A new separation scheme has been developed to concentrate nickel and vanadium complexes in selected fractions before attempting a molecular characterization of these organometallic species in heavy crude oils. It involves a combination of solubility- (n-heptane deasphalting and pyridine/water extraction of asphaltenes) and polarity- (SO3-modified silica gel column) based steps. The proposed method has been applied to Belayim crude oil. The nature of pyridine/water extract from asphaltenes has been investigated using both preparative gel permeation chromatography (GPC) and reverse phase liquid chromatography (RP-HPLC) combined (off-line) to graphite furnace atomic absorption (GFAA). The fractions obtained after the separation of the extract on a SO3-modified silica gel column have been submitted to a series of analytical techniques such as: GFAA, mass spectrometry using fast atom bombardment ionization technique (MS-FAB), UV-VIS spectroscopy and electron paramagnetic resonance (EPR). All the analytical evidences, risen from the present study, indicate that vanadium, present in pyridine/water extract obtained from asphaltenes, is mostly bound in porphyrin structures, although the coordination sphere around vanadyl species, present in the most polar fraction obtained from the extract separation by column chromatography, shows different EPR parameter. On the other hand, nickel complexes are smaller and more polar than common Ni-porphyrins, which are concentrated in deasphalted oil.