Modified chitosans for biomedical applications

ABSTRACT: The subject of this thesis is the exploration of the suitability of chitosan and some of its derivatives for some chosen biomedical applications. Chitosan-graft-poly (N-vinyl imidazole), Chitosan-tripolyphosphate and ascorbyl chitosan were synthesized and characterized for specific biomedical applications in line with their chemical functionalities. Chitosan-graft-poly (N-vinyl imidazole), Chi-graft-PNVI, was synthesized by two methods; via an N-protection route and without N-protection to observe the effect of free amine groups on the antibacterial activity. Both chitosan and Chi-graft-PNVI samples demonstrated antibacterial activity against both gram-positive Staphylococcus epidermidis (S. epidermidis), and against gram-negative Escherichia coli (E. coli). Decreased antibacterial activity was measured with increasing grafting percentage of PNVI. The observed trend can be explained by the fraction of free amine groups present in the samples. PNVI grafting onto chitosan proceeds via both OH-bearing carbons and free amine groups which results in a reduced fraction of free amine groups responsible from the antibacterial effect. When the amine group is protected, grafting proceeds mainly from the hydroxyl bearing carbon atoms. Hence, after deprotection there are higher fraction of free amine groups available for antibacterial activity leading to improved antibacterial activity. New combinations of an aminoglycoside antibiotic, gentamicin with the natural aminopolysaccharide chitosan and Chi-graft-PNVI were investigated as antibacterial agents. It was found that a mixture of gentamicin and Chi-graft-PNVI exhibited a higher antibacterial activity against both S. epidermidis and against E. coli when compared to gentamicin alone. Gentamicin produces an inhibition zone of 8.2±0.2 mm against S. epidermidis when undiluted, while the inhibition zone increases to 25.8±0.7 mm in combination with Chi-graft-PNVI prepared via N-protection. These combinations have a potential to form a basis to new formulations of gentamicin with improved antibacterial activity and might allow usage of decreased doses of the antibiotic. Blood contact properties of chitosan and its derivatives were investigated in detail. Chitosan-TPP bioadsorbents and ascorbyl chitosans were prepared. Nonimprinted and Fe3+ imprinted chitosan TPP gel beads were prepared via physical gel formation. A method based on in-situ crosslinking using ethylene glycol diglycidyl ether, EGDE, was developed to imprint the chitosan-TPP gels with Fe3+ without deteriorating the gel beads. The Fe3+ imprinted and in-situ crosslinked beads proved to be durable and effective adsorbents for Fe3+ in solution. The characteristics of the beads as biomedical iron adsorbents were tested by evaluating their serum iron removal capacities from human blood. The preliminary tests carried out showed that Fe3+ imprinted beads were more effective in decreasing serum iron in human blood, when compared to the nonimprinted beads. The decrease in serum iron level accompanied a parallel decrease in the haemoglobin level. The calcium level was also affected upon contact with the beads. The Fe3+ imprinted beads were less effective than the nonimprinted ones in decreasing the calcium level indicating selectivity towards iron containing species. Blood contact properties of ascorbyl chitosans were investigated by testing for changes in total cholesterol, HDL cholesterol, LDL cholesterol and triglyceride levels in addition to total calcium analysis, complete blood count analysis and prothrombin time determination upon contact in human blood with ascorbyl chitosans. The lipid lowering activity increased with ascorbyl substitution. The inherent non specific adsorption capability of chitosan due to its chelating power with several different functional groups was exhibited by ascorbyl chitosans as well. This behaviour was exemplified in a simultaneous decrease in the total iron values of the volunteers together with lower lipid levels. Furthermore, ascorbyl chitosans were observed to have less hemocompatibility but increased anticoagulant activity when compared to chitosan alone. Ascorbyl chitosans performed better blood contact properties as biomaterials compared to ascorbic acid alone but poorer when compared to chitosan alone. In summary, the suitability of chitosan-based materials was examined in different biomedical domains. Our results highlight the importance of this particular polymer in biotechnology and biomedical applications. Keywords: Chitosan, N-vinyl imidazole, antibacterial activity, tripolyphosphate, Fe3+ adsorption, imprinting, blood contact chitosan, cholesterol lowering activity, vitamin C (ascorbic acid), biomedical applications. …………………………………………………………………………………………………………………………