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Investigation of acute toxicity of chlorpyrifos-methyl on guppy Poecilia reticulata
Chemosphere 60 (2005) 93–96 www.elsevier.com/locate/chemosphere
Investigation of acute toxicity of chlorpyrifos-methyl on guppy Poecilia reticulata Mahmut Selvi, Rabia Sarıkaya, Figen Erkoc¸ *, Oner Koc¸ak
1
Department of Biology Education, Gazi University, Teknikokullar, 06500 Ankara, Turkey Received 3 May 2004; received in revised form 19 August 2004; accepted 29 November 2004 Available online 26 January 2005
Abstract Static bioassays were made to determine acute toxicity of chlorpyrifos-methyl, a wide spectrum organophosphorus insecticide and potential toxic pollutant of aquatic ecosystem, Guppy fish (Poecilia reticulata). Bioassays were made at a regulated temperature of 22 ± 1 °C and were repeated three times. Lethal doses of the insecticides were determined using LC50 software programme of U.S. EPA based on FinneyÕs Probit Analysis statistical method. The 96 h LC50 value and 95% confidence limit of chlorpyrifos-methyl for Guppy was estimated as 1.79 (1.47–2.10) mg/l. The fish exposed to chlorpyrifos-methyl exhibited behavioral changes in the form of neurotoxin toxicity: less general activity than control group, loss of equilibrium, erratic swimming and staying motionless at a certain location generally at mid-water level for prolonged periods. The 1 mg/l (lowest) concentration had similar behavior (NOEC) with the control group. Ó 2005 Elsevier Ltd. All rights reserved. Keywords: Chlorpyrifos-methyl; Acute toxicity; Guppy; Poecilia reticulata; Bioassay; FQPA; Behavioral effects
1. Introduction Chlorpyrifos-methyl [CAS 5598-13-0, O,O-dimethyl O-(3,5,6-trichloro-2-pyridinyl) phosphorothioate, formula: C7H7Cl3NO3PS] is a wide spectrum insecticide of the organophosphorus group (URL 1, URL 2). It is poorly soluble in water (4 mg/l), readily soluble in organic solvents such as acetone, benzene and chloroform and stable under normal storage conditions. It is hydrolyzed more rapidly at higher pH and undergoes rapid photodecomposition in UV light. The insecticide is mainly effective against rice stem borer, aphids, cutworms, plant * Corresponding author. Tel.: +90 312 2126470; fax: +90 312 2228483. E-mail address: [email protected] (F. Erkoc¸). 1 Insecticide Test Laboratory, Hacettepe University, Beytepe, Ankara, Turkey.
and leaf hoppers, mole crickets and some moths; stored grain pests. Present main use is on stored grain. Mode of action is cholinesterase inhibition (Boone and Chambers, 1997; Levin et al., 2003). Maximum residue limits of the insecticide have been recommended by the joint FAO/WHO Meeting on Pesticide Residues (URL 2). Chlorpyrifos-methyl was initially registered in 1985 but is being evaluated with other organophosphate pesticides under FQPA Tolerance Reassessment Progress and Interim Risk Management Decision (TRED) by the United States Environmental Protection Agency (U.S. EPA, 2001, 2002). The significant data gaps including developmental neurotoxicity studies are required for all organophosphate pesticides under FQPA to evaluate their safety to children, and chemical-specific occupational exposure studies. Registrants of chlorpyrifos-methyl (Dow Agrosciences) have requested voluntary cancellation of their products rather than develop the additional
0045-6535/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.chemosphere.2004.11.093
94
M. Selvi et al. / Chemosphere 60 (2005) 93–96
data requested by EPA to complete the toxicology data base. EPA will issue its final tolerance reassessment decision for chlorpyrifos-methyl as soon as the cumulative risk assessment for the insecticide is completed. Present international regulatory status is: ‘‘Not Listed’’ under UNEP Persistent Organic Pollutant, UNEP Prior Informed Consent Chemical, WHO Obsolete Pesticide lists and ‘‘Unlikely to be Hazardous’’ under WHO Acute Hazard Ranking where the parent chemical chlorpyrifos is ‘‘II, Moderately Hazardous’’ (URL 3). While EPA and Dow are discussing the phase-out process and alternatives to chlorpyrifos-methyl, for now, it can still be used on stored barley, oats, rice, sorghum and wheat (URL 4; URL 5). Hazard characterization and human health risk assessment including dietary risk can be found in the EPA web sites (URL 6 