Optimum stability conditions of pH and temperature for ligninase and manganese-dependent peroxidase from Phanerochaete chrysosporium. Application to in vitro decolorization of Poly R-478 by MnP S. Rodrı´guez Couto 1,2, *, D. Moldes 1 and Mª A. Sanroma´n 1 1 Department of Chemical Engineering, University of Vigo, E-36200, Vigo, Spain 2 Department of Chemical Engineering, Rovira i Virgili University, 43007, Tarragona, Spain *Author for correspondence: Tel.: +34-977-559617, Fax: +34-977-559667, E-mail: susana.rodriguez@urv.net Received 16 August 2005; accepted 4 October 2005 Keywords: Decolorization, LiP, MnP, pH, P. chrysosporium, Poly R-478, stability, temperature Summary In the present work, the two main factors affecting enzymatic stability, i.e. pH and temperature, were analysed in order to determine the optimum ones to maintain ligninase (LiP) and manganese-dependent peroxidase (MnP) activities for prolonged periods of time. The optimum pH and temperature range obtained was around 4.2 and 34 °C for the former and 4.5 and 32 °C for the latter. Under these conditions LiP and MnP showed a half-life time of about 100 and 500 h, respectively. In addition, extracellular liquid containing mainly MnP (200 U/l) was able to decolorize about 20% of the polymeric dye Poly R-478 in 15 min. The decolorization was carried out at a pH of 4.5 (6 mM sodium malonate) and a temperature of 30 °C. Abbreviations: LiP – lignin peroxidase; MnP – manganese-dependent peroxidase; Poly R-478 – Polyvinylamine sulfonate anthrapyridone; X i – adimensional value of the independent variable Introduction Lignin constitutes one of the most important carbon sources on the earth. Therefore, lignin degradation, as well as its potential utilization, is very important in terms of global carbon cycle. Being a recalcitrant poly- mer, lignin can be selectively degraded in nature only by white-rot fungi (Eriksson et al. 1990). Among them, the basidiomycete Phanerochaete chrysosporium has become the standard microorganism for lignin degradation research due to its powerful ligninolytic system, fast growth and easy handling in culture (Kirk & Farrell 1987; Eriksson et al. 1990). This fungus when cultured on ligninolytic conditions produces mainly two types of extracellular peroxidases, namely lignin peroxidase, LiP (EC 1.11.1.14) and manganese-dependent peroxidase, MnP (EC 1.11.1.13). These extracellular enzymes in addition to lignin are also capable of oxidizing various recalcitrant xenobiotics released to the environment by human activity (Haemmerli et al. 1986; Hammel & Tardone 1988; Bumpus 1989). Among them, there are several synthetic dyes (Swamy & Ramsay 1999). Syn- thetic dyes have been used increasingly in the textile and dyeing industries because of their ease and cost effec- tiveness in synthesis, fastness, high stability to light, temperature, detergent and microbial attack and variety in color compared with natural dyes. However, synthetic dyes are not easily degraded by the conventional wastewater treatments such as activated sludge, trickling filters, etc. (Pagga & Brown 1986; Shaul et al. 1991), which has impelled the search for alternative technolo- gies such as enzymatic treatments. The application of LiP and MnP enzymes to several bio-processes such as wood pulping and wastewater treatment has been proposed (Farrell 1987; Kirk 1987; Tien 1987). Thus, in vitro stability of such enzymes is an important factor in determining both the economic and technical feasibility of application for industrial uses and is also a critical factor in optimizing commercial pro- duction of enzymes. The aim of the present work was to determine the optimum conditions of pH and temperature to main- tain LiP and MnP activities, secreted by P. chrysospo- rium under solid-state conditions, for prolonged periods of time. These data will be of great importance for the application of such enzymes to several biotechnological processes. In addition, to our knowledge, the effect of the above-mentioned variables on LiP and MnP acti- vity loss has formerly not been investigated. Moreover, decolorization of the polymeric model dye Poly R-478 by the main enzyme produced (MnP) was also assessed. World Journal of Microbiology & Biotechnology (2006) 22: 607–612 Ó Springer 2005 DOI 10.1007/s11274-005-9078-0