Physical properties of type I collagen extracted from fish scales of Pagrus major and Oreochromis niloticas

Abstract

Type I collagens were extracted from fish scales of Pagrus major and Oreochromis niloticas as a possible underutilized resource for medical materials. The fish scales were demineralized with EDTA and digested by pepsin. The resultant type I collagens contained more than 33.6% of glycine as the most abundant amino acid. The denaturation temperatures of the collagens from P. major and O. niloticas were 303 and 308 K, respectively, both of which were relatively lower than that of porcine dermis collagen (314 K). CD spectra indicated that the denaturation temperatures were dependent on the amount of hydroxyproline, rather than proline residues. Raman spectra also indicated that the relative intensities of Raman lines at 879 and 855 cm⁻¹ assigned to Hyp and Pro rings were changed due to the contents of the imino acids. Significantly, the content of sulphur-containing methionine was higher in the fish scales than in porcine dermis. The enthalpy and entropy estimated from thermal analyses could be correlated to amino acid sequences (Gly-Pro-Hyp) of type I collagens and the number of methionine amino acid residues.

Introduction

Type I collagen is the main component of extracellular matrix and has functions that include mechanical protection of tissues and organs or physiological regulation of the cell environment [1]. The use of type I collagen in industry for health foods, cosmetics, and biomaterials is expanding. The benefits of type I collagen for the use of biomaterials are its low antigenic and direct cell adhesion properties. At present, the main sources of type I collagen in many fields are limited to those of bovine or porcine dermis.

Type I collagens have been extracted from the skin of aquatic species, e.g. jellyfish [2], [3], starfish [4], octopus [5], paper nautilus [6], cuttlefish [7], purple sea urchin [8], and others [9], [10], [11], [12], [13]. On the other hand, extraction of type I collagen from fish scales, which are biocomposites of highly ordered type I collagen fibers and hydroxyapatite (Ca₅(PO₄)₃OH) [14], [15], [16], [17], [18], has been described only by Nomura et al. [19]. They extracted collagen from the fish scale of sardine and reported that the acid-solubilized collagen had a denaturation temperature of 300 K, which like the marine skin collagens was lower than the denaturation temperature for porcine dermis (314 K).

In general, pepsin-solubilized collagen (PSC) degenerates at lower temperature than SC and has little antigenic property compared with SC because of the loss of N- and C-terminus domains in PSC. The times and temperatures used for pepsin digestion treatments are important for the production of lower molecular weight fragments than the intact alpha I (I) chains [20]. Raman spectroscopy is particularly useful for analyzing collagen and associated degradation products as shown by Fruchour and Koenig [20], [21], [22], who were able to assign specific functional groups associated with the amino acid backbone of collagen, and thereby identify natural collagens from other proteins.

In this paper, we extract pure native type I collagens from fish scales of Pagrus major (seawater fish) and Oreochromis niloticas (freshwater fish) by a simple method and elucidate the denaturation temperatures with circular dichroism (CD) spectroscopy. We use FT-Raman spectroscopy to correlate these temperatures with changes in the amino acid components.