New Initiative Preliminary evidence of the role of hydrogen peroxide in the degradation of benzo[a]pyrene by a non-white rot fungus Fusarium solani Etienne Veignie a , Catherine Rafin a, *, Patrice Woisel b , Fabrice Cazier c a Universite ´ du Littoral—Cote d’Opale (ULCO), 50 rue Ferdinand Buisson, BP 699, F-62228 Calais, France b ULCO, Laboratoire de Synthe `se Organique et Environnement, MREID, 145 Avenue Maurice Schumann, F-59140 Dunkerque, France c ULCO, CCM, MREID, 145 Avenue Maurice Schumann, F-59140 Dunkerque, France Received 26 July 2003; accepted 14 November 2003 ‘‘Capsule’’: An alternative metabolic pathway was demonstrated. Abstract In order to study the enzymatic mechanisms involved in the successive steps of BaP degradation by a Deuteromycete fungus Fusarium solani, we developed an indirect approach by using inhibitors of enzymes. We used either specific inhibitors of peroxidases (i.e. salicylhydroxamic acid) and of cytochrome P-450 (i.e. piperonyl butoxyde) or inhibitors of both enzymes (i.e. potassium cyanide). Surprisingly, no expected decrease of BaP degradation was observed with most inhibitors tested. On the contrary, more BaP was degraded. Only butylated hydroxytoluene, which acts as a free radical scavenger, inhibited BaP degradation. The inhibition of these enzymes, which use H 2 O 2 as a cosubstrat, might have resulted in an increase of hydrogen peroxide availability in the fungal cultures. This enhancement could induce formation of reactive oxygen species (ROS) which might be the agents that initiate benzo[a]pyrene oxidation. This study proposed a hypothetic alternative metabolic pathway involved in PAH metabolism by Fusarium solani. # 2004 Elsevier Ltd. All rights reserved. Keywords: Benzo[a]pyrene; Fusarium solani; Degradation; Hydrogen peroxide 1. Introduction Polycyclic aromatic hydrocarbons (PAHs) represent an important class of environmental pollutants, of which some are known to be mutagenic and carcino- genic. Due to their hydrophobicity, most PAHs espe- cially high-molecular-weight such as benzo[a]pyrene (BaP) persist in soils and sediments and therefore are less available for biological uptake. Among the pro- cesses whereby these compounds are removed from the environment, microbial degradation plays a major role in the remediation of contaminated sites (Cerniglia, 1992; Alexander, 1999). In previous studies, we reported that telluric fungal strains isolated from fuel-con- taminated compost showed interesting potential for soil bioremediation. In particular, we isolated a Deutero- mycete fungus Fusarium solani able to grow in liquid medium with BaP as sole carbon source and to minera- lize this PAH (Rafin et al., 2000). In a batch fermentor, [7,10- 14 C]benzo[a]pyrene mineralization occurred rapidly at early stages of fermentation (15 h) during the germination of fungal spores (Veignie et al., 2002). Moreover, F. solani showed an average rate of minera- lization of about 65 mg/g dry weight/day within 11 days of incubation, comparable to mineralization rate obtained with the white rot fungus Phanerochaete chry- sosporium (Sanglard et al., 1986). Our objective in the research described herein was to elucidate the enzymatic mechanisms involved in the successive steps of BaP degradation by Fusarium solani. Therefore we developed an indirect approach by using inhibitors of degradative enzymes. As is well known, two main enzyme groups are involved in the initial 0269-7491/$ - see front matter # 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.envpol.2003.11.007 Environmental Pollution 129 (2004) 1–4 www.elsevier.com/locate/envpol * Corresponding author. Tel.: +33-(0)3-21-46-36-00; fax: +33- (0)3-21-46-36-69. E-mail address: rafin@univ-littoral.fr (C. Rafin).