血液低温保存意义重大，其中红细胞长期冻存是急需解决的难题，冷冻和复苏过程中细胞内外结冰导致细胞膜损伤是其致命要害。使用甘油保护剂效果较好，但其洗脱步骤繁琐；从仿生抗冻糖肽角度合成抗冻聚合物尚不能满足需求。本项目拟通过RAFT和NCA-ROP聚合方法，合成几种典型的海藻糖氨基酸共聚物，调控其分子链和侧基结构，使其对红细胞具有冻存保护功能，同时具有生物相容性和抗菌性。研究海藻糖甲基丙烯酸酯共聚物的海藻糖侧基与类氨基酸亲/疏水基团的组成、含量、分子量及分布等因素对大分子链低温自组装行为的影响，分析其与红细胞质膜蛋白和磷脂层及血红蛋白的相互作用原理，进一步探索聚氨基酸接枝海藻糖共聚物对红细胞的透膜性保护作用和抗细菌感染能力。预期所合成的海藻糖氨基酸共聚物链结构会随温度降低具有自响应性，能够保护红细胞免受低温损伤。开展海藻糖肽共聚物对细胞和蛋白低温保护功能的研究，可为该领域提供新的理论依据和新方法。

Long term preservation of blood products for transfusion purposes has been of great scientific and practical interests over the past decade. Cryopreservation of red blood cells (RBCs) is meaningful for human survival. During cryopreservation (freezing and thawing), formation and growth of ice crystals, especially the intracellular ice formation, are lethal to cells. A conventional approach has been adopted by using glycerol as a cryoprotective agent at a high concentration for vitrification. However, followed tedious multistep washing is required to completely remove the highly toxic, cell membrane-permeable cryoprotectant after thawing for uses. Therefore, it is essential to achieve cell cryopreservation with nontoxic cryoprotectants. By a biomimetic manner similar to antifreeze (glyco)proteins, synthetic polymers have been developed by depressing the growth of ice crystals to enhance the cryopreservation of human RBCs, but they did not meet the cell cryopreservation requirements yet. In this project, we propose to synthesize trehalose-amino acid copolymers with thermal responsive behaviors by control of their backbone and side chain structures for cryoprotection and stabilization of RBCs at low temperatures.. Trehalose (meth)acrylate copolymers will be synthesized via reversible addition-fragmentation chain transfer (RAFT) polymerization by using dithioester such as 4-cyanopentanoic acid dithiobenzoate as a chain transfer agent. Trehalose related monomers, for example 4,6-O-(3-propyl methacrylate)-α,α-trehalose (TreMA), will be synthesized from α,α-trehalose and 3,3’-diethoxypropyl methacrylate via an acetal linkage according to the references. The comonomers containing unusual amino acid groups, including cationic 2-(dimethylamino)ethyl methacrylate, hydrophobic methyl methacrylate, hydrophilic oligoethylene glycol methylether acrylate etc. will be introduced to adjust the copolymer chain structure, and the polymeric self-assembly at subzero and room temperatures. . Characterizations of the prepared copolymers containing side-tethered α,α-trehalose and the unusual amino acid groups will be carried out, especially at low temperatures. Effects of the chain structures of the block or random copolymers, the molecular weight and distribution, the single-chain folding of amphiphilic copolymers on the cryoprotective abilities will be investigated. Recovery of RBCs after freezing-thawing under the shield of the trehalose copolymers will be studied. Especially, the interactions of them with plasma membrane proteins, phospholipids and hemoglobin will be highly emphasized. . Furthermore, biodegradable poly(amino acid)s-g-trehalose copolymers as intracellular cryoprotectants will be prepared by ring-opening polymerization (ROP) of α-amino acid N-carboxyanhydride (NCA) monomers. The thehalose-NCA monomer will be synthesized via thiol-ene reaction of allyl functionalized peracetylated thehalose (protected TreMA) to L-cysteine, and followed by converted to thehalose-L-cysteine NCA (Tre-Cys NCA). The copolymerization of Tre-Cys NCA with L-lysine NCA and valine NCA etc. will be carried out to obtain poly(amino acid)s-g-trehalose copolymers. Control of the backbone and side chains of the copolymers at low temperatures for the membrane-permeable cryopreservation, as well as their biocompatibility and antimicrobial activity will be systematically investigated. For comparison, block copolymers of trehalose (meth)acrylate-b-poly(amino acid)s will be prepared via ROP by using an amino macrointiator of trehalose (meth)acrylate copolymers, which is synthesized by RAFT polymerization.. The investigation of trehalose peptide copolymers for cell cryopreservation could provide new strategies and fundamental theories for modern regenerative medicine to improve the storage of proteins, cells and tissues.