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Correlation of dispersant effectiveness and toxicity of oil dispersants towards the alga Chlamydomonas reinhardti
Volume 13/Number 10/October 1982 Carpenter, E. J. & Smith, K. L. (1972). Plastic on the Sargasso Sea surface. Science, 175, 1240-1241. Carpenter, E. J., Anderson, S. J., Harvey, G. R., Miklas, H. P. & Peck, B. P. (1972). Polystyrene spherules in coastal waters. Science, 178, 749-750. Colton, J. B., Knapp, F. D. & Burns, B. R. (1974). Plastic particles in the surface water of the northwestern Atlantic. Science, 185,491-497. Cordes, C., Atkinson, L., Lee, R. & Blanton, J. (1980). Pelagic tar off Georgia and Florida in relation to physical processes. Mar. Pollut. Bull., 11,315-317. Dixon, T. R & Dixon, T. J. (1981). Marine litter surveillance. Mar. Pollut. Bull., 12, 289-295. Failer, A. J. & Woodcock, A. H. (1964). The spacing of windrows of sargassum in the ocean. J. mar. Res., 22, 22-29. Fowler, S. W. & Elder, D. L. (1978). PCB and DDT residues in a Mediterranean pelagic food chain. Bull. envir. Contain. Toxic., 19, 244-249. Friedrich, H. (1969). Marine Biology. Universityof Washington Press, Seattle. Giam, C. S., Chan, H. S. & Neff, G. S. (1978). Phthalate ester plasticizers, DDT, DDE and polychlorinated biphenyls in biota from the Gulf of Mexico. Mar. Pollut. Bull., 9, 249-251. Hincks, T. (1880). Contributions towards a generalhistory of the marine Polyzoa. II. Foreign Membraniporina (cont.) Ann. Mag. Nat. Hist., ser. 5, 6, 376-381. Marcus, E. (1937). Bryozoarios marinhos brasileiros, I. Bol. Fac. Fil. Ci~nc. Letr., Univ. Sao Paulo, Vol. l, Zoologia, No. l, 1-224.
~larinePollutionBullelin,
Morris, R. J. (1970). Phthalic acid in the deep sea jellyfish Atolla. Nature, Lond. 227, 1264. Morris, R. J. (1980a). Floating plastic debris in the Mediterranean. Mar. Pollut. Bull., 11,125. Morris, R. J. (1980b). Plastic debris in the surface waters of the South Atlantic. Mar. Pollut. Bull., 11, 164-166. Osburn, R. D. (1940). Bryozoa of Puerto Rico with a resume of the West Indian bryozoan fauna. N.Y. Acad. ScL, Sci. Survey Porto Rico and Virgin Islands, 16, 321--486. Ryland, J. R. (1974). Observations on some epibionts of gulf-weed, Sargassum natans (L.) Meyen, J. exp. mar. Biol. Ecol., 14, 17-25. Van Dolah, R. F., Burrell. V. G. & West, S. B. (1980). The distribution of pelagic tars and plastics in the South Atlantic Bight. Mar. Pollut. Bull., 11,352-356. Venrick, E. L., Backman, T. W., Bartram, W. C., Platt, C. J., Thornhill, M. S. & Yeates, R. E. (1973). Man-made objects on the surface of the Central North Pacific Ocean. Nature, Lond., 241,271. Weiss, C. M. (1948). The seasonal occurrence of sedentary marine organismsin BiscayneBay, Florida. Ecology, 29, 153-172. Winston, J. E. (1982). Marine bryozoans (Ectoprocta) of the Indian River Area, Florida. Am. Mus. Nat. Hist. Bull. 173, 99-176. Winston, J. E. & Eiseman, N. J. (1980). Bryozoan-algal associationsin coastal and continental shelf waters of eastern Florida. Fla. ScL, 43, 65-74. Wrist, G. (1964). Stratification and circulation in the Antillean-Caribbean Basins. Part 1. Columbia Univ. Press, New York.
Vol. 13, No. 10. pp. 351-353~ 1982
0025-326X/S2 / 100351-03 $03.00/0 (~) 1982 Pergamon Press I td.
