Preparation And Properties Of Chitosan And Its Derivative Grafted Peptides

Due to the excellent biological activity, chitosan and carboxymethylated chitosan, as the biomedical materials, have attracted more and more attention. Collagen peptides own favorable biocompatibility, biodegradable property, antioxidation, moisture absorption and retention properties and so on. They are becoming a hot trend among the field of cosmetic, medicine, food. In the food field, nisin, as the international licensing of biological preservatives, is non-toxic. In recent years, many scholars study nisin as the antimicrobial agent in the field of medicine. In this paper, collagen-chitosan, collagen-carboxymethylated chitosan, nisin-chitosan were synthesized with Microbia transglutaminase( MTGase) as biocatalyst. The enzyme-catalyzed reactions avoided unwanted products. The derivatives showed good water solubility, moisture absorption and retention properties, antioxidation, and promote cell proliferation. The aim is to provide a theory for developing novel biomaterials. The main research contents and results are as follows:(1) Collagen-chitosan was synthesized with MTGase as biocatalyst. And the reaction conditions were optimized through one-factor-at-a-time experiment. The optimal conditions were found to be as follows: the reaction time was 1 h, the reaction temperature was 40 oC, the mass ratio of collagen to chitosan was 1.0, the mass ratio of MTGase to chitosan was 0.1. The moisture absorption and retention test showed that the moisture-retention ability of collagen-chitosan was improved with the DS increased. In vitro antioxidant test, the DS of collagen-chitosan had no evident effect on the H2O2 scavenging activity. However, with the concentration increased, the H2O2 scavenging activity of collagen-chitosan was increased, especially with high DS. With the increased concentration of collagen-chitosan, the DPPH scavenging activity slowly enhanced at first stage, and then sharply increased at later stage. The results of cell viability of fibroblast suggested that collagen-chitosan could efficiently promote the proliferation of fibroblast. With the increase of concentration, the cell viability increased. But the DS of collagen-chitosan had little effect on cell viability.(2) Collagen- carboxymethylated chitosan was synthesized with MTGase as biocatalyst. And the reaction conditions were optimized through one-factor-at-a-time experiment. The optimal conditions were found to be as follows: the reaction time was 6 h, the reaction temperature was 40 oC, the mass ratio of collagen to carboxymethylated chitosan was 1.0, the pH was 1.0. The moisture absorption and retention test showed that the moisture-retention ability of collagen-chitosan was improved with the DS increased. In vitro antioxidant test, the DS of collagen-chitosan had no evident effect on the H2O2 scavenging activity. However, with the concentration increased, the H2O2 scavenging activity of collagen-chitosan was increased, especially with high DS. With the increased concentration of collagen-chitosan, the DPPH scavenging activity enhanced obviously at first stage, and then slowly increased at later stage. At high concentration, the DPPH scavenging activity of collagen- carboxymethylated chitosan increased with the increase of DS. The results of cell viability of fibroblast suggested that collagen-chitosan could efficiently promote the proliferation of fibroblast. With the increase of concentration, the cell viability increased at first, and then reduced. At the same concentration, the cell viability increased with the increase of DS.(3) Nisin-chitosan was synthesized with MTGase as biocatalyst. And the reaction conditions were optimized through one-factor-at-a-time experiment. The optimal conditions were found to be as follows: the reaction time was 1.5 h, the reaction temperature was 30 oC, the mass ratio of nisin to chitosan was 1.2. When the DS were 0.098 and 0.473, the transparent of nisin-chitosan were over 90%. Even when the concentration was 20 mg/ml, the transparent of nisin-chitosan were over 90%. At pH 6.0, nisin-chitosan showed good antibacterial property. In vitro cytotoxicity test, nisin-chitosan with the concentration from 0.005 to 0.5 mg/ml, nisin-chitsoan showed no toxicity.