Development Of Sample Preparation Method Based On Molecularly Imprinted Polymers And Its Application In The Field Of Complex Samples Analysis

Objective:（1）Facile preparation of a novel deep eutectic solvent（DESs）for the first step extraction of oxymatrine and matrine from sophora flavescens.Novel magnetic molecularly imprinted polymers（MMIPs）were prepared and used as adsorbent for dispersive solid-phase extraction（D-SPE）to separate and enrich matrine and oxymatrine from DESs extracts.（2）By using carbon dots（CDs）as detection unit and MIPs as enrichment unit,novel CDs@MIPs composite materials were constructed for the enrichment and detection of 4-Nitrophenol（4-NP）in environmental samples.Methods:（1）Firstly,a series of DESs using choline chloride（ChCl）and tetrabutylammonium bromide（TBAB）as hydrogen bond receptors（HBA）with different hydrogen bonds（HBDs,organic acids and polyols）were successfully prepared.Then,by using DESs as extraction solvents,oxymatrine and matrine in sophora flavescens were effectively extracted by ultrasonic-assisted extraction method.Furthermore,by using the extraction efficiencies of oxymatrine and matrine as optimized indexes,the types of DESs,water content in DESs,solid-liquid ratio,extraction time and extraction temperature were systematically optimized,so as to obtain the best extraction conditions of oxymatrine and matrine.Quantitative analysis of oxymatrine and matrine in DESs extracts was conducted by using HPLC-DAD.Further,magnetic molecularly imprinted polymers（MMIPs）were successfully synthesized by using oxymatrine as template,acrylic acid as functional monomer,ethylene glycol dimethyl acrylate as cross-linker,AIBN as initiator,acetonitrile-toluene（3:1,v/v）as porogen and Fe₃O₄ as nuclear.The physical and chemical properties of MMIPs were characterized by transmission electron microscopy（TEM）,Fourier transform infrared spectroscopy（FT-IR）,vibrating sample magnetometer（VSM）and X-ray Diffraction（XRD）.Static and dynamic experiments were used to evaluate the adsorption capacity of MMIPs.Besides,the desorption conditions and reusability of MMIPs were investigated in detail.Finally,MMIPs were used as adsorbents for D-SPE for the selective separation and enrichment of oxymatrine and matrine from DESs extracts.（2）CDs with blue fluorescnece were synthesized through hydrothermal method by using anhydrous citric acid as carbon source and AEAPMS as surface modification agent.Then,using 4-NP as template,（3-aminopropyl）triethoxysilane（APTES）as functional monomer,tetraethoxysilane（TEOS）as cross-linker and CDs as detection signal source,CDs@MIPs was prepared through a sol-gel method.The CDs@MIPs was characterized by TEM,DLS（Dynamic light scattering）,FT-IR,Zeta potential and XRD.After that,the fluorescent detection conditions of 4-NP were optimized,and the selectivity and sensitivity of CDs@MIPs were also explored.Fluorescent life test and fluorescence quenching rate constants at different temperatures were further applied to explore the quenching mechanism of polymers.At last,CDs@MIPs were applied to the analysis of 4-NP in real water samples.Results:（1）The optimum extraction conditions for oxymatrine were as followed:extraction solvent,DESs-2;water content in DESs,50%;solid-liquid ratio,5:1;extraction time,40min;extraction temperature,50 ℃.The optimum extraction conditions for matrine were as followed:extraction solvent,DESs-8;water content in DESs,30%;solid-liquid ratio,25:1;extraction time,40 min;extraction temperature,50 ℃.The extraction efficiencies of oxymatrine and matrine under each optimum extraction conditions were caluculated as 21.04 and 1.53mg·g^-1,respectively,which were significantly higher than those of traditional extraction solvents.In addition,MMIPs were synthesized under the optimum conditions.The results of SEM indicated that the prepared MMIPs presented typical core–shell structure and the layer of MIPs was approximately 50 nm.FT-IR spectra revealed that Fe-O,Si-O-Si,C=C and C=O existed on MMIP,indicating that MMIPs were successfully prepared.The results of VSM showed that MMIPs had good magnetic properties.Furthermore,the results of static adsorption experiments showed that MMIPs had good adsorption performance for oxymatrine and matrine,and the maximum adsorption capacities of them were calculated as 110.8 and 63.4 