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Biodegradation of crude oil in an experimentally polluted peaty mangrove soil
Marine Pollution Bulletin Bourne, W. R. R (1983). Birds, fish and offal in the North Sea. Mar., Pollut. Bull. 14,294-296. Clarke, R. B., Dunnet, G. & Addy, J. M. (1984). Seabirds and North Sea oil. Mar. Pollut. Bull. 15,272-274. Coulson, J. C. & Thomas, C. S. (1985). Changes in the biology of the kittiwake Rissa tridactyla: a 31 year study of a breeding colony. J. Anita. Ecol. 54, 9-26. Cramp, S., Bourne, W. R. R & Saunders, D. (1974). The Sea-birds of Britain and Ireland. Collins, London. Furness, R. W. (1982a). Modelling relationships among fisheries, seabirds and marine mammals. In Marine Birds: Their Feeding Ecology and Commercial Fisheries Relationships (D. N. Nettleship, G. A. Sanger & P. E Springer, eds), pp. 117-126. Proc. Pacific Seabird Group Symp., Seattle, Washington,6-8 Jan. 1982. Can. Wildl. Serv. Spec. Pbl. Furness, R. W. (1982b). Seabird-fisheriesrelationships in the northeast Atlantic and North Sea. In Marine Birds: Their Feeding Ecology and Commercial Fisheries Relationships (D. N. Nettleship, G. A. Sanger & P. E Springer, eds), pp. 162-169. Proc. Pacific Seabird Group Symp., Seattle, Washington, 6-8 Jan. 1982. Can. Wildl. Serv. Spec. Publ. Furness, R. W. (1984). Seabird biomass and food consumption in the
North Sea. Mar. Pollut. Bull. 15, 244-248. Furness, R. W. & Birkhead, T. R. (1984). Seabird colony distributions suggest competition for food supplies during the breeding season. Nature, Lond., 311,655-656. Harris, M. E & Hislop, J. R. G. (1978). The food of young puffins Fratercula arctica. J. ZooL, Lond. 185,213-236. Hislop, J. R. G. & Harris, M. P. (1985). Recent changes in the food of young puffins Fratercula arctica on the Isle of May in relation to fish stocks. Ibis 127,234-239. Jones, R. (1982). Species interactions in the North Sea. Can. Spec. PubL Fish Aquat. Sci. 59, 48-63. Kunzlik, P. A. (1989). Small fish around Shetland. In Sandeels and Seabirds (Heubeck, M., ed.). Proceedings of a seminar held in Lerwick, Shetland, 15-16 October 1988. Shetland Bird Club, Lerwick. Lid, G. (1981). Reproduction of the puffin on Rest in the Lofoten Islands in 1964-1980. Fauna noru. Ser. C. Cinclus 4, 30-39. Ollason, J. C. & Dunnet, G. M. (1983). Modelling annual changes in numbers of breeding fulmars, Fulmarus glacialis, at a colony in Orkney. J. Anita. Ecol. 52,185-198. Tasker, M. L., Webb, A., Hall, A. J., Pienkowski,M. W. & Langslow,D. R. (1987). Seabirds in the North Sea. Nature Conservancy Council, Peterborough.
0025-326X/89$3.00+0.00 © 1989PergamonPressplc
MarinePollutionBulletin,Volume2(),No.9. pp.430-432,1989. PrintedinGreatBritain.
Biodegradation of Crude Oil in an Experimentally Polluted Peaty Mangrove Soil P. S C H E R R E R * a n d G. M I L L E t * lnstitut de la Carte Internationale de la V@~tation, Universitd Paul Sabatier, 39 allOes J. Guesde, 31062 Toulouse COdex; t Centre de Spectroscopic Mol~culaire, Facultd des Sciences et Techniques de St JOrOme, A v e n u e Escadrille N o r m a n d i e - N i e r n e n 13397 Marseille COdex 13, France
Biodegradation of oil hydrocarbons trapped in a peaty mangrove substratum is a very slow process. In spite of soil periodical oxygenation, aerobic microorganism activity seems to be limited. This may be accounted for by a nutrient deficiency related to a slow organic matter mineralization. This hypothesis was confirmed by stimulating normal alkanes biodegradation with an oleophilic fertilizer. To resort to such a product may compensate the slowness of oil biodegradation in mangrove soils.
