Study On The Preparation And Biocompatibility Of N-Carboxymethyl Chitosan

Purpose:Chitosan(CTS)[(1→4)-2-amino-2-deoxy-β-D-glucanl,a unique polysaccharide derived from deacetylation of chitin,has been widely used in biomedical field owing to its obvious alkaline and positive charges.However,the development and utilization of chitosan is limited by its insoluble behavior except in acid solution.Carboxymethyl chitosan(CMC) is one of the water soluble derivatives of chitosan which has been widely studied.CMC has been a research focus in the field of pharmaceutical and biomedical industry for its unique physicochemical and biological properties such as biocompatibility,biodegradability,degradation -controllability and antimicrobial activity.The C6 hydroxyl,C3 hydroxyl and C2 amino of chitosan can be grafted by carboxymethyl.So,we can derive O- Carboxymethyl chitosan, N-Carboxymethyl chitosan and N,O- Carboxymethyl chitosan in different reaction conditions.Studies find that N,O-CMC always has a lower relative molecular weight and a significant reduction in viscosity for a long period reaction condition in stronger alkaline and higher temperature,which is not suitable as a medical polymer material. The purpose of this study is to prepare N-CMC with different degrees of carboxylation (D.C.) in mild condition,and preliminary study the physicochemical and biological properties.Methods:1.Water-soluble N-carboxymethyl chitosans(N-CMC) is prepared from chitosan and glyoxylic acid in weak acid condition.And the chemical and physical properties are measured.The structure of N-CMC is characterized by FTIR and X-RAY.The D.C.is measured by acid-base titration.The moisture,ash,viscosity, molecular weight,isoelectric point and moisture-absorption and moisture-retention properties are also determined.2.Biocompatibility of N-CMC with two different D.C.is evaluated in this study.By means of cytotoxicity test,hemolysis test,coagulation test and histocompatibility test.Results:1.The D.C.of N-CMC products are between 25%and 78%.New peak at 1590 cm-1 in FTIR spectrum assigned to -COONa groop appears,which indicates that the substation reaction takes place specifically on the -NH₂ of CTS.The crystal structure is changed obviously after carboxymethylation which is manifested by X-ray diffraction results.2.At the concentration range from 62.5μg/mL to 5000μg/mL,N-CMC with high D.C. shows very good cell compatibility with L929 fibroblast.But the N-CMC with low D.C.inhibits the cell growth when the concentration is higher than 500μg/mL.The hemolysis rates of the five N-CMCs were less than 5%,and they all have no influence in coagulation time.Animal tissue compatibility experiments showed that N-CMC in colloidal solution shape was degraded and absorbed more easily in vivo than that in film shape.N-CMC in muscle tissue had a better degradation and absorption than in subcutaneous tissue.This part of the study indicates that N-CMC material has good biocompatibility.Conclusion:N-CMC prepared by this method can avoid rupturing of molecular chains and maintains high molecular weight.N-CMC has good cell compatibility, good blood compatibility and good tissue compatibility.The results in this study provide a theoretical foundation for the research and application of N-CMC as a biomaterial.