Facile Fabrication Of Glyconanoparticles And Their Applications For Carbohydrate-protein Interaction

Carbohydrates play crucial roles in biological processes as recognition sites for cells and various binding partners.Efficient analysis and study of these important biological processes are,however,largely hampered by the weak affinities often associated with carbohydrate-protein interactions.One promising way to overcome these obstacles is the application of various “glycocluster” tools.Glyconanoparticle,which can effectively combine the glycocluster effect with the special chemico-physical properties of various nanoparticles,allowing them to have a wide range of potential applications in glycobiology and biomedicine.Developing new glyconanoparticle fabrication method which has wide applicability,high expansibility and good operability is necessary and beneficial to enhance the application potential of glycomamoparticles.In this thesis,two facile and versatile glyconanoparticle fabrication methods based on the photochemically induced insertion reaction of perfluorophenylazide(PFPA)and the self-polymerization of dopamine are put forwarded.Some different glyconanoparticles are fabricated via these two methods and applicated in detecting carbohydrate-protein interaction,targeted optical fluorescence imaging of cancer cell and targeted cancer therapy.The content of this paper includes the following two aspects:1.Photochemical fabrication of glyconanoparticles and their application for carbohydrate-lectin interaction.PFPA can be converted to nitrene under irradiation or heating that can insert C-H,N-H or C=C bond to form new covalent bond.Here,the PFPA photochemically induced C-H insertion reaction is used for introducing alkynyl group to dextran or polyving akohol coated on iron oxide nanoparticle surface,the amount of alkynyl groups on alkynyl functionalized dextran iron oxide nanoparticles and polyving akohol iron oxide nanoparticles was 1.22 mmol per g NPs and 1.56 mmol per g NPs,respectively.After that,four kinds of glyconanopzrticles,galactose modified dextran iron oxide nanoparticles,galactose modified polyving akohol iron oxide nanoparticles,mannose modified polyving akohol iron oxide nanoparticles and fluorescent glactose-nanoparticles are fabricated via the azide-alkyne click reaction.DLS analysis shows that the diameters of these glyconanoparticles are 300-600 nm.Three kinds of as-fabricated glyconanoparticles are applied to detect their interaction with Con A,PNA,galectin-9,Jurkat cells and KM12 cells via a QCM set.The results show that all the glyconanoparticles can specific interact with corresponding lectins.The fluorescent glactose-nanoparticles are used for targeted optical fluorescence imaging of HepG2 cell(human hepatoblastoma cell line),which over-expressing asialoglycoprotein receptors(ASGP-R)on its surface,and ASGP-R can specific interact with glactose and lactose.The He La cells(Human cervical carcinoma cell line)which express few ASGP-R are used as non-targeted control cell.The results detected by flow cytometer and laser scanning confocal microscope confirmed that the fluorescent glactose-nanoparticles can target HepG2 cell and image it by the conjugated rhodamine B ligand.2.Fabrication of hypericin loaded glyconanoparticle via the self-polymerization of dopamine and its application in targeted anti-cancer research.In alkaline aqueous media,dopamine can undergo oxygen-induced self-polymerization on the surfaces of lots of materials to form thin adherent polydopamine(PDA)films,and PDA surface can be modified via Schiff base reaction or Michael addition reaction between amino or sulfhydryl compound and the abundant catechol in PDA.Here,we add acetone solution of hypericin to the media in which self-polymerization of dopamine on Fe3O4 nanoparticle surface is in progress,thus making hypericin loaded into the PDA film on the Fe3O4 nanoparticle surface.Then,amino-modified carbohydrate is conjugated to the PDA film surface to obtain the hypericin loaded lactose-nanoparticles.The PDA coating and hypericin loading are confirmed via TEM and IR analysis.The amount of htpericin in hypericin loaded lactose-nanoparticles is 33.1 ?g/mg NPs,and DLS analysis shows that the average diameter of this glyconanoparticle is 546 nm.Hypericin can be promoted under 600 nm light irradiation to produce singlet oxygen,which causing cell damage or death.However,the application of hypericin in biomedicine is greatly limited by its poor water solubility.In this part,HepG2 cell is chosen to study the targeted anti-cancer property of as-fabricated hypericin loaded lactose-nanoparticles,which disperses well in water.The cell uptake research shows that the hypericin loaded lactose-nanoparticles can target HepG2 cells;the reactive oxygen species generation research and the cytotoxicity research shows that the hypericin loaded lactose-nanoparticles can targeted kill HepG2 cells via the photodynamic effect of hypericin.