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Escuela Técnica Superior De Ingenieros De Minas ESCUELA TÉCNICA SUPERIOR DE INGENIEROS DE MINAS PROYECTO FIN DE CARRERA Departamento de Ingeniería Química y Combustibles BIOREMEDATION OF OIL SPILLS YOLANDA SANCHEZ-PALENCIA GONZALEZ OCTUBRE 2011 i ÍNDICE Abstract .............................................................................................................................. vi 1 AIM AND SCOPE ...................................................................................................... 8 2 BACKGROUND ......................................................................................................... 9 2.1 Petroleum as source of hydrocarbons in the environment: pollution of marine environments.......................................................................................................................... 9 3 DEFINITIONS .......................................................................................................... 10 4 PETROLEUM AND MICROBIAL DEGRADATION OF HIDROCARBONS IN MARINE ENVIRONMENTS ................................................................................... 12 4.1 Microbial degradation of hydrocarbons ................................................................ 12 4.2 Microbial hydrocarbon-degrading communities................................................... 13 4.3 Obligate hydrocarbonoclastic marine bacteria...................................................... 14 5 BIOAUGMENTATION AND BIOSTIMULATION .......................................... 16 5.1.1 Bacteria consortia .......................................................................................... 19 5.2 Biostimulation ....................................................................................................... 33 5.2.1 Using guano as fertilizer ................................................................................ 35 5.3 Bioaugmentation versus biostimulation................................................................ 38 5.4 Commercial products: ongoing scenario .............................................................. 39 5.5 Confined systems and real-case studies: bridging the gap.................................... 40 6 PRODUCTION OF LIPIDS AND STORAGE COMPOUNDS BY HCB BACTERIA........................................................................................................................ 43 6.1 Production of lipids by hydrocarbonoclastic marine bacteria............................... 43 6.2 Storage compound accumulation in hydrocarbonoclastic bacteria....................... 44 6.3 Production of biosurfactants by oil-degrading bacteria ........................................ 44 ii 6.4 Production and export of lipids by Alcanivorax strains ........................................ 45 6.5 Concluding remarks and future perspectives ........................................................ 45 7 DISPERSANTS AND OIL MINERAL AGREGATES ...................................... 47 7.1 Use of dispersants ................................................................................................. 47 7.2 Oil–mineral interaction in particle image velocimetry (PIV) ............................... 48 8 COLD ENVIRONMENTS ...................................................................................... 58 8.1 Properties of spilled oil ......................................................................................... 59 8.2 Fate of oil spilled in the cold environments .......................................................... 59 8.3 Cold-adapted, oil-degrading microorganisms....................................................... 60 8.4 Factors affecting biodegradation........................................................................... 61 8.5 Techniques, successful cases, challenges and constraints .................................... 65 9 CONCLUSIONS ....................................................................................................... 69 10 REFERENCES .......................................................................................................... 70 iii ÍNDICE DE FIGURAS Figure 1: A. borkumensis ..................................................................................................... 15 Figure 2: CO2 Evolution during degradation of phenanthrene (NaCl: 30 g/l) .................... 25 Figure 3: Degradation of different concentrations phenanthrene ........................................ 26 Figure 4: Degradation of different concentrations fluorene ................................................ 26 Figure 5: Degradation of different concentrations pyrene................................................... 27 Figure 6: Degradation of different concentrations benzo(e)pyrene..................................... 27 Figure 7: Gas chromatography analysis for residual aliphatics through biodegradation (DMCB) ............................................................................................................................... 31 Figure 8: Residual concentrations in independent assays (DMC B) ................................... 32 Figure 9: Oil degradation using guano or ammonium sulfate as the nitrogen source ......... 38 Figure 10: ANS oil droplet with different types of minerals ............................................... 51 Figure 11: Mineral–oil interactions (green area= crude oil and red spots = minerals)........ 51 Figure 12: Effect of dispersants ........................................................................................... 52 Figure 13: Effect of dispersants ........................................................................................... 52 Figure 14: Vector field at the surface of an oil droplet of ANS .......................................... 53 Figure 15: Vector field around a moving oil droplet at different positions ......................... 54 Figure 16: Vector field of modified Kaolin and a static oil droplet (MESA)...................... 55 Figure 17: Comparison of interaction of hydrophilic minerals with different oil types...... 56 Figure 18: Effect of hydrophobic property on oil–mineral interaction ............................... 56 iv ÍNDICE DE TABLAS Table 1: Biodegradation of crude petroleum oil in soil by bacterial consortium. ............... 21 Table 2: Nutrient (NPK) and pH status of the bioremediated and normal soils ................. 22 Table 3: Factors influencing bioaugmentation and biostimulation processes ..................... 41 Table 4: Bioaugmentation and biostimulation: field and real case studies ......................... 42 Table 5: Composition of oils ............................................................................................... 50 Table 6: Physical properties of minerals ............................................................................. 50 v Abstract Due to the increasing demand of petroleum everywhere, and the great amount of spills, accidents and disasters, there is an urgent need to find an effective, non-cost and harmless method to clean up the affected areas. There are microorganisms in nature (bacteria and fungi, mainly) that feed on hydrocarbons and transform them into others harmless chemical substances. These bacteria produce enzymes that degrade oil very effectively. This natural process can be accelerated by adding more bacteria or providing nutrients and oxygen to facilitate their growth, which is called ―bioaugmentation and biostimulation‖. Through this project we discover that these processes can be affected by different factors making difficult the biodegradation execution and opening a gap between the laboratory experiments and the real cases. Therefore, there is much remain to be done and a lot of study ahead to make this technique available in a great scale. vi BIOREMEDIATION OF OIL SPILLS 8 1 AIM AND SCOPE This Project is a review of the last advances and discovers related to biodegradation of petroleum hydrocarbons both in water and in soil during last years. It is included temperate and cold environments, bioaugmentation (focusing on consortia rather than a single strain) and biostimulation, the use of dispersants and oil mineral aggregates and the production of lipids and storage compounds by hydrocarbonoclastic bacteria as a way of feeding. 9 2 BACKGROUND 2.1 Petroleum as source of hydrocarbons in the environment: pollution of marine environments The world demand for oil in 2008 was 85.62 million barrels per day (OPEC 2009). Petroleum-based products are the major source of energy for industry as well as daily life and represent the raw material for many chemical processes. The global transport and use of both petroleum and its derivatives have made petroleum hydrocarbons (PHCs) major contaminants in both prevalence and quantity in the environment. Along with transport, processing of crude oil represent an enormous potential for oil spills caused by tanker accidents exemplified by the one of the tanker Amoco Cadiz, which polluted the French coast of Brittany in 1978 or the Exxon Valdez tanker, which severely affected the Alaska coast in 1989 (Syutsubo et al., 2001). It was estimated that about 2 to 10 millions of tons of crude petroleum oil enter marine environments annually, the majority of which is generated from anthropogenic sources but also from natural sources like plants, algae, and bacteria (Tyagi, 2011). Another major part of oil enters the environment from municipal and industrial wastes and runoffs (Head and Swanell, 1999). In the case of petrol stations, refineries and storage tanks, leakage may be small, but continuous and prolonged
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