Biodegradation Of Phenanthrene By Indigenous Bacterial Isolates From Contaminated Soil
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Date
2015-03
Authors
Ariffin, Fazilah
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Abstract
The present study focused on phenanthrene because it is a toxic pollutant and a major component of the total content of Polycyclic Aromatic Hydrocarbon (PAH) compounds in the environment. Therefore, this study was conducted to evaluate the potential of bacteria isolated from contaminated soils to biodegrade phenanthrene and to study the biodegradation potential of selected bacteria in shake flask and tray systems. The metabolites produced during degradation processes were determined and the enzyme responsible to degrade phenanthrene was characterized. Fourteen bacteria were succesfully isolated using a spread plate method from seven different sites: Cargill Palm Oil Industry, Neram Palm Oil Industry, Kuantan Port, Tanjung Api Port and Taman Gelora Beach in Pahang, Tok Bali Port in Kelantan and Petronas Penapisan Terengganu Sdn. Bhd in Terengganu. These 14 bacterial isolates were screened by inoculation on an enrichment medium containing phenanthrene as the sole source of carbon and energy in order to detect the potential phenanthrene-degrading bacteria. Out of 14 bacterial isolates, three bacterial isolates (Isolate P3d, Isolate P4a and Isolate P6) showed the best degradation activity. Based on 16S rRNA analysis, these Isolate P3d, Isolate P4a and Isolate P6 were identified as Acinetobacter sp. P3d, Bacillus sp. P4a and Pseudomonas sp. P6, respectively. The biodegradation study in shake flask system showed that Acinetobacter sp. P3d, Bacillus sp. P4a and Pseudomonas sp. P6 monocultures were capable to degrade 93.58%, 83.00% and 78.72% of phenanthrene, respectively. Meanwhile, for the
mixed culture system, Consortium A (Acinetobacter sp. P3d + Bacillus sp. P4a + Pseudomonas sp. P6) showed 95.64% degradation activity, followed by Consortium B (Acinetobacter sp. P3d + Bacillus sp. P4a; 87.19%), Consortium D (Bacillus sp. P4a + Pseudomonas sp. P6; 79.37%) and Consortium C (Acinetobacter sp. P3d + Pseudomonas sp. P6; 79.21%) for 20 days of cultivation. Seven phenanthrene metabolites were identified; 9,10-phenanthrene dihydrodiol, 1-naphthalenecarboxylic acid, 1-hydroxy-2-napthoic acid, coumarin-3-carboxylic acid, 1,4-dihyroxy-2-napthoic acid, salicylic acid and catechol from all three bacterial cultures.The biodegradation study in tray system showed that monocultures of Acinetobacter sp. P3d, Bacillus sp. P4a and Pseudomonas sp. P6 were capable to degrade 78.08%, 68.14% and 79.21% of phenanthrene, respectively. Meanwhile, for the mixed culture system, Consortium A showed 100% activity, followed by Consortium B (87.45%), Consortium D (76.58%) and Consortium C (76.13%) after 30 days of cultivation. Enzyme catechol 1, 2 dioxygenase was detected during cultivations of Acinetobacter sp. P3d, Bacillus sp. P4a and Pseudomonas sp. P6 in shake flask system. Catechol 1,2 dioxygenase was purified to 7.5-fold with 3.18 % yield for Acinetobacter sp. P3d, 5- fold with 1.63% yield for Bacillus sp. P4a and 1.88-fold with 0.72% yield for Pseudomonas sp. P6 with the molecular weights of 32.4, 41 and 32kDa, respectively. The highest activity for bacterial catechol 1,2-dioxygenase for all three identified bacterial cultures was at 30oC and pH 7.5. The enzyme activity was stable in the presence of Ca2+, Mg2+ and Co2+ and completely inhibited by Cu2+ and Fe3+ metal ion.
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Keywords
Phenanthrene , Indigenous Bacterial