A Novel Enzymatic Method for Preparation and Characterization of Collagen Film from Swim Bladder of Fish Rohu (Labeo rohita)


A novel enzymatic method for extraction and preparation of fish collagen from swim bladder revealed the occurrence of α, β and γ bands with approximately 12.1 g/100g collagen corresponding to 89% of collagen and thus confirmed the nativity and purity of the fish collagen. FT-IR studies confirmed the retention of all three amide bands of I, II and III, and triple helixcity. UN-crosslinked and UV-crosslinked fish collagen membrane records a very high temperature of helix denaturation at 197℃ and 215℃, shrinkage temperature at 50℃ ± 3.2℃ and 62℃ ± 2.7℃ and tensile strength at 16.89 ± 2.5 and 120.02 ± 1.0 Kg/cm2 respectively. Fish collagen matrix promoted NIH 3T3 and L6 cellular growth and proliferation. The study indicates that availability of pure fish collagen could replace bovine collagen in tissue engineering applications.

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Sripriya, R. and Kumar, R. (2015) A Novel Enzymatic Method for Preparation and Characterization of Collagen Film from Swim Bladder of Fish Rohu (Labeo rohita). Food and Nutrition Sciences, 6, 1468-1478. doi: 10.4236/fns.2015.615151.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Muyonga, J.H., Cole, C.G.B. and Duodu, K.G. (2004) Characterisation of Acid Soluble Collagen from Skins of Young and Adult Nile Perch (Lates niloticus). Food Chemistry, 85, 81-89.
[2] Usha, R., Maheshwari, R., Dhathathreyan, A. and Ramasami, T. (2006) Structural Influence of Mono and Polyhydric Alcohols on the Stabilization of Collagen. Colloid Surface B, 48, 101-105.
[3] Pinazo, A. (1996) Effect of Surfactant Structure on Diffusion through a Collagen Membrane. Colloid Surface B, 8, 63-72.
[4] Reid, G., Lam, D., Policova, Z. and Neumann, A.W. (1993) Adhesion of Two Uropathogens to Silicone and Lubricious Catheters: Influence of pH, Urea and Creatinine. Journal of Materials Science: Materials in Medicine, 4, 17-22.
[5] Ogawa, M., et al. (2004) Biochemical Properties of Bone and Scale Collagens Isolated from the Subtropical Fish Black Drum (Pogonis cromis) and Sheepshead Seabream (Archosargus probatocephalus). Food Chemistry, 88, 495-501.
[6] Fernandes, R.M., Couto Neto, R.G., Paschoal, C.W., Rohling, J.H. and Bezerra, C.W. (2008) Collagen Films from Swim Bladders: Preparation Method and Properties. Colloid Surface B, 62, 17-21.
[7] Sadowska, M., Kolodziejska, I. and Niecikowska, C. (2003) Isolation of Collagen from the Skins of Baltic Cod (Gadus morhua). Food Chemistry, 81, 257-262.
[8] Nagai, T. and Suzuki, N. (2000) Isolation of Collagen from Fish Waste Material—Skin, Bone and Fins. Food Chemistry, 68, 277-281.
[9] Nagai, T. and Suzuki, N. (2002) Preparation and Partial Characterization of Collagen from Paper Nautilus (Argonauta argo, Linnaeus) Outer Skin. Food Chemistry, 76, 149-153.
[10] Senaratne, L.S., Park, P.J. and Kim, S.K. (2006) Isolation and Characterization of Collagen from Brown Backed Toadfish (Lagocephalus gloveri) Skin. Bioresource Technology, 97, 191-197.
[11] Shahidi, F. (1994) Seafoods: Chemistry, Processing, Technology and Quality. Blackie Academic and Professional, Glasgow.
[12] Sano, A., Maeda, M., Nagahara, S., Ochiya, T., Honma, K., Itoh, H., et al. (2003) Atelocollagen for Protein and Gene Delivery. Advanced Drug Delivery Reviews, 55, 1651-1677.
[13] Huang-Lee, L., Cheung, D.T. and Nimni, M.E. (1990) Biochemical Changes and Cytotoxicity Associated with the Degradation of Polymeric Glutaraldehyde Derived Crosslinks. Journal of Biomedical Materials Research, 24, 1185-1201.
