins in existing magnetic composite materials.

[0006] To solve the above-mentioned technical problems, the technical solution of the present invention is as follows: A magnetic composite material is prepared by the following method: (1) Preparation of Fe3O4 nanoparticles FeCl3·6H2O and FeSO4·7H2O were dissolved in water, concentrated hydrochloric acid was added to prevent hydrolysis, and the solution was added dropwise to NaOH solution preheated to 80℃. After stirring and reacting, sodium citrate was added for modification, followed by magnetic separation and washing, and drying to obtain Fe3O4 nanoparticles. (2) Preparation of UiO-66 Zirconium tetrachloride and 2-aminoterephthalic acid were dissolved in N,N-dimethylformamide (DMF), and acetic acid and hydrochloric acid were added as modifier and crystallizing agent, respectively. After homogenization, the solution was transferred to a polytetrafluoroethylene autoclave and subjected to a solvothermal reaction under microwave conditions. The product was washed and dried to obtain UiO-66. (3) Preparation of magnetic composite materials Fe3O4 nanoparticles, ionic liquid, UiO-66 and chitosan are dissolved in acetic acid, and then added dropwise to an alkaline solution using a syringe to form a hydrogel.

[0007] In step (1), the molar ratio of FeCl3·6H2O and FeSO4·7H2O is 2:1, and concentrated hydrochloric acid is added to make the pH of the mixed solution less than 7; the total amount of FeCl3·6H2O and FeSO4·7H2O has a molar ratio of 1:1 to sodium citrate.

[0008] In step (2), the molar ratio of zirconium tetrachloride to 2-aminoterephthalic acid is 1:1, the molar ratio of the total amount of zirconium tetrachloride and 2-aminoterephthalic acid to DMF is 1:(450~500); the molar ratio of zirconium tetrachloride to acetic acid is 1:(8~12), and the molar ratio of zirconium tetrachloride to hydrochloric acid is 1:(0.5~2). In step (2), the temperature of the solvothermal reaction is 120°C and the time is 3-4 hours. The product is washed with DMF.

[0009] In step (3), the ionic liquid is prepared by the following method: 0.1 mol of 1-methylimidazole, 0.11 mol of 1-bromohexane and 50 ml of acetonitrile are placed in a three-necked flask and refluxed at 80 °C under nitrogen protection for 24 h. After cooling to room temperature, the acetonitrile is removed by rotary evaporation at 45 °C. The residue is washed three times with ethyl acetate, and the ethyl acetate is removed by rotary evaporation at 55 °C. The residue is then dried under vacuum at 40 °C for 24 h to obtain the ionic liquid.

[0010] In step (3), the mass ratio of Fe3O4 nanoparticles, ionic liquid, UiO-66 and chitosan is 1:(1~2):(0.8~1.2):(2~3), and the alkaline solution is a 2mol / L NaOH aqueous solution.

[0011] The magnetic composite material is used in the rapid detection of mycotoxins in grains by using magnetic solid-phase extraction for sample pretreatment.

[0012] Specifically, the steps include the following: Extraction: The pulverized grain sample was extracted by shaking with a methanol-water mixture; Enrichment: Add magnetic composite material to the extract and shake to adsorb; Separation: Apply an external magnetic field to adsorb the material, then discard the supernatant; Washing: Rinse with water or a low-concentration organic solvent to remove impurities; Elution: Elute with an eluent and collect the eluent; Filtration: Passed through a 0.22 μm organic filter membrane, to be tested.

[0013] The detection method was high performance liquid chromatography-mass spectrometry, and the chromatographic conditions were as follows; Chromatographic column: C18 column (e.g., 2.1 mm × 50 mm, 1.8 μm); Mobile phase: Methanol-water or acetonitrile-water gradient elution; Mass spectrometry conditions: electrospray ionization (ESI), multiple reaction monitoring (MRM) mode; Quantitative methods: external standard method or internal standard method; The gradient elution program is as follows: 0–5 min, 10% methanol / acetonitrile held for 5 min; 5–30 min, methanol / acetonitrile linearly increased from 10% to 95%, held for 5 min.

[0014] The fungal toxins mentioned are aflatoxin (AFTB1, AFTB2, AFTG1, AFTG2), deoxynivalenol (DON), zearalenone (ZEN), ochratoxin A (OTA), fumonisin (FB1, FB2), T-2 toxin, or HT-2 toxin.

[0015] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. Achieved highly selective and high-affinity simultaneous detection and enrichment of multiple mycotoxins. This composite material ingeniously integrates the regular porous structure of UiO-66, the abundant amino / hydroxyl groups of chitosan, the tunable polarity and electrostatic interactions of ionic liquids, and the magnetism of Fe3O4, thereby forming multiple recognition sites including π-π stacking, hydrogen bonding, hydrophobic interactions, electrostatic attraction, and coordination. These sites can simultaneously and efficiently capture and adsorb fungal toxins with different polarities and molecular structures, including aflatoxins (AFTB1, AFTB2, AFTG1, AFTG2), deoxynivalenol (DON), zearalenone (ZEN), ochratoxin A (OTA), fumonisins (FB1, FB2), T-2 toxin, and HT-2 toxin, achieving broad-spectrum and simultaneous pretreatment enrichment.

