Environ Chem Lett (2005) 3: 74–77 DOI 10.1007/s10311-005-0006-1 ORIGINAL PAPER C. Held · A. Kandelbauer · M. Schroeder · A. Cavaco-Paulo · G. M. Guebitz Biotransformation of phenolics with laccase containing bacterial spores Received: 5 May 2005 / Accepted: 9 August 2005 / Published online: 22 October 2005 C  Springer-Verlag 2005 Abstract Treatment of effluents containing phenols such as textile dyes with fungal laccases is usually limited to the acid to neutral pH range and moderate temperatures. Here we demonstrate for the first time that spore-bound laccases which are stable at high temperatures and pH values can be used for phenolic dye decolorisation. Laccase contain- ing spores from Bacillus SF were immobilized on alumina pellets. Both immobilized and free spores were able to completely decolorize the common textile dyes Mordant Black 9, Mordant Brown 96/Mordant Brown 15, and Acid Blue 74 within 90 min of incubation time and decolorized solutions were successfully used in re-dyeing. Keywords Bacillus . Spores . Biodegradation of phenolics . Effluent treatment Introduction Laccases (benzenediol:oxygen oxidoreductase, EC 1.10.3.2) have found several applications in enzyme re- mediation. Laccases as well as immobilized laccases have been used for the treatment of phenolic effluents, PAHs and PCBs. The enzymes render phenolic compounds less toxic via degradation or polymerization reactions and/or cross- coupling of pollutant phenols with naturally occurring phenols (Bohmer et al. 1998; D’Annibale et al. 2000; Ullah et al. 2000). Besides microbial processes (Fischer-Colbrie et al. 2005), laccases have also a potential for the decol- orization of textile dyes (Kandelbauer and Guebitz 2005). C. Held · A. Kandelbauer · M. Schroeder · G. M. Guebitz () Department of Environmental Biotechnology, Graz University of Technology, Petersgasse 12, 8010 Graz, Austria e-mail: guebitz@tugraz.at Tel.: +43-316-873-8312 Fax: +43-316-873-8815 A. Cavaco-Paulo Department of Textile Engineering, University of Minho, 4800 Guimaraes, Brazil Laccases catalyze the removal of a hydrogen atom from the hydroxyl group of ortho and para-substituted mono- phenolic and poly-phenolic substrates and from aromatic amines by one-electron abstraction, to form free radicals, capable of undergoing further depolymerization, repoly- merization, demethylation or quinone formation. For dye decolorization, the laccases from a number of fungi such as Sclerotium rolfsii, Trametes modesta, T. hir- suta, T. versicolor, Pleurotus pulmonarius among others have been investigated (Kandelbauer and Guebitz 2005). However, the potential of bacterial laccases for this pur- pose has not been assessed so far. In contrast to fungal laccases which are known for many years, laccase activity in bacteria has only been described in 1993 for Azospir- illum lipoferum and has been characterized later in more detail (Diamantidis et al. 2000). Interestingly, the spore coat protein CotA of Bacillus subtilis was identified as laccase (Hullo et al. 2001). Since spores serve microorganisms to survive drastic conditions, spore coat enzymes might also withstand high temperatures or extreme pH values which would be advantageous for industrial applications. It has previously been shown that residual enzyme after enzymatic decolorization of dyeing effluents can restrict the reuse of water (Abadulla et al. 2000). Enzyme protein can adsorb to fabrics and thus lead to unwanted color differ- ences in dyeing with enzymatically decolorized effluents. Therefore, for continuous applications in dye decoloriza- tion and also in other applications immobilization of lac- cases is required and several successful examples have been reported (Abadulla et al. 2000; Kandelbauer et al. 2004). In a previous study higher decolorization rates were re- ported with an immobilized laccase from the white-rot fun- gus Trametes modesta when compared to the free enzyme. Despite the promising efficiency of dye decolorization in general, no decolorization activity at all was observed at pH values higher than pH 7 (Kandelbauer et al. 2004). This limits the industrial potential since many dyeing processes (e.g., cotton) are performed in the alkaline pH range. Alter- natively, spore laccases which are also active in the alka- line pH range could be used for dye decolorization without liberation from spores allowing simple immobilization of