Printed in Great Britain
Correlation of Dispersant Effectiveness and Toxicity of Oil Dispersants Towards the Alga Chlamydomonas reinhardti G U N N A R BRATBAK, M I K A L H E L D A L , G J E R T K N U T S E N , T O R L E I V L I E N and SVEIN N O R L A N D D e p a r t m e n t o f M i c r o b i o l o g y a n d P l a n t Physiology, University o f Bergen, All~gt. 70, 5 0 0 0 Bergen, N o r w a y
Using synchronous cultures of the unicellular green alga Chlamydomonas reinhardti, the toxicifies of mixtures of Ekofisk crude oil and oil dispersants were measured. Sixteen so-called concentrates and I0 solvent-based dispersants were tested. The dispersing effectiveness of these com. pounds with respect to the Ekofisk crude oil was also measured. The concentrates were tested undiluted as well as diluted using algal growth medium (2%o safinity) and artificial sea water (33%o salinity) as dispersing fiquid. The solvent-based compounds were tested in algal medium. For all compounds we found significant correlations between their toxicity and their effectiveness in dispersing the Ekofisk oil, such that the more effective the compound, the more toxic it was. To control the use of chemical oil dispersants in Norwegian waters, the Norwegian authorities have put forward regulations including toxicity testing with the green alga C h l a m y d o m o n a s reinhardti ( A n o n . , 1980). For several years we have been testing the toxicity of dispersants, both water-soluble concentrates a n d solvent-based types, with this alga and its close relative, the marine flagellate Dunaliella bioculata. During these studies we have noticed that the
most toxic c o m p o u n d s seemed to be those which were the most effective in dispersing oil. This impression stems from visual observations of their effectiveness as seen in the algal cultures during testing, a n d from the observations that the mixture of oil a n d dispersant was usually more toxic than the two tested alone. W e here report the results of experiments u n d e r t a k e n to test whether or n o t such a correlation exists between toxicity and effectiveness.
Methods The test oil was Ekofisk crude oil, a n d the dispersants were commercially available ones obtained from the manufacturers. I n this report they are not identified. Toxicity tests were performed with synchronous cultures of C h l a m y d o m o n a s reinhardti, as described previously (Heldal et al., 1977). In short, appropriate a m o u n t s of oil/ dispersant mixture were added to the algal cultures contained in 25 ml test tubes, to give a concentration series. These cultures consisted of zoospores at a density of ca 1.5 million cells ml-1, and they were cultivated in parallel with control cultures at 35 ° C a n d at 20 000 lux, with intermittent 351
Marine Pollution Bulletin
aeration with air containing 2% CO2. The effects of oil/dispersant mixtures and dispersants on this alga are such that a fraction of the population is killed within half an hour after their addition to zoospores, the fraction being concentration dependent. The surviving cells grow and develop normally. After 4 h of cultivation the cell volume distributions of the dead cells and those growing on normally are completely separated, thus facilitating the numerical sizing of the two subpopulations. LCso values are calculated from probit plots of fractions of dead cells versus concentration. Cell number and volume distribution were recorded with a Coulter ZB/Channelyzer particle analysing system connected to a Digital Equipment Corporation MINC laboratory computer. The test for effectiveness was based on the one used by the Warren Spring Laboratory, UK (Martinelli & Cormac, 1979). We used a 500 ml test flask with a screw cap having two inlet tubes, one sampling tube extending to a point 1.5 cm above the bottom, and one reaching just into the neck of the flask. As test liquids we used 250 ml of algal growth medium (20/0osalinity) and artificial sea water (Rila Marine Mix, 33%0 Rila Products, N.J., USA). The liquid was sampled by applying low air pressure to the flask via the short inlet tube. The concentrates were tested undiluted, as well as diluted with 9 parts of water. The undiluted concentrates were mixed with oil in an 1 to 1 ratio. Oil and dispersant were blended by thorough shaking in a screwcapped 50 ml flask. Five hundred microliters was pipetted onto the surface of the test water by letting the mixture flow down the flask wall from a point 1 cm above the liquid surface. This method of application was the most reproducible of several tested. The oil/dispersant sample and the test water were mixed in the test flask by shaking the flask in its upright position on a shaking table with circular motion at 160 rpm for 10 min before sampling. After shaking, the mixture was left standing for 10 rain before sampling. The first 10 ml sampled was discarded, and the next 50 ml was collected in a volumetric flask. The sample was quantitatively transferred to a 250 ml separatory funnel and 10 ml of spectrograde methylene dichloride (MD) was added. In the tests where algal medium was used, the salt concentration was increased by adding 5 ml saturated NaCI before adding MD to prevent foaming. Immediately after extraction the extract was centrifuged in a capped tube to remove foam and water droplets. The amount of oil in the extract was determined by measuring absorption at 580 nm with a Shimadzu 50L Spectrophotometer, using MD as reference, and expressed as per cent effectiveness, % E. All glassware was thoroughly cleaned by rinsing in hot water, followed by an acetone wash, and a final rinsing with hot water.
