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Abstract Investigations of abiotic and biotic contributions to dissolved organic carbon (DOC) are required to constrain microbial habitability in continental subsurface fluids. Here we investigate a large (101–283 mg C/L) DOC pool in an ancient (>1Ga), high temperature (45–55 °C), low biomass (10²−10⁴cells/mL), and deep (3.2 km) brine from an uranium-enriched South African gold mine. Excitation-emission matrices (EEMs), negative electrospray ionization (–ESI) 21 tesla Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR MS), and amino acid analyses suggest the brine DOC is primarily radiolytically oxidized kerogen-rich shales or reefs, methane and ethane, with trace amounts of C₃–C₆hydrocarbons and organic sulfides. δ²H and δ¹³C of C₁–C₃hydrocarbons are consistent with abiotic origins. These findings suggest water-rock processes control redox and C cycling, helping support a meagre, slow biosphere over geologic time. A radiolytic-driven, habitable brine may signal similar settings are good targets in the search for life beyond Earth. 

Abstract Deep within the Precambrian basement rocks of the Earth, groundwaters can sustain subsurface microbial communities, and are targets of investigation both for geologic storage of carbon and/or nuclear waste, and for new reservoirs of rapidly depleting resources of helium. Noble gas-derived residence times have revealed deep hydrological settings where groundwaters are preserved on millions to billion-year timescales. Here we report groundwaters enriched in the highest concentrations of radiogenic products yet discovered in fluids, with an associated⁸⁶Kr excess in the free fluid, and residence times >1 billion years. This brine, from a South African gold mine 3 km below surface, demonstrates that ancient groundwaters preserved in the deep continental crust on billion-year geologic timescales may be more widespread than previously understood. The findings have implications beyond Earth, where on rocky planets such as Mars, subsurface water may persist on long timescales despite surface conditions that no longer provide a habitable zone.