CN111269454A

CN111269454A - Preparation method of magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin

Info

Publication number: CN111269454A
Application number: CN202010085445.XA
Authority: CN
Inventors: 杨鑫; 赵倩玉
Original assignee: Harbin Institute of Technology
Current assignee: Harbin Institute of Technology
Priority date: 2020-01-30
Filing date: 2020-01-30
Publication date: 2020-06-12

Abstract

A method for preparing a magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin. The invention belongs to the field of molecular imprinting polymers. The invention aims to solve the technical problem that the traditional separating filler for separating anthocyanin has low selectivity. The method comprises the following steps: one, aminated magnetic nano Fe₃O₄‑NH₂Synthesizing; secondly, synthesizing 7 magnetic covalent organic framework molecular engrams (MCPMIPs) by using C3G as a template molecule and adopting a room temperature synthesis method. The MCMIPs have magnetic responsiveness, high adsorption performance and excellent specificity, and are suitable for quickly and specifically fishing C3G from a complex sample matrix. After the MCMIPs are purified for one time, high purity can be obtained>88% of anthocyanin. The separating medium simplifies the steps of separating and purifying anthocyanin, shortens the time of separating and purifying and greatly improves the efficiency of separating and purifying.

Description

Preparation method of magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin

Technical Field

The invention belongs to the field of molecularly imprinted polymers, and particularly relates to a preparation method of a magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin.

Background

Anthocyanin (AOC) belongs to flavonoid compounds, is composed of anthocyanin and sugar, and is a safe and nontoxic water-soluble pigment. Anthocyanins have many important biological activities in addition to their use as pigments in food products. Research shows that, up to now, anthocyanin has the strongest antioxidant activity and can treat body injury and various diseases caused by oxidative stress. In addition, it also has radioprotective, antiinflammatory, anticancer, diabetes preventing, and eyesight protecting effects. However, because the anthocyanin is various in types, has cis-trans isomers and poor stability, the separation and purification of anthocyanin monomers are always difficult. Monomers are difficult to obtain and the price is high, so that many pharmacological experiments only adopt crude anthocyanin as an experimental material, which greatly influences the research progress of the structure-activity relationship of anthocyanin.

The traditional separating filler for separating anthocyanin at present has low selectivity and is difficult to meet the existing requirements, so that a novel separating and purifying medium which can simplify the purifying process of anthocyanin and improve the yield of anthocyanin is urgently needed to be developed, the speed-limiting link influencing the deep research of anthocyanin is solved, and the effective, efficient and quick-acting separating and purifying of anthocyanin is realized.

Disclosure of Invention

The invention provides a preparation method of a magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin, which aims to solve the technical problem of low selectivity of the traditional separating filler for separating anthocyanin.

The preparation method of the magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin provided by the invention comprises the following steps:

mono, superparamagnetic amino functional Fe₃O₄Preparing magnetic nanoparticles: FeCl is added₃·6H₂Adding O into ethylene glycol, performing ultrasonic treatment to completely dissolve, adding anhydrous sodium acetate and 1, 6-hexanediamine, mechanically stirring to obtain reddish brown solution, and mixing with red pigmentReacting the brown solution at the temperature of 150-250 ℃ for 6-12 h, then performing magnetic separation and sedimentation, washing the separated solid matter, removing unreacted matter, and performing vacuum drying to obtain superparamagnetic amino functionalized Fe₃O₄Magnetic nanoparticles, noted Fe₃O₄-NH₂Magnetic nanoparticles;

secondly, preparing magnetic covalent organic framework molecularly imprinted polymers (MCMIPs), namely ① adding cyanidin-3-O-glucoside (C3G) and functional monomers into absolute ethyl alcohol, carrying out ultrasonic treatment until the mixture is completely dissolved, carrying out prepolymerization reaction for 3 to 5 hours at room temperature to obtain a prepolymerization solution, and ② adding Fe obtained in the first step into the prepolymerization solution₃O₄-NH₂Magnetic nanoparticles, sonicated to Fe₃O₄-NH₂Uniformly dispersing magnetic nanoparticles in the prepolymerization solution, adding 1,3, 5-trialdehyde phloroglucinol (Tp), mechanically stirring, and adding scandium trifluoromethanesulfonate (Sc (OTf)₃) The preparation method comprises the steps of reacting at room temperature to obtain a polymer, ③ washing the obtained polymer to obtain a clear solution, repeatedly eluting the clear solution by using a mixed solution of methanol and acetic acid to remove C3G until no C3G is detected in an eluent, ④ finally washing off residual acetic acid by using double distilled water, and drying in vacuum to obtain the magnetic covalent organic framework molecularly imprinted polymer, which is marked as MCMIPs.

Further defining, the FeCl in step one₃·6H₂The mass ratio of O to the volume of ethylene glycol is (12-15) g: 500 mL.

Further defining, the FeCl in step one₃·6H₂The ratio of the mass of O to the volume of ethylene glycol is (13-14) g: 500 mL.

Further defining, the FeCl in step one₃·6H₂The ratio of the mass of O to the volume of ethylene glycol was 13.5 g: 500 mL.

Further defining, the FeCl in step one₃·6H₂The mass ratio of the O to the anhydrous sodium acetate is (12-15) g: 27 g.

