阅读说明：本技术 一种用于选择性分离富集马兜铃酸i的温敏型磁性分子印迹聚合物的制备方法 (Preparation method of temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I ) 是由 万益群 熊辉煌 郭岚 万昊 范庸 郭娴 于 2021-02-23 设计创作，主要内容包括：本发明公开了一种用于选择性分离富集马兜铃酸I的温敏型磁性分子印迹聚合物的制备方法,其由以下步骤制备而成：(1)制备氧化石墨烯；(2)制备磁性氧化石墨烯；(3)制备二氧化硅包覆的磁性氧化石墨烯；(4)将上述二氧化硅包覆的磁性氧化石墨烯作为载体,以马兜铃酸I为模板分子,N-异丙基丙烯酰胺为温敏型功能单体,甲基丙烯酸为功能单体,二乙烯基苯为交联剂,偶氮二异丁腈为引发剂,在载体表面制备对马兜铃酸I具有特异识别能力的温敏型磁性分子印迹聚合物。本发明制备的温敏型磁性分子印迹聚合物在外界磁场下快速分离,对目标化合物有较高的特异性吸附能力和识别速率,可重复使用,其吸附/解吸过程随温度变化可逆、可控,对环境友好。(The invention discloses a preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I, which is prepared by the following steps: (1) preparing graphene oxide; (2) preparing magnetic graphene oxide; (3) preparing magnetic graphene oxide coated by silicon dioxide; (4) the magnetic graphene oxide coated by the silicon dioxide is used as a carrier, aristolochic acid I is used as a template molecule, N-isopropylacrylamide is used as a temperature-sensitive functional monomer, methacrylic acid is used as a functional monomer, divinylbenzene is used as a cross-linking agent, and azodiisobutyronitrile is used as an initiator, so that the temperature-sensitive magnetic molecularly imprinted polymer with the specific recognition capability on the aristolochic acid I is prepared on the surface of the carrier. The temperature-sensitive magnetic molecularly imprinted polymer prepared by the invention can be rapidly separated under an external magnetic field, has higher specific adsorption capacity and recognition rate on a target compound, can be repeatedly used, has reversible and controllable adsorption/desorption process along with temperature change, and is environment-friendly.)

1. A preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I is characterized by comprising the following steps:

(1) preparing graphene oxide;

(2) preparing magnetic graphene oxide;

(3) preparing magnetic graphene oxide coated by silicon dioxide;

(4) and (3) preparation of the temperature-sensitive magnetic molecularly imprinted polymer.

2. A preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I is characterized by comprising the following steps:

(1) preparation of graphene oxide

Weighing 1.0 g of graphite and 1.2 g of potassium nitrate, uniformly mixing, adding 46 mL of iced 98% concentrated sulfuric acid, stirring for 30 min under the condition of an ice-water bath, slowly adding 6.0 g of potassium permanganate into the mixed dispersion solution, continuously stirring for 15 min in the ice-water bath, removing the ice-water bath, raising the temperature of a reaction system to 35 ℃, stirring for 1 h, then slowly adding 90 mL of deionized water, raising the temperature of the reaction system to 98 ℃, continuously stirring for 30 min, then slowly pouring into 200 mL of hot deionized water, controlling the reaction temperature to be 98 ℃, then dropwise adding 6 mL of 30% hydrogen peroxide until the color of the solution is changed from dark brown to bright yellow, centrifuging while hot, washing a centrifugal precipitate with 20 mL of 5% hydrochloric acid and 200 mL of deionized water for three times to obtain a neutral supernatant without SO, and sequentially washing the precipitate with 20 mL of 5% hydrochloric acid and 200 mL of deionized water for₄ ^2- Residual and vacuum drying to obtain graphene oxide;

(2) preparation of magnetic graphene oxide

Weighing 0.45 g of graphene oxide, adding the graphene oxide into 70 mL of ethylene glycol, performing ultrasonic dispersion for 3 hours, sequentially adding 1.4 g of ferric chloride hexahydrate and 2.8 g of anhydrous sodium acetate, stirring for 30 minutes, transferring the mixture into a 100 mL polytetrafluoroethylene and stainless steel autoclave, heating for 8 hours at 200 ℃, separating under the action of an external magnetic field, sequentially leaching the product with ultrapure water and ethanol, and performing vacuum drying to obtain magnetic graphene oxide;