Results As a preliminary we tested different methods for mixing oil and dispersant, for the addition of this mixture to the test water, for dispersion, for sampling and measuring the amount of dispersed oil. Based on the results, we established the test procedure described in the Methods section. It was also important to keep the test conditions as close to the conditions of the toxicity test as possible. Since Chlamydomonas is a fresh water alga, the main testing of effective352
150
100
50
f. x I
I
213
411
I
I
88
EFFECTIVENESS,
Fig. 1 LCso values as function of effectiveness of diluted concentrates tested in algal medium.
ness was done in algal medium of 2 0 0 salinity. However, some compounds were also tested for effectiveness in artificial sea water at 330/0osalinity. For twelve of the concentrates tested in diluted form in algal medium, the effectiveness values used in the plot of Fig. 1 are averages of 2-5 separate measurements; the values for the other four represent single measurements. The standard deviation for the results of one compound tested five times was + 17 %, which might indicate the precision of the effectiveness measurements as a whole. All except one of the concentrates showed consistent effectiveness values when tested in diluted form in algal medium. The remaining compound, denoted A, showed rather high effectiveness in some tests and almost zero in the rest. Figure 1 is a plot of LCs0 values versus effectiveness in algal medium, and with both the low and high effectiveness values for compound A. The relationship between toxicity and effectiveness seems to be hyperbolic, as indicated with the drawn curve. Statistical correlation evaluation of log transformed values showed significant correlation at the 95% level. Thus one can conclude that high effectiveness is associated with high toxicity. In these experiments with algal medium the test conditions of the effectiveness test were closest to those of the toxicity test. From the plot it can be seen that the compound marked B clearly lies outside the drawn curve, and obviously conforms less to the general trend. It displays relatively low toxicity at relatively high effectiveness. Figure 2 shows a similar plot for the 10 solvent-based dispersants. A similar significant correlation (95°70 level) was also found for this group of compounds, but with none of the members deviating from the general trend. In addition to the experiments with diluted concentrates performed under conditions pertinent to the toxicity test, some concentrates were also tested (1) in the undiluted form with algal medium as dispersing liquid, (2) undiluted and (3) diluted, and with artificial sea water as dispersing liquid. The results of these experiments corroborated those described in Figs 1 and 2, also with respect to B, deviating under all three testing conditions. The highest correlation between toxicity and effectiveness was found under conditions (1) and (2). It is important to note that the correlations are all done with the same set of LCs0 values. Obviously we could not do toxicity tests with the fresh-water alga in sea water.
Volume 13/Number 10/October 1982
400
r: I
0 13
20
40
60
ill2
EFFECTIVENESS,
Fig. 2 LCs0 values as function of effectiveness of solvent-based dispersants tested in algal medium.