Further defining, the FeCl in step one₃·6H₂The mass ratio of O to anhydrous sodium acetate is (1)3～14)g：27g。

Further defining, the FeCl in step one₃·6H₂The mass ratio of O to anhydrous sodium acetate is 13.5 g: 27 g.

Further defining, the FeCl in step one₃·6H₂The ratio of the mass of O to the volume of 1, 6-hexanediamine is (12-15) g: 172 mL.

Further defining, the FeCl in step one₃·6H₂The ratio of the mass of O to the volume of 1, 6-hexanediamine is (13-14) g: 172 mL.

Further defining, the FeCl in step one₃·6H₂The ratio of the mass of O to the volume of 1, 6-hexanediamine was 13.5 g: 172 mL.

Further defined, the ultrasound parameters in step one are: the frequency is 35 kHz-45 kHz, and the time is 20 min-40 min.

Further limiting, in the step one, the mechanical stirring speed is 100 rpm-200 rpm, and the time is 20 min-40 min.

Further limiting, the reddish brown solution obtained in the step one is reacted for 6 hours at the temperature of 180-200 ℃.

Further defined, the washing process in the first step is as follows: washing with double distilled water for 3-6 times, and then washing with ethanol for 3-6 times.

Further limited, the vacuum drying parameters in the first step are as follows: the temperature is 40-60 ℃.

Further defined, in step two ①, the functional monomer is p-phenylenediamine (Pa-1), p-diaminobiphenyl (BD), 3' -dihydroxybenzidine (DHBD), 4 ' -diaminodiphenyl ether (ODA), 4 ' -diaminop-terphenyl (DT), 2, 6-Diaminoanthraquinone (DAAQ), or 1, 5-diamino-4, 8-dihydroxy-9, 10-anthracenedione (DADHAQ).

Further, the ratio of the C3G to the amount of the functional monomer in step two ① is 0.12 (0.4-0.55).

Further, the ratio of the C3G to the amount of the functional monomer in step two ① is 0.12 (0.45-0.5).

Further defined, the ratio of the amount of C3G to the amount of functional monomer material in step two ① is 0.12: 0.48.

Further limiting, the ratio of the amount of the C3G substance to the volume of the absolute ethyl alcohol in the second step ① is 0.12mmol (5-15) mL.

Further defined, the ratio of the amount of C3G in step two ① to the volume of absolute ethanol was 0.12 mmol: 10 mL.

Further limiting, in the second step ①, the ultrasonic parameters are that the frequency is 35 kHz-45 kHz, and the time is 15 min-25 min.

Further limiting, the prepolymerization in step two ① was 4 h.

Further limiting, the amount of C3G and Fe in step two ②₃O₄-NH₂The mass ratio of the magnetic nanoparticles is 0.12 mmol: (40-60) mg.

Further limiting, the amount of C3G and Fe in step two ②₃O₄-NH₂The mass ratio of the magnetic nanoparticles is 0.12 mmol: (45-55) mg.

Further limiting, the amount of C3G and Fe in step two ②₃O₄-NH₂The mass ratio of the magnetic nanoparticles is 0.12 mmol: 50 mg.

Further limiting, in the second step ②, the ultrasonic parameters are that the frequency is 35 kHz-45 kHz, and the time is 15 min-25 min.

Further limiting, the concentration of the 1,3, 5-trialdehyde phloroglucinol in step two ② is 0.045mmol/mL to 0.05 mmol/mL.

Further defined, the concentration of 1,3, 5-trialdehyde phloroglucinol in step two ② is 0.048 mmol/mL.

Further limit, the ratio of the amount of the substance of the tri-C3G to the volume of the 1,3, 5-trialdehyde-m-benzene in the step two ② is 0.12mmol (5-15) mL.

Further defined, the ratio of the amount of tri-C3G species to the volume of 1,3, 5-trialdehyde-m-benzene in step two ② is 0.12 mmol: 10 mL.

Further, in the step two ②, the concentration of scandium trifluoromethanesulfonate is from 0.005mmol/mL to 0.01 mmol/mL.

Further limiting, the concentration of scandium trifluoromethanesulfonate in step two ② is 0.008 mmol/mL.

Further, in the step two ②, the ratio of the amount of the substance of the tri-C3G to the volume of the scandium trifluoromethanesulfonate is 0.12mmol (0.5-1.5) mL.

Further defining the ratio of the amount of tri-C3G species to the volume of scandium trifluoromethanesulfonate in step two ② to be 0.12 mmol: 1 mL.

Further limiting, in the second step ②, the mechanical stirring speed is 100 rpm-250 rpm, and the time is 20 min-40 min;

further, the reaction time at room temperature in the step two ② is 20-24 h.

Further limiting, in the second step ③, the washing process includes washing with N, N' -Dimethylformamide (DMF) for 3-6 times, and then washing with methanol for 3-6 times.

Further, in the step two ③, the volume ratio of methanol to acetic acid in the mixed solution of methanol and acetic acid is 9: 1.

Further limiting, in the second step ④, the vacuum drying parameter is that the temperature is 40-60 ℃.

