Biodegradation of the Recalcitrant Endocrine Disruptor Nonylphenol

University dissertation from Biotechnology, Lund University

Abstract: Currently, over 100 high-volume chemicals have been classified as endocrine disrupters. One of these is nonylphenol, which was used as the model compound in this study. This substance originates mainly from the degradation of non-ionic surfactants (nonylphenol ethoxylates) which are widely used in industrial and domestic applications. Hence, nonylphenol is widespread in the environment. Under anaerobic conditions the degradation of nonylphenol with 10 ethoxylate groups was observed to take place through sequential removal of ethoxylate groups, resulting in the formation of nonylphenol (up to 11 mg/kg dw) and nonylphenol ethoxylate with 1-3 ethoxylate groups that were accumulated on the sludge. Due to its harmful effects and recalcitrance, the investigation of organisms that can metabolize nonylphenol is important. This was performed by acclimatization and enrichment of selected inocula with nonylphenol as the only carbon source. Suitable bacteria were only found in soil contaminated with ethoxylated non-ionic surfactants collected in an industrial area in Gothenburg, Sweden. The bacteria isolated were classified as Stenotrophomonas sp., Pseudomonas mandelii and Pseudomonas veronii, and were able to grow at low temperatures, with an optimum at 10?C for the two Pseudomonas spp. and they were thus classified as cold-adapted microorganisms. These are, to the best of our knowledge, the first reported cold-adapted organisms that can degrade nonylphenol.. The enriched culture was used to inoculate a lab-scale packed-bed bioreactor fed with mineral salt medium saturated with nonylphenol in order to simulate contaminated water/groundwater. Nonylphenol was degraded by 99-100%, at rates of 43 mg/l?day at 10?C and 22 mg/l?day at temperatures of 5.5?C and 15?C. Removal of endocrine-disrupting activity was also observed. The same type of bioreactor was inoculated with a pure strain of Sphingomonas sp. TTPN3, known to biodegrade nonylphenol and classified as a mesophilic organism. The degradation rate achieved at reactor removal yields of 99-100% was 33 mg nonylphenol/l?day at room temperature (22?2?C), which was lower than the rate obtained at 10?C. These results indicate that the low degradation rates usually observed at low temperatures, common in most of the industrialized countries, can be overcome by the use of adapted organisms. This underlines the relevance of studying cold-adapted organisms which are poorly known, according to FISH analysis performed, in favour of mesophilic organisms. Besides bacteria, white-rot fungi, Trametes versicolor and Bjerkandera sp. BOL13 were screened, among other fungi, for their ability to remove nonylphenol from liquid matrices. Removal rates of 6.6 mg nonylphenol/l?day and 6.1 mg nonylphenol/l?day were observed after 25 days of incubation by Trametes versicolor and Bjerkandera sp. BOL13, respectively. These rates could be increased to 9.7 mg nonylphenol/l?day for Bjerkandera sp. BOL13 by raising the concentration of active biomass in the inoculum and reducing mass transfer limitations by agitating the cultures. However, the removal of nonylphenol was not clearly correlated with the production of extracellular enzymes. For Trametes versicolor, nonylphenol removal was closely related with laccase production. This fungus was inhibited at nonylphenol concentrations above 15 mg/l, indicating the toxic effects of nonylphenol on this fungal strain. Nonylphenol removal from soil was also attempted by washing artificially contaminated soil at 40?C. The leachate was then fed into a packed-bed bioreactor where nonylphenol was successfully degraded. However, when real contaminated soil was used, the removal of nonylphenol from the soil decreased, showing the limitation of this method in aged soils where adsorption restricted the extraction process. White-rot fungi are potential soil colonizers with high surface growth rates, and are able to reach pollutants in soil in ways bacteria can not. Trametes versicolor and Bjerkandera sp. BOL13 degraded approximately 430 mg nonylphenol/kg in artificially contaminated soil after 5 weeks of incubation. These results could not be repeated with real contaminated soil due to the high pH of the soil used and the presence of other contaminants which interfered with the growth of the fungi.