An improved method to evaluate the degradation of polycyclic aromatic hydrocarbons (PAHs) for filamentous fungi is hereby proposed. The PAH adsorption through fungal mycelia implies the use of an exhaustive extraction, which is usually performed through a combination of different techniques applied separately to the liquid medium and to the mycelia. This work proposes a simplified extraction procedure with reduced analysis time and sample manipulation, thereby guaranteeing its precision and accuracy. The selected PAHs, pyrene and benzo[a]pyrene, were quantified by gas chromatography using a mass selective detector operating in single ion monitoring mode. The method was validated presenting accuracy within the scope of this study and it was used for evaluating the potential of PAHs degradation by thirteen fungi derived from marine environment. This study selected the fungus Aspergillus sclerotiorum CBMAI 849 for being able to degrade 85% of pyrene and 61% of benzo[a]pyrene after 4 and 8 days, respectively, and the fungus Mucor racemosus CBMAI 847 for being able to degrade 44% of benzo[a]pyrene after 8 days.