(3) preparation of magnetic graphene oxide coated with silicon dioxide

Weighing 0.5 g of magnetic graphene oxide, dispersing in 49.5 mL of absolute ethyl alcohol, adding 6.3 mL of deionized water and 2.2 mL of ethyl orthosilicate in an ice-water bath, stirring for 10 min, dropwise adding 2 mL of concentrated ammonia water, continuously stirring for 12 h, separating the mixture in an external magnetic field, washing the product with absolute ethyl alcohol for three times, and performing vacuum drying to obtain silicon dioxide coated magnetic graphene oxide;

(4) preparation of temperature-sensitive magnetic molecularly imprinted polymer

The method is characterized in that aristolochic acid I is taken as a template molecule, methacrylic acid is taken as a functional monomer, N-isopropyl acrylamide is taken as a temperature-sensitive functional monomer, divinylbenzene is taken as a cross-linking agent, azobisisobutyronitrile is taken as an initiator, and the corresponding molar ratio is as follows: the aristolochic acid I is N-isopropylacrylamide methacrylic acid divinylbenzene = 1: 6: 6: 20, and the addition amount of azobisisobutyronitrile is about 5% of the total mass of reactants, and the specific steps are as follows: 0.01365 g aristolochic acid I is weighed into a 100 mL round-bottom flask, 2.4 mL of N-isopropylacrylamide solution and methacrylic acid solution with the concentration of 100 mmol/L are added in sequence, 35.2 mL of acetonitrile is added, prepolymerization was carried out at 4 ℃ for 12 hours, 0.13019 g of divinylbenzene were mixed with 5 mL of acetonitrile solution and added to the above prepolymerization solution, then 150 mg of magnetic graphene oxide coated by silicon dioxide is added, nitrogen is introduced by ultrasonic, the reaction system is heated to 60 ℃, then, 10 mg of azobisisobutyronitrile is mixed with 5 mL of acetonitrile solution, the mixture is slowly added into the mixed solution drop by drop, the mixture is stirred for 24 hours under the nitrogen atmosphere, eluting the polymer obtained by separating and collecting the external magnetic field with acetonitrile, eluting with an ethanol-acetic acid solution with the volume ratio of 9:1, washing off redundant acetic acid with methanol, and drying in vacuum to obtain the temperature-sensitive magnetic molecularly imprinted polymer;

the blank temperature-sensitive magnetic molecularly imprinted polymer is prepared by the same method without adding template molecules.

Technical Field

The invention relates to a preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I, belonging to the technical field of functional material preparation and endogenous pollutant analysis.

Background

Aristolochic Acid I (AAI) is a nitrophenanthrene compound, and is widely present in plants of Aristolochia and Asarum of Aristolochiaceae. AAI-containing plants have been used for adjuvant treatment of arthritis, rheumatism, diuresis, venomous snake bite, cancer, etc. In recent years, various toxic and side effects of AAI are reported successively, the renal toxicity of AAI is most concerned, and multiple groups of genotoxicity experiments also prove that AAI has mutagenic and carcinogenic effects. At present, plants containing aristolochic acid are listed in the category 1 carcinogen list in the carcinogen primary list published by the world health organization international agency for research on cancer. Although the use of Chinese herbal medicines containing AAI is forbidden in many countries, the national food and drug administration of China also cancels the medicinal standards of Chinese herbal medicines such as caulis Aristolochiae Manshuriensis, radix Aristolochiae, etc., but improper use of the medicines still exists. Reports have demonstrated that some vegetal food supplements contain genotoxic and carcinogenic components and that risk assessment of these components, including AAI, has been performed. It has also been found that the common field aristolochia debilis weed releases free AAI upon decay, then enters soil and water, is absorbed by food crops (such as corn and wheat) growing nearby, and is transferred to their edible parts, which may ultimately pose a threat to human health. Therefore, it is necessary to establish an effective recognition and detection system for the endogenous pollutant aristolochic acid I.

