In this study, preconcentration and separation of Cu(II) and Pb(II) ions by using Fe3O4@SiO2@Bacillus pumilis before their determinations by flame atomic absorption spectrometry (FAAS) were investigated. The thermophilic Bacillus pumilis were isolated from Meyremderesi spring, Sirnak, Turkey. Effects of important parameters such as pH, adsorbent amount, eluent type, concentration and volume of eluent and sample volume on magnetic solid phase extraction (MSPE) were examined in details. The preconcentration factors for Cu(II) and Pb(II) ions were calculated as 30 and 40, respectively. The accuracy of the proposed extraction procedure was validated analysing certificated reference materials and addition - recovery tests. The concentration of copper and lead were determined in water samples from Turkey by Flame AAS after application developed preconcentration-separation method.