Influence of acidification of CCB (Cu/Cr/B) impregnated wood on fungal copper tolerance Miha Humar * , Marjeta S ˇ entjurc, Samuel A. Amartey, Franc Pohleven Biotechnical faculty, Institute Jozef Stefan, Buckinghamshire Chilterns University College, UK Received 4 March 2004; received in revised form 27 August 2004; accepted 20 September 2004 Abstract Copper tolerant fungi are known for more than 60 years but the complete mechanisms of copper tolerance by these fungi are still not fully understood. Copper tolerance has previously been linked to oxalic acid excretion by copper tol- erant brown rot fungi. The oxalic acid then reacts with copper in the wood to form an insoluble and therefore less toxic copper oxalate. It has been suggested that copper tolerance could be due to lowering of the pH of the medium rather than the low solubility of copper oxalate. In order to elucidate this presumption, copper/chromium/boron (CCB) trea- ted wood specimens were acidified with organic (oxalic, acetic, lactic, formic) and inorganic (sulphuric) acids and exposed to copper tolerant (Antrodia vaillantii, Leucogyrophana pinastri) and copper sensitive (Poria monticola, Gloe- ophyllum trabeum) brown rot fungal strains according to the mini block procedure. After eight weeks of exposure, the wood specimens were isolated and their mass losses determined. Additionally, electron paramagnetic resonance (EPR) measurements on the exposed specimens were performed. The EPR spectra of the specimens decayed by A. vaill- antii were very similar to the EPR spectra of the specimens acidified with oxalic acid. Additionally, acidification of the CCB impregnated specimens made them significantly more susceptible to decay by both the copper tolerant and copper sensitive brown rot fungi. Ó 2004 Elsevier Ltd. All rights reserved. Keywords: Copper toxicity; Copper tolerance; Wood; Brown rot fungi; Oxalic acid; Wood preservatives 1. Introduction Copper based solutions have been widely and suc- cessfully used for wood preservation for more than a century (Richardson, 1997; Humar et al., 2003). One of the most important issues regarding copper treated wood is the occurrence of copper tolerant fungi (Humar et al., 2002a; Clausen and Green, 2003). In addition to the scientific interest in copper tolerant fungi, there is also a commercial interest in the development of new wood preservatives that would protect wood against these organisms. A further motive for understanding copper tolerance is the possibility of using copper toler- ant fungal stains for bioremediation of copper contain- ing waste wood (Stephan et al., 1996; Humar et al., 2004). Despite these interests, the mechanism of copper tolerance by these fungi is still not completely understood. 0045-6535/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.chemosphere.2004.09.031 * Corresponding author. Address: Biotechnical faculty, Department of Wood Science and Technology, Rozna dolina, C.VIII/34, SI-1000 Ljubljana, Slovenia. Tel.: +38 614 231161; fax: +38 642 35035. E-mail address: miha.humar@bf.uni-lj.si (M. Humar). Chemosphere 58 (2005) 743–749 www.elsevier.com/locate/chemosphere