Compared with the prior art, the invention has the remarkable effects as follows:

the method of the invention firstly dopes 1, 6-hexamethylene diamine in the magnetic nano to prepare the aminated magnetic carrier; and then forming a covalent organic framework with imprinting holes on the surface of the magnetic carrier by Schiff base reaction. The method utilizes the molecular recognition capability of the molecularly imprinted polymer (similar to enzyme) to directly extract cyanidin-3-O-glucoside (C3G) from a complex crude extract, thereby improving the separation efficiency. Covalent Organic Frameworks (COFs) with porous structures (providing more identification dots) and large pi-electron systems (forming strong pi-x interactions with C3G) are introduced to improve the adsorption capacity of Molecularly Imprinted Polymers (MIPs). In addition, the COFs are a natural protective barrier, and provide a hydrophobic environment for C3G, so that the C3G is not hydrolyzed in the separation process, and the biological activity of C3G is ensured to a great extent. The rigid framework structure of COFs effectively prevents the collapse of a molecular imprinting cavity and ensures the accuracy of the spatial configuration of the recognition site. The magnetic responsiveness of magnetic nano-particles (MNPs) makes the rapid separation of solid and liquid in the extraction process possible. In consideration of the stability of AOC, a Room Temperature Synthesis Strategy (RTSS) is adopted to replace a virtual template molecular imprinting technology, and a molecular imprinting polymer of a thermosensitive substance is successfully synthesized. The technology can realize the specific identification and the rapid separation of the AOC, and is a breakthrough progress for preparing the AOC monomer.

7 kinds of magnetic covalent organic framework molecular engrams (MCMIPs) synthesized by the invention; the MCMIPs have magnetic responsiveness, high adsorption performance and excellent specificity, and are suitable for quickly and specifically fishing C3G from a complex sample matrix. After the first purification of MCMIPs, the anthocyanin with high purity (> 88%) can be obtained. The novel separation medium simplifies the steps of separating and purifying anthocyanin, shortens the time of separating and purifying and greatly improves the efficiency of separating and purifying.

