Deep (>0.8 km depth) fracture water with residence time estimates on the order of several Ma from the Witwatersrand Basin, South Africa contains up to 40 mu M of NO3-, up to 50 mM N-2 (90 times air saturation at surface) and 1 to -400 mu M NH3/NH4+ To determine whether the oxidized N species were introduced by mining activity, by recharge of paleometeoric water, or by subsurface geochemical processes, we undertook N and 0 isotopic analyses of N species from fracture water, mining water, pore water, fluid inclusion leachate and whole rock cores.|The NO2-, NO3- and NH3/NH4+ concentrations of the pore water and fluid inclusion leachate recovered from the low porosity quartzite, shale and metavolcanic units were similar to 10(4) times that of the fracture water. The delta N-15-NO3- and delta O-18-NO3- of the pore water and fluid inclusion leachate, however, overlapped that of the fracture water with the delta N-15-NO3- ranging from 2 to 7 parts per thousand and the delta N-18-NO3- ranging from 20 to 50 parts per thousand. The delta N-15-NO3- of the mining water ranged from 0 to 16 parts per thousand and its delta N-18-NO3- from 0 to 14 parts per thousand making the mining water NO3- isotopically distinct from that of the fracture, pore and fluid inclusion water. The delta N-15-N-2 of the fracture water and the delta N-15-N from the cores ranged from -5 to 10 parts per thousand and overlapped the delta N-15-NO3-. The delta N-15-NO4+ of the fracture water and pore water NH3/NH4+ ranged from -15 to 4 parts per thousand. Although the NO3- concentrations in the pore water and fluid inclusions were high, mass balance calculations indicate that NO3- accounts for <= 10% of the total rock N, whereas NH3/NH4+ trapped in fluid inclusions or NH4+ present in phyllosilicates account for >= 90% of the total N.|Based on these findings, the fluid inclusion NO3- appears to be the source of the pore water and fracture water NO3- rather than paleometeoric recharge or mining contamination. Irradiation experiments indicate that radiolytic oxidation of NH3 to NO3- can explain the fluid inclusion NO3- concentrations and, perhaps, its isotopic composition, but only if the NO3- did not attain isotopic equilibrium with the hydrothermal fluid 2 billion years ago. The delta N-15-N, delta N-15-N-2 and delta N-15-NO4+ suggest that the reduction of N-2 to NH4+ also must have occurred in the Witwatersrand Basin in order to explain the abundance of NH4+ throughout the strata. Although the depleted NH3- concentrations in the fracture water relative to the pore water are consistent with microbial NH3- reduction, further analyses will be required to determine the relative importance of biological processes in the subsurface N cycle and whether a complete subsurface N cycle exists. (C) 2011 Elsevier B.V. All rights reserved.