Bioremediation of Libyan soil contaminated with crude oil tank bottom sludge

Mansur, A 2015, Bioremediation of Libyan soil contaminated with crude oil tank bottom sludge, Doctor of Philosophy (PhD), Applied Science, RMIT University.

Document type: Thesis
Collection: Theses

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Title Bioremediation of Libyan soil contaminated with crude oil tank bottom sludge
Author(s) Mansur, A
Year 2015
Abstract Petroleum crude oil represents about 50% of the world’s energy and heating services and secures raw materials for many industries. The oil industry generates massive amounts of crude oil tank bottom sludge (COTBS), a complex mixture of hydrocarbons. Disposal of untreated COTBS represents a significant economic loss and threatens human and environment health. This research project was designed to promote a safe and environmentally bioremediation technology which involved extraction of oil from COTBS followed by utilization of efficient bioremediation technologies including bioaugmentation (inoculating indigenous hydrocarbon degrading agents), biostimulation (supplying with the needed nutrients and air) and natural attenuation where the contaminants were left to be degraded naturally.

In Libya, currently chemical and physical treatment methods are used to treat the contaminated soil, but both techniques are unsafe and expensive. Although bioremediation is a widely used methodology for petrogenic hydrocarbon contaminated soils, to date its application to the treatment of COTBS contaminated soils is limited especially in Mediterranean countries such as Libya. The aim of this project was to develop a cost effective, efficient, environment friendly and sustainable alternative treatment method that offers an alternative technology to current expensive, non-environmental friendly or sustainable physical and chemical approaches. In the first part of this study, the availability of oil within the COTBS for recycling purposes was investigated. Dichloromethane was used to extract the oil from the COTBS. The extracted oil was evaluated, characterised and compared to the parent oil (Hamada petroleum crude oil). The results indicated that COTBS contained a significant amount of recoverable oil (42.08 ± 1.1%). Gas Chromatography Mass Spectrometry detected 139 different hydrocarbon fractions within COTBS, composed of light hydrocarbons (30.7 ± 0.07%), heavy hydrocarbons (69.3 ± 0.4%), water (2.9 ± 0.2%) and solids (55.02 ± 0.6%).

The API gravity of the extracted oil was 33.03 which classified the oil as Brent, similar to the parent oil. The benefits of oil reclamation is two-fold; firstly to improve oil utilization efficiency and secondly to reduce the environmental contamination from the oil industry. In the second part of the study, hydrocarbonclastic bacteria were isolated from COTBS, COTBS contaminated soil and treated COTBS contaminated soil and then characterized in terms of their hydrocarbonoclastic potential. The results indicated the presence of 49 different bacterial phenotypes capable of growth on weathered Hamada crude oil. Evaluation of the substrate-degrading abilities of the individual isolates confirmed the growth of the 49 isolates on at least one substrate from the six chosen. Seven organisms were able to degrade 5 out of the 6 substrates. Amongst the 6 substrates, phenanthrene was the most utilized and octadecane was the least utilized. Cluster analysis divided the hydrocarbon degraders into two separated clusters. Cluster 2 represented the highly hydrocarbon degrading group. They represented 7 bacterial isolates from four phylogenic groups (Gammaproteobacteria, Firmicutes, Actinobacteria and Alphaproteobacteria).

These results confirmed that the isolation media was highly selective for hydrocarbon degrading organisms and the treated COTBS contaminated soil contained large numbers of hydrocarbonoclastic bacterial isolates. In the third part of the research, three bacterial isolates from cluster 2 (Pseudomonas sp, Pseudomonas xanthomarina and Arthrobacter nitroguajacolicus) were used in microcosm slurry phase bioremediation trials (through bioaugmentation, biostimulation and natural attenuation) of soil contaminated with COTBS. After 30 days, the degradation rate ranged from 97.8 to 99.4% where the total petroleum hydrocarbon (TPH) concentration was reduced from 30703 mg kg-1 to 170 to 664 mg kg-1, accompanied by a substantial reduction in PAH concentration, from 13816 mg kg-1 to below detection limit. In addition, the complete biodegradation of the carcinogenic and mutagenic fractions occurred. DNA-PCR-DGGE confirmed that no detectable changes in bacterial community between day 0 to day 30 were observed, with UPGMA analysis showing up to 100% similarity between day 0 and day 30 and between amended and control microcosms.

In the final part of the research, larger scale slurry phase mesocosms (500 ml) were used to further assess the potential of the hydrocarbonoclastic isolates. The ability of two bacterial isolates (Pseudomonas sp and Pseudomonas xanthomarina) to bioremediate contaminated Libyan soils was investigated using three strategies; (i) (bioaugmentation) (BA), (ii) (biostimulation) (BS), (iii) both biostimulation-bioaugmentation (BA-BS). The results showed that BS-BA was the most efficient bioremediation option, with the greatest reduction in TPH for both isolates (97.19 and 96.67 %) observed (from 30703 mg kg-1 to 860 mg kg-1 and 1020 mg kg-1). In contrast, the control showed only a 17.15% reduction in TPH concentration. After 90 days, BS-BA mesocosms also showed the highest rates of soil respiration (0.07 mg day-1 g soil-1), some 2.5-fold higher than that observed in the control soils (0.021 mg day-1 g soil-1), confirming that increased microbial activity correlated with degradation of the contaminant. Denaturing Gradient Gel Electrophoresis showed very little change in the bacterial community within the BS and BAS mecocosms while analysis of the BS-BA mesocosms showed an increase in microbial diversity. This study provides further understanding and knowledge about the bioremediation of COTBS contaminated soil and confirms the potential and benefit of indigenous hydrocarbonoclastic bacteria and the potential of BS-BA remediation technology. The research suggests that this sustainable remediation technology can substitute for the currently used physical and chemical treatment methods applied in Libya.
Degree Doctor of Philosophy (PhD)
Institution RMIT University
School, Department or Centre Applied Science
Keyword(s) Bioremediation
Crude Oil Tank Bottom Sludge
Bacterial consortia
Biolog and Eco plates
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