Drawings

FIG. 1 shows Fe obtained in the first step of the present embodiment₃O₄-NH₂SEM images of magnetic nanoparticles; FIG. 2 is an SEM image of MCMIPs-Pa-1 according to a first embodiment; FIG. 3 is an SEM diagram of MCMIPs-BD of the second embodiment; FIG. 4 is an SEM image of MCMIPs-DHBD according to embodiment three; FIG. 5 is an SEM image of MCMIPs-ODA according to a fourth embodiment; FIG. 6 is an SEM diagram of MCMIPs-DT according to a fifth embodiment; FIG. 7 is an SEM image of MCMIPs-DAAQ of a sixth embodiment; FIG. 8 is an SEM image of MCMIPs-DADHAQ according to embodiment seven; FIG. 9 shows Fe obtained in the first step of the present embodiment₃O₄-NH₂Infrared spectra of magnetic nanoparticles, MCMIPs-Pa-1 in the first embodiment, MCMIPs-BD in the second embodiment, and MCMIPs-ODA in the fourth embodiment; FIG. 10 is an infrared spectrum of MCMIPs-DHBD of embodiment three, MCMIPs-DT of embodiment five, MCMIPs-DAAQ of embodiment six, and MCMIPs-DADHAQ of embodiment seven; FIG. 11 shows Fe obtained in step one of the embodiments₃O₄-NH₂N of magnetic nanoparticles₂Adsorption isotherm plot; FIG. 12 shows N of MCMIPs-Pa-1 in accordance with a first embodiment₂Adsorption isotherm plot; FIG. 13 shows N of MCMIPs-BD in the second embodiment₂Adsorption isotherm plot; FIG. 14 shows the N of MCMIPs-DHBD according to the third embodiment₂Adsorption isotherm plot; FIG. 15 shows N of MCMIPs-ODA according to the fourth embodiment₂Adsorption isotherm plot; FIG. 16 shows N of MCMIPs-DT according to a fifth embodiment₂Adsorption isotherm plot; FIG. 17 is a diagram of N of MCMIPs-DAAQ according to a sixth embodiment₂Adsorption isotherm plot; FIG. 18 is a diagram of N of MCMIPs-DADHAQ according to the seventh embodiment₂Adsorption isotherm plot; FIG. 19 is an isothermal graph of equilibrium adsorption amounts of MCMIPs-Pa-1 and corresponding MCNIPs at different initial concentrations of C3G, according to a first embodiment; FIG. 20 is an isothermal graph of equilibrium adsorption amounts of MCMIPs-BD and corresponding MCNIPs of embodiment two in C3G at different initial concentrations; FIG. 21 is an isothermal plot of equilibrium adsorption amounts of MCMIPs-DHBD and corresponding MCNIPs at different initial concentrations of C3G, according to a third embodiment; FIG. 22 is an isothermal plot of equilibrium adsorption amounts of MCMIPs-ODA and corresponding MCNIPs of the fourth embodiment at different initial concentrations of C3G; FIG. 23 is an isothermal plot of equilibrium adsorption amounts of MCMIPs-DT and corresponding MCNIPs at different initial concentrations of C3G for the fifth embodiment; FIG. 24 is an isothermal graph of equilibrium adsorption amounts of MCMIPs-DAAQ and corresponding MCNIPs of embodiment six in C3G at different initial concentrations; FIG. 25 is an isothermal graph of equilibrium adsorption amounts of MCMIPs-DADHAQ and corresponding MCNIPs of embodiment seven in C3G at different initial concentrations; FIG. 26 is a graph of the rate of adsorption equilibrium for MCMIPs-Pa-1 and the corresponding MCNIPs of the first embodiment; FIG. 27 is a graph of the adsorption equilibrium rates for MCMIPs-BD and the corresponding MCNIPs of embodiment two; FIG. 28 is a graph of the adsorption equilibrium rates for MCMIPs-DHBD and the corresponding MCNIPs in accordance with the third embodiment; FIG. 29 is a graph of the adsorption equilibrium rates for MCMIPs-ODA and corresponding MCNIPs for the fourth embodiment; FIG. 30 is a graph of the rate of adsorption equilibrium for MCMIPs-DT and corresponding MCNIPs for the fifth embodiment; FIG. 31 shows MCMIPs-DAAQ and MCMIPs-DAAQ of a sixth embodimentCorresponding rate profile of the MCNIPs adsorption equilibrium; FIG. 32 is a graph of the rate of adsorption equilibrium for MCMIPs-DADHAQ and corresponding MCNIPs for embodiment seven; FIG. 33 is a bar graph of the adsorption selectivity of MCMIPs-Pa-1 and the corresponding MCNIPs of the first embodiment; FIG. 34 is a bar graph of adsorption selectivity of MCMIPs-BD and corresponding MCNIPs of embodiment two; FIG. 35 is a bar graph of adsorption selectivity of MCMIPs-DHBD and corresponding MCNIPs of embodiment three; FIG. 36 is a bar graph of the adsorption selectivity of MCMIPs-ODA and the corresponding MCNIPs of the fourth embodiment; FIG. 37 is a bar graph of the adsorption selectivity of MCMIPs-DT and the corresponding MCNIPs of embodiment five; FIG. 38 is a bar graph of the adsorption selectivity of MCMIPs-DAAQ and the corresponding MCNIPs for the sixth embodiment; FIG. 39 is a bar graph of the adsorption selectivity of MCMIPs-DADHAQ and the corresponding MCNIPs of embodiment seven; FIG. 40 is a histogram of the reuse efficiency of MCMIPs-Pa-1 and the corresponding MCNIPs of the first embodiment; FIG. 41 is a histogram of the reuse efficiency of MCMIPs-BD and corresponding MCNIPs of embodiment two; FIG. 42 is a histogram of the reuse efficiency of MCMIPs-DHBD and corresponding MCNIPs for the third embodiment; FIG. 43 is a bar graph of the recycling rates of MCMIPs-ODA and corresponding MCNIPs of the fourth embodiment; FIG. 44 is a bar graph of the reuse efficiency of MCMIPs-DT and corresponding MCNIPs for the fifth embodiment; FIG. 45 is a bar graph of the reuse efficiency of MCMIPs-DAAQ and corresponding MCNIPs for the sixth embodiment; FIG. 46 is a bar graph of the reuse efficiency of MCMIPs-DADHAQ and corresponding MCNIPs for embodiment seven; FIG. 47 is a high performance liquid chromatogram of Sorbus commixta extract, in which a is the indigo honeysuckle extract, b is the residue of the extract after adsorption by MCMIPs-DAAQ, c is the residue of the extract after adsorption by MCNIPs-DAAQ, d is the MCMIPs-DAAQ eluate, and e is the MCNIPs-DAAQ eluate; FIG. 48 is a high performance liquid chromatogram of a Vaccinium myrtillus extract, in which a is the Lonicera edulis extract, b is the residue of the extract after adsorption by MCMIPs-DAAQ, c is the residue of the extract after adsorption by MCNIPs-DAAQ, d is the MCMIPs-DAAQ eluate, and e is the MCNIPs-DAAQ eluate; FIG. 49 is a high performance liquid chromatogram of a blueberry extract, where a is the indigo honeysuckle extract, b is the residue of the extract after adsorption by MCMIPs-DAAQ, and c is the residue of the extract after adsorption by MCNIPs-DAAQD is MCMIPs-DAAQ eluate, e is MCNIPs-DAAQ eluate.

Detailed Description

The first embodiment is as follows: in the embodiment, the preparation method of the magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin comprises the following steps:

mono, superparamagnetic amino functional Fe₃O₄Preparing magnetic nanoparticles: 13.50g of FeCl₃·6H₂Adding O into 500mL of ethylene glycol, performing ultrasonic treatment for 30min under the condition of 40kHz frequency to completely dissolve the O, then adding 27g of anhydrous sodium acetate and 172mL of 1, 6-hexanediamine, mechanically stirring for 30min under the condition of 150rpm stirring speed to obtain a reddish brown solution, reacting the obtained reddish brown solution at the temperature of 200 ℃ for 6h, then performing magnetic separation and sedimentation by strong magnet, washing the separated solid matter, washing the solid matter with double distilled water for 5 times, then washing the solid matter with ethanol for 5 times to remove unreacted matters, and then performing vacuum drying at the temperature of 60 ℃ to obtain the superparamagnetic amino functionalized Fe₃O₄Magnetic nanoparticles, noted Fe₃O₄-NH₂Magnetic nanoparticles;

