Background: Cobalt is one of the most important essential trace metals of human nutrition. Low doses of cobalt are needed for many humans and animals to stay healthy. Cobalamine (named vitamin B-12) is a cobalt-containing essential vitamin. The direct detection of metal ions in various matrices by AAS may be difficult due to matrix interferences and extremely low levels of metal ions. Thus, a preconcentration and separation step is normally demanded. In this report, a two-step microextraction technique, combining room temperature ionic liquid based dispersive liquid-liquid microextraction (IL-DLLME) and dispersive magnetic solid-phase microextraction (MSPME) was prepared for the flame atomic absorption spectrometric determination (FAAS) of trace cobalt ions in water samples.