Isolation, Characterization and Biological Evaluation of Collagen from Rhizostoma pulmo Jellyfish from the Sea of Azov for Biomedical Applications

Collagen is a major extracellular-matrix protein widely used in regenerative medicine, yet conventional terrestrial sources raise biosafety and acceptability concerns, motivating the search for marine alternatives. This study evaluates the jellyfish <i>Rhizostoma pulmo</i> (<i>R. pulmo</i>) from the Azov Sea as a sustainable collagen source and assesses its suitability for biomedical materials. Acid-soluble collagen was extracted using 0.5 M acetic acid and purified by salt precipitation and dialysis, followed by physicochemical/structural characterization (sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Limulus amebocyte lysate (LAL) endotoxin testing, transmission electron microscopy (TEM), and immunofluorescence with type I collagen antibodies) and biological evaluation <i>in vitro</i> (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity on MRC5 fibroblasts; adhesion and proliferation assays on HeLa cells). The extracted collagen showed a high yield (~26.2%), a type I-like electrophoretic profile with α-, β-, and γ-components, fibrillar ultrastructure by TEM, and positive type I collagen immunoreactivity; endotoxin levels were low (0.461 EU/µL), and no cytotoxicity was detected under the tested conditions. Porous collagen sponges/scaffolds were fabricated by lyophilization, displaying interconnected pores with an average size of ~80 µm and pH-dependent swelling, and they supported 3D cell growth and tumor-cell dissemination in an <i>in vitro</i> breast carcinoma scaffold model. Overall, Azov Sea <i>R. pulmo</i> collagen demonstrates promising structural quality, low endotoxin burden, and cytocompatibility, supporting its potential as a marine biomaterial for sponge/scaffold-based tissue engineering and wound-related applications.