secondly, preparing magnetic covalent organic framework molecularly imprinted polymers (MCMIPs-Pa-1), namely ① adding 0.12mmol of C3G and 0.48mmol of p-phenylenediamine (Pa-1) into 10mL of absolute ethyl alcohol, carrying out ultrasonic treatment for 20min under the condition of 40kHz frequency until the p-phenylenediamine is completely dissolved, carrying out prepolymerization reaction for 4h at room temperature to obtain a prepolymerization solution, and ② adding 50mg of Fe obtained in the first step into the prepolymerization solution₃O₄-NH₂Magnetic nanoparticles, sonicated at a frequency of 40kHz for 20min to Fe₃O₄-NH₂The magnetic nanoparticles were uniformly dispersed in the pre-polymerization solution, 10mL of 1,3, 5-trialdehyde phloroglucinol (Tp) with a concentration of 0.048mmol/mL was added, mechanical stirring was performed at a stirring speed of 150rpm for 30min, and then 1mL of scandium trifluoromethanesulfonate (Sc (OTf))₃) Reacting at room temperature for 24 hr to obtain polymer, ③ washing the polymer with N, N' -Dimethylformamide (DMF) for 5 times, then with methanol for 5 times, and washing to obtain clarified solutionAnd (2) cleaning the solution, repeatedly eluting the clear solution by using a mixed solution of methanol and acetic acid (wherein the volume ratio of the methanol to the acetic acid is 9:1) to remove C3G until no C3G is detected in the eluent, ④, finally washing away residual acetic acid by using double distilled water, and drying in vacuum at the temperature of 60 ℃ to obtain the magnetic covalent organic framework molecularly imprinted polymer, which is marked as MCMIPs-Pa-1.

The second embodiment is as follows: the first difference between the present embodiment and the specific embodiment is: the functional monomer is Benzidine (BD), and the obtained magnetic covalent organic framework molecularly imprinted polymer is MCMIPs-BD. Other steps and parameters are the same as those in the first embodiment.

The third concrete implementation mode: the first difference between the present embodiment and the specific embodiment is: the functional monomer is 3,3' -dihydroxy benzidine (DHBD), and the obtained magnetic covalent organic framework molecularly imprinted polymer is MCMIPs-DHBD. Other steps and parameters are the same as those in the first embodiment.

The fourth concrete implementation mode: the first difference between the present embodiment and the specific embodiment is: the functional monomer is 4, 4' -diaminodiphenyl ether (ODA), and the obtained magnetic covalent organic framework molecularly imprinted polymer is MCMIPs-ODA. Other steps and parameters are the same as those in the first embodiment.

The fifth concrete implementation mode: the first difference between the present embodiment and the specific embodiment is: the functional monomer is 4, 4' -diamino-p-terphenyl (DT) to obtain the magnetic covalent organic framework molecularly imprinted polymer MCMIPs-DT. Other steps and parameters are the same as those in the first embodiment.

The sixth specific implementation mode: the first difference between the present embodiment and the specific embodiment is: the functional monomer is 2, 6-Diaminoanthraquinone (DAAQ) to obtain the magnetic covalent organic framework molecularly imprinted polymer which is MCMIPs-DAAQ. Other steps and parameters are the same as those in the first embodiment.

The seventh embodiment: the first difference between the present embodiment and the specific embodiment is: the functional monomer is 1, 5-diamino-4, 8-dihydroxy-9, 10-anthracenedione (DADHAQ) to obtain the magnetic covalent organic framework molecularly imprinted polymer which is MCMIPs-DADHAQ. Other steps and parameters are the same as those in the first embodiment.

And (3) detection by a scanning electron microscope: a. fe obtained in step one of the embodiments₃O₄-NH₂And (3) carrying out scanning electron microscope detection on the magnetic nanoparticles to obtain an SEM image shown in figure 1. As can be seen from FIG. 1, Fe was produced by hydrothermal method₃O₄-NH₂The magnetic nanoparticles are spherical and aggregate. The diameter is about 80nm, and the particle size distribution is relatively uniform.

b. Scanning electron microscope detection is performed on the magnetic covalent organic framework molecularly imprinted polymers (MCMIPs) obtained in the first to seventh embodiments to obtain SEM images of the magnetic covalent organic framework molecularly imprinted polymers shown in fig. 2 to 8, wherein fig. 2 is MCMIPs-Pa-1; FIG. 3 is MCMIPs-BD; FIG. 4 is a diagram of MCMIPs-DHBD; FIG. 5 is a diagram of MCMIPs-ODA; FIG. 6 is MCMIPs-DT; FIG. 7 is a diagram of MCMIPs-DAAQ; FIG. 8 is MCMIPs-DADHAQ. As can be seen from fig. 2 to 8, after covalent organic framework molecularly imprinted polymers (CMIPs, i.e., MCMIPs remove portions of magnetic nanoparticles), the morphology thereof is significantly changed, such as particle size increase, resulting in a new morphology consisting of short rods and sheets.

Elemental analysis: the surface elemental analysis was performed on the magnetic covalent organic framework molecularly imprinted polymers (MCMIPs) obtained in embodiments one to seven using a Vario Micro cube elemental analyzer, and the results are shown in table 1.

TABLE 1 content of C, N, H in MCMIPs

CMIPs are covalent organic framework molecularly imprinted polymers, i.e., MCMIPs with the magnetic nano-sized removed

And (4) conclusion: from Table 1, it can be seen that Fe₃O₄-NH₂The magnetic nano-material contains C, N, H elements but has a relatively low content, and is mainly derived from 1, 6-hexamethylene diamine, so that the magnetic nano-material is successfully doped with the 1, 6-hexamethylene diamine through coordination bonds in the formation process, and the grafting ratio of amino groups is 0.51mmol/g calculated by the content of N elements.