Discussion The effectiveness of an oil dispersant does not depend only on its own properties, but also on a series of chemical and physical factors which characterize the environment, and the oil which it is applied to. Some of the factors defining the environment or the field conditions, such as temperature and salinity, and others, such as oil type and dispersant/oil ratio, can be controlled in the laboratory. Others, like wave and wind action, and water turbulence, can hardly be simulated and controlled for testing purposes. The testing apparatus and procedures also create physical conditions which have no parallels under natural conditions. The choice of testing conditions with respect to physical and chemical factors and testing procedures will therefore to a great extent determine the outcome of the test. We aimed at an effectiveness test with conditions not very different from those of the toxicity test. Whilst the toxicity test was done at 35°C, 20000 lux, 2%0 salinity, and with aeration for 10 s every min for 4 h, the effectiveness test was performed at 20°C, room illumination, 2 and 33%0 salinity, and with circular movements at 160 rpm for 10 min. Since effectiveness data were not available in the literature, we had to measure effectiveness for the dispersants in question. In accordance with the procedure for the toxicity test the dispersants and oil were also mixed before their application to the water. The preliminary experiments clearly showed that the mode of application of the mixture to the water influenced the results of the effectiveness test. This influence was strong for some
compounds whereas others were little affected. The simplest and most reproducible method was found to be a gentle addition down the vessel wall. The samples of dispersed oil were taken from analysis 10 min after shaking was stopped, and from near the bottom. They are therefore representing a rather stable oilin-water emulsion consisting of small oil droplets. Thus dispersion effectiveness measured in this test mainly refers to the ability of the dispersants to form such stable emulsions. Comparing toxicity data with dispersion effectiveness, the clearest relationship was found with effectiveness data from the experiments performed under conditions being closest to those of the toxicity test. This holds both for concentrates and solvent-based dispersants. It is important to note that our studies with concentrates indicate that a similar correlation exists with effectiveness in sea water. This indication stems from comparison of toxicity data obtained from tests on fresh water with effectiveness data from tests in sea water. However, we have unpublished data of toxicity gained from experiments with the green marine flagellate Dunaliella bioculata supporting the indication: The LCs0 values obtained with this alga are very similar to those found with Chlamydomonas. The main conclusion is that there is a clear association between effectiveness and toxicity under the present test conditions, such that with increasing effectiveness the toxicity increases for concentrates as well as solvent-based dispersants. The results indicate two exceptions from this relationship among the concentrates, the compounds in question being clearly less toxic than should be expected from their effectiveness. The general validity of these observations will be further explored in experiments with a series of other oils, other dispersants and algae.
We wish to thank The Norwegian State Oil Company, The Norwegian Research Council for Scientific and Industrial Research, and the Norwegian Fisheries Research Council for financial support, and Miss Siri Ottesen for skilled technical assistance.
Anon (1980). Norweigna Environmental Protection Agency. Forskrifter for sammensetting og bruk av oljedispergeringsmidler for bekjempelse av oljes¢l. Oslo. Heldal, M., Norland, S., Lien, T. & Knutsen, G. (1977). Acute toxicity of several oil dispersants towards the green algae Chlamydomonas and Dunaliella. Chemosphere, 6, 247-255. Martinelli, F. N. & Cormac, D. (1979). Investigation of the effects of oil viscosity and water-in-oil emulsion formation on dispersant efficiency. Report from Warren Spring Laboratory, Department of Industry, Stevenage, England.
353
~larinePollutionBullelin,
Morris, R. J. (1970). Phthalic acid in the deep sea jellyfish Atolla. Nature, Lond. 227, 1264. Morris, R. J. (1980a). Floating plastic debris in the Mediterranean. Mar. Pollut. Bull., 11,125. Morris, R. J. (1980b). Plastic debris in the surface waters of the South Atlantic. Mar. Pollut. Bull., 11, 164-166. Osburn, R. D. (1940). Bryozoa of Puerto Rico with a resume of the West Indian bryozoan fauna. N.Y. Acad. ScL, Sci. Survey Porto Rico and Virgin Islands, 16, 321--486. Ryland, J. R. (1974). Observations on some epibionts of gulf-weed, Sargassum natans (L.) Meyen, J. exp. mar. Biol. Ecol., 14, 17-25. Van Dolah, R. F., Burrell. V. G. & West, S. B. (1980). The distribution of pelagic tars and plastics in the South Atlantic Bight. Mar. Pollut. Bull., 11,352-356. Venrick, E. L., Backman, T. W., Bartram, W. C., Platt, C. J., Thornhill, M. S. & Yeates, R. E. (1973). Man-made objects on the surface of the Central North Pacific Ocean. Nature, Lond., 241,271. Weiss, C. M. (1948). The seasonal occurrence of sedentary marine organismsin BiscayneBay, Florida. Ecology, 29, 153-172. Winston, J. E. (1982). Marine bryozoans (Ectoprocta) of the Indian River Area, Florida. Am. Mus. Nat. Hist. Bull. 173, 99-176. Winston, J. E. & Eiseman, N. J. (1980). Bryozoan-algal associationsin coastal and continental shelf waters of eastern Florida. Fla. ScL, 43, 65-74. Wrist, G. (1964). Stratification and circulation in the Antillean-Caribbean Basins. Part 1. Columbia Univ. Press, New York.
