The study aimed to determine circulatory endotoxin concentration, cytokine profile, and gastrointestinal symptoms of ultra-endurance runners (UER, n=17) in response to a 24-h continuous ultra-marathon competition (total distance range: 122–208 km) conducted in temperate ambient conditions (0–20°C) in mountainous terrain. Body mass and body temperature were measured, and venous blood samples were taken before and immediately after competition. Samples were analysed for gram-negative bacterial endotoxin, C-reactive protein, cytokine profile, and plasma osmolality. Gastrointestinal symptoms were also monitored throughout competition. Mean exercise-induced body mass loss was (mean±SD) 1.7±1.8% in UER. Pre- and post-competition plasma osmolality in UER was 286±11 mOsmol·kg−1 and 286±9 mOsmol·kg−1, respectively. Pre- to post-competition increases (p0.05). Gastrointestinal symptoms were reported by 75% of UER, with no symptoms reported by CON. IL-10 (r=0.535) and IL-8 (r=0.503) were positively correlated with gastrointestinal symptoms. A 24-h continuous ultra-marathon competition in temperate ambient conditions resulted in a circulatory endotoxaemia and pro-inflammatory cytokinaemia, counteracted by a compensatory anti-inflammatory response.