DEGRADATION OF MIXED AROMATIC POLLUTANTS BY TRAMETES VERSICOLOR STRAIN 1 Husein Yemendzhiev1, Anna Terziyska1, Jordan Manasiev, Zlatka Alexieva1 1 Bulgarian Academy of Sciences, The Stephan Angeloff Institute of Microbiology, Sofia, Bulgaria Correspondence to: Hussein Yemendzhiev E-mail: [email protected]
One of the most distributed groups of industrial pollutants consists of aromatic hydrocarbons. Because of the complex nature of such wastes the obtaining of new strains able to degrade different toxic aromatic compounds in mixtures is a challenge to many scientists exploring the area of environmental microbiology. In this study the strain of white rot fungus Trametes versicolor was cultivated in a medium comprising a mixture of phenol (0.2 g/l), resorcinol (0.1 g/l), p-cresol (0.1 g/l), and o-nitrophenol (0.14 g/l). Phenol, resorcinol and p-cresol were degraded in 24 hours and then a slow o-nitrophenol degradation was observed. There was about 50% decrease of o-nitrophenol concentration in 144 hours. The degradation of the tested concentrations of phenol, resorcinol and p-cresol in a mixture showed similar characteristics as in the experiments with each of them used as a single carbon substrate. The data showed that the presence of o-nitrophenol does not influence the biodegradation rate of other compounds included in the mixture. The capability of the investigated strain demonstrated its potential for future application in bioremediation technologies oriented to cleaning and preservation of industrially polluted water. Biotechnol. & Biotechnol. Eq. 2011, 25(4), Suppl., 39-40 Keywords: Trametes biodegradation
Introduction Wastewaters from paper, chemical and petroleum industries, coal mining, and a variety of other industrial productions contain many aromatics such as phenol, cresols, nitrophenols, etc (1, 5). The methylated phenol derivatives are one of the most toxic compounds for the environment. The Environmental Protection Agency of the USA has classified phenol derivates as a pollutant of group C (possible human carcinogens) (7). The potential of white rot fungi to remove a variety of toxic organo-pollutants from wastewaters has recently attracted the attention of scientists. This group of microorganisms is capable of degrading polymers of phenolic origin, including lignin (8). The biotransformation activity toward some wide-spread phenolic pollutants has been studied together with expression of specific enzymes (2). The ability of Trametes versicolor 1 to utilize phenol as a sole carbon source was described for the first time in our recent publication (9). The simultaneous presence of different organic mixtures in most of the industrial wastes makes the investigations on the microbial destruction of composite substrates of great importance. The removal or degradation of one or all components can be delayed and/or ceased depending on the composition of the studied mixture (3, 4). WHO WE ARE AND WHAT WE ACHIEVED
The aim of this study was to investigate the potential of Trametes versicolor strain 1 toward degradation of a model mixture of phenol, resorcinol, p-cresol and o-nitrophenol.
Materials and Methods Microorganisms and growth conditions The investigated strain of Trametes versicolor was grown on slants on a medium of the following composition: malt extract 3.0 g/dm3, yeast extract 3.0 g/dm3, peptone 5.0 g/dm3, glucose 10 g/dm3 and agar 20 g/dm3. The organism on the slants was allowed to grow for 72 h at 30 °C and then stored at 4 °C. Biodegradation was conducted on the carbon-free CzapekDox medium containing phenolic compounds as sole carbon and energy sources (9) The flasks containing 50 ml of inoculated culture medium were agitated on a New Brunswick rotary shaker (240 rpm) at 28 °C. Samples were taken at a 24 h interval and centrifuged at 5000 rpm for 20 min to settle the cells down. The dry weight of the cells was determined by ULTRA X apparatus for drying (6). Analytical methods The content of phenols in the supernatant was determined by the HPLC analyses performed in C18 10 μm Bondapac Column (3.9 mm × 300 mm) and Waters 484UV detector (260 nm). The mobile phase was methanol – water (70:30), the flow rate was 0.2 ml/min and the temperature, 22 °C. The experiments for determination of biodegradation capacity of the investigated strain were performed in triplicate. 39
Results and Discussion
The capacity of Trametes versicolor 1 to utilize phenol was previously demonstrated (9). It was shown that the strain had mineralized phenol (0.5 g/l).The activities of key enzymes of the ortho-fission phenol catabolic pathway: phenol hydroxylase [EC 188.8.131.52], catechol 1,2-dioxigenase [EC 184.108.40.206] and cis,cis-muconate lactonizing enzyme [EC 220.127.116.11], were established (9). These results gave us a reason to investigate the strain ability to degrade a model mixture of phenolic industrial pollutants. The degradation ability of the investigated strain was examined in a medium not containing carbon components except the phenolic derivatives. The initial concentration of the investigated compounds in the mixture was as follows: phenol – 0.2 g/l; resorcinol – 0.1 g/l; p-cresol – 0.1 g/l, and o-nitrophenol – 0.14 g/l. The results obtained form these experiments are shown in Fig. 1.
Fig. 1. Growth and degradation of strain Trametes versicolor 1 in a synthetic medium complemented with phenolic compounds mixture. Phenol (■); Resorcinol (▲); p-Cresol (♦); o-Nitrophenol (•); Cell growth (○).
The complete assimilation of the investigated mixture was observed in 120-144 h. We established that phenol, p-cresol and resorcinol in the mixture were utilized in 48 hours. The degradation of o-nitrophenol was much slower especially in the first 24-48 hours of the process. These data in combination with the growth observed show that the preferred substrates in the mixture are phenol, resorcinol and p-cresol.
The complete utilization of this complex mixture of phenols is an evidence for the potential of the investigated Trametes versicolor strain for future application in bioremediation technologies oriented to purification and preservation of industrially polluted water.
This work was supported by grant BG051PO001-3.3.04/32 financed by Operational Programme Human Resources Development (2007 – 2013) and co-financed by the European Social Fund of the European Union.
1. Aggelis G., Ehaliotis C., Nerud F., Stoychev I., Lyberatos G., Zervakis G.I. (2002) Appl. Microbiol. Biotechnol., 59, 353-360. 2. Aktas N. and Tanyolac A. (2003) Biores. Technol., 87, 209-214. 3. Okpokwasili G. C. and Nweke C. O. (2006) African J. Biotechnol., 5, 305-317. 4. Saravanan P., Pakshirajan K., Saha P. (2008) J. Environ. Sci., 20, 1508-1513. 5. Sikkema J., De Bont J., Poolman B. (1995) Microbiol. Rev., 59, 201-222. 6. Stoilova I., Krastanov A., Stanchev V., Daniel D., Gerginova M., Alexieva Z. (2006) Enz. Microb. Technol., 39, 1036-1041. 7. U.S. EPA. Current National Recommended Water Quality Criteria, http://www.epa.gov/waterscience/ criteria/ wqcriteria.html (Accessed Aug 23, 2007) 8. Xavier A. M. R. B., Tavares A. P. M., Ferreira R., Amado F. (2007) Electron. J. Biotechnol., 10, 444-451. 9. Yemendzhiev H., Gerginova M., Krastanov A., Stoilova I., Alexieva Z. (2008) J. Ind. Microbiol. Biotechnol., 35, 1309-1312.
Biotechnol. & Biotechnol. Eq. 25/2011/4, Suppl.