When magnetic carrier (Fe)₃O₄-NH₂) C, after the molecularly imprinted polymer is formed on the surface,N, H, because the amino and aldehyde monomers used to construct the CMIPs are composed of C, N, H, further illustrating the successful preparation of MCMIPs. And the content of CMIPs in the MCMIPs is 29.11-55.27%. CMIPs in MCMIPs-DADHAQ accounts for only 29.11%, because DADHAQ has larger steric hindrance, which affects the sheet stacking of COFs and inhibits the axial growth of polymers.

Infrared spectrum detection: adopting an AVATAR360 type Fourier infrared spectrometer and adopting a KBr tabletting method at 4000-400 cm^-1In the scope of Fe obtained in the first step of the first embodiment₃O₄-NH₂Performing infrared spectrum detection on the magnetic nanoparticles and the magnetic covalent organic framework molecularly imprinted polymers (MCMIPs) obtained in the first to seventh embodiments to obtain an infrared spectrum as shown in figures 9 to 10.

And (4) conclusion: from Fe₃O₄-NH₂Is visible in the infrared spectrum of 591cm^-1Near by the absorption peak of the stretching vibration of Fe-O, which is Fe₃O₄Characteristic absorption peak of (1), 3400cm^-1The absorption peak corresponds to the absorption peak of N-H stretching vibration, 1621cm^-1And 1394cm^-1Nearby are N-H and CH respectively₂Bending vibration peak of (2), proving Fe₃O₄-NH₂The synthesis of the magnetic particles was successful.

With Fe₃O₄-NH₂Compared with the infrared spectrum of MCMIPs, 1609-1621cm^-1The newly added absorption peak is a characteristic absorption peak of C ═ O, and is derived from the crosslinking agent Tp. In addition, the characteristic absorption of C as the benzene ring in the functional monomer (1457-1472 cm)^-1). Tp and the functional monomer are connected by condensation through Schiff base reaction, but absorption peaks of C ═ N and C-C are not observed, but C ═ C appears (1575-1583 cm)^-1)、C-N(1264-1339cm^-1) And N-H (3400-^-1) The absorption peak of (a) is mainly due to enol-keto interconversion. The pH-responsive C-N is converted into C-N, so that the MCMIPs framework is more stable and can be used in an acid-base environment. In addition, ether linkages (1239 cm) were also observed in MCMIPs-ODA^-1) Characteristic absorption peak of (1). Absorption peaks of each characteristicThe successful synthesis of MCMIPs is demonstrated.

N₂And (3) detection of adsorption quantity: determination of Fe obtained in step one of the embodiments by 3H-2000PS1 specific surface and pore size analyzer₃O₄-NH₂Magnetic nanoparticles, N of MCMIPs and MCNIPs obtained in embodiments one to seven₂Adsorption isotherms are shown in FIGS. 11 to 18; wherein the MCNIPs are non-molecularly imprinted polymers, and the synthesis steps and parameters are the same as those of embodiments one to seven except that the template molecule C3G is not added.

And (4) conclusion: as can be seen from FIGS. 11 to 18, Fe₃O₄-NH₂Is of a solid sphere structure, so the specific surface area is small and is only 39m²(ii) in terms of/g. The specific surface area is remarkably improved (S) due to the surface being coated with the porous material_BET(MCMIPs)＝294-105m²/g，S_BET(MCNIPs)＝173-76m²/g), more adsorption sites are provided, and a basis is provided for high loading capacity. Compared with MCPIPs, the MCPIPs have higher specific surface area, because the addition of the template molecules enables the functional monomers to be aligned in the formation process of the MCPIPs, and recognition holes matched with the template molecules are formed, so that the MCPIPs have higher specific surface area.

Adsorption isotherm curve detection-some macroscopic characteristics of the adsorption phenomena (adsorption capacity, adsorption strength, adsorption state) can be derived from the isotherm adsorption curve: the equilibrium adsorption amounts of MCMIPs and corresponding MCNIPs of embodiments one to seven were determined at 25 ℃ using a static adsorption method in C3G at different initial concentrations, and the results are shown in fig. 19 to 25. As can be seen from FIGS. 19-25, 7 MCMIPs have similar adsorption behaviors, and when the initial concentration of C3G is low, the adsorption amounts of the MCMIPs and the MCNIPs are continuously increased along with the increase of the concentration of anthocyanin because the surface has a large number of unoccupied binding sites. When the adsorption sites are all bound by C3G, the adsorption amount of the imprinted polymer can not increase with the increase of the concentration of anthocyanin, and the adsorption reaches saturation. The initial concentration and the maximum adsorption quantity of C3G corresponding to the equilibrium of adsorption of MCMIPs formed by different functional monomers are different according to the number of binding sites of the adsorbent and the adsorbateThe magnitude of the indirect force. Maximum adsorption of MCMIPs-DAAQ (112.44mg g)^-1) Because the three rings of DAAQ are in one plane, the degree of conjugation is high and the pi-interaction with C3G is strong.

