Annals of Microbiology, 58 (3) 427-432 (2008) Ligninolytic activity from newly isolated basidiomycete strains and effect of these enzymes on the azo dye orange II decolourisation Maira Corteline ABRAHÃO 1 , Adriana de Mello GUGLIOTTA 2 , Roberto Da SILVA 1 , Roberto Joanne Yashihiro FUJIEDA 1 , Maurício BOSCOLO 3 ; Eleni GOMES 1 * 1 Laboratório de Bioquímica e Microbiologia Aplicada, IBILCE, UNESP, São Paulo State University, Rua Cristóvão Colombo, 2265, Jd Nazareth, São José do Rio Preto, SP, Brazil, CEP: 15054-000; 2 Section of Mycology and Lichenology, Botanical Institute, São Paulo, Brazil; 3 Laboratório de Fisico Química e Sucro Derivados, IBILCE, UNESP, São Paulo State University, Rua Cristóvão Colombo, 2265, Jd Nazareth, São José do Rio Preto, SP, Brazil, CEP: 15054-000 Received 18 February 2008 / Accepted 9 May 2008 Abstract - Wood-rotting fungi have the ability to degrade lignin by secreting ligninases, a promising enzyme for degradation of envi- ronmental pollutants. Nine basidiomycete strains collected just outside the city of São José do Rio Preto, upstate São Paulo, Brazil, were evaluated for their manganese peroxidase (MnP), lignin peroxidase (LiP) and laccase production by solid-state fermentation on wheat bran. Datronia caperata SP381992, Polyporus tenuiculus SP381977 and Pycnoporus sanguineus SP381968 were the highest producers of laccase, while Polyporus tenuiculus SP381971, Datronia caperata SP381992, Coriolopsis polyzona SP381989 and Hexagonia hirta SP382026 produced the most MnP and LiP activity. The majority of strains secreted laccase with optimum activity at 70 °C and, when maintained at 60 °C, in the absence of substrate, the crude enzymes preserved 100% of their initial activity for periods of 30 min up to 8 h. Enzymes from D. caperata SP381992, P. tenuiculus SP381977, P. sanguineus SP381968 and H. hirta SP382026 were tested for activity on the azo dye orange II and afforded 96-100% decolourisation of the dye in 1 to 48 h. Since this reaction depended on the presence of ABTS and there was no decolourisation when H 2 O 2 or MnSO 4 was present, it was attributed to the laccase activity. Key words: ligninases, basidiomycetes, azo dye, orange II. INTRODUCTION Wood-decomposing fungi causing white-rot degrade lignin because they secrete oxidoreductases including lignin peroxidase (1,2-bis (3,4-dimethoxylphenyl) propene-1,3- diol:hydrogen-peroxide-Lip EC 1.1.1.14) (Paszczynski et al., 1986), manganese peroxidase (Mn(II):hydrogen-peroxide oxidoreductase EC 1.11.1.13) (Glenn et al., 1986) and laccase (benzenediol:oxygen reductase EC 1.10.3.2) (Mayer and Harel, 1979). These enzymes oxidise, in a nonspecific way, both phenolic and non-phenolic lignin derivatives and thus are promising candidates for the degradation of environmental pollutants e.g. phenols, anilines, dyes (Fahr et al., 1999; Ferreira-Leitão et al., 2007) and highly recalcitrant compounds such as polychlorinated biphenyls (PCBs) and polycyclic aromatic hydrocarbons (PAHs) (Svobodová et al., 2008). Biodegradation of several persistent compounds such as dyes, pesticides and lignin derivatives has been attributed to the oxidative enzymes, especially laccase (Robinson et al., 2001; Riccardi et al., 2005; Revankar and Lele, 2007). Azo dyes are used for textile dyeing and paper printing and represent toxic content in the effluents because their partial degradation results in aromatic amines that are toxic to aquatic life and mutagenic and carcinogenic to humans (Pinheiro et al., 2004). Biodegradation of dyes has been the focus of several studies in which a wide range of physical, chemical and biological methods have been proposed to remove dyes from wastewater (Forgacs et al., 2004). Among these, microbiological and enzymatic decomposition have received much attention in recent years (Chagas and Durrant, 2001; Iranzo et al., 2001; Baggi et al., 2002; Zheng and Obbard, 2002; Murugesan et al., 2007; Susla et al., 2007). However, the use of enzymes in such biotechnological processes will require a reduction in production costs to become viable. Solid-state fermentation (SSF) reproduces natural microbiological living conditions and has proved to be suitable for enzyme production by filamentous fungi. In addition, it permits the use of agricultural by-products, which could reduce costs. In this study, wheat bran was used as substrate to assess the ligninase production by strains of basidiomycete fungi in SSF. The thermostability of the laccase produced and ligninolytic enzyme secreted was studied by decolourisation of an azo dye. MATERIAL AND METHODS Isolation of microorganisms. Wood-rotting basidiomycete polypores were collected on the outskirts of São José do Rio Preto, State of São Paulo, Brazil. The strains were isolated and purified as described previously (Xavier-Santos et al., 2003). * Corresponding author. Phone: +55 017 32212393, Fax: +55 017 32212390. E-mail: eleni@ibilce.unesp.br