Lyophilized samples of human liver ferritin and chicken liver and spleen were measured at room temperature using Mössbauer spectroscopy with high velocity resolution. An increase in the velocity resolution of Mössbauer spectroscopy permitted us to increase accuracy and decrease experimental error in determining the hyperfine parameters of human liver ferritin and chicken liver and spleen. Mössbauer spectroscopy with high velocity resolution may be very useful for revealing small differences in hyperfine parameters during biomedical research.Keywords Mössbauer spectroscopy · High velocity resolution · Human liver ferritin · Chicken liver and spleen