Vol. 13, No. 10. pp. 351-353~ 1982
0025-326X/S2 / 100351-03 $03.00/0 (~) 1982 Pergamon Press I td.
Printed in Great Britain
Correlation of Dispersant Effectiveness and Toxicity of Oil Dispersants Towards the Alga Chlamydomonas reinhardti G U N N A R BRATBAK, M I K A L H E L D A L , G J E R T K N U T S E N , T O R L E I V L I E N and SVEIN N O R L A N D D e p a r t m e n t o f M i c r o b i o l o g y a n d P l a n t Physiology, University o f Bergen, All~gt. 70, 5 0 0 0 Bergen, N o r w a y
Using synchronous cultures of the unicellular green alga Chlamydomonas reinhardti, the toxicifies of mixtures of Ekofisk crude oil and oil dispersants were measured. Sixteen so-called concentrates and I0 solvent-based dispersants were tested. The dispersing effectiveness of these com. pounds with respect to the Ekofisk crude oil was also measured. The concentrates were tested undiluted as well as diluted using algal growth medium (2%o safinity) and artificial sea water (33%o salinity) as dispersing fiquid. The solvent-based compounds were tested in algal medium. For all compounds we found significant correlations between their toxicity and their effectiveness in dispersing the Ekofisk oil, such that the more effective the compound, the more toxic it was. To control the use of chemical oil dispersants in Norwegian waters, the Norwegian authorities have put forward regulations including toxicity testing with the green alga C h l a m y d o m o n a s reinhardti ( A n o n . , 1980). For several years we have been testing the toxicity of dispersants, both water-soluble concentrates a n d solvent-based types, with this alga and its close relative, the marine flagellate Dunaliella bioculata. During these studies we have noticed that the
most toxic c o m p o u n d s seemed to be those which were the most effective in dispersing oil. This impression stems from visual observations of their effectiveness as seen in the algal cultures during testing, a n d from the observations that the mixture of oil a n d dispersant was usually more toxic than the two tested alone. W e here report the results of experiments u n d e r t a k e n to test whether or n o t such a correlation exists between toxicity and effectiveness.