Because the aristolochia plants and related food substrates are complex and the content of AAI in a sample is low, the sample pretreatment technology is particularly critical to realize rapid and accurate analysis. The sample pretreatment technology for extracting and separating AAI mainly comprises liquid-liquid extraction (LLE), solid-phase extraction (SPE) and the like. The liquid-liquid extraction has long time consumption, low efficiency and large consumption of organic solvent, is easy to cause secondary pollution to the environment and is not beneficial to the health of operators; the solid adsorbent commonly used in the solid phase extraction technology has poor selectivity to target compounds, low reusability and can not completely eliminate coexisting interferents. In order to further match with a high-sensitivity chromatographic analysis technology and adapt to the requirements of rapid and accurate detection of trace components in a complex matrix, the development of a high-selectivity separation and enrichment material is necessary. Compared with the traditional adsorption material, the intelligent molecularly imprinted polymer (SR-MIPs) prepared by combining the molecularly imprinted technology and the stimulus response technology has high affinity and high selectivity on target analytes, can realize controllable adsorption/desorption by changing external conditions (such as temperature, light, magnetism, pH and the like), has the advantages of good stability, simplicity in preparation, low cost, wide application range and the like, and is particularly suitable for selective separation and enrichment of trace analytes in complex matrix samples. In the prior published literature, the research of intelligent molecularly imprinted polymers for AAI has been rarely reported or is limited to magnetic molecularly imprinted polymers. Therefore, the development of other novel intelligent molecularly imprinted polymers has important significance for efficient separation and enrichment of AAI in a complex matrix, and a new research idea is provided for separation and analysis of endogenous pollutants.

Disclosure of Invention

The invention aims to overcome the defects and shortcomings in the prior art and provides a preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I.

The technical scheme of the invention is as follows: a preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I comprises the steps of taking aristolochic acid I as a template molecule, N-isopropylacrylamide as a temperature-sensitive functional monomer, methacrylic acid as a functional monomer, divinylbenzene as a cross-linking agent and azodiisobutyronitrile as an initiator, and preparing the temperature-sensitive magnetic molecularly imprinted polymer with specific recognition capability on the surface of a magnetic graphene oxide carrier coated with silicon dioxide.

The invention mainly comprises the following steps: (1) preparing Graphene Oxide (GO); (2) preparing magnetic graphene oxide (Mag @ GO); (3) magnetic graphene oxide coated with silicon dioxide (Mag @ GO @ SiO)₂) Preparing; (4) and (3) preparation of temperature-sensitive magnetic molecularly imprinted polymers (TMMIPs).

A preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I mainly comprises the following steps:

(1) preparation of Graphene Oxide (GO)

1.0 g of graphite and 1.2 g of potassium nitrate were weighed, mixed well, and 46 mL of iced concentrated sulfuric acid (98%) was added and stirred for 30 min under ice-water bath conditions. 6.0 g of potassium permanganate was slowly added to the above mixed dispersion solution, and stirring was continued in an ice-water bath for 15 min. Removing the ice water bath, raising the temperature of the reaction system to 35 ℃, stirring for 1 h, then slowly adding 90 mL of deionized water, raising the temperature of the reaction system to 98 ℃, and continuously stirring for 30 min; then slowly poured into 200 mL of hot deionized water, and the reaction temperature was controlled at 98 ℃. Then, 6 mL of hydrogen peroxide (30%) was added dropwise until the solution changed from dark brown to bright yellow in color; the mixture was centrifuged while hot, and the centrifuged precipitate was washed three times with 20 mL of hydrochloric acid (5%) and 200 mL of deionized water in that order until the supernatant was neutral and free of SO₄ ^2- And (4) residual and vacuum drying to obtain Graphene Oxide (GO).

(2) Preparation of magnetic graphene oxide (Mag @ GO)

0.45 g of GO is weighed and added into 70 mL of ethylene glycol, ultrasonic dispersion is carried out for 3 h, and 1.4 g of ferric chloride (III) hexahydrate and 2.8 g of anhydrous sodium acetate are sequentially added. After stirring for 30 min, the mixture was transferred to a 100 mL Teflon and stainless steel autoclave and heated at 200 ℃ for 8 h. Separating under the action of an external magnetic field, sequentially leaching the product with ultrapure water and ethanol, and drying in vacuum to obtain the magnetic graphene oxide (Mag @ GO).

(3) Magnetic graphene oxide coated with silicon dioxide (Mag @ GO @ SiO)₂) Preparation of

0.5 g of Mag @ GO is weighed and dispersed in 49.5 mL of absolute ethyl alcohol, 6.3 mL of deionized water and 2.2 mL of ethyl orthosilicate are added in an ice-water bath, after stirring for 10 min, 2 mL of strong ammonia water is added dropwise, and stirring is continued for 12 h. Separating the mixture in an external magnetic field, washing the product with absolute ethyl alcohol for three times, and drying in vacuum to obtain the magnetic graphene oxide (Mag @ GO @ SiO) coated with silicon dioxide₂）。

(4) Preparation of temperature-sensitive magnetic molecularly imprinted polymers (TMMIPs)