[0016] 2. Possesses excellent adsorption performance and high specific surface area. By combining UiO-66 (a metal-organic framework material) with chitosan hydrogel, the specific surface area of ​​the material is significantly increased, providing more effective adsorption sites. Simultaneously, the introduction of ionic liquid further modulates the polarity and charge distribution of the material surface, enhancing its affinity for toxins of different polarities, enabling the material to maintain high adsorption capacity and rapid mass transfer rate even in complex grain matrices.

[0017] 3. Simple to operate, time-saving and efficient, greatly simplifying the sample pretreatment process. The Fe3O4 in the material endows it with good magnetic responsiveness, and it can be rapidly separated under an external magnetic field. There is no need for the cumbersome steps of column packing, activation, and rinsing in traditional solid phase extraction. The entire adsorption-elution process can be completed within minutes, which greatly shortens the pretreatment time, reduces the complexity of operation, and is suitable for high-throughput sample analysis.

[0018] 4. Environmentally friendly, in line with the concept of green analytical chemistry. Using chitosan, a biodegradable, non-toxic, and widely available natural polymer, as the matrix not only reduces the burden on the environment but also improves the safety of the material.

[0019] 5. Low cost, easy to mass-produce and reuse The raw materials used, such as FeCl3·6H2O, FeSO4·7H2O, chitosan, and sodium citrate, are all conventional chemicals with low cost. The synthesis process of UiO-66 and ionic liquids is easy to prepare in batches. The material has certain mechanical and chemical stability and can be reused multiple times, which significantly reduces the cost of consumables for a single test.

[0020] 6. High detection sensitivity, suitable for trace analysis Combining magnetic solid-phase extraction with high-performance liquid chromatography-fluorescence detection (HPLC-FLD) significantly improves the method's sensitivity by leveraging the material's excellent enrichment effect. Specific examples demonstrate that this material exhibits good linearity and a low detection limit for aflatoxin detection in the HPLC-FLD system, making it suitable for the precise detection of trace and ultra-trace mycotoxins in grains.

[0021] 7. Applicable to a wide range of substrates, with strong method robustness. This material and method are applicable not only to wheat but can also be extended to the detection of mycotoxins in various grains and their processed products, such as corn, rice, barley, and oats. The material exhibits strong resistance to interference in complex matrices, stable recovery rates, and good RSD control, demonstrating excellent method robustness and practical application potential. 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 description of the embodiments or the prior art 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 The infrared absorption spectrum (FI-IR) of the magnetic composite material prepared in Example 1 of this invention; Figure 2 This is a scanning electron microscope (SEM) image of the magnetic composite material prepared in Example 1 of the present invention. Figure 3 The image shows the energy dispersive spectroscopy (EDS) analysis of the magnetic composite material prepared in Example 1 of this invention. Figure 4 The hysteresis loop diagram is shown for the magnetic composite material obtained in Example 1 of this invention. Figure 5 The infrared absorption spectrum (FI-IR) of the magnetic composite material prepared in Example 4 of this invention; Figure 6 This is a scanning electron microscope (SEM) image of the magnetic composite material prepared in Example 4 of the present invention. Figure 7 The image shows the energy dispersive spectroscopy (EDS) analysis of the magnetic composite material prepared in Example 4 of this invention. Figure 8 The hysteresis loop diagram of the magnetic composite material obtained in Example 4 of the present invention; Figure 9 Chromatograms for rapid detection of mycotoxins in grains; Detailed Implementation

[0024] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0025] In the following embodiments, unless otherwise specified, the raw materials or processing techniques are conventional commercial products or conventional processing techniques in the art. Example 1

[0026] This embodiment provides a magnetic composite material, which is prepared by the following method: ①Preparation of Fe3O4 Weigh 5.4 g FeCl3·6H2O and 2.8 g FeSO4·7H2O into a 50 mL beaker, and add 0.85 mL concentrated HCl (to prevent Fe from being absorbed). 2+ and Fe 3+ Hydrolysis occurs to bring the pH of the mixed solution to 6. Then, 25 mL of deionized water is added, and the mixture is stirred with a glass rod until completely dissolved to obtain a standard iron solution. 250 mL of a 1.5 mol / L NaOH solution (to co-precipitate ferrous and ferric iron to form Fe3O4) is added to a 500 mL three-necked flask. The flask is heated to 80°C in a water bath. Under N2 protection, the prepared iron standard solution is added dropwise to the NaOH solution at a rate of 10 d / min, with mechanical stirring for 1 hour. Heating is then stopped, and the solution is cooled to room temperature. The nitrogen protection device is removed, and 100 mL of a 0.3 mol / L sodium citrate solution is added to a constant pressure funnel. Mechanical stirring continues for 1 hour. After the reaction is complete, the product is first washed with deionized water to neutralize it, then washed with anhydrous ethanol, and finally washed 2-3 times with deionized water. The product is separated and collected using a magnet, the supernatant is discarded, and the final product is freeze-dried.

