Prediction Of Natural Products Biomembrane Permeability By Fluorescent Liposome

Natural products show good pharmacological activities in many fields.Between 2001 and 2005,23 new drugs derived from natural products were introduced for the treatment of disorders such as bacterial and fungal infections,cancer,diabetes, dyslipidemia,atopic dermatitis and Alzheimer's disease.Getting good pharmacological activities compounds from natural products is an important way to get new drugs.There are numerous natural products found every year.In order to find the good pharmacological activity drug in the numerous natural products,the high-throughput screening assay is very important.Traditional screening assays such as animals,cells,receptors and enzymes are time-costing and require many types of equipment.So,they are not quite suitable in high-throughput screening.Herein,we proposed a fluorescent liposome to high-throughput screening natural products.This model is used to estimate the drug permeability in biomembrane. Natural products with good biomembrane permeability are supposed to have good pharmacological activity.The following work is carried out in our research.1.The fluorescent liposome model was established.We used the fluorescent liposomes to simulate the biomembrane,and predicted the permeability of natural products by fluorescence quenching of liposome.We chose three different fluorescence probes which lay in different regions in liposome.When natural products gathered in membrane,the fluorescence intensity of fluorescence probe will be quenched.We estimated the permeability of natural products by fluorescence quenching.According to the zeta potential,particle size and fluorescence intensity,we chose the best formulation of fluorescent liposome is DSPG:PC 0.1:1.2.We deter mined the liposomes fluorescent quenching caused by natural products.We selected the best reaction time and drug concentration in experiments:4μM drug reacting 10 min with fluorescent liposome.In this condition,fluorescence signals were stable,high-sensitivity,and cost a few samples.35 kinds of natural products were chose to react with fluorescent liposomes. Fluorescence quenching was recorded.Hydrophobic drug showed strong fluorescence quenching,while hydrophilic drug showed slight fluorescence quenching.The fluorescent polarization of liposomes was deter mined.Natural products can increase the P and decreased the fluidity of membrane.The more natural products gathered in the liposome,the more P increased.Natural products reacted with CF liposome and didn't change the fluorescence intensity obviously,which indicated that drug hadn't affects the structures of fluorescent liposomes.3.We deter mined the erythrocyte membrane fluorescent quenching caused by natural products.We labelled the erythrocyte membrane by DPH and DPH-TMA.35 kinds of natural products were chose to react with fluorescent erythrocyte membrane. Fluorescence quenching was recorded.Hydrophobic drug showed strong fluorescence quenching,while hydrophilic drug showed slight fluorescence quenching.It was found good correlation between fluorescence liposome and erythrocyte membrane by comparing fluorescence quenching percentage of these two models. In 2μM-16μM concentration,fluorescence liposome and erythrocyte membrane had good correlation.Fluorescence liposome can substitute erythrocyte membrane to estimate drug biomembrane permeability in some extent.4.We deter mined P_app and P_eff of some natural products.P_app and P_eff were deter mined using Caco-2 cell and rat intestinal perfusion separately.It was found that P_app had good correlation with erythrocyte membrane and liposome fluorescence quenching.It was also found P_eff had good correlation with erythrocyte membrane and liposome fluorescence quenching.Fluorescent liposome not only can predict drug permeability in biomembrane,but also can estimate drug non-protein passive absorption.5.We researched the reaction of natural products with BSA.We firstly used fluorescent liposome to research drug partition between target cell and BSA.We found competitive binding existed between target cell and BSA.In summary,fluorescent liposome had good correlation with erythrocyte membrane, Caco-2 cell and rat intestinal perfusion.Fluorescence liposome can substitute erythrocyte membrane,Caco-2 cell and rat intestinal perfusion to estimate drug biomembrane permeability and non-protein passive absorption in some extent.Fluorescence liposome is quick,accurate,easy to detect,and need only a few samples.So,it is quite suitable for fluorescent liposome to use in high-throughput screening natural products.