and URL 7). Recently in a review on the microbiological, biological, and chemical weapons of warfare and terrorism, Greenfield et al. (2002) estimated lethal dose of the parent compound chlorpyrifos liquidas approximately 15 000 000 lg/person, drawing attention to health risks of this group of agricultural insecticides. It is toxic to non-target organisms such as shrimps, crabs, fish (96 h LC50 for rainbow trout: 12.6 ppb) and Daphnia (48 h LC50 for daphnia: 1.11 ppb) (URL 8). Although most of the information on possible ecological risk of chlorpyrifos exist in the open literature, there is very little published on the aquatic toxicity of chlorpyrifos-methyl. The Columbia Environmental Research Center Acute Toxicity Database (URL 9) reports 96 h chlorpyrifos-methyl toxicity to various standard fish test species in static test system as: Rainbow trout (1.40 g) technical grade 301 lg/l at 13 °C Brook trout (1.15 g) technical grade 200 lg/l at 12 °C Brook trout (1.15 g) 61% emulsifiable concentrate 100 lg/l at 12 °C Fathead minnow (0.90 g) technical grade 678 lg/l at 18 °C. The widespread use of these pesticides consequently leads to the exposure of manufacturing workers, field applicators, the ecosystem and finally the public to the possible toxic effects of these pesticides. The aim of this study is to provide fish acute toxicity data for chlorpyrifos-methyl to contribute to the significant ecological risk data gap. Therefore, static bioassays were made to determine LC50 value of chlorpyrifos-methyl to Guppy (Poecilia reticulata).
2. Materials and methods Male, adult specimens of Guppy were obtained from a local breeder in Ankara and brought to the laboratory within 30 min in plastic bags with sufficient air. The plas-
tic bags were placed into the maintenance aquarium for about 30–35 min for acclimatization. Then the bags were cut open and the fish were allowed to swim into the aquarium water. Test chambers were glass aquaria of about 25 l capacity. Temperature was regulated at 22 ± 1 °C by using heaters. At the time of dosing air was turned off; it was on at all times otherwise. The water was continuously aerated for several days before putting the fish in, to remove chlorine. Test chambers were filled with 20 l of tap water. Characteristics of this aquarium water were as follows; temperature 22 ± 1 °C, dissolved oxygen 6.3–6.6 mg/l and conductivity 0.189–0.207 mS, French hardness 21– 24 FS°, total ammonia 0.017–0.021 mg/l, nitrite 0.008– 0.009 mg/l, nitrate 0.11–0.16 mg/l. Technical grade (92%) chlorpyrifos-methyl was obtained from the Insecticide Testing Laboratory of Hacettepe University, Ankara (source: Shenzen Co. Ltd., China) and was stored at +4 °C until stock solution preparation. Stock solution was prepared by bringing chlorpyrifos-methyl to room temperature then weighing a certain amount and diluting it in acetone to give the stock material. Dosing solutions were prepared from this stock by diluting with acetone to give the dosing concentrations of 1, 1.5, 2, 2.5, 3 mg/l. Groups of experimental animals, each consisting of 10 individuals, were selected at random and placed into aerated aquaria. After 48 h of adaptation, the different concentrations of chlorpyrifos-methyl in acetone were added to the experimental aquaria. During the last 24 h of adaptation, and throughout the duration of the experiment, animals were not fed. Mortality was assessed at 24, 48, 72 and 96 h after the start of the tests. Dead individuals were removed immediately. Behavioral changes were followed closely. The dosing volume never exceeded 0.2 ml. Control group received acetone at the maximum acetone volume used in the dilution of the dosing concentrations. Each bioassay was repeated three times. The bioassay system was as described in standardized methods (OECD, 1993; APHA, 1998) and the national regulation (Turkish Official Gazette, 1991). The selected species is also as recommended in these references. LC50 and 95% confidence limits were calculated by a computer program (U.S. EPA, 1999).
3. Results The calculated 96 h acute LC50 value (95% confidence limits) of technical chlorpyrifos-methyl, dissolved in acetone, using a static bioassay system to adult, male Guppy Poecilia reticulata was 1.79 mg/l (1.47–2.10). Control mortality was zero. Results are in Table 1 and Fig. 1.