Fitting an adsorption isothermal curve by adopting Langmuir and Freundlich models, analyzing the action mode of surface molecules when the MCMIPs adsorb target molecules, and researching whether the MCMIPs are adsorbed by monomolecular layers or polymolecular layers and whether binding sites distributed on the surface of the polymer are uniform or not, thereby knowing the adsorption mechanism of the imprinted polymer. The Langmuir model means that the surface-uniform adsorbent is in dynamic equilibrium with the adsorbate during the action process, and the whole adsorption process is monolayer adsorption. The Freundlich model represents a heterogeneous surface adsorbent, the adsorption process for the target molecule is a multi-molecular layer adsorption, and the ease of the whole adsorption process can be revealed. By comparing R²(see table 2), the Freundlich model fitted better, indicating that the adsorption process of MCMIPs to C3G is a multi-molecular layer adsorption and its surface recognition sites are not uniformly distributed. In addition, the ratio of 1/n to 0.5 shows that the MCPIPs are easy to adsorb C3G.

TABLE 2 isothermal adsorption Curve model parameters

Adsorption kinetics and its model: the rates of adsorption equilibrium of MCMIPs and the corresponding MCNIPs of embodiments one to seven were evaluated by adsorption kinetics, as shown in fig. 26 to 32. As can be seen from FIGS. 26-32, all 7 MCMIPs have a fast adsorption rate for C3G, and 80% of the maximum adsorption amount can be achieved at 240 min. The adsorption equilibrium of MCNIPs is reached before MCMIPs due to the fact that MCMIPs have more binding sites. The research mainly analyzes the adsorption kinetics of the MCMIPs by constructing an adsorption kinetics quasi-first-stage model and an adsorption kinetics quasi-second-stage model, so that the adsorption mechanism of the MCMIPs is discussed. The adsorption dynamics quasi-first-level model is suitable for an ideal single-factor environment, and if the reaction rate and the concentration of a reactant are in a linear relation, the adsorption process of the adsorbent is expandedThe bulk step control is a physical adsorption process. The absorption kinetic quasi-second order model reflects the chemical absorption process of electron transfer or electron sharing between the absorbent and the adsorbate. Comprehensively comparing the two dynamic models, and fitting a curve R through a dynamic quasi-secondary model²Closer to 1, the calculated theoretical maximum adsorption was closer to the actual value (see table 3), indicating that the MCMIPs adsorption rate is determined by the square of the number of adsorption sites that are unoccupied on the adsorption surface.

TABLE 3 adsorption kinetics model parameters

Adsorption selectivity: the adsorption selectivity of the MCMIPs and the corresponding MCNIPs of the first to seventh embodiments is detected, as shown in fig. 33 to 39, and as can be seen from fig. 33 to 39, the adsorption amount of the MCMIPs to C3G is significantly higher than that of other structural analogs, which indicates that the MCMIPs have the specific recognition capability for anthocyanin. The analysis reason is that when the MCMIPs adsorb C3G, C3G as a template molecule perfectly matches in shape and size with the imprinted pores of the polymer, while adsorption of other substances is mainly by ordinary physical action and has no chemical binding sites matching with it. Furthermore, the selectivity factor of MCMIPs for competitors (see tables 4-10) is in order: ru, Qu < p-HPA CA, SAA, SIA < Nar, Amy. The reason for this is that Ru, Qu and C3G have similar aglycones. The structures of p-HPA, CA, SAA and SIA are similar to those of C3G, but can enter the hole though not matching with the imprinted hole due to their small sizes. The minimum adsorption of MCMIPs to both Nar and Amy is due to greater steric hindrance. In addition, the conjugation degree of Nar and Amy structures is small, and the acting force between the Nar and Amy structures and the MCMIPs is weak.

TABLE 4 partition coefficients (K) of MCMIPs-ODA and MCNIPs-ODA for different competitors_d) Selecting a factor (k)^sel) Relative selection factor (k)^rel)

TABLE 5 partition coefficients (K) of MCMIPs-BD and MCNIPs-BD for different competitors_d) Selecting a factor (k)^sel) Relative selection factor (k)^rel)

TABLE 6 partition coefficients (K) of MCMIPs-Pa-1 and MCNIPs-Pa-1 for different competitors_d) Selecting a factor (k)^sel) Relative selection factor (k)^rel)

TABLE 7 partition coefficients (K) of MCMIPs-DT and MCNIPs-DT for different competitors_d) Selecting a factor (k)^sel) Relative selection factor (k)^rel)

TABLE 8 partition coefficients (K) of MCMIPs-DADHAQ and MCNIPs-DADHAQ for different competitors_d) Selecting a factor (k)^sel) Relative selection factor (k)^rel)

TABLE 9 partition coefficients (K) of MCMIPs-DHBD and MCNIPs-DHBD for different competitors_d) Selecting a factor (k)^sel) Relative selection factor (k)^rel)

TABLE 10 partition coefficients (K) of MCMIPs-DAAQ and MCNIPs-DAAQ for different competitors_d) Selecting a factor (k)^sel) Relative selection factor (k)^rel)

The repeated utilization rate is as follows: ten adsorption elution cycle experiments are performed on the MCMIPs of the first to seventh embodiments, the adsorption amount of the MCMIPs is calculated after each adsorption, and a reuse factor graph of the MCMIPs is drawn, as shown in fig. 36 to 42.