[0027] ② Preparation of ionic liquids 0.1 mol of 1-methylimidazolium, 0.11 mol of 1-bromohexane, and 50 mL of acetonitrile were placed in a three-necked flask and refluxed at 80 °C for 24 h under nitrogen protection. After cooling to room temperature, the acetonitrile was removed by vacuum distillation at 45 °C. The residue was washed three times with ethyl acetate, and then the ethyl acetate was removed by vacuum distillation at 55 °C. Finally, the residue was dried under vacuum at 40 °C for 24 h to obtain the ionic liquid 1-hexyl-3-methylimidazolium bromide.

[0028] ③Preparation of UiO-66 Zirconium tetrachloride and 2-aminoterephthalic acid were dissolved in 480 molar amounts of DMF (N,N-dimethylformamide) at a molar ratio of 1:1. Acetic acid and hydrochloric acid were added as modifiers and crystallizing agents, respectively, with a molar ratio of acetic acid to zirconium tetrachloride of 1:10 and a molar ratio of hydrochloric acid to zirconium tetrachloride of 1:1. After sonication, the solution was magnetically stirred for 15 min, transferred to a polytetrafluoroethylene autoclave, and then continuously heated to 120 °C under microwave conditions. After reacting for 3.5 h, crystals were obtained, washed with DMF solvent, and finally dried in an oven at 120 °C.

[0029] ④ Preparation of magnetic composite hydrogels The prepared 0.5g Fe3O4, 0.8g ionic liquid, 0.5g UiO-66 and 1.3g chitosan were first dissolved in acetic acid, and then added dropwise to NaOH aqueous solution (2mol / L) with a syringe to form a hydrogel. The hydrogel was washed with deionized water and freeze-dried to obtain the final product.

[0030] The infrared absorption spectrum of the magnetic composite hydrogel prepared in this embodiment is shown below. Figure 1 As shown in the figure, 570 cm -1 A distinct characteristic absorption peak appears at 1100 cm⁻¹, corresponding to the stretching vibration peak of the Fe-O bond in Fe₃O₄, indicating the successful formation of the magnetic core. -1 The nearby absorption peaks are attributed to the stretching vibrations of COC in chitosan. (1400–1650 cm⁻¹) -1 The characteristic peaks within the range correspond to the symmetric and asymmetric stretching vibrations of the carboxyl group in the phthalic acid ligand of NH2-UiO-66, indicating the successful construction of the MOF structure. (1460–1550 cm⁻¹) -1 The absorption peak at the specified location is related to the characteristic vibrations of the imidazole ring in the ionic liquid, indicating that the ionic liquid was successfully introduced into the material surface. The common presence of the above characteristic peaks indicates that Fe3O4, UiO-66, the ionic liquid, and CS were successfully composited, and the magnetic composite material was completed.

[0031] The scanning electron microscope image of the magnetic composite material prepared in this embodiment is shown below. Figure 2 As shown in the figure, the composite material exhibits an irregular spherical structure with a relatively rough surface and an average particle size of approximately 900 μm, indicating that chitosan coating forms a stable composite structure.

[0032] The energy dispersive spectroscopy (EDS) analysis diagram of the magnetic composite material prepared in this embodiment is shown below. Figure 3As shown in the figure, C, O, N, Fe, and Br elements are uniformly distributed in the material. Fe originates from the Fe3O4 magnetic core, while C, N, and O mainly come from chitosan and MOF organic ligands, and Br originates from the ionic liquid anion. This uniform elemental distribution indicates the successful composite formation of Fe3O4, UiO-66, the ionic liquid, and CS, thus completing the construction of the magnetic composite adsorbent.

