HAM PETROL İLE KİRLENMİŞ TOPRAKLARIN BİYOREMİDASYON SÜRECİNDE YAĞ ASİTLERİNDEKİ (FLYA-FLME) DEĞİŞİMLER

Yıl 2014, Cilt: 2 Sayı: 3, 25 - 37, 01.09.2014

Esin Eraydın Erdoğan Fikrettin Şahin Ayten Namlı

Öz

Bu çalışmada amaç ülkemizde petrol bulaşmasından kaynaklanan toprak kirliliği sorunlarına karşı “Biyoremidasyon” olarak bilinen ve biyolojik yöntemler ile toprağın yerinde iyileştirilmesi prensibine dayanan yaklaşımlar geliştirmek ayrıca başlangıç ve final topraklarında bulunan yağ asidi proflini ortaya koymaktır. Bu amaca yönelik olarak laboratuvar koşullarında oluşturulan üç temel biyoremidasyon uygulamasının (biyolojik çoğalım, biyolojik uyarım ve bu iki yaklaşımın birleşik uygulaması) ham petrolden kaynaklanan kirliliğin giderilmesindeki etkinliği test edilerek toprakların yağ asidi profili ortaya konmuştur. Biyolojik çoğalım uygulamaları altında, Adana, Batman ve Adıyaman’nın petrol ile kirlenmiş topraklarından izole edilen ham petrollü ortamda en iyi gelişmeyi gösteren ve ham petrolü parçalama düzeyleri en yüksek olan 6 bakteri seçilmiştir. (Pseudomons aeruginosa, Pseudomonas putida biotype A, Citrobacter amalonaticus-GC subgroup A, Acinetobacter genomospecies). Biyolojik uyarım uygulamaları altında hümik-fülvik asit olmak üzere organik materyalin ve birleşik uygulamalarda ise bakteri karışımı ile organik materyallin farklı birleşimlerinin 120 günlük bir inkübasyon sürecinde ne kadar ham petrol ayrıştırdığı niceliksel hidrokarbon analizleri ile belirlenmiştir. En yüksek petrol ayrışmasının %56 ile bakteri karışımı uygulanmış biyolojik çoğalım uygulamaları altında meydana gelmiştir. Değişik organik materyallerin kirli toprağa karıştırıldığı biyolojik uyarım koşullarında ise %18 düzeyinde bir petrol ayrışması görülmüştür. Birleşik uygulamalarda petrol ayrışması açısından %30’luk bir başarı sağlanmıştır. Temiz topraklarda ve petrol ile kirlenmiş topraklarda en fazla 15:0 iso, 15:0 anteiso, 16:0, 16:1 w7c, 17:0ai, 18:2w6,9, 18:1w9c yağ asitleri tespit edilmiştir. 18:1w9c yağ asidinin yüksek değerler gösterdiğinden Pseudomonas spp. bakterilerine özgü yağ asidi olduğunu vurgulayabiliriz. 15:0 anteiso yağ asidi değerleri temiz topraklara kıyasla petrol ile kirlenmiş topraklarda daha fazla olduğu tespit edilmiştir. Petrollü ortamda gram pozitif bakterilerin varlığının arttığını vurgulayabiliriz.

Anahtar Kelimeler

Toprak, ham petrol, bakteri, biyoremidasyon, FLYA-FLME

PLFA-FAME Changes During Bioremediation of Crude Oil Contamination Soil

Öz

This study aims to develop certain perspectives based on the principle of on-site remediation of the soil through biological means which is known as "bioremediation" against soil pollution issues resulting from fuel contamination in our country and to reveal the fatty acid profile in the final soils. The fatty acid profile of the soils were pointed out by testing the activity of three basic bioremediation applications (biological multiplication, biological excitation and the combined application of these two approaches) established in the laboratory environment for this aim. Under biological multiplication applications, 6 strains of bacteria were selected which exhibit the highest growth in crude oil environment isolated from oil-contaminated soils of Adana, Batman and Adıyaman and which have the highest levels of crude oil degradation. (Pseudomons aeruginosa, Pseudomonas putida biotype A, Citrobacter amalonaticus-GC subgroup A, Acinetobacter genomospecies). Under biological excitation applications, the organic materials being humic-fulvic acid and, in combined applications, different combinations of bacteria mixture and organic materials were examined as to the amount of crude oil they degrade in an incubation period of 120 days by qualitative hydrocarbon-type analyses. The highest level of oil degradation, being %56, occurred under biological multiplication applications where the bacteria mixture was applied. Under biological excitation conditions where various organic materials were applied to the contaminated soil, degradation to %18 was observed. In combined applications, oil degradation was achieved to %30. In unpolluted and oil-contaminated soils, max. 15:0 iso, 15:0 anteiso, 16:0, 16:1 w7c, 17:0ai, 18:2w6,9, 18:1w9c fatty acids were detected. Because the fatty acid 18:1w9c exhibited high levels, we may emphasize that it is a fatty acid typical to the bacteria Pseudomonas spp. It was determined that the levels of the fatty acid 15:0 anteiso is higher in oil-contaminated soils than in unpolluted soils. We may emphasize that the existence of gram positive bacteria increases in oil-contaminated environment.

