The sustainable production of cell-cultivated meat requires the use of environmentally friendly nutrients. Culture media, which provides the necessary nutrients for cell proliferation and differentiation is a major contributor to the environmental impacts of cultivated meat production. Fetal bovine serum (FBS) is problematic due to animal sourcing, ethical concerns, and high cost. Beefy-R and Beefy-9 are experimentally validated, serum-free cell culture media designed for bovine myoblast culture. While both formulations include basal media, ascorbic acid, sodium selenite, recombinant proteins, and growth factors, Beefy-9 and Beefy-R contain recombinant albumin and rapeseed protein isolates (RPI) to replace FBS, respectively. While these culture media do not include animal-sourced components, their environmental impacts have not yet been investigated. In this study, life cycle assessment (LCA) was performed to estimate the environmental burdens of Beefy-9 and Beefy-R culture media production and identify hotspots for optimization. Uncertainty was assessed through sensitivity and scenario analyses, as well as Monte Carlo simulation. Our results demonstrate that Beefy-R has significantly lower environmental impacts compared to Beefy-9 in 13 out of 18 evaluated impact categories. For instance, the global warming potential (GWP) of Beefy-R was almost five times lower when compared against that of Beefy-9 production. In Beefy-R, despite the absence of recombinant albumin, recombinant proteins and growth factors were the primary hotspots, accounting for 41 %, 5.9 %, and 12 % of GWP, land use, and water consumption impacts, respectively. This study confirms that RPI can be a more sustainable substitute for recombinant albumin in bovine myoblast serum-free media formulations. Additional research is required to identify low-impact alternatives for other recombinant proteins and growth factors in culture media.