J. Basic Microbiol. 46 (2006) 2, 126 – 134 DOI: 10.1002/jobm.200510014 © 2006 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim 0233-111X/06/0204-0126 (Departamento de Bioquímica, Universidade Estadual de Maringá, 87.020-900, Maringá, PR, Brazil) Co-production of ligninolytic enzymes by Pleurotus pulmonarius on wheat bran solid state cultures DANIELA FARANI DE SOUZA, GIOVANA KIRST TYCHANOWICZ, CRISTINA GIATTI MARQUES DE SOUZA and ROSANE MARINA PERALTA* (Received 14 April 2005/Returned for modification 10 May 2005/Accepted 15 July 2005) In this paper, the production of biomass and ligninolytic enzymes by the white-rot fungus Pleurotus pulmonarius (FR.) QUÉLET cultured on wheat bran at high initial moisture was evaluated. When the initial moisture was lower than 86%, the fungal hyphae penetrated into and bound tightly to the solid- substrate particles. When the moisture was equal or higher than 86%, the growth was characterized by formation of a large mycelial mass above the substrate (surface growth). In this case, the mycelial mass could be easily separated from the residual solid substrate and quantified by gravimetric analysis. The fungus produced 0.32 g of dry mycelial mass per g of dry substrate after 15 days of cultivation. Analysis of the residual substrate showed that growth was mainly due to the consumption of soluble proteins and carbohydrates. The condition of high initial moisture strongly promoted the expression of laccase (up to 24,000 U per g of substrate or 78,000 U per g of dry fungal biomass), while the production of manganese peroxidase was negatively affected. In fact, manganese peroxidase was maximally produced when the initial moisture was 75% (2,000 U per g of substrate). The production of other enzymes, such as polysaccharidases and proteases, was not significantly affected by the initial moisture. White-rot fungi, belonging mostly to the basidiomycetes, are characterised by their ability to quickly and efficiently degrade the lignin moiety of woody tissues. They produce several oxidative enzymes including lignin peroxidase (LiP), manganese peroxidase (MnP), laccase and oxidases that generate the H 2 O 2 needed for peroxidase activity. Some fungi lack one or more of these enzymes, suggesting that there is more than one mechanism for fungal degra- dation of lignin (HATAKKA 1994). Extensive studies of the white-rot fungus Phanerochaete chrysosporium have revealed that its ligninolytic activity is closely correlated with secretion of LiP and MnP (TIEN and KIRK 1983). However, the white-rot fungi Dichomitus squalens (LANG et al. 1998) and Ceriporiopsis subvermispora (URZÚA et al. 1995) can efficiently degrade lignin without expressing detectable LiP. The combination of laccase and MnP is much more common in white-rot fungi than the combination of LiP and MnP found in P. chrysosporium (LEONOWICZ et al. 2001). Furthermore, the white-rot fungus Pycnoporus cinnabarinus appears to have only one of these enzymes (laccase), yet it degrades lignin effectively (EGGERT et al. 1996). Pleurotus spp. are among the easiest mushrooms to cultivate (COHEN et al. 2002). The two most important species cultivated on an industrial scale are P. ostreatus and P. pulmon- arius (P. sajor-caju, P. sapidus). In nature they grow on wood, usually on dead, standing trees or on fallen logs. Various substrates that contain lignin and cellulose can be used for Pleurotus cultivation, such as wood chips, corn wheat, rice straw, cotton stalks, waste hulls, and other agricultural wastes, some of which can be recycled and upgraded for use as ani- mal feed or for preparation of other products (COHEN et al. 2002). These substrates are * Corresponding author: Dr. R. M. PERALTA; e-mail: rmperalta@uem.br