M a n g r o v e is a coastal ecosystem regularly polluted by a c c i d e n t a l oil spills ( G e t t e r et al., 1981). T h e forest cover of these low energy tropical areas is particularly v u l n e r a b l e to oil pollution, since oil is very likely to a c c u m u l a t e a n d stagnate in their s u b s t r a t u m (Scherrer, 1988). Therefore, the long range future of a p o l l u t e d mangrove, d e p e n d s directly o n the t r a n s f o r m a t i o n rate of the oil t r a p p e d in the soil. T h e r e are few precise data 430
available on this subject as only a very small n u m b e r of chemical studies have been carried out in mangrove, where experimental conditions are particularly difficult. In the present paper we report a study on the role of environmental factors on oil biodegradation in a typical peaty mangrove soil.
Material and Methods I n March, 1986, two plots, side by side, each 2 m 2 were delimited for controlled c o n t a m i n a t i o n in a peaty m a n g r o v e of Petit-Bourg, G u a d e l o u p e (France). Both plots were polluted with 5 1 m -2 of Light A r a b i a n C r u d e Oil (LAC). O n e plot received c r u d e oil, the other, oil p r e m i x e d with 10% b i o a c t i v a t o r (an oleophilic fertilizer) in o r d e r to assess the possible effect of this p r o d u c t o n the b i o d e g r a d a t i o n process. T h e soil was c o m p o s e d of a b o u t 6 0 % peat a n d 4 0 % clay (in weight). Peat, which was very c o m p a c t , was
Volume 2 0 / N u m b e r 9/September 1989
produced by the roots of Rhizophora mangle trees which populated the station. This station was generally covered with water, except for some emergence periods during the dry season (December to April). When we applied the oil, the water was 30 cm deep. Redox potentials were markedly negative, only the uppermost cm were likely to be oxygenated. For each plot, samples were collected, with a graduated mud shovel, after periods of 3 days, 3 months and I1 months. Samples were stored at - 2 0 ° C . We analysed separately the 0-5 cm, 5-10 cm, and 10-20 cm levels. Hydrocarbons were extracted using an alkaline hydrolysis scheme (Farrington & Tripp, 1975), isolated by chromatography and analysed by high resolution capillary gas chromatography. A more detailed description of the experimental section has been published elsewhere (Scherrer, 1988).
30 cm were contaminated with decreasing concentrations from top to bottom. Observations during the first year indicate that oil was well trapped in the substratum. The addition of bioactivator to the oil favoured its accumulation at the soil surface. This phenomenon might be explained by bond formation between the soil organic matter and the bioactivator polar molecules. A similar effect has been reported by Oudot (1984) who used an oleophilic fertilizer associated to LAC spilled in a sand-mud substratum. At 3 and 11 months, we calculated for each sample the nC17/Pr and the nC~8/Ph ratios (Table 1). Their decrease reflects normal alkanes biodegradation since these hydrocarbons are more biodegradable than isoprenoids like pristane and phytane (Atlas et al., 1981). The degradation of oil without bioactivator was extremely limited over the time of this study. During the first three months (dry season), almost no biodegradation occurred in the uppermost 20 cm. During the following 8 months, although the station was under water, a slight biodegradation could be observed at the soil surface. Addition of bioactivator obviously helped the oil
Results and Discussion Figure 1 shows oil distribution in the soil of the two related plots. Biogenic organic matter corresponded to a concentration of 10 g kg -t sediment dry wt. The top
Concentration (g/drykg) 60 40
A
25 months ) Depth
(cm)
Concentration ( g ] dry kg ) 120
" ..
25
B
3
""""
( months )
Depth (cm)
Fig. 1 Evolution of the extracted organic matter concentration with depth and with time at the peaty station. A, plot polluted with 5 I m -2 oil. B, plot polluted with 5 I m -z oil premixed with 10% bioactivator.