The method is characterized in that Aristolochic Acid I (AAI) is used as a template molecule, methacrylic acid (MAA) is used as a functional monomer, N-isopropylacrylamide (NIPAM) is used as a temperature-sensitive functional monomer, Divinylbenzene (DVB) is used as a cross-linking agent, Azobisisobutyronitrile (AIBN) is used as an initiator, and the corresponding molar ratio is N (AAI), N (NIPAM), N (MAA), N (DVB) = 1: 6: 6: 20, and the addition amount of AIBN is about 5% of the total mass of reactants. The method comprises the following specific steps: 0.01365 g AAI was weighed into a 100 mL round-bottomed flask, and 2.4 mL of a NIPAM solution and a MAA solution each having a concentration of 100 mmol/L were added in this order, and 35.2 mL of acetonitrile was added, followed by prepolymerization at 4 ℃ for 12 hours. 0.13019 g DVB was mixed with 5 mL acetonitrile and added to the pre-polymerization solution, then 150 mg Mag @ GO @ SiO₂And introducing nitrogen by ultrasonic waves, and heating the reaction system to 60 ℃. Subsequently, 10 mg of AIBN was mixed with 5 mL of acetonitrile solution, slowly added dropwise to the mixture, and stirred under nitrogen atmosphere for 24 hours. And leaching the polymer obtained by separating and collecting the external magnetic field with acetonitrile, eluting with an ethanol-acetic acid solution (9: 1, v/v), washing away excessive acetic acid with methanol, and drying in vacuum to obtain the temperature-sensitive magnetic molecularly imprinted polymer (TMMIPs).

The blank temperature-sensitive magnetic molecularly imprinted polymers (TMNIPs) are prepared by the same method without adding template molecules.

The invention has the beneficial effects that: the invention designs a preparation method of a temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I. The temperature-sensitive magnetic molecularly imprinted polymer prepared by the invention can be rapidly separated under an external magnetic field, has higher specific adsorption capacity and recognition rate on a target compound, can control the adsorption/desorption process of the target compound by adjusting the temperature, has simple preparation method and good recycling effect, and has great application prospect in the fields of separating and enriching endogenous pollutants and the like.

Drawings

FIG. 1 is a flow chart of the synthesis process of the temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I according to the present invention;

FIG. 2 shows GO (a), Mag @ GO (b), Mag @ GO @ SiO @, prepared in example 1₂(c) TMMIPs (d), TMNIPs (e);

FIG. 3 is GO, Mag @ GO @ SiO prepared in example 1₂TMMIPs and TMNIPs;

FIG. 4 is GO, Mag @ GO @ SiO prepared in example 1₂TMMIPs and TMNIPs;

FIG. 5 is GO, Mag @ GO @ SiO prepared in example 1₂Thermogravimetric plots of TMMIPs and TMNIPs;

FIG. 6 is a plot of nitrogen adsorption and desorption isotherms (A) and pore size distribution curves (B) for TMMIPs and TMNIPs prepared in example 1;

FIG. 7 is a graph of the temperature response performance of TMMIPs prepared in example 1, where (A) is the effect of temperature on TMMIPs adsorption and (B) is the effect of temperature on TMMIPs AAI release behavior;

FIG. 8 shows the static adsorption (A), dynamic adsorption (B) and specific adsorption (C) of TMMIPs and TMNIPs prepared in example 1;

FIG. 9 is an adsorption fit analysis of TMMIPs and TMNIPs prepared in example 1, wherein (A) is a static adsorption isotherm fit and (B) is an adsorption kinetics fit;

fig. 10 is a reusability study of TMMIPs prepared in example 1.

Detailed Description

Example 1:

the preparation method of the temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I mainly comprises the following steps: (1) preparing Graphene Oxide (GO); (2) preparing magnetic graphene oxide (Mag @ GO); (3) magnetic graphene oxide coated with silicon dioxide (Mag @ GO @ SiO)₂) Preparing; (4) and (3) preparation of temperature-sensitive magnetic molecularly imprinted polymers (TMMIPs). The preparation process is shown in figure 1.

