Preparation Of Carbohydrate-functionalized Gold Nanoclusters And Their Applications In Biomedical Fields

The interactions between carbohydrates and carbohydrate-binding proteins（CBPs）participate in and mediate many important physiological and pathological processes in living organisms.Therefore,it is of great significance to study and utilize the biological functions of carbohydrates for developing new strategies of disease diagnosis and treatment.However,individual carbohydrate-based interaction with the corresponding receptor is very weak,bringing great challenges to the research and applications of the biological functions of carbohydrates.With the rapid development of nanotechnology,the applications of nanoparticles in biomedical fields are deepening day by day.In the meantime,glyconanoparticles（G-NPs）emerged as the times require.G-NPs can effectively utilize the multivalent effect of carbohydrate and thus greatly enhance the binding affinity between carbohydrate and its protein receptor.In the last twenty years,G-NPs have become powerful tools for the study and application of biological function of carbohydrates.Notably,as a new-emerged class of gold nanomaterials,fluorescent gold nanoclusters（Au NCs）are easy to be prepared and functionalized,as well as exhibiting excellent fluorescence properties and low biological toxicity,which makes them promising multivalent scaffolds for carbohydrate loading.In this study,a variety of carbohydrate-functionalized Au NCs were constructed by using different coupling methods.Based on the biological functions of carbohydrate,the as-prepared carbohydrate-Au NCs were then applied to different biomedical fields.The main research contents were as follows:（1）By using a natural sialoglycoprotein（transferrin）rich in cysteine and tyrosine as template,sialic acid-functionalized Au NCs（Tf-Au NCs）with strong near-infrared fluorescence emission were rapidly prepared within 5 min with the assistance of microwave radiation.Due to the specific recognition of sialic acid to the neurotransmitter 5-hydroxytryptamine（5-HT）associated with mental diseases,5-HT triggered the aggregation and fluorescence enhancement of Tf-Au NCs.Furthermore,the fluorescence enhancement percentage of Tf-Au NCs had a good linear relationship with the concentration of 5-HT.Thus,the fluorescence detection of 5-HT was achieved by Tf-Au NCs with good selectivity and high sensitivity.（2）In general,natural glycoproteins are expensive and limited in variety.The carbohydrate amount on individual protein is also difficult to regulate,greatly limiting the extensive preparation of various carbohydrate-functionalized Au NCs.In order to reduce the preparation cost and increase the loading amount of carbohydrate on Au NCs to further enhance their biological recognition ability to receptors,the complex of amino-functionalized mannose and bovine serum albumin was firstly used as template for the preparation of Au NCs with high mannose loading.Based on the specific recognition of mannose to the lectin Concanavalin A,this nanosystem was applied to the fluorescence detection of Concanavalin A with high sensitivity.Besides,targeted fluorescence imaging of breast cancer cells was also achieved due to the interaction between mannose on Au NCs and the overexpressed mannose receptor on breast cancer cells.The strategy of preparing carbohydrate-functionalized Au NCs with the complex of functionalized mannose and protein as template not only significantly reduced the cost,but also increased the carbohydrate loading amount on Au NCs.More importantly,this strategy could be easily extended to other carbohydrates containing reactive groups.（3）Due to the excellent targeted imaging effect of mannose-functionalized Au NCs for breast cancer cells,mannose-functionalized nanotheranostic system was further designed and prepared for targeted therapy of breast cancer.Through the simple and efficient biomineralization process,a composite probe of mannose-functionalized Au NCs-Mn O₂nanosheet was prepared using the complex of amino functionalized mannose and bovine serum albumin as template.The composite probe could not only efficiently decompose hydrogen peroxide to oxygen,but also deplete reduced glutathione.As a nano-drug carrier,the composite probe exhibited high loading efficiency and loading capacity for the near-infrared photosensitizer indocyanine green（ICG）.The stability of ICG on probe was also significantly enhanced.In addition,the composite probe with ICG revealed good photothermal and photodynamic effects under laser（808 nm）irradiation.By virtue of the targeting effect of mannose,the composite probe was successfully applied to the effective treatment of breast cancer.This system provided a momentous reference for the construction of multifunctional nano-therapeutic system based on Au NCs and the biological recognition of carbohydrates.（4）In order to further propose a versatile strategy of preparing carbohydrate-functionalized Au NCs using reducing carbohydrate,a bifunctional compound S-（3-（methoxyamino）propyl）thioacetate（MPTA）was designed and synthesized.Using MPTA as a linker,a variety of monosaccharides,disaccharides and oligosaccharides with reducing ends were simply and efficiently immobilized on Au NCs,thereby constructing different G-NPs systems.Notably,structure of the reducing end of carbohydrate could be completely preserved by this method,which maintained the biological function of carbohydrate to the maximum extent.Preliminary results showed that the binding strength of reducing carbohydrate-Au NCs to receptor proteins was 1300 times higher than that of free carbohydrates（Taking mannose as an example）.In addition,the achievement of mannose-based lectin detection and cancer cell targeted imaging further confirmed the receptor recognition ability of the carbohydrate-Au NCs prepared by MPTA.This study provided a new and efficient method for the preparation of G-NPs applicable to all reduced carbohydrates,paving a new avenue for the construction of various G-NPs.In conclusion,a variety of carbohydrate-functionalized Au NCs systems were constructed through different coupling methods and applied to different biomedical fields based on the recognition capability of carbohydrates.This study provided an effective reference for the construction of G-NPs and their applications in biomedical fields.