431
Marine Pollution Bulletin
biodegradation process. During the dry season, oil transformation occurred down to 20 cm depth. When the soil was f l o o d e d , b i o d e g r a d a t i o n still p r o c e e d e d in the u p p e r m o s t level (Table 1) b u t it was s t o p p e d d e e p e r d o w n . T h e i n c r e a s e o f the c h a r a c t e r i s t i c b i o d e g r a d a t i o n ratios b e t w e e n 5 a n d 20 c m c o u l d b e d u e to the d o w n w a r d migration, f r o m the superficial a c c u m u l a t i o n level, of p a r t of the n o n - b i o d e g r a d e d oil (Fig. 1). TABLE 1
nC,7/Pristane and nC~8/Phytane ratios calculated from the chromatograms of the saturated fractions. Light Arabian Crude Oil (LAC) in the peaty soil after 3 and 11 months, at different depths. LAC + Bioactivator
LAC Date
Depth
nCyPr
nCls/Ph
nCyPr
nC~s/Ph
18 June 86 (3 months)
0- 5cm 5-10 cm 10-20 cm
4.2 4.6 4.5
2.5 2.7 2.5
3.7 3.9 3.0
1.9 2.1 1.9
17 Feb. 87 (11 months)
0- 5 cm 5-10 cm 10-20 cm
3.8 4.8 4.6
2.4 2.8 2.6
2.7 4.6 4.6
1.4 2.5 2.5
over the nearly continuous flooding period (JuneFebruary). A limited water oxygenation, could have supported an aerobic microbial activity at the watersoil interface. The lack of biodegradation deeper down, could indicate that during the flooding period, oxygen was the main biodegradation limiting factor. Conclusion The results collected at this station suggest that an oil p o l l u t e d m a n g r o v e is likely to r e m a i n p o l l u t e d for a long time. Oil m e t a b o l i s m , which is the m a i n d e g r a d a tion p a t h w a y for c r u d e oil in soil, was limited b y the availability of o x y g e n a n d nutrients in the peat. O l e o philic fertilizer might b e u s e d to f a v o u r b i o d e g r a d a t i o n in a nutrient deficient e n v i r o n m e n t . Nevertheless, it is a b s o l u t e l y n e c e s s a r y to test the toxicity of such p r o ducts on the fauna a n d flora b e f o r e to r e c o m m e n d their
use in mangrove. The authors are very grateful to ELF Aquitaine Company for their logistic and analytical help. They also thank professor J. Portecop and Dr. D. Imbert for the field cooperation.
(Ref. LAC: nClT/Pr = 5.1 ; nCts/Ph = 2.9) To s u m up, oil m e t a b o l i s m s e e m e d to o c c u r very slowly in b o t h plots, w h a t e v e r the depth. T h e oil t r a p p e d in the p e a t was n o t w a s h e d d o w n by the u n d e r g r o u n d c u r r e n t s a n d its b i o d e g r a d a t i o n was difficult. T h e p e a t y soil of this station turns out to b e a very g o o d oil trap. D u r i n g the e m e r g e n c e p e r i o d , r e d o x p o t e n t i a l s were positive a b o v e the w a t e r level b e c a u s e of o x y g e n diffusion in the soil. T h e r e f o r e , the m a i n b i o d e g r a d a t i o n limiting f a c t o r was n o t oxygen. It might b e a n u t r i e n t deficiency, related to the s l o w n e s s of the o r g a n i c m a t t e r m i n e r a l i z a t i o n p r o c e s s e s . This is s u p p o r t e d b y the stimulating a c t i o n of the b i o a c t i v a t o r n u t r i e n t s o n m i c r o o r g a n i s m s activity. A t soil surface, the b i o d e g r a d a t i o n of the oil a n d b i o a c t i v a t o r mixture k e p t going
432
Atlas, R. M., Boehm, P. D. & Calder, J. A. (1981). Chemical and biological weathering of oil, from the Amoco Cadiz Spillage, within the littoral zone. Estuar. coast. ShelfSci. 12,589-608. Farrington, J. W. & Tripp, B. W. (1975). A comparison of analysis methods for hydrocarbons in surface sediments. In Marine Chemist O' in the Coastal Environment (T. M. Church, ed.), ACS Symposium series. Getter, C. D., Scott, G. I. & Michel. J. (1981). The effects ofoil spills on mangrove forests: a comparison of five oil spill sites in the Gulf of Mexico and the Caribbean Sea. In Proceeding of the 198l Oil Spill Conference, pp. 535-540. American Petroleum Institute, Washington, D.C. Oudot, J. (1984). La bioddgradation microbienne des hydrocarbures. Etude du potentiel de bioddgradation et de son expression dans le milieu. Th~se de Doctorat en Sciences. Universitd Paris VII, France. Scherrer, P. (1988). La r6gdndration de la mangrove apr~s un ddversement accidentel d'hydrocarbures: phytotoxicitd et dvolution physicochimique du pdtrole brut pidg~ dans le substrat. Th~se de Doctorat en Sciences. Universit6 Paul Sabatier, Toulouse, France.