[0033] The hysteresis loop of the magnetic composite material prepared in this embodiment is as follows: Figure 4 As shown in the figure, its saturation magnetization value is approximately 15.0 emu / g, which indicates that the adsorbent has sufficient magnetic response to meet the requirements of magnetic separation (generally, a saturation magnetization value greater than 10 emu / g is considered "sufficient for magnetic separation"). Example 2

[0034] This embodiment provides a magnetic composite material, which is prepared by the following method: ①Preparation of Fe3O4 Weigh 5.4 g FeCl3·6H2O and 2.8 g FeSO4·7H2O into a 50 mL beaker, and add 0.85 mL concentrated HCl (to prevent Fe from being absorbed). 2+ and Fe 3+ Hydrolysis occurs, bringing the pH of the mixed solution to 6. Then, 25 mL of deionized water is added, and the mixture is stirred with a glass rod until completely dissolved, yielding a standard iron solution. 250 mL of a 1.5 mol / L NaOH solution (to co-precipitate ferrous and ferric iron to form Fe3O4) is added to a 500 mL three-necked flask. The flask is heated to 80°C in a water bath. Under N2 protection, the prepared iron standard solution is added dropwise to the NaOH solution at a rate of 10 d / min, with mechanical stirring for 1 hour. Heating is then stopped, and the solution is cooled to room temperature. The nitrogen protection device is removed, and 100 mL of a 0.3 mol / L sodium citrate solution is added to a constant pressure funnel. Mechanical stirring continues for 1 hour. After the reaction is complete, the product is first washed with deionized water to neutralize, then washed with anhydrous ethanol, and finally washed 2-3 times with deionized water. The product is separated and collected using a magnet, the supernatant is discarded, and the final product is freeze-dried.

[0035] ② Preparation of ionic liquids 0.1 mol of 1-methylimidazolium, 0.11 mol of 1-bromohexane, and 50 mL of acetonitrile were placed in a three-necked flask and refluxed at 80 °C for 24 h under nitrogen protection. After cooling to room temperature, the acetonitrile was removed by vacuum distillation at 45 °C. The residue was washed three times with ethyl acetate, and then the ethyl acetate was removed by vacuum distillation at 55 °C. Finally, the residue was dried under vacuum at 40 °C for 24 h to obtain the ionic liquid 1-hexyl-3-methylimidazolium bromide ([HMIM][Br]).

[0036] ③Preparation of UiO-66 Zirconium tetrachloride and 2-aminoterephthalic acid were dissolved in 480 molar amounts of DMF (N,N-dimethylformamide) at a molar ratio of 1:1. Acetic acid and hydrochloric acid were added as modifiers and crystallizing agents, respectively, with a molar ratio of acetic acid to zirconium tetrachloride of 1:8 and a molar ratio of hydrochloric acid to zirconium tetrachloride of 1:0.5. After sonication, the solution was magnetically stirred for 15 min, transferred to a polytetrafluoroethylene autoclave, and then continuously heated to 120 °C under microwave conditions. After reacting for 3 h, crystals were obtained, washed with DMF solvent, and finally dried in an oven at 120 °C.

[0037] ④ Preparation of magnetic composite hydrogels The prepared 0.5g Fe3O4, 0.8g ionic liquid, 0.5g UiO-66 and 1.3g chitosan were first dissolved in acetic acid, and then added dropwise to NaOH aqueous solution (2mol / L) with a syringe to form a hydrogel. The hydrogel was washed with deionized water and freeze-dried to obtain the final product. Example 3

[0038] This embodiment provides a magnetic composite material, which is prepared by the following method: ①Preparation of Fe3O4 Weigh 5.4 g FeCl3·6H2O and 2.8 g FeSO4·7H2O into a 50 mL beaker, and add 0.85 mL concentrated HCl (to prevent Fe from being absorbed). 2+ and Fe 3+ After hydrolysis, add 25 mL of deionized water and stir with a glass rod until completely dissolved to obtain a standard iron solution. Add 250 mL of 1.5 mol / L NaOH solution (to co-precipitate ferrous and ferric iron to form Fe3O4) to a 500 mL three-necked flask, heat in a water bath to 80°C, and under N2 protection, add the prepared iron standard solution dropwise to the NaOH solution at a rate of 10 d / min, mechanically stir for 1 hour, stop heating, cool the solution to room temperature, remove the nitrogen protection device, add 100 mL of 0.3 mol / L sodium citrate to a constant pressure funnel, and continue mechanical stirring for 1 hour. After the reaction is complete, wash the product with deionized water to neutralize it, then wash with anhydrous ethanol, and finally wash with deionized water 2-3 times. Separate and collect the product using a magnet, discard the supernatant, and freeze-dry the final product.

[0039] ② Preparation of ionic liquids Take 0.1 mol of 1-methylimidazole, 0.11 mol of 1-bromohexane and 50 mL of acetonitrile in a three-necked flask, reflux at 80 °C under nitrogen protection for 24 h, cool to room temperature, remove acetonitrile by rotary evaporation at 45 °C, wash the residue three times with ethyl acetate, remove ethyl acetate by rotary evaporation at 55 °C, and dry under vacuum at 40 °C for 24 h to obtain an ionic liquid.