(1) Preparation of Graphene Oxide (GO)

1.0 g of graphite and 1.2 g of potassium nitrate were weighed, mixed well, and 46 mL of iced concentrated sulfuric acid (98%) was added and stirred for 30 min under ice-water bath conditions. 6.0 g of potassium permanganate was slowly added to the above mixed dispersion solution, and stirring was continued in an ice-water bath for 15 min. Removing the ice water bath, raising the temperature of the reaction system to 35 ℃, stirring for 1 h, then slowly adding 90 mL of deionized water, raising the temperature of the reaction system to 98 ℃, and continuously stirring for 30 min; then slowly poured into 200 mL of hot deionized water, and the reaction temperature was controlled at 98 ℃. Then, 6 mL of hydrogen peroxide (30%) was added dropwise until the solution changed from dark brown to bright yellow in color; the mixture was centrifuged while hot, and the centrifuged precipitate was washed three times with 20 mL of hydrochloric acid (5%) and 200 mL of deionized water in that order until the supernatant was neutral and free of SO₄ ^2- And (4) residual and vacuum drying to obtain Graphene Oxide (GO).

(2) Preparation of magnetic graphene oxide (Mag @ GO)

0.45 g of GO is weighed and added into 70 mL of ethylene glycol, ultrasonic dispersion is carried out for 3 h, and 1.4 g of ferric chloride (III) hexahydrate and 2.8 g of anhydrous sodium acetate are sequentially added. After stirring for 30 min, the mixture was transferred to a 100 mL Teflon and stainless steel autoclave and heated at 200 ℃ for 8 h. Separating under the action of an external magnetic field, sequentially leaching the product with ultrapure water and ethanol, and drying in vacuum to obtain the magnetic graphene oxide (Mag @ GO).

(3) Magnetic graphene oxide coated with silicon dioxide (Mag @ GO @ SiO)₂) Preparation of

0.5 g of Mag @ GO is weighed and dispersed in 49.5 mL of absolute ethyl alcohol, 6.3 mL of deionized water and 2.2 mL of ethyl orthosilicate are added in an ice-water bath, after stirring for 10 min, 2 mL of strong ammonia water is added dropwise, and stirring is continued for 12 h. Separating the mixture in an external magnetic field, washing the product with absolute ethyl alcohol for three times, and drying in vacuum to obtain the magnetic graphene oxide (Mag @ GO @ SiO) coated with silicon dioxide₂）。

(4) Preparation of temperature-sensitive magnetic molecularly imprinted polymers (TMMIPs)

The method is characterized in that Aristolochic Acid I (AAI) is used as a template molecule, methacrylic acid (MAA) is used as a functional monomer, N-isopropylacrylamide (NIPAM) is used as a temperature-sensitive functional monomer, Divinylbenzene (DVB) is used as a cross-linking agent, Azobisisobutyronitrile (AIBN) is used as an initiator, and the corresponding molar ratio is N (AAI), N (NIPAM), N (MAA), N (DVB) = 1: 6: 6: 20, and the addition amount of AIBN is about 5% of the total mass of reactants. The method comprises the following specific steps: 0.01365 g AAI was weighed into a 100 mL round-bottomed flask, and 2.4 mL of a NIPAM solution and a MAA solution each having a concentration of 100 mmol/L were added in this order, and 35.2 mL of acetonitrile was added, followed by prepolymerization at 4 ℃ for 12 hours. 0.13019 g DVB was mixed with 5 mL acetonitrile and added to the pre-polymerization solution, then 150 mg Mag @ GO @ SiO₂And introducing nitrogen by ultrasonic waves, and heating the reaction system to 60 ℃. Subsequently, 10 mg of AIBN was mixed with 5 mL of acetonitrile solution, slowly added dropwise to the mixture, and stirred under nitrogen atmosphere for 24 hours. And leaching the polymer obtained by separating and collecting the external magnetic field with acetonitrile, eluting with an ethanol-acetic acid solution (9: 1, v/v), washing away excessive acetic acid with methanol, and drying in vacuum to obtain the temperature-sensitive magnetic molecularly imprinted polymer (TMMIPs).

The blank temperature-sensitive magnetic molecularly imprinted polymers (TMNIPs) are prepared by the same method without adding template molecules.

Effect embodiment:

characteristics of temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I

For GO, Mag @ GO @ SiO used in synthesis₂And TMMIPs and TMNIPs obtained by preparation are subjected to a series of material characterization, including transmission electron microscopy, Fourier infrared spectroscopy, X-ray diffraction, thermogravimetric analysis, nitrogen adsorption-desorption isotherms, pore size distribution and the like. The characterization results are as follows:

1. transmission electron microscope scanning analysis

From fig. 2 (a) it can be clearly seen that GO exhibits a wrinkled, lamellar structure. In FIG. 2 (b), Fe₃O₄The magnetic nanoparticles are in a regular spherical shape, are well assembled on the GO sheet layer, and have the particle size of about 173-205 nm. Through SiO₂After coating, the surface of the magnetic nanoparticles on GO in FIG. 2 (c) becomes smoother, and the particle size distribution is 205-259 nm. And Mag @ GO @ SiO₂In contrast, TMMIPs in FIG. 2 (d) consist of a solid core layer of dark color and a less colored imprinting layer, with a significant increase in size, approximately 307-329 nm. In FIG. 2 (e), TMNIPs and TMMIPs have similar morphology, and the particle size is about 236-309 nm. This indicates that the imprinted polymer has been modified at Mag @ GO @ SiO₂From the surface, it can be estimated that the thickness of the imprinted layer is about 35-51 nm.

