## 6 Modifications of Chitosan
### Graft Copolymerization of Chitosan  
Grafting copolymerization, which can change the physicochemical properties of chitosan, is an effective method to broaden the practical use of chitosan. The molecular structure, number and length mainly affect the properties of the final graft copolymerization products. These methods include free radical graft copolymerization, condensation copolymerization, oxidative coupling copolymerization, ring-opening copolymerization of ring mounted monomers, polymer grafting, etc. Chitosan C6 primary hydroxyl, C3 secondary hydroxyl and C2 amino groups can all be grafted points. The side chain of chitosan C6 can be grafted selectively to obtain the branched polysaccharide with unique immuno promoting activity, that is, the polysaccharide with the side chain of \(\alpha\)-(1\(\rightarrow\)6) on the main chain of \(\alpha\)-(1\(\rightarrow\)4). Glycosyl, polypeptide, polyester and alkyl chain can be introduced into chitosan by the grafting reaction, which can improve the affinity of chitosan to the solvent, as well as the antibacterial and immune activity. They are mainly used as drug membranes, gelatin, microspheres (microapsules), nanoparticles, slow-release materials, gene delivery vectors, polymer drugs, drug-loaded micelles, etc. [PERSON] et al. [53] reported the synthesis of chitosan grafted polyvinyl alcohol (PVA) and its release of prednisolone. As a drug release coating, chitosan has some disadvantages, such as poor solubility, too strong dependence on pH and poor mechanical properties, but grafting PVA with good water solubility and biocompatibility can greatly improve the drug release behavior. [PERSON] et al. [54] prepared chitosan with three different molecular weights through the p-coumaric acid method to improve their antioxidant property and water solubility. Further, they reported that the water solubility and antioxidant property of modified products decrease with an increased molecular weight of corresponding native chitosan. [PERSON] et al. [55] used chitosan alkalization and carboxymethylation reactions to prepare carboxymethyl chitosan followed by polyethyleneimine (PEI) grafting through an amidation reaction. Figure 5 shows the steps involved in carboxymethylation-PEI copolymer synthesis. The carboxymethylation-PEI copolymer formed nanoparticles through high complexation capability with DNA and attained high transfection efficiency and minimum cytotoxicity than 25 kDa PEI against 3T3 and 293T cells. Furthermore, the carboxymethylation-PEI copolymer with \(<\)0.05 mg/mL concentration showed a minimum effect on the morphology, lysis of human red blood cells or aggregation or on blood coagulation, demonstrating excellent blood compatibility. Therefore, the copolymer serves as an alternative, safe and effective non-viral vector for practical application. Two varying forms of nanostructured carbon were grafted by functionalized single-walled carbon nanohorns to the amine group of chitosan and covalently linking the carboxylic group existing on graphene oxide, respectively (Figure 6). Compared to unmodified chitosan, the nanostructured carbon significantly accelerated the biological functions using a comparative analysis of substrate-osteoblast cell line (MC3T3-E1) communication. Moreover, protein adsorption and nanostructured carbon have a synergistic effect to create favorable modulating biological functions such as cell proliferation, viability and adhesion, with a pronounced effect on nanostructured carbon-modified scaffolds. A study also highlighted that nanostructured carbon-modified scaffolds favored protein adsorption, bioactivity and other biological tasks [56]. Figure 5: Scheme of the preparation of the CMCS–PEI copolymer [55]. In a word, various side chains can be grafted on the primary, secondary hydroxyl and amino groups of chitosan and new properties endowed.