Preparation Of Molecularly Imprinted Polymers And Their Applications In Pulsatilla Saponin And Formononetin Study

Molecularly imprinted polymers?MIPs?has been widely applied in natural products pretreatment,due to their predetermined,specific recognition property,low costs and easy preparation.Core-shell structured MIPs,designing by surface molecular imprinting technology,has gained popularity,since the technology not only provide fast adsorption kinetics and high densities of recognition sites,but also facilitate the introduction of various properties?e.g.magnetic-sensing?into MIPs.However,the pretreatment of natural products always requires multi-step enrichment and purification procedures.The aim of this thesis is to explore and develop new complex sample pretreatment methods based on MIPs.The main contents are listed as follows:?1?In this study,the metabolic profile of anemoside B4?AB4?with microflora in rat small and large intestines in vitro was investigated.Gut microflora was collected from different intestinal segments and anaerobically incubated with AB4 at 37°C for 24,48,72 and 96 h,respectively.A total of 10 metabolites were detected and identified by ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry?UPLC-Q-TOF-MS/MS?,involving the products of oxygenation and deglycosylation reactions.Gut microflora in the large intestine generated more comprehensive metabolic pathways,which appears to be attributable to the wider range of bacterial types and numbers of bacteria.?2?Polymers were prepared by using Fe₃O₄ nanoparticles as the supporting matrix,D-glucose as fragment template,and dopamine?DA?as the functional monomer and cross-linker.The MMIPs for glucoside-specific adsorption was used to identify metabolites of Pulsatilla chinensis in rat feces.Results showed that MMIPs exhibited large adsorption capacity?5.65 mg/g?,fast kinetics?60 min?.Furthermore,the MMIPs coupled with UPLC-Q-TOF-MS were successfully utilized for the identification of 17compounds including 15 metabolites from the pulsatilla saponin metabolic pool.This study provides a reliable protocol for the separation and identification of saponin metabolites in a complex biological sample.?3?Thermo-responsive magnetic molecularly imprinted polymers?TMMIPs?were prepared by simple surface molecular imprinting polymerization for selective adsorption and enrichment of formononetin from Trifolium pretense L.by temperature regulation.Using formononetin as the template,N-isopropylacrylamide as the thermo-responsive functional monomer,methacrylic acid as an assistant functional monomer,the polymers were synthesized on the surface of the magnetic substrate.The results show that TMMIPs attained controlled adsorption of formononetin in response to the temperature change,with large adsorption capacity?16.43 mg/g?,fast kinetics?60 min?and good sensitivity at 35°C.The selectivity experiment indicated that the materials had excellent recognition ability for formononetin and the selectivity factors were between1.32 and 2.98 towards genistein and daidzein.Under the optimal conditions,the low detection limits and low quantitation limits are 0.017 and 0.063?g/mL,respectively.Furthermore,the TMMIPs were successfully utilized for enriching and purifying formononetin from Trifolium pretense L.The analytical results indicate that the TMMIPs are promising materials for selective identification and enrichment of formononetin in complicated herbal medicines by simple temperature-responsive regulation.