2. Fourier infrared spectral analysis

3424 cm in FIG. 3^-1The vibration peak is determined to be-OH stretching vibration peak, 1720 cm^-1And 1619 cm^-1The peaks were C = O and skeletal carbon (C = C), 1225 cm^-1、1052 cm^-1Then the peak is the stretching vibration peak of C-O and alkoxy, and the existence of the peaks proves that the graphite is successfully changed into GO; 588 cm^-1Is the stretching vibration peak of Fe-O, which indicates that Fe₃O₄The magnetic nano particles cover the surface of the graphene oxide by 1082 cm^-1、957 cm^-1、801 cm^-1And 463 cm^-1The peak points are respectively the stretching vibration peaks of Si-O-Si, Si-O-H, Si-O and Si-O-Fe, which shows that the SiO₂The magnetic graphene oxide is successfully coated on the surface of the magnetic graphene oxide. 1652 cm^-1The absorption peak at (A) is the stretching vibration peak of C = C, 1729 cm^-1And 1439 cm^-1The peaks are absorption vibration peaks of C = O and C-N respectively, which shows that N-isopropylThe existence of two functional monomers of acrylamide and methacrylic acid. Furthermore, relative to Mag @ GO @ SiO₂Infrared spectra of (1), 1082 cm of TMMIPs and TMNIPs^-1And 957 cm^-1The intensity of the absorption peak is obviously reduced, which is caused by Mag @ GO @ SiO₂The surface is formed by polymer.

3. X-ray diffraction pattern analysis

As can be seen from fig. 4, a sharp diffraction peak near 11 ° 2 θ corresponds to the characteristic peak of the (001) crystal plane in GO, indicating that the graphite structure has been destroyed and the crystal structure of GO has been formed. After GO is modified, Fe appears near the 2 theta of 30 degrees, 35 degrees, 43 degrees, 53 degrees, 57 degrees and 63 degrees₃O₄Five characteristic diffraction peaks of the magnetic nano particles respectively correspond to Fe₃O₄The (220), (311), (400), (422), (511) and (440) crystal planes of (A) illustrate the Mag @ GO, Mag @ GO @ SiO prepared₂TMNIPs and TMMIPs all contain Fe₃O₄A crystalline phase.

4. Thermogravimetric analysis

As can be seen from FIG. 5, the thermogravimetric curve of GO shows a certain weight loss in the range of 55-150 deg.C, which is mainly due to the evaporation of the adsorbed water in the sample. While a large mass loss continues to occur around 218 ℃, due to the removal of a large number of oxygen-containing functional groups from the GO surface, it is also laterally demonstrated that the surface of the synthesized GO contains abundant oxygen-containing groups. Compared with GO, MaG @ GO and Mag @ GO @ SiO₂A large mass loss occurs around 500 deg.c due to the introduction of Fe in the GO lamellar structure₃O₄Magnetic nanoparticles, which enhance their stability; through SiO₂After coating, Mag @ GO @ SiO₂Has increased organic component content and has a mass loss greater than MaG @ GO. TMMIPs have greater loss in the range of 470-680 ℃ and then tend to be smooth, probably due to SiO₂The layer and the molecularly imprinted layer are caused by pyrolysis. The thermogravimetric curve trend of TMNIPs is substantially consistent with that of TMMIPs, but the thermal stability of the TMMIPs sample is better.

5. Nitrogen adsorption-desorption isotherm and pore size distribution

FIG. 6A shows TMMIPs and TMNIPsNitrogen adsorption-desorption isotherms. It can be seen that when the relative pressure P/P is reached₀When the value is less than 0.9, the adsorption amount of the gas is low, and the slope of the curve is low. However, when the relative pressure P/P₀When the value is more than 0.9, the gas adsorption amount is remarkably increased and the slope of the curve is remarkably increased. Meanwhile, the desorption curve is close to the adsorption curve, but is slightly higher than the adsorption curve, and the TMNIPs also have similar nitrogen adsorption and desorption isotherms. As can be seen from fig. 6B, the pore size distributions of TMMIPs and TMNIPs are relatively concentrated, and the average pore size is small. TMMIPs have a larger pore size than TMNIPs, demonstrating that specific recognition sites formed by the template molecules are formed in TMMIPs. These narrow pore sizes play an important role in the specific recognition of target compounds by TMMIPs.

