S. Soleymani; A. Lakzian; A. Fottovat
Abstract
Introduction: Environmental contamination by crude oil and its various processing products is becoming a common phenomenon which severely damages soil and groundwater resources. Among the constituents of oil waste, polycyclic aromatic hydrocarbons (PAHs) are of environmental concern because of their ...
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Introduction: Environmental contamination by crude oil and its various processing products is becoming a common phenomenon which severely damages soil and groundwater resources. Among the constituents of oil waste, polycyclic aromatic hydrocarbons (PAHs) are of environmental concern because of their toxic, mutagenic and/or carcinogenic effects. Bioremediation involves the use of living microorganisms, bacteria or fungi, for detoxification of soil and water organic pollutants by biodegradation, biotransformation, and/or mineralization. Collaboration between different microbes under co-culture conditions such as co-metabolism or antagonism makes the system to perform better than a single microorganism. Total petroleum degradation is a result of a microbial consortium action, which is composed of different species with specific biochemical roles. On the other hand, the majority of components of petroleum products has low solubility in water and tends to bind to soil particles reducing their availability to microorganisms for degradation. This has been well described as a major limitation to the bioremediation of hydrocarbon contamination. The surfactants can be employed to enhance hydrocarbon biodegradation by mobilization, solubilization, or emulsification. Some microorganisms synthesize a wide range of surface-active compounds, generally called biosurfactants, which increases the bioavailability of these compounds. The application of these microbial surfactants in the remediation of hydrocarbons aims to increase their bioavailability or mobilize and remove the contaminants by pseudo-solubilization and emulsification in a treatment process. This work aimed to investigate the impact of the biosurfactant producing consortium on the benzo(a)pyrene biodegradation.
Materials and Methods: Four gasoline contaminated soils were enriched in Bushnell-Hass mineral medium with Benzo(a)pyrene (200 mg/l) for three months at 30°C. After this time, to obtain Benzo(a)pyrene-degrading isolates, 0.1 ml of soil suspensions were plated on BH agar plates containing pollutant. Three colonies with different morphological distinct properties were purified on LB agar plates. The screening of the most potent surfactant strain was assayed quantitatively using measurement of surface tension by the Du Nouy ring method. For increasing the production of biosurfactant, medium conditions including pH (6, 7, 8), temperature (25, 30, 35) and carbon source (glucose, sucrose and ribose) were optimized with fractional factorial based on Taguchi. The capability of the isolates and consortium in hydrocarbon biodegradation was investigated in liquid medium of Bushnell-Hass with 150 ppm of Benzo(a)pyrene, during 14 days. Treatments included inoculation of isolates AP3 and BM1 and their consortium in presence and absence of extracted isolates biosurfactants and control (no isolate and biosurfactant). Based on the results of Benzo(a)pyrene degradation in the liquid medium, AP3 isolate, consortium and biosurfactant extracted from AP3 were selected for soil experiment. Four sets of biodegradation experiments were carried out with soil contaminated by 150 ppm of benzo(a)pyrene for 45 days, as follows: set 1: soil + AP3 isolate; set 2: soil + consortium; set 3: soil + consortium + AP3 biosurfactant and set 4: blank (soil). The residual concentrations of contaminant were extracted on days 15, 30 and 45 by dichloromethane solvent and analyzed using GC-FID.
Results and Discussion: The results revealed that strains AP3 and BM1 showed a significant potential to produce surface-active agents in the presence of Benzo(a)pyrene as substrate, reducing the surface tension to 43 and 46 mN/m, respectively. Taguchi experimental design method was applied in order to optimize the biosurfactant production by isolates. Results of experiments indicated that the optimum biosurfactant production conditions were found to be temperature of 35º C and pH of 7, and glucose as water soluble carbon source. The produced biosurfactant reduced surface tension to 31/52 mN/m and 30/81 mN/m for BM1 and AP3, respectively. Biodegradation experiments of Benzo(a)pyrene in liquid cultures showed that the overall biodegradation efficiency of the individual isolates after 14 days was lower than consortium. Bacterial consortium enhanced degradation of contaminant to 87.3% (with addition of biosurfactant) compared to 27.6% of removal in presence of BM1 isolate. However, there was no statistically significant change in the degradation rates of contaminant in consortium with addition of AP3 and BM1 surfactant and surfactant free (87.3, 85.6 and 86.8%, respectively). The degradation of Benzo(a)pyrene was significantly enhanced in presence of AP3 biosurfactant at individual BM1 treatments (28.3 and 44.5 to 74.8%). Maximum degradation of Benzo(a)pyrene in contaminated soil was found (100%) in set 3: soil + consortium + AP3 biosurfactant. Based on GC-MS analyses, it degraded around 100% of penzo(a)pyrene, used as the sole carbon and energy source, at an initial concentration of 150 mg L-1, after 45 days of incubation, while alone consortium and isolate were able to remove 86% and 68% of hydrocarbon, respectively. Overall, these results provide evidence that consortium and AP3 biosurfactant could be potential candidates for further bioremediation.
