The cultivation of cyanobacteria by exploiting available in situ resources represents a possible way to supply food and oxygen to astronauts during long-term crewed missions on Mars. Here, we evaluated the possibility of cultivating the extremophile cyanobacterium Chroococcidiopsis thermalis CCALA 050 under operating conditions that should occur within a dome hosting a recently patented process to produce nutrients and oxygen on Mars. The medium adopted to cultivate this cyanobacterium, named Martian medium, was obtained using a mixture of regolith leachate and astronauts’ urine simulants that would be available in situ resources whose exploitation could reduce the mission payload. The results demonstrated that C. thermalis can grow in such a medium. For producing high biomass, the best medium consisted of specific percentages (40%vol) of Martian medium and a standard medium (60%vol). Biomass produced in such a medium exhibits excellent antioxidant properties and contains significant amounts of pigments. Lipidomic analysis demonstrated that biomass contains strategic lipid classes able to help the astronauts facing the oxidative stress and inflammatory phenomena taking place on Mars. These characteristics suggest that this strain could serve as a valuable nutritional resource for astronauts.