[14] Koide, M., Osaki, K., Konishi, J., Oyamada, K., Katakura, T., Takahashi, A. and Yoshizato, K. (1993) A New Type of Biomaterial for Artificial Skin: Dehydrothermally Cross-Linked Composites of Fibrillar and Denatured Collagen. Journal of Biomedical Materials Research, 27, 79-84.
[15] Safandowska, M. and Pietrucha, K. (2013) A New Method of Determination of Collagen Conjugated with Keratin. Autex Research Journal, 13, 37-39.
[16] Bama, P., Vijayalakshimi, M., Jayasimman, R., Kalaichelvan, P.T., Deccaraman, M. and Sankaranarayanan, S. (2010) Extraction of Collagen from Cat Fish (Tachysurus maculatus) by Pepsin Digestion and Preparation and Characterization of Collagen Chitosan Sheet. International Journal of Pharmacy and Pharmaceutical Sciences, 2, 133-137.
[17] Sripriya, R., Ahmed, M.R., Sehgal, P.K. and Jayakumar, R. (2003) Influence of Laboratory Ware Related Changes in Conformational and Mechanical Properties of Collagen. Journal of Applied Polymer Science, 87, 2186-2192.
[18] Neuman, R.E. and Logan, M.A. (1950) Determination of Collagen and Elastin in Tissues. The Journal of Biological Chemistry, 186, 549-556.
[19] Leammli, U.K. (1970) Discontinuous Buffer System for Slab Gel. Nature, 227, 680-681.
[20] Mosmann, T. (1983) Rapid Colorimetric Assay for Cellular Growth and Survival: Application to Proliferation and Cytotoxicity Assays. Journal of Immunological Methods, 65, 55-63.
[21] Marshall, N.J., Goodwin, C.J. and Holt, S.J. (1995) A Critical Assessment of the Use of Microculture Tetrazolium Assays to Measure Cell Growth and Function. Growth Regulation, 5, 69-84.
[22] Spackman, D.H., Stein, W.H. and Moore, S. (1958) Automatic Recording Apparatus for Use in the Chromatography of Amino Acids. Analytical Chemistry, 30, 1190-1205.
[23] Oliver, R.F., Barker, H., Cooke, A. and Grant, R.A. (1982) Dermal Collagen Implants. Biomaterials, 3, 38-40.
[24] Liu, D., Liang, L., Joe, M. and Zhou, R.P. (2012) Extraction and Characterisation of Pepsin-Solubilised Collagen from Fins, Scales, Skins, Bones and Swim Bladders of Bighead Carp (Hypophthalmichthys nobilis). Food Chemistry, 133, 1441-1448.
[25] Goes, J.D., Figueiro, S.D., de Paiva, J.A.C., de Vasconcelos, I.F. and Sombra, A.S.B. (2002) On the Piezoelectricity of Anionic Collagen Films. Journal of Physics and Chemistry of Solids, 63, 465-470.
[26] Silva, C.C., Pinheiro, A.G., Thomazini, D., Góes, J.C., Figueiró, S.D., de Paiva, J.A.C. and Sombra, A.S.B. (2001) Effect of the pH on the Piezoelectric Properties of Collagen Films. Materials Science and Engineering: B, 83, 165-172.
[27] Wong, D.W.S. (1989) Mechanism and Theory in Food Chemistry. Van Nostrand Reinhold Company, New York.
[28] Cooper, D.R. and Davidson, R.J. (1965) The Effect of Ultraviolet Irradiation on Soluble Collagen. Biochemical Journal, 97, 139-147.
[29] Fujimori, E. (1965) UV Light-Induced Change in Collagen Macromolecules. Biopolymers, 3, 115-119.
[30] Singer, A.J. and Clark, R.A. (1999) Cutaneous Wound Healing. The New England Journal of Medicine, 341, 738-746.
[31] Sefton, M.V. and Woodhouse, K.A. (1998) Tissue Engineering. Journal of Cutaneous Medicine and Surgery, 3, 18-23.
[32] Schor, S.L. (1980) Cell Proliferation and Migration on Collagen Substrata in Vitro. Journal of Cell Science, 41, 159-175.
[33] Chen, F., Yoo, J.J. and Atala, A. (1999) Acellular Collagen Matrix as a Possible “Off the Shelf” Biomaterial for Urethral Repair. Urology, 54, 407-410.
[34] El-Kassaby, A.W., Retik, A.B., Yoo, J.J. and Atala, A. (2003) Urethral Structure Repair with an Off-the-Shelf Collagen Matrix. The Journal of Urology, 169, 170-173.

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