Anahtar Kelimeler

Soil, crude oil, bacteria, bioremediation, FLYA-FLME

Kaynakça

• Atlas, R.M., 1981,”Microbial Degradation of Petroleum Hydrocarbons”, An Environmental Perspective Microbial Rev., vol: 45:180-209.
• Atlas, R.M. and Bartha, R., 1992, “Hydrocarbon Biodegradation and Oil Spill Bioremediation” – Adv. Microb. Ecol., 12:287 – 338.
• Bailey, V.L., Peacock, A.D., Smith, J.L. and Bolton, H., 2002, “Relationships between soil microbial biomass determined by cloroform fumigation-extraction, substrate-induced respiration, and phospholipid fatty acid analysis”, Soil Biology and Biochemistry, Vol.34, Issue.9, 1385-1389.
• Bligh E.G. and Dyer, W.J., 1959, “ A rapid method of total lipid extraction and purification”, Canadian Journal of Biochemistry and Physiology, 37, 911–917.
• Bossio, D.A., Scow, K.M., Gunapala, N. and Graham, K.J., 1998, “Determinants of soil microbial communities: Effects of agricultural management, season, and soil type on phospholipid fatty acid profiles”. Microb. Ecol., 36:1-12
• Bundy, J.G., Paton, G.I. and Campbell, C.D., 2002, “Microbial communities in different soil types do not converge after diesel contamination”, Journal of Applied Microbiology, vol: 92: 276-288.
• Erdogan E.E. 2010: Petrol İle Kirlenmiş Toprakların Biyolojik Olarak İyileştirilmesinin Laboratuvar Koşullarında Denenmesi, Ankara Üniversitesi, Fen Bilimleri Enstitüsü, Toprak Anabilim Dalı, (Ph. D. Thesis, p:236), Doktora Tezi, 236 sayfa
• Erdoğan, E.E., Şahin, F. and Karaca, A., 2011, “Determination of petroleum-degrading bacteria isolated from crude oil-contaminated soil in Turkey”, African Journal of Biotechnology Vol. 11(21), pp. 4853-4859, 13 March, 2012 .Available online at http://www.academicjournals.org/AJB DOI: 10.5897/AJB10.2239 ISSN 1684–5315 © 2012 Academic Journals
• Fierer, N., Schimel, J.P. and Holden, P.A., 2003,”Variations in microbial community composition through two soil depth profiles”, Soil Biology & Biochemistry, 35; 167–176.
• Filauro, G.G., Andreotti, G., Arlotti, D. and Reisinger, H.J., 1998, “Blow out of Trecate 24 crude oil well: how bioremediation techniques are soluing a major environmental emergency in a valuable agricultural area in contaminated soil 98”, Thomas Telford, pp. 403- 412.
• Frostegard, A., Tunlid, A. and Baath, E., 1993, “Phospholipid fatty acid composition, biomass, and activity of microbial communities from two different soil types experimentally exposed to different heavy metals”, Applied and Environmental Microbiology, 59, 3605-3617.
• Ghazali, F.M., Zaliha, R.N., Rahman, A., Salleh, A.B and Basri, M., 2004, “Biodegradation of hydrocarbons in soil by microbial consortium”, International Biodeterioration & Biodegradation, 54; 61 – 67.
• Guckert, J.B, Antworth C.P., Nichols, P.D. and White, D.C., 1985, “Phospholipid, ester-linked fatty acid profiles as reproducible assays for changes in prokaryotic community structure of estuarine sediments”, FEMS Microbiol Ecol, 31:147–158
• Gürbüz, F., Başpınar, E., Çamdeviren, H. ve Keskin, S., 2003, Tekrarlanan ölçümlü deneme düzenlerinin analizi, Van. 120s.Baskı: Yüzüncü Yıl Üniversitesi, ISBN: 975-92253-0-1.
• Harwood, J.L. and Russell, N.J., 1984, “Lipids in plants and microbes”. George, Allen and Unwin,London, England. Hayes, M.H.B. and Wilson, W.S. (1997). Humic substances, peats and sludges. The Royal Society of Chemistry Cambridge, pp 496.