(II) temperature stimulation responsiveness for selectively separating temperature-sensitive magnetic molecularly imprinted polymer enriched in aristolochic acid I

In order to examine the temperature stimulation responsiveness of TMMIPs, the specific experimental steps are as follows: (i) dispersing 3 mg TMMIPs in 3 mL of AAI methanol solution with the concentration of 30 mg/L, oscillating for 12 h under different temperature conditions (25-50 ℃), filtering by using a 0.22 mu m organic filter membrane after magnetic attraction, and measuring the AAI concentration in the supernatant by HPLC-DAD, wherein the adsorption amount of the TMMIPs to the AAI with the specific concentration is the initial amount minus the residual amount; (ii) dispersing 3 mg TMMIPs in 3 mL of AAI methanol solution with the concentration of 30 mg/L, oscillating for 12 h at the temperature of 35 ℃, separating by an external magnetic field, dispersing the TMMIPs adsorbed with the AAI in 3 mL of ethanol-acetic acid (9: 1, v: v) solution again, eluting for 12 h at different temperature conditions (25-50 ℃), filtering by a 0.22 mu m organic filter membrane after magnetic attraction, measuring the AAI concentration in the supernatant by HPLC-DAD, and calculating the desorption capacity.

Temperature stimulus responsiveness results as shown in fig. 7, as the temperature increases, the affinity of tmpmis for the target compound AAI increases, and when the temperature is 35 ℃, tmpmis shows a higher adsorption amount, and when the temperature is higher than 35 ℃, the recognition ability of tmpmis for the target compound is greatly reduced. This shows that TMMIPs has thermal switch effect on AAI along with temperature change, because the temperature-sensitive functional monomer NIPMA is added in the polymerization process, the lower critical temperature (LCST) is about 33 ℃, and under the temperature stimulation, the hydrophilic amide group and the critical hydrophobic isopropyl group in the NIPMA can cause the TMMIPs to generate reversible volume transition. That is, when the temperature is higher than the LCST (e.g., 45 ℃ and 50 ℃), the AAI is prevented from entering the imprinted cavity due to shrinkage of the polymer by the NIPAM unit, thereby exhibiting weak reactivity. On the contrary, when the temperature is close to LCST (such as 35 ℃), the NIPAM unit causes the swelling of the polymer, so that the number of binding sites with AAI is increased, and the imprinting effect is obviously improved. Therefore, under the stimulation of different temperatures, the controllable identification and controllable elution of the template molecule AAI can be realized, 35 ℃ is used as the optimal adsorption temperature of TMMIPs on the target compound AAI, and 45 ℃ is used as the optimal desorption temperature.

(III) adsorption performance of temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I

The adsorption performance of the temperature-sensitive magnetic molecularly imprinted polymer is mainly investigated through a static adsorption experiment, a dynamic adsorption experiment and a selective adsorption experiment, and each group of experiments are measured in parallel for three times. (i) The procedure for the static adsorption experiment was as follows: 3 mg TMMIPs/TMNIPs were dispersed in 3 mL of AAI methanol solutions of different concentrations, shaken at 35 ℃ for 12 h, magnetically absorbed, filtered through a 0.22 μm organic filter, and the AAI concentration in the supernatant was determined by HPLC-DAD. (ii) The dynamic adsorption experiment was performed as follows: dispersing 40 mg TMMIPs/TMNIPs in 40 mL AAI methanol solution with concentration of 15 mg/L, oscillating at 35 deg.C, taking out a certain amount of solution at different time intervals, magnetically adsorbing, filtering with 0.22 μm organic filter membrane, and quantifying with HPLC-DAD. (iii) The procedure for the selective adsorption experiment was as follows: 3 mg TMMIPs/TMNIPs were dispersed in 3 mL of a mixed solution of AAI and its structural analogs (aristolochic acid II (AAII), aristololactam I (ALI), benzoic acid, p-nitrophenol, and tanshinone IIA) at a concentration of 15 mg/L, shaken at 35 ℃ for 12 h, magnetically adsorbed, filtered through a 0.22 μm organic filter membrane, and the AAI concentration in the supernatant was determined by HPLC-DAD.

