Isolation,Extraction And Study Of Flavonoids Based On Molecular Imprinted Technology

Myricetin is a major representative of the flavonol,which has multiple activities,including: elimination free radicals,anti-oxidation,anti-tumor,reduce neurotoxicity,affect lymphocyte activation and proliferation,anti-platelet activating factor,antagonism hypoglycemic effect,relieve delight alcohol intoxication,prevention of alcoholic liver,prevention of fatty liver,inhibit the deterioration of liver cells,reduce the incidence of liver cancer,hepatoprotective effect,anti-inflammatory and other pharmacological activities.In recent years,researchers have been researching flavonoids and the flavonoids compounds have received much attention of the researchers.In the previous research methods,most of them were detected and peparated by high performance liquid chromatography,solid phase extraction,high performance liquid chromatography-mass spectrometry.Because of the complexity similarity of the structure of flavonoids,which leads to some obstacles in the detection and separation.Molecular imprinted polymers(MIPs)are a newtype of polymer materials,which are crosslinked synthetic macromolecules with template molecular and functional monomer polymerized by crosslinking agent and initiator.The three-dimensional cavities are matched with the size and shape of the molecular imprinted polymer,and these cavities have the capacity of selective recognition for the template molecular.The polymers have selective recognition capacity,high reservation,high mass transfer rate and wide application range,and they had been widely appled for various fields,which includes chromatographic separation,solid-phase extraction,chemical bionic sensor,natural antibody simulation,simulated enzyme catalysis,clinical drug analysis,membrane separation and so on.Therefore,it is of great significance to detect and separate myricetin by applying of molecular imprinted polymers.In this study,we comprehensively researched the preparation principle and application of molecular imprinted polymer,and we successfully extracte and separate the myricetin from the actual sample.In the experiment,the myricetin molecular imprinted polymers were prepared using myricetin as template molecular by molecular imprinted technology.The polymers were analyzed by the characterization measures of Fourier transform infrared spectroscopy(FT-IR),thermo-gravimetric analysis(TGA)and scanning electron microscopy(SEM)and so on.Besides,we also studied the isotherms adsorption properties,binding kinetic capacity,chromatographic properties.The specific research contents are as follows:1.In chapter one,the research progress of molecular imprinted polymers(MIPs)has been reviewed.The principle of molecular imprinted polymers prepared by molecular imprinted technique and the factors that affect the synthesis of polymer materials are systematically introduced.In this paper,the methods of separation and analysis of myricetin and other flavonoids were reviewed.The purpose and significance of this work were put forward.2.In the second chapter,a rapid method for the specific adsorption of myricetin was established.The myricetin was selected as the template molecular,4-vinyl pyridine(4-Vpy)was selected as functional monomer,ethylene glycol dimethacrylate(EGDMA)was selected as cross linking agent,2,2-azobisisobutyronitrile(AIBN)was selected as initiator,which could prepare the conventional myricetin molecular imprinted polymers(MIPs).The influence of the template molecular,the ratio of different functional monomers and porogen solvent on the properties of the materials were investigated.The imprinted efficiency was explored by isotherms adsorption,binding kinetic adsorption,Scatchard analysis and selective recognition.The experimental results show that this polymer material has specific adsorption on the target flavonoids myricetin.3.In the third chapter,the myricetin surface molecular imprinted polymer(SMIPs)was successfully prepared by surface molecular imprinting with a particle size of 5 ?m silica microspheres,and the imprinted polymer material was manually packed as high performance liquid chromatography column.The sieved uniform SMIP powders were packed in stainless steel columns of 250 mm?4.6 mm by a slurry packing method.Scanning electron microscopy(SEM),Fourier transform infrared spectroscopy(FT-IR)and thermo gravimetric analysis(TGA)were used as characterization measures to analysize the structure of the imprinted polymers.We also studied the imprinted efficiency of the polymers by isotherms adsorption,binding kinetic adsorption and selective recognition.The obtained imprinted polymers were grinded into uniform size powders packed into high performance liquid chromatography column,then the packed column was connected to the Agilent 1100 series HPLC,and the column was investigated its chromatographic evaluation.Compared to the Thermo Scientific C18 column,the imprinted polymers chromatography stationary phase possesses a better adsorption capacity.Therefore,the surface molecular imprinted polymers stationary phase was used to detecte and separate four kinds of flavonoid mixture,the experiment results show that the surface molecular imprinted polymers stationary phase separated the flavonoid mixture completely and the imprinted polymers indeed a potentially promising stationary phase for the separation of natural complex compounds.4.In the fourth chapter,the solid phase extraction adsorbent was prepared by surface molecular imprinted technology on the 40 ?m silica microspheres.In this experiment,the myricetin was selected as the template molecular,4-vinyl pyridine(4-Vpy)was selected as functional monomer,the myricetin surface molecular imprinted polymers were successfully synthesized on the surface of 40 ?m silica microspheres,then the polymers were packed as new solid phase extraction adsorbent combined with high performance liquid chromatography(HPLC)to enrich and separate flavonoid compound in the fox grape.The surface molecular imprinted polymers(SMIPs)were characterized by scanning electron microscopy(SEM),Fourier transform infrared spectroscopy(FT-IR)and thermo gravimetric analysis(TGA),and the results show that SMIPs was successfully synthesized on the surface of 40 ?m silica.Then the surface molecular imprinted polymers solid phase extraction adsorbent(SMISPE)was used for the separation and enrichment of fox grape samples of myricetin,the eluent solution was analyzed by Agilent 1100 series HPLC-DAD system.The experiments showed good linearity with correlation coefficients(R)of 0.9995,good precision(RSD)of 5.4 % and good sensitivity(LOD(S/N=3))of 15.9 ng·L-1.The average recovery of the solution treatment with SMISPE was 98.8 %,the content of myricetin in fox grape was 0.0067 mg/g.The results indicated that the SMISPE have better adsorption capacity than commercial C18-SPE,and it indeed a promising method to extract and enrich flavonoid-structure substance.