Extraction of acid soluble collagen from Coryphaenahippurus(Mahimahi) and its application in tissue engineering

Abstract

Collagen is a structural protein in the extracellular matrix which provides mechanical support and rigidity to the tissues in the skin. Many extraction techniquesprevail for the extraction of collagen of from Fish skin having a wide availability and largely discarded as waste byproduct. Also, skin are the major sites of fibroblasts cells that aid in tissue injury and they have excellent wound healing capacities. Therefore, Current study focuses on extraction of collagen from the skin of Coryphaenahippurus(Mahimahi) fish by acid assisted method. After extraction, the sample was characterized using different spectral methods such as UV, FTIR, SEM, TGA, DSC, MALDI and SDS-PAGE. UV peaks at 270 – 280 nm confirmed the protein peaks of collagen in the sample. TGA analysis showed the poor thermal stability of the collagen. ATR analysis confirmed the presence of functional groups of amide present in the sample. MALDI TOF protein sequencing showed the presence of collagen in the sample. SDS PAGE analysis showed the presence of bands of type I α1, α2 and β chains. Fibril assay and staining using safranin and van gieson dyes showed the fibrillar structure of the collagen under microscope. Hydroxyproline analysis was done to detect the presence of hydroxyproline, the major amino acid in collagen and estimated to be 0.62 µg/mg of sample. After conformation of presence of collagen, wound scratch assays was carried for checking its wound healing applications as well as MTT assays was carried out to estimate its cytotoxicity level. At a higher concentration of 100 µg/ml, the sample showed 88.36% of migration rate and93.2% of cell viability against S3T6 cell line.  In conclusion, we extracted thermostable, bioactive, non-toxic variety of collagen from the Coryphaenahippurus(Mahimahi) fish.