[0040] ③Preparation of UiO-66 Zirconium tetrachloride and 2-aminoterephthalic acid were dissolved in 480 molar amounts of DMF (N,N-dimethylformamide) at a molar ratio of 1:1. Acetic acid and hydrochloric acid were added as modifiers and crystallizing agents, respectively, with a molar ratio of acetic acid to zirconium tetrachloride of 1:12 and a molar ratio of hydrochloric acid to zirconium tetrachloride of 1:2. After sonication, the solution was magnetically stirred for 15 min, transferred to a polytetrafluoroethylene autoclave, and then continuously heated at 120°C under microwave conditions for 4 h to obtain crystals. The crystals were washed with DMF solvent. Finally, the obtained product was dried in an oven at 120°C.

[0041] ④ Preparation of magnetic composite hydrogels The prepared 0.5g Fe3O4, 1.0g ionic liquid, 1.2g UiO-66 and 1.5g chitosan were first dissolved in acetic acid, and then added dropwise to NaOH aqueous solution (2mol / L) with a syringe to form a hydrogel. The hydrogel was washed with deionized water and freeze-dried to obtain the final product. Example 4

[0042] This embodiment provides a magnetic composite material, which is prepared by the following method: ①Preparation of Fe3O4 Weigh 5.4 g FeCl3·6H2O and 2.8 g FeSO4·7H2O into a 50 mL beaker, and add 0.85 mL concentrated HCl (to prevent Fe from being absorbed). 2+ and Fe 3+After hydrolysis, add 25 mL of deionized water and stir with a glass rod until completely dissolved to obtain a standard iron solution. Add 250 mL of 1.5 mol / L NaOH solution (to co-precipitate ferrous and ferric iron to form Fe3O4) to a 500 mL three-necked flask, heat in a water bath to 80°C, and under N2 protection, add the prepared iron standard solution dropwise to the NaOH solution at a rate of 10 d / min, mechanically stir for 1 hour, stop heating, cool the solution to room temperature, remove the nitrogen protection device, add 100 mL of 0.3 mol / L sodium citrate to a constant pressure funnel, and continue mechanical stirring for 1 hour. After the reaction is complete, wash the product with deionized water to neutralize it, then wash with anhydrous ethanol, and finally wash with deionized water 2-3 times. Separate and collect the product using a magnet, discard the supernatant, and freeze-dry the final product.

[0043] ② Preparation of ionic liquids 0.1 mol of 1-methylimidazolium, 0.11 mol of 1-bromohexane, and 50 mL of acetonitrile were placed in a three-necked flask and refluxed at 80 °C for 24 h under nitrogen protection. After cooling to room temperature, the acetonitrile was removed by vacuum distillation at 45 °C. The residue was washed three times with ethyl acetate, and then the ethyl acetate was removed by vacuum distillation at 55 °C. Finally, the residue was dried under vacuum at 40 °C for 24 h to obtain the ionic liquid 1-hexyl-3-methylimidazolium bromide. 1 g of the obtained 1-hexyl-3-methylimidazolium bromide and 5 g of potassium hexafluorophosphate (KPF6) were stirred in 100 mL of water. The mixture was sonicated for 10 min and stirred at room temperature for 24 h. The obtained product was washed with water and dried to obtain 1-hexyl-3-methylimidazolium hexafluorophosphate.

[0044] ③Preparation of Fe3O4@NH2-UiO-66 0.5 g of Fe3O4 was weighed and dispersed in 50 mL of DMF, and sonicated for 20 min. 0.6 g of ZrCl4 and 0.45 g of 2-aminoterephthalic acid were added, and the mixture was magnetically stirred for 30 min before being transferred to a reaction vessel. The reaction was carried out at 120 °C for 24 h. After the reaction was completed, the mixture was cooled to room temperature, and the product was separated under an external magnetic field. The product was repeatedly washed with DMF and ethanol, and dried to obtain the Fe3O4@NH2-UiO-66 magnetic composite material.

[0045] ④ Preparation of magnetic composite hydrogels Weigh 1.0 g of chitosan and dissolve it in 5% acetic acid solution, stirring until completely dissolved. Then add 0.9 g of Fe3O4@UiO-66 and 0.9 g of 1-hexyl-3-methylimidazolium hexafluorophosphate, and sonicate for 20 min. Next, use a syringe to dropwise add the mixture to a 2 mol / L sodium hydroxide solution to form a hydrogel. Wash with deionized water and freeze-dry to obtain the final product.

[0046] The infrared absorption spectrum of the magnetic composite material prepared in this embodiment is shown below. Figure 5 As shown in the figure, 567cm -1 A distinct characteristic absorption peak appears at 3470 cm⁻¹, corresponding to the stretching vibration peak of the Fe-O bond in Fe₃O₄, indicating the successful formation of the magnetic core. -1 and 1080cm -1 The nearby absorption peaks are attributed to the stretching vibrations of -OH and COC in chitosan. (1380 cm⁻¹) -1 and 1621 cm -1 The characteristic peak at 1630 cm⁻¹ corresponds to the symmetric and asymmetric stretching vibrations of the carboxyl group in the phthalic acid ligand of UiO-66, indicating that the UiO-66 structure was successfully constructed. -1 The absorption peak at this location is related to the characteristic vibrations of the imidazole ring in the ionic liquid, indicating that the ionic liquid was successfully introduced into the material surface. The common presence of the above characteristic peaks demonstrates the successful preparation of the magnetic composite hydrogel.