North Sea. Mar. Pollut. Bull. 15, 244-248. Furness, R. W. & Birkhead, T. R. (1984). Seabird colony distributions suggest competition for food supplies during the breeding season. Nature, Lond., 311,655-656. Harris, M. E & Hislop, J. R. G. (1978). The food of young puffins Fratercula arctica. J. ZooL, Lond. 185,213-236. Hislop, J. R. G. & Harris, M. P. (1985). Recent changes in the food of young puffins Fratercula arctica on the Isle of May in relation to fish stocks. Ibis 127,234-239. Jones, R. (1982). Species interactions in the North Sea. Can. Spec. PubL Fish Aquat. Sci. 59, 48-63. Kunzlik, P. A. (1989). Small fish around Shetland. In Sandeels and Seabirds (Heubeck, M., ed.). Proceedings of a seminar held in Lerwick, Shetland, 15-16 October 1988. Shetland Bird Club, Lerwick. Lid, G. (1981). Reproduction of the puffin on Rest in the Lofoten Islands in 1964-1980. Fauna noru. Ser. C. Cinclus 4, 30-39. Ollason, J. C. & Dunnet, G. M. (1983). Modelling annual changes in numbers of breeding fulmars, Fulmarus glacialis, at a colony in Orkney. J. Anita. Ecol. 52,185-198. Tasker, M. L., Webb, A., Hall, A. J., Pienkowski,M. W. & Langslow,D. R. (1987). Seabirds in the North Sea. Nature Conservancy Council, Peterborough.
0025-326X/89$3.00+0.00 © 1989PergamonPressplc
MarinePollutionBulletin,Volume2(),No.9. pp.430-432,1989. PrintedinGreatBritain.
Biodegradation of Crude Oil in an Experimentally Polluted Peaty Mangrove Soil P. S C H E R R E R * a n d G. M I L L E t * lnstitut de la Carte Internationale de la V@~tation, Universitd Paul Sabatier, 39 allOes J. Guesde, 31062 Toulouse COdex; t Centre de Spectroscopic Mol~culaire, Facultd des Sciences et Techniques de St JOrOme, A v e n u e Escadrille N o r m a n d i e - N i e r n e n 13397 Marseille COdex 13, France
Biodegradation of oil hydrocarbons trapped in a peaty mangrove substratum is a very slow process. In spite of soil periodical oxygenation, aerobic microorganism activity seems to be limited. This may be accounted for by a nutrient deficiency related to a slow organic matter mineralization. This hypothesis was confirmed by stimulating normal alkanes biodegradation with an oleophilic fertilizer. To resort to such a product may compensate the slowness of oil biodegradation in mangrove soils.
M a n g r o v e is a coastal ecosystem regularly polluted by a c c i d e n t a l oil spills ( G e t t e r et al., 1981). T h e forest cover of these low energy tropical areas is particularly v u l n e r a b l e to oil pollution, since oil is very likely to a c c u m u l a t e a n d stagnate in their s u b s t r a t u m (Scherrer, 1988). Therefore, the long range future of a p o l l u t e d mangrove, d e p e n d s directly o n the t r a n s f o r m a t i o n rate of the oil t r a p p e d in the soil. T h e r e are few precise data 430
available on this subject as only a very small n u m b e r of chemical studies have been carried out in mangrove, where experimental conditions are particularly difficult. In the present paper we report a study on the role of environmental factors on oil biodegradation in a typical peaty mangrove soil.
