Biodegradation of Pentachlorophenol in Soil Slurry Cultures by Bjerkandera adusta and Anthracophyllum discolor O. Rubilar, ‡ G. Feijoo, ² C. Diez, ‡ T. A. Lu-Chau, ² M. T. Moreira,* ,² and J. M. Lema ² Department of Chemical Engineering, School of Engineering, UniVersity of Santiago de Compostela, E-16782 Santiago de Compostela, Spain, and Doctorate Programme Natural Resources, UniVersity of La Frontera, Temuco, Chile Pentachlorophenol (PCP) belongs to a class of environmental pollutants which have a high toxicity and are resistance to degradation. White-rot fungi are potential candidates for treating PCP-contaminated soils because of their high capability of degrading a wide range of xenobiotics. A series of laboratory-based studies was developed to determine the range of PCP concentration in soil (100, 250, and 350 mg of PCP/kg of soil) which could be degraded by two white-rot fungi: Bjerkandera adusta and Anthracophyllum discolor. Both fungi were grown on Kirk medium and on distillers dried grains with solubles (DDGS). The highest PCP degradation was attained by A. discolor using DDGS (95% after 28 days). The use of DDGS increased both PCP degradation and the production of ligninolytic enzymes, although no correlation was found between them. The process was scaled up to a 5 L reactor, in which 81% PCP was degraded after 28 days. The absence of ligninolytic activity of this culture indicates that other enzymatic systems may be participating in the degradation of PCP. 1. Introduction Pentachlorophenol (PCP) is a pollutant that causes significant environmental impacts not only on the ecosystem but also on human health. 1 The most commonly used methods for biore- mediation of PCP-polluted soils are incineration, disposal in landfills, soil washing, and chemical extraction. 2 Alternatively, the development of biological approaches to remediation has generated great interest. White-rot fungi are microorganisms with a well-known capacity for degrading a wide range of organic compounds, including PCP. This degradation capacity has been attributed to their ability to secrete enzymes involved in the degradation of lignin, including lignin peroxidase (LiP), manganese peroxidase (MnP), and laccase. Phanerochaete chrysosporium 3,4 and Trametes Versicolor 5,6 have been the most widely used fungi for PCP degradation. Nevertheless, diverse studies have been performed to evaluate new fungal strains with a high ability for the degradation of recalcitrant organic compounds. 7-9 PCP degradation by white-rot fungi has been widely studied in liquid media 6,10,11 and soil. 12-14 However, the process had not been studied in detail under soil slurry conditions. The use of these fungi in the bioremediation of contaminated soils requires the determination of more adequate conditions for enhancing their growth and activation of the ligninolytic system at conditions different from those found in their natural environment. One of these conditions is related to the nitrogen level of the culture medium. For instance, Leatham and Kirk 15 observed that a low nitrogen level, similar to levels found in wood, stimulates the growth of white-rot fungi. A strategy which favors fungal growth and confers a selective pressure against soil native microflora would be the addition of a lignocellulosic material, such as straw, wood chips, or wood shavings, to the culture medium. 16,17 Soil bioremediation is effective only on the bioavailable fraction while the nonavailable fraction remains unaltered due to the strong adsorption and/or polymerization of the pollutant to the organic and inorganic fractions of soil. The agitation and mixing of the culture medium with soil in suspension facilitates the particle separation and the exposure of the contaminant to the fungal action. This can be achieved by applying a soil slurry phase system, which is a recent technological development of great interest to environmental scientists and specifically for the bioremediation of contaminated soils. The basis for the slurry culture is described here. The soil is mixed with water and culture medium to a concentration between 5 and 40% determined by the proportions of the contaminants in soils, the biodegradation rates, and the physical nature of the soils. In agitated cultures the slurry is incubated with the inoculum to allow microbial growth. The overall biodegradation rate is controlled by microbial activity. One drawback to this technique is that it requires strong agitation to keep the soil in suspension. Most of the soil slurry cultures conducted for the degradation of different contaminants have been conducted by bacterial cultures 18-21 which are more resistant to mechanical stress. The successful application of soil slurry cultures in a stirred tank reactor have been recently conducted using the white-rot fungus Bjerkandera adusta for the degradation of four polycyclic aromatic hydrocarbons from marsh soil. 22 In this work the degradation of PCP from spiked soil by two white-rot fungi, B. adusta and Anthracophyllum discolor (a strain obtained from Chilean forests), was studied in slurry phase cultures. Three different effects on PCP degradation and activation of the ligninolytic system were assayed with both fungi: (a) effect of initial PCP concentration in soil, (b) effect of substrate (synthetic Kirk medium or solid lignocellulosic material (distillers dried grain with solubles)), and (c) influence of native microflora. Moreover, the degradation process is scheduled to be scaled up from 125 mL flasks to a 5 L stirred tank reactor. 2. Materials and Methods 2.1. Microorganisms. The fungal strains used were Bjer- kandera adusta (ATTC 90940), which was obtained from the * To whom correspondence should be addressed. Tel.: +34981563100, ext. 16776. Fax.: +34981528050. E-mail tmoreira@usc.es. ² University of Santiago de Compostela. ‡ University of La Frontera. 6744 Ind. Eng. Chem. Res. 2007, 46, 6744-6751 10.1021/ie061678b CCC: $37.00 © 2007 American Chemical Society Published on Web 04/03/2007