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We designed graphene oxide composites with increased morphological and structural variability using fatty acid-coupled polysaccharide co-polymer as the continuous phase. The matrix was synthesized by N, O-acylation of chitosan with palmitic and lauric acid. The obtained co-polymer was crosslinked with genipin and composited with graphene oxide. FTIR spectra highlighted the modification and multi-components interaction. DLS, SEM, and contact angle tests demonstrated that the conjugation of hydrophobic molecules to chitosan increased surface roughness and hydrophilicity, since it triggered a core-shell macromolecular structuration. Nanoindentation revealed a notable durotaxis gradient due to chitosan/fatty acid self-organization and graphene sheet embedment. The composited building blocks with graphene oxide were more stable during in vitro enzymatic degradation tests and swelled less. In vitro viability, cytotoxicity, and inflammatory response tests yielded promising results, and the protein adsorption test demonstrated potential antifouling efficacy. The robust and stable substrates with heterogeneous architecture we developed show promise in biomedical applications.