Methods The test oil was Ekofisk crude oil, a n d the dispersants were commercially available ones obtained from the manufacturers. I n this report they are not identified. Toxicity tests were performed with synchronous cultures of C h l a m y d o m o n a s reinhardti, as described previously (Heldal et al., 1977). In short, appropriate a m o u n t s of oil/ dispersant mixture were added to the algal cultures contained in 25 ml test tubes, to give a concentration series. These cultures consisted of zoospores at a density of ca 1.5 million cells ml-1, and they were cultivated in parallel with control cultures at 35 ° C a n d at 20 000 lux, with intermittent 351
Marine Pollution Bulletin
aeration with air containing 2% CO2. The effects of oil/dispersant mixtures and dispersants on this alga are such that a fraction of the population is killed within half an hour after their addition to zoospores, the fraction being concentration dependent. The surviving cells grow and develop normally. After 4 h of cultivation the cell volume distributions of the dead cells and those growing on normally are completely separated, thus facilitating the numerical sizing of the two subpopulations. LCso values are calculated from probit plots of fractions of dead cells versus concentration. Cell number and volume distribution were recorded with a Coulter ZB/Channelyzer particle analysing system connected to a Digital Equipment Corporation MINC laboratory computer. The test for effectiveness was based on the one used by the Warren Spring Laboratory, UK (Martinelli & Cormac, 1979). We used a 500 ml test flask with a screw cap having two inlet tubes, one sampling tube extending to a point 1.5 cm above the bottom, and one reaching just into the neck of the flask. As test liquids we used 250 ml of algal growth medium (20/0osalinity) and artificial sea water (Rila Marine Mix, 33%0 Rila Products, N.J., USA). The liquid was sampled by applying low air pressure to the flask via the short inlet tube. The concentrates were tested undiluted, as well as diluted with 9 parts of water. The undiluted concentrates were mixed with oil in an 1 to 1 ratio. Oil and dispersant were blended by thorough shaking in a screwcapped 50 ml flask. Five hundred microliters was pipetted onto the surface of the test water by letting the mixture flow down the flask wall from a point 1 cm above the liquid surface. This method of application was the most reproducible of several tested. The oil/dispersant sample and the test water were mixed in the test flask by shaking the flask in its upright position on a shaking table with circular motion at 160 rpm for 10 min before sampling. After shaking, the mixture was left standing for 10 rain before sampling. The first 10 ml sampled was discarded, and the next 50 ml was collected in a volumetric flask. The sample was quantitatively transferred to a 250 ml separatory funnel and 10 ml of spectrograde methylene dichloride (MD) was added. In the tests where algal medium was used, the salt concentration was increased by adding 5 ml saturated NaCI before adding MD to prevent foaming. Immediately after extraction the extract was centrifuged in a capped tube to remove foam and water droplets. The amount of oil in the extract was determined by measuring absorption at 580 nm with a Shimadzu 50L Spectrophotometer, using MD as reference, and expressed as per cent effectiveness, % E. All glassware was thoroughly cleaned by rinsing in hot water, followed by an acetone wash, and a final rinsing with hot water.
Results As a preliminary we tested different methods for mixing oil and dispersant, for the addition of this mixture to the test water, for dispersion, for sampling and measuring the amount of dispersed oil. Based on the results, we established the test procedure described in the Methods section. It was also important to keep the test conditions as close to the conditions of the toxicity test as possible. Since Chlamydomonas is a fresh water alga, the main testing of effective352
150
100
50
f. x I
I
213
411
I
I
88
EFFECTIVENESS,
Fig. 1 LCso values as function of effectiveness of diluted concentrates tested in algal medium.
ness was done in algal medium of 2 0 0 salinity. However, some compounds were also tested for effectiveness in artificial sea water at 330/0osalinity. For twelve of the concentrates tested in diluted form in algal medium, the effectiveness values used in the plot of Fig. 1 are averages of 2-5 separate measurements; the values for the other four represent single measurements. The standard deviation for the results of one compound tested five times was + 17 %, which might indicate the precision of the effectiveness measurements as a whole. All except one of the concentrates showed consistent effectiveness values when tested in diluted form in algal medium. The remaining compound, denoted A, showed rather high effectiveness in some tests and almost zero in the rest. Figure 1 is a plot of LCs0 values versus effectiveness in algal medium, and with both the low and high effectiveness values for compound A. The relationship between toxicity and effectiveness seems to be hyperbolic, as indicated with the drawn curve. Statistical correlation evaluation of log transformed values showed significant correlation at the 95% level. Thus one can conclude that high effectiveness is associated with high toxicity. In these experiments with algal medium the test conditions of the effectiveness test were closest to those of the toxicity test. From the plot it can be seen that the compound marked B clearly lies outside the drawn curve, and obviously conforms less to the general trend. It displays relatively low toxicity at relatively high effectiveness. Figure 2 shows a similar plot for the 10 solvent-based dispersants. A similar significant correlation (95°70 level) was also found for this group of compounds, but with none of the members deviating from the general trend. In addition to the experiments with diluted concentrates performed under conditions pertinent to the toxicity test, some concentrates were also tested (1) in the undiluted form with algal medium as dispersing liquid, (2) undiluted and (3) diluted, and with artificial sea water as dispersing liquid. The results of these experiments corroborated those described in Figs 1 and 2, also with respect to B, deviating under all three testing conditions. The highest correlation between toxicity and effectiveness was found under conditions (1) and (2). It is important to note that the correlations are all done with the same set of LCs0 values. Obviously we could not do toxicity tests with the fresh-water alga in sea water.
