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Collagen dressings have been widely used as effective treatments for chronic wounds acting as barrier, protecting the area from infections and participating in the healing process. Collagen from fish skin is biocompatible, presents low immunogenicity and is able of stimulating wound healing. In this scenario, skin of flounder fish (Paralichthys sp.) may constitute a promising source for collagen. Then, our hypothesis is that fish collagen is able of increasing cell proliferation, with no cytotoxicity. In this context, the aim of the present study was to investigate the physicochemical and morphological properties of collagen using scanning electron microscopy (SEM), Energy dispersive X-ray spectroscopy (EDS), mass loss and pH. Moreover, the cytotoxicity and genotoxicity of collagen were studied using in vitro studies (cell viability, comet assay and micronucleus assay). Fish collagen showed no variation of pH and mass weight, with characteristic peaks of collagen in FTIR. Furthermore, all the extracts presented cell viability at least over 50% and no cytotoxicity was observed. Regarding genotoxicity data, the results showed that only the extract of 100% showed higher values in comparison with negative control group for CHO-K1 cell line as depicted by comet and micronucleus assays. Based on the results, it is suggested that fish collagen is biocompatible and present non-cytotoxicity in the in vitro studies, being considered a suitable material for tissue engineering proposals., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)