[0047] The scanning electron microscope image of the magnetic composite material prepared in this embodiment is shown below. Figure 6 As shown in the figure, the composite material exhibits an irregular spherical structure with a relatively rough surface and an average particle size of approximately 1400 μm, indicating that chitosan coating forms a stable composite structure.

[0048] The energy dispersive spectroscopy (EDS) analysis diagram of the magnetic composite material prepared in this embodiment is shown below. Figure 7 As shown, the elements C, O, N, Fe, Zr, F, and P are uniformly distributed in the material. Fe originates from the Fe3O4 magnetic core, while C, N, O, and Zr mainly come from chitosan and MOF organic ligands. F and P originate from ionic liquid anions. This uniform elemental distribution indicates the successful composite of Fe3O4, UiO-66, the ionic liquid, and CS, signifying the successful preparation of the magnetic composite material.

[0049] The hysteresis loop of the magnetic composite material prepared in this embodiment is as follows: Figure 8 As shown in the figure, its saturation magnetization value is approximately 17.0 emu / g, which indicates that the adsorbent has sufficient magnetic response to meet the requirements of magnetic separation (generally, a saturation magnetization value greater than 10 emu / g is considered "sufficient for magnetic separation"). Example 5

[0050] This embodiment provides a magnetic composite material, which is prepared by the following method: ①Preparation of Fe3O4 Weigh 5.4 g FeCl3·6H2O and 2.8 g FeSO4·7H2O into a 50 mL beaker, and add 0.85 mL concentrated HCl (to prevent Fe from being absorbed). 2+ and Fe 3+ After hydrolysis, add 25 mL of deionized water and stir with a glass rod until completely dissolved to obtain a standard iron solution. Add 250 mL of 1.5 mol / L NaOH solution (to co-precipitate ferrous and ferric iron to form Fe3O4) to a 500 mL three-necked flask, heat in a water bath to 80°C, and under N2 protection, add the prepared iron standard solution dropwise to the NaOH solution at a rate of 10 d / min, mechanically stir for 1 hour, stop heating, cool the solution to room temperature, remove the nitrogen protection device, add 100 mL of 0.3 mol / L sodium citrate to a constant pressure funnel, and continue mechanical stirring for 1 hour. After the reaction is complete, wash the product with deionized water to neutralize it, then wash with anhydrous ethanol, and finally wash with deionized water 2-3 times. Separate and collect the product using a magnet, discard the supernatant, and freeze-dry the final product.

[0051] ② Preparation of ionic liquids 1-Methylimidazolium, 1-bromohexane, and acetonitrile were added to a three-necked flask and refluxed at 80 °C for 24 h under nitrogen protection. After cooling to room temperature, the acetonitrile was removed by vacuum distillation at 45 °C. The residue was washed three times with ethyl acetate, and then the ethyl acetate was removed by vacuum distillation at 55 °C. Finally, the residue was dried under vacuum at 40 °C for 24 h to obtain the ionic liquid 1-hexyl-3-methylimidazolium bromide. 1 g of the obtained 1-hexyl-3-methylimidazolium bromide and 7 g of potassium hexafluorophosphate (KPF6) were stirred in 100 mL of water. The mixture was sonicated for 10 min and stirred at room temperature for 24 h. The obtained product was washed with water and dried to obtain 1-hexyl-3-methylimidazolium hexafluorophosphate.

[0052] ③Preparation of Fe3O4@NH2-UiO-66 0.5 g of Fe3O4 was weighed and dispersed in 50 mL of DMF, and sonicated for 20 min. 0.9 g of ZrCl4 and 0.9 g of 2-aminoterephthalic acid were added, and the mixture was magnetically stirred for 30 min before being transferred to a reaction vessel. The reaction was carried out at 120 °C for 24 h. After the reaction was completed, the mixture was cooled to room temperature, and the product was separated under an external magnetic field. The product was repeatedly washed with DMF and ethanol, and dried to obtain the Fe3O4@NH2-UiO-66 magnetic composite material.

[0053] ④ Preparation of magnetic composite hydrogels Weigh 1.0 g of chitosan and dissolve it in 5% acetic acid solution, stirring until completely dissolved. Then add 1.8 g of Fe3O4@UiO-66 and 0.9 g of 1-hexyl-3-methylimidazolium hexafluorophosphate, and sonicate for 20 min. Next, use a syringe to dropwise add the mixture to a 2 mol / L sodium hydroxide solution to form a hydrogel. Wash with deionized water and freeze-dry to obtain the final product.