mg·g^-1,respectively.The dynamic adsorption experiments showed that the saturation adsorption time of oxymatrine and matrine was about at 60 min.Moreover,the maximum adsorption capacities of MMIPs for oxymatrine and matrine were much higher than MNIPs,indicating that MMIPs had specific binding sites for two alkaloids.Besides,the optimal desorption solvent for oxymatrine was methanol-acetic acid（9:1,v/v）,and the optimal desorption time was 90 min.The optimal desorption solvent of matrine was methanol,and the optimal desorption time was 60 min.The MMIPs were still maintained good adsorption capacity after six adsorption-desorption cycles,indicated that the MMIPs had good reusability.Finally,MMIPs were used as D-SPE adsorbents for the separation and enrichment of oxymatrine and matrine in DES extracts,and the extraction efficiencies of them ranged between 80.21～89.15%and 85.33～95.28%,respectively.These results showed that the prepared MMIPs could be effectively used for the enrichment of oxymatrine and matrine in actual samples.（2）Blue fluorescence CDs with quantum yield of 51.8%and CDs@MIPs composites were successfully prepared.The results of TEM demonstrated that the CDs are spherical and the average diameter was about 2.2 nm.The CDs@MIPs had a spherical nanostructure and the average particle size were about 20 nm.Compared the results of FT-IR spectra of CDs@MIPs before and after removing the templates,it can be easily found that without removing the template,the strong peak at 1495cm^-1 was attributed to the symmetric stretching vibration of the-NO₂ for the 4-NP,which proved that the templatewas successfully imprinted in CDs@MIPs.However,this characteristic peak disappeared after elution of the template,indicating that 4-NP was successfully eluted.The strong and broad peak of CDs around1069 cm^-1 was assigned to Si-O-Si asymmetric stretching,the bands of CDs at 3454 cm^-1（N-H bond）and 2932 cm^-1（C-H stretching band）were attributed to the aminopropyl groups.These all results indicated that CDs existed in the polymers.Moreover,it can be easily obtained from the results of fluorescence analysis that the maximum excitation wavelength was 370 nm while the corresponding maximum emission wavelength was about 470 nm.Moreover,CDs@MIPs showed good fluorescence stability.In order to obtain the optimal detection condition for 4-NP,several parameters were optimized and the results indicated that the optimum detection condition for 4-NP detection were as followed:p H=5;Na Cl with 0 M;water as solvent;optimal concentration of CDs@MIPs was10 mg·m L^-1;optimum incubation time was 4 min.Under the optimal conditions,the fluorescence detection response range of CDs@MIPs for 4-NP was 0.025μg﹒m L^-1-5μg﹒m L^-1,and the LOD was 5 ng·m L^-1（35 n M）.The imprinting factor（IF）of CDs@MIPs and CDs@NIPs was 6.69.The above results showed that the composite materials combined with molecular imprinting technology could significantly improve the sensitivity and selectivity to 4-NP.Moreover,the fluorescence response of CDs@MIPs toward 4-NP was much higher than those of other five analogs and metal ions,demonstrating that CDs@MIPs had a better binding ability and selectivity for 4-NP.Furtehrmore,the result of quenching mechanism showed that 4-NP can quench the fluorescence of composite materials through static quenching.Finally,the CDs@MIPs were applied to the detection and enrichment of4-NP in real water samples,and the relative recoveries of 4-NP ranged from 88.0%～103.4%.This result revealed that the prepared CDs@MIPs could be effectively used for the analysis of 4-NP in actual samples.Conclusion:（1）DESs had outstanding properties and can be used to extract alkaloids from TCMs instead of traditional organic solvents.MMIPs can be used for the selectively separation and enrichment of oxymatrine and matrine.The first extraction based on DESs combined with secondary enrichment method based on MMIPs can be applied for the efficient purification and enrichment of oxymatrine and matrine in complex TCMs samples.（2）The new fluorescent composites（CDs@MIPs）which were constructed by using CDs as the detection unit and MIPs as the specific recognition site showed good fluorescence properties and selectivity.CDs@MIPs can be effectively used for the enrichment and sensitivity detection of 4-NP in environmental samples.