Material and Methods I n March, 1986, two plots, side by side, each 2 m 2 were delimited for controlled c o n t a m i n a t i o n in a peaty m a n g r o v e of Petit-Bourg, G u a d e l o u p e (France). Both plots were polluted with 5 1 m -2 of Light A r a b i a n C r u d e Oil (LAC). O n e plot received c r u d e oil, the other, oil p r e m i x e d with 10% b i o a c t i v a t o r (an oleophilic fertilizer) in o r d e r to assess the possible effect of this p r o d u c t o n the b i o d e g r a d a t i o n process. T h e soil was c o m p o s e d of a b o u t 6 0 % peat a n d 4 0 % clay (in weight). Peat, which was very c o m p a c t , was
Volume 2 0 / N u m b e r 9/September 1989
produced by the roots of Rhizophora mangle trees which populated the station. This station was generally covered with water, except for some emergence periods during the dry season (December to April). When we applied the oil, the water was 30 cm deep. Redox potentials were markedly negative, only the uppermost cm were likely to be oxygenated. For each plot, samples were collected, with a graduated mud shovel, after periods of 3 days, 3 months and I1 months. Samples were stored at - 2 0 ° C . We analysed separately the 0-5 cm, 5-10 cm, and 10-20 cm levels. Hydrocarbons were extracted using an alkaline hydrolysis scheme (Farrington & Tripp, 1975), isolated by chromatography and analysed by high resolution capillary gas chromatography. A more detailed description of the experimental section has been published elsewhere (Scherrer, 1988).
30 cm were contaminated with decreasing concentrations from top to bottom. Observations during the first year indicate that oil was well trapped in the substratum. The addition of bioactivator to the oil favoured its accumulation at the soil surface. This phenomenon might be explained by bond formation between the soil organic matter and the bioactivator polar molecules. A similar effect has been reported by Oudot (1984) who used an oleophilic fertilizer associated to LAC spilled in a sand-mud substratum. At 3 and 11 months, we calculated for each sample the nC17/Pr and the nC~8/Ph ratios (Table 1). Their decrease reflects normal alkanes biodegradation since these hydrocarbons are more biodegradable than isoprenoids like pristane and phytane (Atlas et al., 1981). The degradation of oil without bioactivator was extremely limited over the time of this study. During the first three months (dry season), almost no biodegradation occurred in the uppermost 20 cm. During the following 8 months, although the station was under water, a slight biodegradation could be observed at the soil surface. Addition of bioactivator obviously helped the oil
Results and Discussion Figure 1 shows oil distribution in the soil of the two related plots. Biogenic organic matter corresponded to a concentration of 10 g kg -t sediment dry wt. The top
Concentration (g/drykg) 60 40
A
25 months ) Depth
(cm)
Concentration ( g ] dry kg ) 120
" ..
25
B
3
""""
( months )
Depth (cm)
Fig. 1 Evolution of the extracted organic matter concentration with depth and with time at the peaty station. A, plot polluted with 5 I m -2 oil. B, plot polluted with 5 I m -z oil premixed with 10% bioactivator.
