Nanosized iron oxide powders are materials considered with regard to its application in medical therapy called hyperthermia. Magnetite nanopowders with crystallite size varying from 6.6 to 11.8 nm have been prepared by the co-precipitation method. In this study a change of a crystallite size is driven mainly by varying of initial pH of water ammonia solution in which a process of magnetite precipitation runs. Crystallographic structures and phase composition obtained samples and the size of magnetite nanoparticles were determined by X-ray diffraction method. Positron lifetime spectroscopy has been used to assess defectiveness of microstructure. Experimental positron annihilation spectra were successfully resolved into three lifetime components. It appears that from point of view of microstructure the defects concentrations in studied nanopowder samples are very high which causes a saturation of positron trapping.