Exercise-associated hyponatremia (EAH) has mainly been investigated in runners and triathletes. In mountain bikers, EAH was studied in two multi-stage races, but not in a single stage race. The aim of this study was to investigate the prevalence of EAH in a single-stage mountain bike ultra-marathon. In the 'Swiss Bike Masters' over 120 km with a climb of ~ 5,000 m in altitude, we determined pre and post race body mass, hematocrit, plasma sodium concentration ([Na⁺]), and urinary specific gravity in 37 cyclists. Athletes recorded their fluid intake while racing. No athlete developed EAH. The cyclists drank on average (means ± SD) 0.7 ± 0.2 l/h. Fluid intake was significantly and negatively related to race time (r = -0.41, P < 0.05), but showed no association with post race plasma [Na⁺], the change in plasma [Na⁺], post race body mass, or the change in body mass. The athletes lost 1.4 kg body mass (P < 0.05), plasma [Na⁺] decreased by 0.7% (P < 0.05), plasma volume increased by 1.4% and urinary specific gravity increased by 0.4% (P < 0.05). The change in body mass was neither related to post race plasma [Na⁺] nor to the change in plasma [Na⁺]. The decrease in plasma [Na⁺] was not related to fluid intake. The change in plasma [Na⁺] was related to post race plasma [Na⁺] (r = 0.40, P < 0.01). Ad libitum fluid intake showed no case of EAH in a single-stage mountain bike ultra-marathon. In contrast to previous findings, the faster athletes drank more than the slower ones.