431
Marine Pollution Bulletin
biodegradation process. During the dry season, oil transformation occurred down to 20 cm depth. When the soil was f l o o d e d , b i o d e g r a d a t i o n still p r o c e e d e d in the u p p e r m o s t level (Table 1) b u t it was s t o p p e d d e e p e r d o w n . T h e i n c r e a s e o f the c h a r a c t e r i s t i c b i o d e g r a d a t i o n ratios b e t w e e n 5 a n d 20 c m c o u l d b e d u e to the d o w n w a r d migration, f r o m the superficial a c c u m u l a t i o n level, of p a r t of the n o n - b i o d e g r a d e d oil (Fig. 1). TABLE 1
nC,7/Pristane and nC~8/Phytane ratios calculated from the chromatograms of the saturated fractions. Light Arabian Crude Oil (LAC) in the peaty soil after 3 and 11 months, at different depths. LAC + Bioactivator
LAC Date
Depth
nCyPr
nCls/Ph
nCyPr
nC~s/Ph
18 June 86 (3 months)
0- 5cm 5-10 cm 10-20 cm
4.2 4.6 4.5
2.5 2.7 2.5
3.7 3.9 3.0
1.9 2.1 1.9
17 Feb. 87 (11 months)
0- 5 cm 5-10 cm 10-20 cm
3.8 4.8 4.6
2.4 2.8 2.6
2.7 4.6 4.6
1.4 2.5 2.5
over the nearly continuous flooding period (JuneFebruary). A limited water oxygenation, could have supported an aerobic microbial activity at the watersoil interface. The lack of biodegradation deeper down, could indicate that during the flooding period, oxygen was the main biodegradation limiting factor. Conclusion The results collected at this station suggest that an oil p o l l u t e d m a n g r o v e is likely to r e m a i n p o l l u t e d for a long time. Oil m e t a b o l i s m , which is the m a i n d e g r a d a tion p a t h w a y for c r u d e oil in soil, was limited b y the availability of o x y g e n a n d nutrients in the peat. O l e o philic fertilizer might b e u s e d to f a v o u r b i o d e g r a d a t i o n in a nutrient deficient e n v i r o n m e n t . Nevertheless, it is a b s o l u t e l y n e c e s s a r y to test the toxicity of such p r o ducts on the fauna a n d flora b e f o r e to r e c o m m e n d their
use in mangrove. The authors are very grateful to ELF Aquitaine Company for their logistic and analytical help. They also thank professor J. Portecop and Dr. D. Imbert for the field cooperation.
(Ref. LAC: nClT/Pr = 5.1 ; nCts/Ph = 2.9) To s u m up, oil m e t a b o l i s m s e e m e d to o c c u r very slowly in b o t h plots, w h a t e v e r the depth. T h e oil t r a p p e d in the p e a t was n o t w a s h e d d o w n by the u n d e r g r o u n d c u r r e n t s a n d its b i o d e g r a d a t i o n was difficult. T h e p e a t y soil of this station turns out to b e a very g o o d oil trap. D u r i n g the e m e r g e n c e p e r i o d , r e d o x p o t e n t i a l s were positive a b o v e the w a t e r level b e c a u s e of o x y g e n diffusion in the soil. T h e r e f o r e , the m a i n b i o d e g r a d a t i o n limiting f a c t o r was n o t oxygen. It might b e a n u t r i e n t deficiency, related to the s l o w n e s s of the o r g a n i c m a t t e r m i n e r a l i z a t i o n p r o c e s s e s . This is s u p p o r t e d b y the stimulating a c t i o n of the b i o a c t i v a t o r n u t r i e n t s o n m i c r o o r g a n i s m s activity. A t soil surface, the b i o d e g r a d a t i o n of the oil a n d b i o a c t i v a t o r mixture k e p t going
432
Atlas, R. M., Boehm, P. D. & Calder, J. A. (1981). Chemical and biological weathering of oil, from the Amoco Cadiz Spillage, within the littoral zone. Estuar. coast. ShelfSci. 12,589-608. Farrington, J. W. & Tripp, B. W. (1975). A comparison of analysis methods for hydrocarbons in surface sediments. In Marine Chemist O' in the Coastal Environment (T. M. Church, ed.), ACS Symposium series. Getter, C. D., Scott, G. I. & Michel. J. (1981). The effects ofoil spills on mangrove forests: a comparison of five oil spill sites in the Gulf of Mexico and the Caribbean Sea. In Proceeding of the 198l Oil Spill Conference, pp. 535-540. American Petroleum Institute, Washington, D.C. Oudot, J. (1984). La bioddgradation microbienne des hydrocarbures. Etude du potentiel de bioddgradation et de son expression dans le milieu. Th~se de Doctorat en Sciences. Universitd Paris VII, France. Scherrer, P. (1988). La r6gdndration de la mangrove apr~s un ddversement accidentel d'hydrocarbures: phytotoxicitd et dvolution physicochimique du pdtrole brut pidg~ dans le substrat. Th~se de Doctorat en Sciences. Universit6 Paul Sabatier, Toulouse, France.