And (4) conclusion: from fig. 40-46, it can be seen that after 10 times of repeated adsorption, the adsorption amount of the MCMIPs is reduced by only 10.57% -17.83%, and the experimental results prove that the binding sites of the MCMIPs are reproducible and reusable. In addition, the MCMIPs have good stability, and the framework structure cannot be degraded and collapsed under an acidic condition.

The decrease in adsorption capacity is mainly the plugging and loss of sites.

Purification of crude anthocyanin extract: C3G in the sorbus nigricans, Vaccinium myrtillus and blueberry samples is respectively purified by using MCMIPs-DAAQ obtained in the sixth specific embodiment, the results are shown in FIGS. 47-49, and it can be seen from FIGS. 47-49 that after MCMIPs-DAAQ purification, the purities of C3G are 94%, 88% and 90% respectively. The experimental result shows that the novel adsorption medium MCMIPs designed according to the C3G structure can intelligently recognize and capture C3G, and the rapid and efficient separation of anthocyanin is realized.

Claims

1. A preparation method of a magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin is characterized by comprising the following steps:

mono, superparamagnetic amino functional Fe₃O₄Preparing magnetic nanoparticles: FeCl is added₃·6H₂Adding O into ethylene glycol, performing ultrasonic treatment to completely dissolve, adding anhydrous sodium acetate and 1, 6-hexanediamine, and mechanically stirring to obtain redBrown solution, reacting the obtained reddish brown solution at the temperature of 150-250 ℃ for 6-12 h, then performing magnetic separation and sedimentation, washing the separated solid matter, removing unreacted matter, and performing vacuum drying to obtain superparamagnetic amino functionalized Fe₃O₄Magnetic nanoparticles, noted Fe₃O₄-NH₂Magnetic nanoparticles;

secondly, preparing the magnetic covalent organic framework molecularly imprinted polymer, namely ① adding C3G and functional monomers into absolute ethyl alcohol, performing ultrasonic treatment until the monomers are completely dissolved, performing prepolymerization reaction for 3-5 h at room temperature to obtain a prepolymerization solution, and ② adding the Fe obtained in the step one into the prepolymerization solution₃O₄-NH₂Magnetic nanoparticles, sonicated to Fe₃O₄-NH₂Uniformly dispersing magnetic nanoparticles in a prepolymerization solution, adding 1,3, 5-trialdehyde phloroglucinol, mechanically stirring, adding scandium trifluoromethanesulfonate, reacting at room temperature to obtain a polymer, ③ washing the obtained polymer to obtain a clear solution, repeatedly eluting the clear solution by using a mixed solution of methanol and acetic acid to remove C3G until no C3G is detected in an eluent, ④ finally washing away residual acetic acid by using double distilled water, and performing vacuum drying to obtain the magnetic covalent organic framework molecularly imprinted polymer, which is marked as MCMIPs.

2. The method for preparing an anthocyanin-isolated magnetic covalent organic framework molecularly imprinted polymer as claimed in claim 1, wherein FeCl is added in the first step₃·6H₂The mass ratio of O to the volume of ethylene glycol is (12-15) g: 500 mL.

3. The method for preparing an anthocyanin-isolated magnetic covalent organic framework molecularly imprinted polymer as claimed in claim 1, wherein FeCl is added in the first step₃·6H₂The mass ratio of the O to the anhydrous sodium acetate is (12-15) g: 27 g.

4. The magnetic covalent organic framework of claim 1 for separating anthocyaninsThe preparation method of the molecularly imprinted polymer is characterized in that FeCl is adopted in the step one₃·6H₂The ratio of the mass of O to the volume of 1, 6-hexanediamine is (12-15) g: 172 mL.

5. The method for preparing the magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin according to claim 1, wherein the reddish brown solution obtained in the first step is reacted for 6 hours at the temperature of 180-200 ℃.

6. The method of claim 1, wherein in step two ①, the functional monomer is p-phenylenediamine, p-diaminobiphenyl, 3' -dihydroxybenzidine, 4 "-diaminodiphenyl ether, 4" -diamino-p-terphenyl, 2, 6-diaminoanthraquinone, or 1, 5-diamino-4, 8-dihydroxy-9, 10-anthracenedione.

7. The method for preparing a magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin of claim 1, wherein the ratio of the amount of the C3G to the amount of the functional monomer in the second step ① is 0.12 (0.4-0.55).

8. The method for preparing a magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin of claim 1, wherein the ratio of the amount of C3G substance to the volume of absolute ethyl alcohol in the second step ① is 0.12mmol (5-15) mL.

9. The method for preparing an anthocyanin-isolated magnetic covalent organic framework molecularly imprinted polymer as claimed in claim 1, wherein the amount of C3G substance and Fe in step two ② are equal₃O₄-NH₂The mass ratio of the magnetic nanoparticles is 0.12 mmol: (40-60) mg.

10. The method for preparing a magnetic covalent organic framework molecularly imprinted polymer for separating anthocyanin according to claim 1, wherein the ratio of the amount of substance of tri-C3G to the volume of 1,3, 5-trialdehyde-m-benzene in step two ② is 0.12mmol (5-15) mL, and the ratio of the amount of substance of tri-C3G to the volume of scandium trifluoromethanesulfonate in step two ② is 0.12mmol (0.5-1.5) mL.