Volume 13/Number 10/October 1982
400
r: I
0 13
20
40
60
ill2
EFFECTIVENESS,
Fig. 2 LCs0 values as function of effectiveness of solvent-based dispersants tested in algal medium.
Discussion The effectiveness of an oil dispersant does not depend only on its own properties, but also on a series of chemical and physical factors which characterize the environment, and the oil which it is applied to. Some of the factors defining the environment or the field conditions, such as temperature and salinity, and others, such as oil type and dispersant/oil ratio, can be controlled in the laboratory. Others, like wave and wind action, and water turbulence, can hardly be simulated and controlled for testing purposes. The testing apparatus and procedures also create physical conditions which have no parallels under natural conditions. The choice of testing conditions with respect to physical and chemical factors and testing procedures will therefore to a great extent determine the outcome of the test. We aimed at an effectiveness test with conditions not very different from those of the toxicity test. Whilst the toxicity test was done at 35°C, 20000 lux, 2%0 salinity, and with aeration for 10 s every min for 4 h, the effectiveness test was performed at 20°C, room illumination, 2 and 33%0 salinity, and with circular movements at 160 rpm for 10 min. Since effectiveness data were not available in the literature, we had to measure effectiveness for the dispersants in question. In accordance with the procedure for the toxicity test the dispersants and oil were also mixed before their application to the water. The preliminary experiments clearly showed that the mode of application of the mixture to the water influenced the results of the effectiveness test. This influence was strong for some
compounds whereas others were little affected. The simplest and most reproducible method was found to be a gentle addition down the vessel wall. The samples of dispersed oil were taken from analysis 10 min after shaking was stopped, and from near the bottom. They are therefore representing a rather stable oilin-water emulsion consisting of small oil droplets. Thus dispersion effectiveness measured in this test mainly refers to the ability of the dispersants to form such stable emulsions. Comparing toxicity data with dispersion effectiveness, the clearest relationship was found with effectiveness data from the experiments performed under conditions being closest to those of the toxicity test. This holds both for concentrates and solvent-based dispersants. It is important to note that our studies with concentrates indicate that a similar correlation exists with effectiveness in sea water. This indication stems from comparison of toxicity data obtained from tests on fresh water with effectiveness data from tests in sea water. However, we have unpublished data of toxicity gained from experiments with the green marine flagellate Dunaliella bioculata supporting the indication: The LCs0 values obtained with this alga are very similar to those found with Chlamydomonas. The main conclusion is that there is a clear association between effectiveness and toxicity under the present test conditions, such that with increasing effectiveness the toxicity increases for concentrates as well as solvent-based dispersants. The results indicate two exceptions from this relationship among the concentrates, the compounds in question being clearly less toxic than should be expected from their effectiveness. The general validity of these observations will be further explored in experiments with a series of other oils, other dispersants and algae.
We wish to thank The Norwegian State Oil Company, The Norwegian Research Council for Scientific and Industrial Research, and the Norwegian Fisheries Research Council for financial support, and Miss Siri Ottesen for skilled technical assistance.
Anon (1980). Norweigna Environmental Protection Agency. Forskrifter for sammensetting og bruk av oljedispergeringsmidler for bekjempelse av oljes¢l. Oslo. Heldal, M., Norland, S., Lien, T. & Knutsen, G. (1977). Acute toxicity of several oil dispersants towards the green algae Chlamydomonas and Dunaliella. Chemosphere, 6, 247-255. Martinelli, F. N. & Cormac, D. (1979). Investigation of the effects of oil viscosity and water-in-oil emulsion formation on dispersant efficiency. Report from Warren Spring Laboratory, Department of Industry, Stevenage, England.
353