Preparation And Evaluation Of Quercetin Nano-emulsion Modified By Poly(2-ethyl-oxazoline)-cholesteryl Methyl Carbonate

Quercetin（QUE）is a natural flavonoid with anti-oxidant,anti-inflammatory,anti-cancer,and has pharmacological effects that can reduce the cardiovascular diseases.However,QUE is poor in water solubility,and it is unstable in light and heat,which limits its clinical application.In this paper,a pH-sensitive polymer,poly（2-ethyl-oxazoline）-cholesteryl methyl carbonate（PEtOz-CHMC）,modified the nano-emulsion was used to encapsulate QUE to increase the solubility and stability of the drug.In this paper,a method for determination of QUE by UV-vis spectrophotometry has been established,which had a good linearity in the concentration range of 10 to 100μg/m L.The precision and recovery rate were in line with the experimental requirements.QUE nano-emulsion（QUE-E）was prepared by an emulsification and low-temperature solidification method.PEtOz-CHMC was modified on the surface of QUE-E by direct insertion method to obtain pH-sensitive PQUE-E.The encapsulation efficiency and loading efficiency of the optimal QUE-E formulation optimized by orthogonal test were 84.0%and5.7%,respectively.In order to improve the stability of the preparation,the nano-emulsions were prepared as freeze-drying preparations,and it was finally determined that the type of the lyoprotectant was lactose,and the ratio of the freeze-drying agent to the oil phase was 3:1（w/w）.The appearance of the preparation was smooth and fine.Transmission electron microscope（TEM）and scanning electron microscope（SEM）were used to determine the morphology of the nano-emulsions.The appearance of the nano-emulsions is spherical or spheroidal.The average particle size of QUE-E is about 110 nm,and the zeta potential is about-47 m V.And the modification of PEtOz has no significant effect on the encapsulation efficiency,loading efficiency and particle size of the nano-emulsion.The stability of the nano-emulsions was evaluated in 10%fetal bovine serums which simulate the environment in vivo.The transmittances of QUE-E and PQUE-E at 24 h were70.8%and 88.7%,respectively.The results showed that the stability of PQUE-E was better than that of QUE-E（P<0.05）.The in vitro release experiment showed that the cumulative release of QUE-E and PQUE-E at 24 h were 43.5%and 29.3%under pH 7.4,respectively,indicating that PEtOz gives good stability to the preparation.And at pH 6.4,the cumulative release of the nano-emulsions at 24 h was 49.5%and 80.1%,respectively.The release of PQUE-E was significantly higher than that of QUE-E（P<0.01）,showing pH sensitivity.The inhibitory effects of the nano-emulsions on human breast cancer cell line MCF-7 and human hepatoma cell line Hep G-2 were investigated by MTT assay.Compared with QUE-E,PQUE-E had a strong inhibitory effect on both cells.At pH 6.4,the half maximal inhibitory concentration(IC₅₀)of PQUE-E was the lowest,indicating that PEtOz has the response to the slightly acidic environment.High performance liquid chromatography（HPLC）was used to determine the intracellular uptake of the nano-emulsions.At pH 6.4,the uptake of PQUE-E was higher than that of QUE-E（P<0.05）,indicating that PEtOz confers certain pH sensitivity to the preparation.The pharmacokinetics and tissue distribution of the nano-emulsions in mice were investigated.The results showed that the t_1/2βof the PQUE-E group was 1.3 times and 1.2times that of the solution group and QUE-E group,respectively,indicating that the half-life of the nano-emulsions was prolonged in vivo.The AUC of PQUE-E group was about 2.2 times and 1.4 times higher than that of solution group and QUE-E group which indicated that the elimination of PQUE-E in vivo was slow.Liver had the highest tissue concentration of quercetin solution,followed by lung,kidney,heart and spleen,while liver also had the highest tissue concentration of QUE-E and PQUE-E,followed by kidney,lung,spleen and heart.The accumulation of QUE in PQUE-E in each tissue was higher than that in the other two groups,indicating that PEtOz increses the stability of QUE-E in vivo.