[0054] Application Example 1: Limit of Detection Test for Rapid Detection of Mycotoxins in Grains The sample pretreatment process is as follows: Extraction: The pulverized grain sample was extracted by shaking with a methanol-water mixture; Enrichment: Add the prepared adsorbent (60 mg) to the sample solution (20 mL) and vortex for 3 minutes to adsorb; Separation: Apply an external magnetic field to adsorb the material, discard the supernatant, and separate the adsorbent from the solution; Washing: Rinse with water or a low-concentration organic solvent to remove impurities; Elution: AFs were eluted with 1 mL of 0.1% hydrochloric acid / acetonitrile (1 / 9, v / v) by sonication for 2 minutes, and the eluent was collected; Filtration: Passed through a 0.22 μm organic filter membrane, to be tested.

[0055] The sample was then injected into the HPLC-FLD system for analysis. The chromatographic conditions of the HPLC-FLD system are as follows: HPLC (Shimadzu RF-20A FD, USA); C18 column (4.6 mm × 250 mm, 5 μm); mobile phase: methanol:acetonitrile:water (2 / 2 / 6, V / V / V); flow rate: 1.0 mL / min; column temperature: 30℃; injection volume: 20 μL; fluorescence detector: excitation wavelength: 365 nm; emission wavelength: 440 nm.

[0056] The results are shown in the table below:

[0057] The data above show that AFTB1 and AFTG1 exhibit good linearity in the range of 2–100 μg / kg, while AFTB2 and AFTG2 show good linearity in the range of 1.5–100 μg / kg, all meeting the requirements for quantitative analysis. Under optimized conditions, the limit of detection (LOD, S / N=3) of the method is 0.1–0.5 μg / kg, and the relative standard deviation (RSD) is 5.7%–13.3% (n=3), indicating that the method has good sensitivity and repeatability.

[0058] Application Example 2: Chromatographic Detection The sample pretreatment process is as follows: Extraction: Add 5g of corn sample to 20ml of acetonitrile:water (84 / 16, volume ratio) and stir for 20 minutes; Enrichment: Add the prepared adsorbent (60 mg) to the sample solution (20 mL) and vortex for 3 minutes to adsorb; Separation: Apply an external magnetic field to adsorb the material, discard the supernatant, and separate the adsorbent from the solution; Washing: Rinse with water or a low-concentration organic solvent to remove impurities; Elution: AFs were eluted with 1 mL of 0.1% hydrochloric acid / acetonitrile (1 / 9, v / v) by sonication for 4 minutes, and the eluent was collected; Filtration: Passed through a 0.22 μm organic filter membrane, to be tested.

[0059] The sample was then injected into the HPLC-FLD system for analysis. The chromatographic conditions of the HPLC-FLD system are as follows: HPLC (Shimadzu RF-20A FD, USA); C18 column (4.6 mm × 250 mm, 5 μm); mobile phase: methanol:acetonitrile:water (2 / 2 / 6, V / V / V); flow rate: 1.0 mL / min; column temperature: 30℃; injection volume: 20 μL; fluorescence detector: excitation wavelength: 365 nm; emission wavelength: 440 nm.

[0060] The chromatogram of the corn sample analysis is shown in the figure. Figure 9 As shown in the figure, (a) the mixed spiked solution was directly injected (AFTB1 and AFTG1 were 20.0 μg / kg, AFTB2 and AFTG2 were 6.0 μg / kg). The mixed solution after MSPE-HPLC analysis: (b) AFTB1 and AFTG1 concentrations were 20.0 μg / kg, AFTB2 and AFTG2 concentrations were 6.0 μg / kg; (c) AFTB1 and AFTG1 concentrations were 40.0 μg / kg, AFTB2 and AFTG2 concentrations were 12.0 μg / kg. Chromatographic peaks: 1. AFTG2, 2. AFTG1, 3. AFTB2, 4. AFTB1.

[0061] As shown in the figure, the four aflatoxins (AFTG2, AFTG1, AFTB2, and AFTB1) achieved baseline separation under the selected chromatographic conditions, with symmetrical peaks and no overlapping interference, indicating that the MSPE-HPLC method has good separation capability. Compared with direct injection (a), the chromatograms after MSPE treatment (b and c) have lower background and significantly fewer impurity peaks, indicating that magnetic solid-phase extraction can effectively remove matrix interference in corn samples and improve the selectivity of detection. Comparing (b) and (c), it can be seen that when the spike concentration increases, the response values ​​of each chromatographic peak increase approximately proportionally, proving that the method has a good linear response within the tested concentration range and can be used for quantitative analysis. This pretreatment method combined with HPLC-FLD can meet the requirements for trace detection of aflatoxins in grain samples, providing a reliable technical means for food safety monitoring.

