In this study, Fe3O4 (magnetite) nanoparticles were synthesized by in situ in polyacrylamide (PAAm) gels. Structural and magnetic properties of magnetite nanoparticles were investigated by X-Ray diffractometry (XRD), transmission electron microscopy (TEM) and vibrating sample magnetometry (VSM) methods. Interaction of pyranine molecules with Fe3O4 nanoparticles was investigated by fluorescence spectroscopy measurements in which pyranine (POH) molecules were used as a flouroprobe. It was observed that magnetite nanoparticles have single domain structure with average grain size of 12 nm which was also supported by magnetization measurements. M-H hysteresis curves revealed the superparamagnetic nature of magnetite nanoparticles synthesized in PAAm gels. Fluorescence measurements depicted that there is an interaction between POH and magnetite nanoparticles which was deduced from the presence of two new peaks at 380 nm and 405 nm in emission spectrum. Besides, it was observed that POH molecules could not diffuse into the gel when it consists of magnetite nanoparticles which also make the gel more homogeneous. As swelling ratio increases from 1 to 13, magnetization of the gel does not change which reveals that magnetic nanoparticles do not diffuse out of the gel during water-intake. This feature makes the gel suitable for applications as waste water treatments. (C) 2013 Elsevier B.V. All rights reserved.