• Ibekwe, A.M. and Kennedy, A.C., 1998, “Fatty acid methyl ester (FAME) profiles as a tool to investigate community structure of two agricultural soils”, Plant Soil. 206:151-161.
• Jùrgensen, K.S., Puustinen, J.A. and Suortti, M., 2000, “Bioremediation of petroleum hydrocarboncontaminated soil by composting in biopiles”, Environmental Pollution 107; 245 - 254.
• Kapley, A., Purohit, H.J., Chhatre, S., Shanker, R., Chakrabati, T. and Khanna, P., 1999, “Osmotolerance and hydrocarbon degradation by a genetically engineered microbial consortium”, Biosource Technology, 67; 241-245.
• Keinanen, M.M., Martikainen, P.J., Korhonen, L.K. and Suutari, M.H., 2003, “Microbial community structure in biofilms and water of a drinking water distribution system determined by lipid biomarkers”, Water Science and Technology, 47; pp. 143–147.
• Kishore, D. and Ashis, M.K., 2007, “Crude petroleum-oil biodegradation eYciency of Bacillus subtilis and Pseudomonas aeruginosa strains isolated from a petroleum-oil contaminated soil from North-East India”, Bioresource Technology, 98; 1339–1345.
• Kneif, C., Kolb, S., Bodelier, P.L.E., Lindri, A. and Dunfield, P.F., 2006, “The active methanotrophic community in hydromorphic soils changes in response to changing methane concentration”, Environmental Microbiology, 8, 321-333.
• Margesin, R. and Schinner, F., 1997, “Bioremediation of diesel-oil contaminated alpine soils at low temperatures”, Appl. Microbiol. Biotechnol., 47,462-468.
• Miller, I., and Berger, T., 1985, “Bacteria identification by gas chromatography of whole cell fatty acids”, Hewlett-Packard Gas Chromatography Application Note, Hewlett-Packard Co., Alto, CA., 228-238.
• Morgan, P. and Watkinson, R.J., 1989, “Hydrocarbon degradation in soils and methods for soils biotreatment”, CRC Critical Reviews in Biotechnology 8, 305-333.
• Obire, O. and Okudo, I. V., 1997, “Effects of Crude Oil on a Freshwater Stream in Nigeria” Discov. Innov., 9: 25-32.
• Olsson, S. and Persson, P., 1999, “The composition of bacterial populations in soil fractions differing in their degree of adherence to barley roots”, Applied Soil Ecology, vol 12-3; 205- 215.
• Parker, R.J. and Taylor, J., 1983, “The relationship between fatty acid distributions and bacterial respiratory types in contemporary marine sediments”, Estuarine Coastal Shelf Sci 16:173– 189.
• Pelz O., A. Chatzinotas, N. Andersen, S.M. Bemasconi, C. Hesse, W.R. Abraham and J. Zeyer, 2001, “Use of isotopic and molecular techniques to link toluene degradation in denitrifying aquifer microcosms to a specific microbial population”, Archives of Microbiology, 175; pp. 270–281.
• Peressuttia, S.R., Alvarez, H.M. and Oscar, H.P., 2003, “Dynamics ofhydrocarbon-degrading bacteriocenosis ofan experimental oil pollution in Patagonian soil”, International Biodeterioration and Biodegradation, 52 (2003) 21 – 30.
• Pokethıtıyook, P., Sungpetch, A., Upathame, S. and Kruatrachue, M., “Enhancement of Acinetobacter Calcoaceticus in biodegradation of Tapis crude oil” 17th WCSS, Symposium no: 42, paper no. 2309.Thailand, 2002.
• Porta, A., Trovato, A., McCarty, K. Uhler, A. and Andreotti, G., 1998, “Degredation of saturated and polycyclic aromatic hydrocarbons and formation of their metabolites in bioremediated crude oil containing sois”, In: Alleman, B.C. , Leeson, A. (Eds), In situ and on site Bioremediation , vol:1. Battelle Press, Columbus, USA, pp. 505-510.