The static adsorption results are shown in FIG. 8A, the static saturation adsorption amounts of TMMIPs and TMNIPs to AAI are respectively 8.51 mg/g and 2.60 mg/g, and the imprinting factor alpha is3.27. The adsorption curves of TMNIPs and TMMIPs are consistent in trend, but the adsorption amount of AAI is lower at each concentration. To further analyze the state of static adsorption, Langmuir and Freundlich isotherm model fitting treatments were performed on the experimental data of TMMIPs/TMNIPs to evaluate the binding uniformity, and the results are shown in FIG. 9A. For TMMIPs, the fitting results of the two models are similar, wherein Freundlich has the best fitting effect and the correlation coefficient R² = 0.8752. In addition, the maximum theoretical adsorption amounts of TMMIPs and TMNIPs to AAI can reach 12.89 mg/g and 3.15 mg/g respectively through calculation of a Langmuir model, and certain reference is provided for analysis of experimental results.

The dynamic adsorption results are shown in fig. 8B, where the adsorption amount of tmpmis to AAI is greater than that of tmpips in each time period. TMMIPs adsorption rate is fast first and then slow, the adsorption is balanced in the first 60 min, TMNIPs can reach an adsorption saturation state within about 210 min, and the adsorption quantity is very low. Therefore, the TMMIPs have more effective recognition sites, and have stronger and rapid binding effect with the template molecule AAI. In order to further research the dynamic recognition capability, four dynamic adsorption models, namely Pseudo-first-order, Pseudo-second-order, Elovich and Intra differential diffusion, are adopted for fitting to obtain relevant parameters. As shown in FIG. 9B, in which the Pseudo-second-order model has the highest fitting effect, the correlation coefficient is the highest (R)² = 0.9886), provides the most suitable adsorption model for the process of adsorption of AAI by tmpims. The theoretical adsorption capacities of TMMIPs and TMNIPs in equilibrium are respectively 9.10 mg/g and 2.53 mg/g through calculation of a Pseudo-second-order model, and the results are consistent with the results of saturated adsorption capacity measured by experiments, so that the kinetic identification process can be well predicted. Therefore, the dynamic adsorption process of TMMIPs conforms to the Pseudo-second-order kinetic model.

The selective adsorption results are shown in fig. 8C, where TMMIPs have the highest AAI adsorption capacity and have a small amount of adsorption to compounds with similar structures, and TMNIPs have lower adsorption capacities to various compounds, and are roughly irregular. For compounds with simpler molecular structures or structures close to AAI, such as AAII and ALI, the adsorption capacity of TMMIPs is relatively higher; for compounds with molecular structures greatly different from AAI, such as benzoic acid, p-nitrophenol, tanshinone IIA and the like, the adsorption quantity of TMMIPs is low, the selectivity coefficient is in the range of 1.49-3.12, and the TMMIPs are proved to have identification specificity on AAI.

(IV) reproducibility of temperature-sensitive magnetic molecularly imprinted polymer for selectively separating and enriching aristolochic acid I

In order to evaluate the reuse capacity of TMMIPs, the specific flow is as follows: 30 mg TMMIPs were dispersed in 30 mL of 10 mg/L AAI methanol solution, shaken at 35 ℃ for 12 hours, magnetically absorbed, filtered through a 0.22 μm organic filter, and the concentration of AAI in the supernatant was determined by HPLC-DAD. TMMIPs adsorbing the target molecule were eluted at 45 ℃ using an ethanol-acetic acid (9: 1, v: v) solution as a solution, dried, and then the above experiment was repeated.

As shown in fig. 10, after seven adsorption-desorption cycles, the adsorption capacity of the mips to the AAI is not significantly reduced, and the good recovery effect is maintained, with a recovery rate of 91.16% and a standard error of less than 1.07%, and after more than seven times of repeated use, the recovery effect is initially less than 90%. The synthetic imprinted polymer has good reusability and regeneration capacity, has high recovery rate and good stability for the template molecule AAI, can meet the requirement of repeated use, has strong economic practicability, and is beneficial to popularization.

The foregoing is a more detailed description of the invention in connection with specific preferred embodiments and it is not intended that the invention be limited to these specific details. It should be understood by those skilled in the art that various changes and substitutions may be made in accordance with the technical solution and the inventive concept of the present invention, and the same properties or uses should be considered as the protection scope of the present invention.