Study On Separation And Analysis Of Active Components From Leaves Of Lonicera Japonica Thunb

Lonicera japonica Thunb is a common medicinal plant,which plays a significant role in traditional Chinese medicine formula for treating various diseases,such as arthritis,diabetes mellitus,fever,infections,sores and swelling etc.Our research have proved that the components such as organic acids,flavonoids,iridoids,alkanes,olefins and sterols in flower bud,leaves and stem parts of Lonicera japonica Thunb have no obvious difference.Furthermore,the production of L.japonica leaves was ten times higher than that of flowers;they can be used as an alternative medicinal resource for the flowers of this plant.Therefore,it is necessary to establish an efficient method for the preparative isolation and purification of active compounds from L.japonica leaves.Therefore,in this thesis,the active components of L.japonica leaves were separated by traditional separation method and new adsorbent.This thesis is divided into two parts: 1.Extraction and separation the components from L.japonica leaves by MCI/Pre-HPLC and its anti-inflammatory activity studyThere are abundant active ingredients in the leaves of Lonicera japonica,and some pharmacological activities are comparable to Lonicera japonica,but they are abandoned in the process of picking.In order to improve the medicinal value of L.japonica leaves,a series of extraction and separation of anti-inflammatory active ingredients in L.japonica leaves were carried out,and the anti-inflammatory activity of each component was further studied with rat foot swelling model.Firstly,using L.japonica leaves as raw materials,N-Butanol phase（RDY-n-B phase）of L.japonica leaves extract was obtained by soxhlet extraction and fractionation extraction.The RDY-n-B phase was separated by MCI GEL CHP20 P column chromatography with different proportion of methanol and ultra-pure water,and 50% methanol elution fraction（M1）was obtained.Secondly,the anti-inflammatory activities of M1 and RDY n-B phase were evaluated by using the rat foot swelling model,and the anti-inflammatory activities of M1 and RDY n-B phase were studied by measuring the swelling rate of rat feet induced by fresh egg white.The results showed that the anti-inflammatory effect of L.japonica leaves extract M1 was better than that of RDY n-B phase,and was comparable to that of positive control group dexamethasone.The results provided a theoretical basis for the application of anti-inflammatory active components of L.japonica leaves in the preparation of anti-inflammatory drugs.Then,the M1 component was separated and prepared by Pre-HPLC,and three unknown compounds were obtained.Analyzed by HPLC-MS,¹H NMR,¹³ C NMR,UV-vis and FT-IR,one of the unknown compound was confirmed to be 5,7,3’,4’-Tetrahydroxyflavone-7-O-sambubioside,which was isolated from L.japonica leaves for the first time.The structures of the other two compounds were still not obtained due to their low purity.2.Selective adsorption of organic acids from L.japonica leaves using zeolitic imidazolate framework-8:experimental study and theoretical investigationIt is challenging to establish an efficient method for the preparative isolation and purification of organic acids from L.japonica leaves.In this work,we used ZIF-8 as adsorbent for selective extraction of organic acids from L.japonica leaves and studied the adsorption mechanism.Firstly,the ZIF-8 was synthesized,and a series of characterizations such as XRD,FT-IR,BET,TGA,TEM,and SEM were performed,which proved the successful synthesis of the material.The main parameters such as the amount of absorbent,adsorption time,extraction temperature and desorption conditions were evaluated.The proposed method was evaluated under the optimum conditions.As a result,good linearity was achieved for compounds 1-6 in the linear range of 0.12-26 μg/m Lwith R²≥0.9967 and LODs in the range of 0.047-0.093 μg/m L.The e-a phase of L.japonica leaves was enriched and extracted by ZIF-8.It was found that ZIF-8 mainly selectively adsorpted the organic acids,which was neochlorogenic acid,chlorogenic acid,caffeic acid,isochlorogenic acid B,isochlorogenic acid A and isochlorogenic acid C,respectively.Then,the recoveries of ZIF-8 for the six compounds at three spiked levels（0.5,5,10 μg/m L）in the e-a phase were detected to be in the range of 70.7-107.7%,with RSDs% between 0.89 and 7.57%.Furthermore,using DFT（Density Functional Theory）,XPS and Zeta potential investigation,we proposed that the adsorption of ZIF-8 for organic acids were mainly attributing to surface adsorption,hydrogen bond and the electrostatic interactions.These results demonstrated the feasibility of ZIF-8 in selective extraction of active components from complex medicinal plants.