• Ratledge, C. and Wilkinson, S.G. (ed)., 1988, “Microbial lipids”, vol:1 Academic Pres. London, England. In. Fessica R. Hanson, Jennifer L. Macalady, David Haris and Kate M. Scow 1999. Linking Toluene degradation with Specific Microbial Populations in soil. Applied and Environmental Microbiology, p:5403-5408.
• Ringelberg, D., Richmond, M., Foley, K. and Reynolds, C., 2008, “Utility of lipid biomarkers in support of bioremediation efforts at army sites”, Journal of Microbiological Methods, 74(1): 17- 25.
• Rojas-Avelizapa N.G., Rodriguez-Vazquez R., Enriquez-Villanueva F., Martinez-Cruz J. and PoggiVaraldo, H.M., 1999, “Transformer oil degradation by an indigenous microflora isolated from a contaminated soil”, Resources, Conservation and Recycling, 27 ;15-26.
• Sasser, M., 1990, “ Identification of bacteria by gas chromatography of cellular fatty acids”, Tech. Note. 101. Microbial ID, Newark, DE.
• Sextone, A.J., Everett, K., Jenkins, T. and Atlas, R., 1978, “Fate of crude and refined oils in North slope soils”, Arctic 31, pp. 339-347.
• Song, H.G. and Bartha, R., 1990, “Effects of jet fuel on the microbial community of soil”, Appl. Environ. Microbiol., 56, 646-651.
• Steffan, R., Mccloy, J. K., Vainberg, S., Condee, C. W. and Zhang, D., 1997, “Biodegradation of the Gasoline Oxygenates Methyle tert-Butyl Ether, Ethyl tert – Butyl Ether and tert – Amyl Methyl Ether by Propane-Oxidizing Bacteria”, Appl. Environ. Microbiol., 63(11):4216-4222.
• Şahin, F., 1997, Detection, identification and characterization of strains of Xanthomonas campestris pv. vesicatoria by traditional and molecular methods, and resistance in Capsicum species to Xanthomonas campestris pv. vesicatoria pepper race6. (Ph. D. Thesis). The Ohio State University, 182 p. Ohio. Şahin, F., Kotan, R. and Dönmez, M.F., 1999, “First report of bacterial blight of Mulberries caused by Pseudomonas syringae pv. mori in the eastern Anatolia Region of Turkey”, Plant Dis., 83, 1176.
• Venosa, A.D., Stephen, J.R., Macnaughton, S.J., Chang, Y. and White, D.C., 2000, “Microbial population changes during bioremediation of an experiment al oil spill”, In Microbial Biosystems: New Frontiers ed. Bell, C.R., Brylinsky, M. and Johnson-Green, P. pp. 767-773. Halifax:Atlantic Canada Society for Microbial Ecology.
• Vestal, R.J. and White, D.C., 1989, “Lipid analysis in microbial ecology”, Bioscience 39: 535-541. Vidali M. 2001. Bioremediation. An overview. Pure Appl. Chem. Vol. 73, No. 7, pp. 1163–1172, 2001.
• White, D.C., Pinkart, H.C., and Ringelberg, D.B., 1996a, “Biomass measurements: biochemical approaches” In Manual of Environmental Microbiology, 1st edn. Hurst, C.J., Knudsen, 169 G.R., McInerney, M.J., Stetzenbach, L.D., and Walter, M.V. (eds). Washington DC: American Society for Microbiology Press, pp. 91–101.
• White, D.C., Stair, J.O. and Ringelberg, D.B., 1996b, “Quantitative comparisons of in situ microbial biodiversity by signature biomarker analysis”, J Ind Microbiol, 17: 185–196.
• Winer, B.J., Brown, D.R. and Michels, K.M., 1991, “Statistical Principles in Experimental Design”, ISBN: 0-07-070982-3. 3 eds.Page: 1057.
• Zelles, L., Bai, Q.Y., Beck, T. and Beese, F., 1992, “Signature fatty acids in phospholipids and lipopolysaccharides as indicators of microbial biomass and community structure in agricultural soils”, Soil Biology & Biochemistry, 24, 317–323.