Quantification of corticosterone metabolites excreted in faeces and urine is increasingly being used for assessment of preceding corticosterone concentrations in the circulation. This is a promising approach to non-invasive stress assessment in laboratory rodents. It is however unknown whether the proportions of corticosterone metabolites excreted in faeces and urine may differ, depending on the concentration of corticosterone in blood. This uncertainty undermines the applicability of urinary and faecal corticosterone metabolite measurements as biomarkers for stress. Therefore, the terminal distribution and time course of corticosterone excretion, after intravenous injection of varying corticosterone concentrations, was investigated in female mice. Female BALB/c mice excreted 60% of all corticosterone in the urine with an approximate delay of 5h from tail vein administration. The remaining 40% were excreted in faeces, with an approximate delay of 9h from administration. The faecal/urinary excretion ratio, as well as time course of excretion, remained unaltered by administration of various doses of corticosterone covering the entire physiological range of serum corticosterone. Although currently untested for other strains of mice and species of animals, these findings add credence to the utility of faecal and urinary corticosterone as non-invasive biomarkers for physiological stress.