Comparative Study Of Three-spot Seahorse Skin And Nile Tilapia Skin Collagen

The preliminary experiment suggested that pepsin-solubilized collagen(PSC)from the collagen-rich skin of three-spot seahorse(Hippocampus trimaculatus)was preliminarily identified as type I collagen while Nile tilapia skin collagen was also the type I.Although two kinds of PSC were both identified as type I collagen,they were different from amino acids composition to molecular structure in some degree.In recent years,H.trimaculatus has been generally considered as a highly valued species in medicinal and aquarium trades,however,rarely research reports H.trimaculatus skin collagen(PSC-S).It is well known that there are plenty of studies about Nile tilapia skin collagen(PSC-T).In order to make the most of H.trimaculatus,this research characterized physicochemical properties of H.trimaculatus skin collagen which was then compared with Nile tilapia skin collagen.In a word,this article provided the theoretical basis for further research and market application in H.trimaculatus skin collagen.The research contents are as follows:1.The extraction and composition analysis of H.trimaculatus and Nile tilapia skin.The contents of the total crude protein from H.trimaculatus skin and Nile tilapia skin were respectively 34.08%and 33.45%(wet weight basis).Through determining the hydroxyproline,it indicated that the contents of collagen in H.trimaculatus skin and Nile tilapia skin were 18.34%and 27.23%(wet weight basis).In order to extract more collagen from H.trimaculatus skin and Nile tilapia skin,PSC extraction method was used to isolate PSC-S and PSC-T.The recovery ratio of PSC-S was 40.41%(dry weight basis),which was lower than that of PSC-T(52.6%).After amino acids analysis,the amino acids compositions of PSC-S and PSC-T basically fitted the characteristics of type I collagen,but the contents of partial amino acids were different in some degree.Through contrasting research,among the amino acids compositions of these two collagens,relatively high contents of glycine,alanine,proline,hydroxyproline,and glutamic acid were detected.It was worth mentioning that imino acid(proline and hydroxyproline)belonged to characteristic amino acid of collagen was less in PSC-S than in PSC-T.2.The examination of fundamental characteristics.For further research into the different properties of PSC-S,this study respectively determined the SDS-PAGE,ATR-FTIR,DSC and ZETA potential.The analysis results of SDS-PAGE and ATR-FTIR suggested that PSC-S and PSC-T were preliminarily identified as type I collagen while the structure of imino acids was different between PSC-S and PSC-T.The DSC index indicated that the denaturation temperature of PSC-S was 35.12 ? which was lower than that of PSC-T(38.66 ?).ZETA potential showed that the isoelectric points of PSC-S and PSC-T were respectively 5.72 and 5.97.3.The gel-forming capacity of PSC-S and PSC-T.It is well known that the gel-forming ability is an important physical index of collagen.Thus,this study determined gel-forming capacity of PSC-S and PSC-T under different conditions(pH and NaCl).The highest gel-forming capacity of PSC-S was observed when gel-forming condition was adjusted at pH 6.8 of phosphate buffer and at an ionic strength of 400 mM NaCl.The highest gel-forming capacitie of collagens from the skin of Nile tilapia was obtained at pH 7.1 in 50 mM NaCl.4.The highest gel-forming capacity of PSC-S and PSC-T after EDC crosslinking.The object of exploring gel-forming capacity was to make PSC-S better applied into the medical biomaterial.1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide(EDC)was introduced into collagen solution during collagen gel-formation.After EDC crosslinking,the thermal stability and mechanical strength of PSC-S gel reached the maximum values at the conditions:pH 6.8,10 mM Na-phosphate buffer,100 mM NaCl and 25 mM EDC.For PSC-T gel after EDC crosslinking,the best gel-forming conditions was Na-phosphate buffer(pH 6.3,40 mM)including 100 mM NaC1 and 50 mM EDC.5.Characterization of PSC-S and PSC-T gel after EDC crosslinking.After EDC crosslinking,the melt temperature,mechanical strength,structural morphology(scanning electron microscopy/SEM)and cell proliferation rates of PSC-S and PSC-T gel were evaluated.By the contrast analysis,the melt temperature and mechanical strength PSC-S gel after EDC crosslinking were improved significantly and were much better than those of PSC-T gel.At the same time,the SEM analysis suggested that the microstructure of PSC-S gel after EDC crosslinking was changed obviously and the collagen fibril bundle become thick and porous.Moreover,compared with PSC-T gel,collagen fibril bundle of PSC-S gel was thicker.The result of cell proliferation rates evaluating suggested that PSC-S gel after EDC crosslinking was more suitable for the proliferation of MC3T3-E1 cell compared to PSC-T and PSC-P gel.