The Preparation Of Nano-EGCG And Its Inhibitory Effects In MCF-7Human Breast Cancer Cells In Vitro

EGCG is a bioactive component of green tea extraction. It is associated with reducted risks of developing cancer and other chronic diseases. In body, the ability of EGCG targeted attacking tumor tissues is weak and the bioavailability is low. Drug delivery systems for EGCG are inefficient. All the disadvantages block the EGCG show its ability. In order to improve the bioavailability of EGCG, we use nanotechnology to optimize EGCG, make up for its disadvantages and achieve dose advantage.The purpose of this study is improve the targeting and sustained-release effect of EGCG on MCF-7tumor cell. We optimize nanoparticle preparation technology taking EGCG as drug model. Study how the EGCG nanoparticles affect the growth of MCF-7. On the basis of previous studies, consider the effect of different drug delivery systems and different modification for EGCG on MCF-7cells. Use chitosan as the material of carrier and modified nano-EGCG with FA and PEG which can improve the targeted ability and sustained release effect. Compare the distinguish of difference modified EGCG-NPs:EGCG-FA-CSNPS、EGCG-PEG-CSNPS、 EGCG-(FA+PEG)-CSNPS-1and EGCG-(FA+PEG)-CSNPS-2.First, the encapsulation of EGCG inside nanoparticles meant for cancer prevention research is a challenging process where several parameters need to be modulated in order to design nanoparticles with specific tailored characteristics. We investigate and improve the formulation parameters of EGCG loaded in chitosan nanoparticles using tripolyphosphate (TPP) as polyanionic cross linker. Nanoparticles morphology and encapsulation efficiency were analyzed as a function of chitosan molecular mass, chitosan concentration, chitosan-TPP ratio and EGCG concentration. The results showed that the average size of nanoparticles ranged from150-300nm, as the viscosity of chitosan was200cps with95%degree deacetylation and the mass ratio of chitosan and TPP was4:1. The manipulation of these parameters influenced not only the particle size but also the encapsulation and release of EGCG. In addition, studies on the release profile of EGCG in vitro demonstrated that the controlled release of EGCG using chitosan-TPP nanoparticles have the potential for use in drug delivery.Second, the modification of FA and PEG used two methods:1) Formation of EGCG-NPs in crosslinking of CS and TPP; grafting FA to EGCG-NPs with the help of EDC, then graft PEG to nanoparticles. EGCG-FA-CSNPS, EGCG-PEG-CSNPS, EGCG-(FA+PEG)-CSNPS-1are achieved.2) Graft CS with FA and PEG to form FA-PEG-CS, then using the ionic crosslink method to for EGCG-(FA+PEG)-CSNPS-2. Characterize the conversion rate of FA by ultraviolet spectrophotometer. The FA conversion rate of EGCG-(FA+PEG)-CSNPS-1and EGCG-(FA+PEG)-CSNPS-2were81.67%and88.97%, respectively. Use SEM to observation morphology of different modified EGCG-NPs. The picture in the SEM showed that nanoparticles all have small particle size and uniform distribution. The results of FT-IR and’HNMR characterization further proved that the modification of EGCG-NPs is successful.Third, inverted microscope was used to observe MCF-7cell morphology; methylene blue method was used to study the effect of EGCG and different modified EGCG on MCF-7cytotoxicity assay and antiproliferative activity. Median-effect plot was used to dose effect analysis of calculating individual and combined FA and PEG modification effect. Cytotoxicity assay showed no cytotoxicity in EGCG-FA-CSNPS、EGCG-PEG-CSNPS、EGCG-(FA+PEG)-CSNPS-1and EGCG-(FA+PEG)-CSNPS-2. EGCG、EGCG-NPs、EGCG-FA-CSNPS、EGCG-PEG-CSNPS EGCG-(FA+PEG)-CSNPS-1and EGCG-(FA+PEG)-CSNPS-2showed significant antiproliferative activities against MCF-7human breast cancer cells in a dose-dependent manner. And the antiproliferative activity toward the growth of MCF-7human breast cancer cells in vitro was increased significantly by nanoparticles which FA and PEG modified together when compare to the FA and PEG modified alone.These studies have shown that nano-EGCG preparation technology was optimized with certain slow-release ability. After modified with targeting ligand FA and hydrophilic polymer PEG, the inhibition ability on MCF-7human breast cancer of modified EGCG-NPs have improved.