Syntheses And Characterizationof Hemicyanine-based Fluorescent PH Probe And Folic Acid-ModifiedCMD-Coated Liposomes

Small molecule fluorescent probe has the advantages of high sensitivity, quick responsibility, facile preparation and in-situ real-time detection, which make it very attractive for cell imaging in life science. Based on the intramolecular charge transfer (ICT) mechanism, a l-ethyl-2,3,3-trimethyl indole based ratiometric pH probe was synthesized using4-hydroxybenzaldehyde as receptor. This ratiometric probe possesses inherent self-calibration and provides a more accurate and reliable way for fluorescent assay. The probe responds to H+with high selectivity and sensitivity in aqueous solution with5%DMSO. The fluorescence intensity ratio (R=I513nm/I553nm) exhibits a good linearity with pH within the range of5.4-7.2at λex=430nm. MTT assay revealed that the probe has low cytotoxicity. Fluorescence imaging of HeLa cells shows that CyP can penetrate the cells membrane by the uptake of HeLa cells, indicating that CyP is potential in analyzing the intracellular pH. Liposomes, as drug delivery system, have good biocompatibility, passive targeting ability and ability to protect the drug activity. However, un-coated liposomes (UC-LIPs) are limited in appplicatiob by lack of active targeting, and instability in the presence of plasma serum which leads to drugs leakage. Carboxymethyl dextran (CMD) is a polymer with non-toxicity, non-immunogenicity, biodegradability, and has been previously used to stabilize liposomes by our group. The CMD-LIPs was proved to have good stability in plasma serum at4℃, and pH-sensitive drug release property. In this thesis, FA-CMD was synthesized by conjugating amphiphilic CMD with folic acid (FA) as targeting ligands, then FA-CMD was used to prepare FA-CMD modified liposomes (FA-CMD-LIPs) via thin-lipid film method. The FA-CMD-LIPs have a narrow distribution with size about200nm, negativeζ-potentials (-18mV) and good stability at4℃for30days. DOX-loaded FA-CMD-LIPs were prepared by remote loading method with (NH4)2SO4gradient with94%encapsulation efficiency. DOX-loaded FA-CMD-LIPs show a sustained and pH-responsive drug release behavior. DOX release rate of FA-CMD-LIPs is slower than that of UC-LIPs. The MTT assay indicated that DOX-loaded FA-CMD-LIPs have higher cell cytotoxicity than DOX-loaded CMD-LIPs to cancerous HeLa cells. Fluorescence images of HeLa cells showed that FA-CMD-LIPs can deliver more DOX into HeLa cells than that delivered by CMD-LIPs, which is due to that the FA on the surface of FA-CMD-LIPs can facilitate the endocytosis of liposomes by targeting the over-expressed FA reeceptor on the membrane of HeLa cells.