| Collagen is a structure protein which exists in organisms widely. In recent years, collagen is used widely in tissue engineering of medicine as a biological material. In this study, we got acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC) extracted from grass carp fish scale by methods of acid and acid- pepsin. On the basis of polymer structure, it has a system study on biological properties compared to pig collagen (PC) including solution behavior, formation and texture of gel, vitro enzymatic degradation and material properties. This provides data base for application in biomedical materials.ASC, PSC and PC were extracted at low temperature. These collagens are typical typeⅠcollagen and high purity by infrared spectroscopy and SDS electrophoresis. Amino acid composition analysis showed that the extraction method and source of collagen may lead to differences in amino acid composition.Using turbidity test method for preliminary study on vitro self-assembly behavior in solution. The results show that under the same conditions, the self-assembly degree of PC is significantly higher than ASC and PSC. Meanwhile, the vitro self-assembly behavior of collagen in the solution is affected by environmental temperature, the pH of system, concentration of collagen and ions. Circular dichroism analysis show that the conformation and behavior of collagen molecules in the solution are affected by many factors. Under the same conditions, the circular dichroism signal value of PSC is the most big, followed by PC and ASC. The treatment of thermal denaturation, the pH closing to the protein isoelectric point and adding calcium can lead to decrease of the value of circular dichroism signal of collagen. It indicates that these factors can affect the behavior of collagen molecules in solution. It analyzes that thermal denaturation of collagen of three collagen by the temperature scan mode of circular dichroism. The results show that in the heat denaturation process, the collagens have two types of denaturation phase, suggesting the existence of domains of thermal stability of two different in the collagen molecules. The initial denaturation temperatures of ASC, PSC, and PC are 33℃, 35℃and 40℃respectively.It gets the formation of and viscoelastic properties of three collagen gel using viscoelastic model of dynamic rheometer ,and texture, influence factors using texture analyzer. The results show that pH and protein concentration are the decisive factor which affect on the conversion from solution into sol (gel). Among them, the critical points of change concentration are all 5mg/mL.The critical pH of ASC is 4.5, but that of PSC and PC are about 5. Variable temperature experiments, deformation rate scan, creep test and texture analysis show that ASC is a kind gel with strong hardness, high brittleness, while PSC and PC gel are soft and good toughness. These three gels are subjected to temperature and heating treatment can lead to the irreversible destruction of ASC structure, showing a sustained decline in modulus of elasticity; but for PSC and PC, heating can promote the orderly rearrangement of collagen molecules below the denaturation temperature, so heating can enhance the performance of PSC and PC gel texture.Enzymatic degradation in vitro of the collagens shows that ASC, PSC, and PC have different enzyme sensitivity. Under the same conditions, PC degradation rate is the slowest, followed by PSC and ASC. The treatment of thermal denaturation would improve the sensitivity of protein, but the self-assembly process before enzymatic degradation is opposite.It gets preliminary evaluation about material properties of three collagen sponges by tensile test, specific surface area measurement and SEM observation. The results show that PSC and ASC from freshwater sources are sponge materials with porous structure and weaker tensile strength. But PC from mammals is a sponge materials with great tensile strength and dense structure. So different collagens have different applications in biomedical materials. |
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