Stable isotope composition varies due to different reactivity or mobility among the isotopes. Various pioneering studies revealed that isotope fractionation is common for many elements, and it is now widely recognized that the stable isotope compositions of biometals can be used as new tracers for element metabolism. In this review, we summarize the recently published isotope compositions of iron (Fe), copper (Cu), zinc (Zn), and calcium (Ca) in various biological samples, including tissues from plants, animals, and humans. Discussions were carried out with respect to age, sex, organ, and the presence or absence of particular diseases for animals and humans. For Fe and Cu isotopes, changes in oxidation states generate large isotopic fractionation through the metabolism of those elements. Isotope composition of Zn greatly fractionates among tissues even without changes in oxidation state. Isotopic composition of Ca is a powerful tracer for the metabolism of Ca in bones. The review results suggest that the stable isotope compositions of the biometals can be used as effective markers for diagnostics of various kinds of diseases related to metabolic disorders.