Conclusion: The results revealed that the hydrocarbon removal efficiency of the consortium was higher than single species, and the final removal efficiency for the consortium could be reached in a considerably shorter time. The results suggest that biosurfactant-assisted bioremediation may be a promising practical bioremediation strategy for aged PAH-contaminated soils. It is evident from the results that the consortium alone and its producer species are both capable of promoting biodegradation to a large extent.
Nasrin Ansari; Mehdi Hassanshahian; MohammadReza Khoshro
Abstract
Introduction: Petroleum hydrocarbons are widespread pollutant that enters to soil by some pathwayssuch as: Transportation of crude oil, conservation of oil compounds, crude oil spill and treatment process on refineries. Oil pollution has some ecological effect on soil that disturbed composition and ...
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Introduction: Petroleum hydrocarbons are widespread pollutant that enters to soil by some pathwayssuch as: Transportation of crude oil, conservation of oil compounds, crude oil spill and treatment process on refineries. Oil pollution has some ecological effect on soil that disturbed composition and diversity of microbial community. Also this pollution has some effects on microbial activity and enzymes of soil. Forests ecosystems may be polluted with petroleum hydrocarbons via different ways such as transportation and spill of crude oil from resource of petroleum storage. Industrial soil defined as the soils that located in industrial area such as petrochemical plant, mine, chemical factories and etc. These soils always contaminated to many pollutant such as: oil, diesel and heavy metals. These pollutants have some effects on the texture of the soil and microbial community. The aim of this research is to understand the effect of oil pollution on two different soils.
Material and Methods: In order to evaluate the effect of crude oil on soil microbial community, two different soil samples were collected from industrial and forest soils. Six microcosms were designed in this experiment. Indeed each soil sample examined inthree microcosms asunpolluted microcosm, polluted microcosm, and polluted microcosm with nutrient supply of Nitrogen and PhosphorusSome factors were assayed in each microcosm during 120 days of experiment. The included study factors were: total heterotrophic bacteria, total crude oil degrading bacteria, dehydrogenase enzyme and crude oil biodegradation. For enumeration of heterotrophic bacteria nutrient agar medium was used. In this method serial dilutions were done from each soil and spread on nutrient agar medium then different colonies were counted. For enumeration of degrading bacteria Bushnel-Hass (BH) medium were used. The composition of this medium was (g/lit): 1 gr KH2PO4, 1gr K2HPO4, 0.2 gr MgSO4.7H2O, 0.02 gr CaCl2, 1 gr NH4NO3, and two drops of FeCl3 60% , the pH was 7. The carbon source of this medium was crude oil (1%). In MPN method microplates (24 well) were utilized and turbidity was calculated as positive index.
Results and Discussion: The results of this study showed that the highest quantity of heterotrophic bacteria was related to forest soil (8 × 108). The quantities of degradative bacteria significantly were lower than heterotrophic bacteria in all soil microcosms. This result may be expected because heterotrophic bacteria can use other carbon sources instead of crude oil such as organic carbon, suger and some nutrients that exist in the soil, but degrading bacteria have some limit in the use of organic carbons and only capable to use crude oil hydrocarbons. Sothe quantity of these bacteria is lower than heterotrophic bacteria. The quantity of degradative bacteria have decrement pattern until 60th day of experiment but after this day these bacteria have increment pattern. This result can be interpreted as from beginning of experiment until 60th day of experiment the bacteria adapted to toxic effect of crude oil and after this time the quantity of bacteria increased and have ability to use pollutant in the soil. The best deydrogenase activity between different microcosms related to polluted microcosm with nutrient. This result confirms that nitrogen and phosphorus can decrease the damage effect of crude oil on soil microbial community. The mechanism of this attenuation of toxicity effect of crude oil on microbial community can be related to enhance bioavailability of essential elements for bacteria in the soil. So after oil pollution of an area, soil supply upto nitrogen and phosphorus demand must be mentioned as a necessary practice to decrease the toxicity effect of pollutants. The highest biodegradation of crude oil in all studied soils belonged to industrial microcosm (95 %). It can be explained by adaptation theory because the bacteria in the industrial soil were better adapted to different pollutants and these bacteria have more capability for biodegradation of crude oil. By this reasonthe rate of degradation of crude oil in the industrial soil were higher than forest soil. Statistical analysis of the results showed that there was a significant correlation between MPN quantity of heterotrophic bacteria and other assayed factors. Also, forest soil seemed to have significant difference with other soils.
Conclusion: according to the obtained results by this study, it can be possibly proposed appropriate strategies for bioremediation of different studied soil types. The selection of best bioremediation strategies belong to specific types of soil. Just as this research confirmed that the type of soil plays significant role in the percentage of degradation.
