5-Fluorouracil (5-FU) is an important chemotherapy that is classified as an antimetabolite and is widely used in cancer treatment, particularly for liver cancer. Yet, administering high doses of 5-FU for liver cancer can result in a range of adverse effects. Therefore, employing a suitable delivery system for this important drug can greatly enhance its safety and effectiveness, leading to improved treatment outcomes and better quality of life for patients. Au-doped Fe3O4 nanoparticles have attracted significant interest in scientific research due to their unique physical and chemical properties. This study presents a straightforward and cost-effective method for synthesizing Au-doped Fe3O4 nanoparticles by using a natural reducing agent. The study involved a comprehensive analysis of the Au-doped Fe3O4 nanoparticles using various techniques such as FT-IR, XRD, XPS, FE-SEM, TEM, UV–Vis, EDS, VSM, and DLS. The delivery experiments indicated a loading capacity of approximately 78 %. The release profile showed slow release profile, dependent on pH. Therefore, 41 % release was detected under physiological pH 7.4, while a much faster release was observed at pH 5.7 (84 %) within 48 h. Moreover, the high crystallinity degree of approximately 84 % was attained for Au-doped Fe3O4, confirming this nanoparticle as an efficient drug delivery system. In another attempt, PDI and hydrated particle size distribution analyses on Au-doped Fe3O4 showed no significant change in size after 72 h. Therefore, it can be concluded that drug loaded Au-doped Fe3O4 is stable enough and maintained its efficacy after a long time of 3 days. Finally, an in vitro assessment of cellular cytotoxicity was performed on a liver cancer cell line (HepG2) to assess the safety and efficacy of the Au-doped Fe3O4 nanoparticles. The results from the MTT assay confirmed the potential of these nanoparticles as a promising vehicle for delivering 5-Fluorouracil.