[0062] The foregoing description is intended to provide detailed embodiments for those skilled in the art, ensuring they can accurately grasp and apply the present invention. Any improvements or modifications to the present invention obtained by those skilled in the art through logical analysis, reasoning, or simple enumeration, without creative work, based on existing technology, should be considered within the scope of protection defined by the claims of this invention.

Claims

1. A magnetic composite material, characterized in that... Prepared by the following method: (1) Preparation of Fe3O4 nanoparticles FeCl3·6H2O and FeSO4·7H2O were dissolved in water, concentrated hydrochloric acid was added to prevent hydrolysis, and the solution was added dropwise to NaOH solution preheated to 80℃. After stirring and reacting, sodium citrate was added for modification, followed by magnetic separation and washing, and drying to obtain Fe3O4 nanoparticles. (2) Preparation of UiO-66 Zirconium tetrachloride and 2-aminoterephthalic acid were dissolved in N,N-dimethylformamide, and acetic acid and hydrochloric acid were added as modifier and crystallizing agent, respectively. After homogenization, the solution was transferred to a polytetrafluoroethylene autoclave and subjected to a solvothermal reaction under microwave conditions. The product was washed and dried to obtain UiO-66. (3) Preparation of magnetic composite materials Fe3O4 nanoparticles, ionic liquid, UiO-66 and chitosan are dissolved in acetic acid, and then added dropwise to an alkaline solution using a syringe to form a hydrogel.

2. The magnetic composite material according to claim 1, characterized in that: In step (1), the molar ratio of FeCl3·6H2O and FeSO4·7H2O is 2:1, and concentrated hydrochloric acid is added to make the pH of the mixed solution less than 7; the total amount of FeCl3·6H2O and FeSO4·7H2O is in a molar ratio of 1:1 to sodium citrate.

3. The magnetic composite material according to claim 1, characterized in that: In step (2), the molar ratio of zirconium tetrachloride and 2-aminoterephthalic acid is 1:1, the molar ratio of the total amount of zirconium tetrachloride and 2-aminoterephthalic acid to DMF is 1:(450-500); the molar ratio of zirconium tetrachloride to acetic acid is 1:(8-12), and the molar ratio of zirconium tetrachloride to hydrochloric acid is 1:(0.5-2).

4. The magnetic composite material according to claim 1, characterized in that: In step (2), the temperature of the solvothermal reaction is 120°C and the time is 3-4 hours. The product is washed with DMF.

5. A magnetic composite material according to claim 1, characterized in that: In step (3), the ionic liquid is prepared by the following method: 0.1 mol of 1-methylimidazole, 0.11 mol of 1-bromohexane and 50 ml of acetonitrile are placed in a three-necked flask and refluxed at 80 °C under nitrogen protection for 24 h. After cooling to room temperature, the acetonitrile is removed by rotary evaporation at 45 °C. The residue is washed three times with ethyl acetate, and the ethyl acetate is removed by rotary evaporation at 55 °C. The residue is then dried under vacuum at 40 °C for 24 h to obtain the ionic liquid.

6. A magnetic composite material according to claim 1, characterized in that: In step (3), the mass ratio of Fe3O4 nanoparticles, ionic liquid, UiO-66 and chitosan is 1:(1-2):(0.8-1.2):(2-3), and the alkaline solution is a 2mol / L NaOH aqueous solution.

7. The application of the magnetic composite material according to any one of claims 1-6 in the rapid detection of mycotoxins in grains, characterized in that: Magnetic solid-phase extraction is used in sample pretreatment.

8. The application according to claim 7, characterized in that... Specifically, the steps include the following: Extraction: The pulverized grain sample was extracted by shaking with a methanol-water mixture; Enrichment: Add magnetic composite material to the extract and shake to adsorb; Separation: Apply an external magnetic field to adsorb the material, then discard the supernatant; Washing: Rinse with water or a low-concentration organic solvent to remove impurities; Elution: Elute with an eluent and collect the eluent; Filtration: Passed through a 0.22 μm organic filter membrane, to be tested.

9. The application according to claim 7, characterized in that: The detection method was high performance liquid chromatography-mass spectrometry, and the chromatographic conditions were as follows; Chromatographic column: C18 column; Mobile phase: Methanol-water or acetonitrile-water gradient elution; Mass spectrometry conditions: electrospray ionization source, multiple reaction monitoring mode; Quantitative methods: external standard method or internal standard method; The gradient elution program is as follows: 0–5 min, 10% methanol / acetonitrile held for 5 min; 5–30 min, methanol / acetonitrile linearly increased from 10% to 95%, held for 5 min.

10. The method for rapid detection of mycotoxins in grains according to claim 7, characterized in that: The fungal toxins mentioned are aflatoxin, deoxynivalenol, zearalenone, ochratoxin A, fumonisin, T-2 toxin, or HT-2 toxin.

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