Complex interplay between the intestinal environment and the host has attracted considerable attention and has been well studied with respect to the gut microbiome and metabolome. Oxygen free radicals such as superoxide and the hydroxyl radical (•OH) are generated during normal cellular metabolism. They are toxic to both eukaryotic and prokaryotic cells and might thus affect intestinal homeostasis. However, the effect of oxygen free radicals on the intestinal environment has not been widely studied. Herein, we applied electron spin resonance spectroscopy with spin trapping reagents to evaluate oxygen free radical production capacity in the intestinal lumen and the faeces of mice. •OH was generated in faeces and lumens of the small and large intestines. There were no remarkable differences in •OH levels between faeces and the large intestine, suggesting that faeces can be used as alternative samples to estimate the •OH production capacity in the colonic contents. We then compared free radical levels in faecal samples among five different mouse strains (ddY, ICR, C57BL/6, C3H/HeJ, and BALB/c) and found that strain ddY had considerably higher levels than the other four strains. In addition, strain ddY was more susceptible to dextran sulphate sodium-induced colitis. These differences were possibly related to the relative abundance of the gut bacterial group Candidatus Arthromitus, which is known to modulate the host immune response. From these results, we suggest that the production capacity of oxygen free radicals in mouse faeces is associated with intestinal homeostasis.