M. Selvi et al. / Chemosphere 60 (2005) 93–96
95
Table 1 Acute 96 h toxicity of technical chlorpyrifos-methyl in adult male Guppy (Poecilia reticulata) Point
Concentration (mg/l)
95% Confidence limits
Slope ± SE
Intercept ± SE
LC LC LC LC LC LC LC LC LC
0.76 0.98 1.12 1.22 1.79 2.62 2.86 3.27 4.20
0.33–1.05 0.52–1.26 0.67–1.39 0.78–1.48 1.47–2.10 2.21–3.72 2.38–4.36 2.63–5.54 3.16–8.78
6.28 ± 1.51
3.41 ± 0.47
1.00 5.00 10.00 15.00 50.00 85.00 90.00 95.00 99.00
Note: Control group (theoretical spontaneous response rate) = 0.0000.
Observations of behavioral response of Guppy were conducted at 1–8, and every 12 h during the acute toxicity tests. The control group showed normal behavior during the test period. The changes in behavioral response started 1 h after dosing. The 1 mg/l (lowest) concentration had similar behavior with the control group. Observed behavioral changes/effects were typical of neurotoxin toxicity: less general activity than control group, loss of equilibrium, erratic swimming and staying motionless at a certain location generally at mid-water level for prolonged periods. Fish exposed to 1.50 mg/l
showed less general activity with occasional loss of equilibrium. Loss of equilibrium become more frequent in the 2 mg/l concentration. The 2.50 mg/l concentration group stayed motionless close to the water surface and later fell to the aquarium bottom in an uncontrolled manner. The highest concentration of 3 mg/l showed all responses at high intensities: the loss of equilibrium, hanging vertically in water, rapid gill movement, erratic swimming, sudden swimming motion in a spiral fashion, after long periods of motionlessness lying down on the aquarium bottom and suddenly starting to move.
Fig. 1. Plot of adjusted probits and predicted regression line for chlorpyrifos-methyl to Guppies (P. reticulata).
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M. Selvi et al. / Chemosphere 60 (2005) 93–96
4. Discussion The results show that chlorpyrifos-methyl is less toxic to Guppy than to most other species. A range of 100– 678 lg/l have been reported for other standard test species (URL 9). It is interesting to note that only a few studies on the acute toxicity of one of the most aquatic toxic organophosphates, namely chlorpyrifos-methyl, to fish exist in the open literature. Chlorpyrifos-methyl is a highly toxic insecticide widely used in agriculture. Here special attention is drawn to itsÕ heavy use in grain storage and other aquatic uses which necessitate in-depth sub-chronic and chronic toxicity tests to fish species and to non-target species to be undertaken. In addition, potential risk from chlorpyrifos-methyl metabolites should be investigated to get a more complete picture in terms of toxicity. The low toxicity of chlorpyrifos-methyl to mammals may be misleading at this point in terms of ecotoxicology and lead to extrapolation problems to aquatic species. Delistraty (2000) in the study of examining relationships among physicochemical properties and acute toxicity endpoints of 231 chemicals in rats and trout concluded that; trout aquatic LC50 was predicted from rat inhalation LC50 with moderate success. Deener et al. (1999) reported log Kow (log of octanol–water partition coefficient) of chlorpyrifos-methyl as 4.37 ± 0.01 and lethal body burdens in Guppy of 9.9 ± 5.6 for low exposure and 7.8 ± 3.6 lmol/g for high exposure levels. Chlorpyrifos-methyl was in the highest lethal body burden group among the 13 organophosphorus chemicals tested. They concluded that this situation may be indicative of less formation of toxic metabolite. Therefore such data are useful in ecological risk assessment but there are limitations and uncertainties. 5. Conclusion Further work with toxicity testing methods directly on fish will be very useful in assessing possible ecological risk assessment of these pesticides. To overcome discrepancies and potential synergistic effects from the components of chlorpyrifos-methyl formulations, toxicity tests with formulations must be included together with active ingredient tests. Using only the active ingredient in the tests may be misleading. Special attention is drawn to the registration status of chlorpyrifos-methyl where availability of missing data is of significance. Using the lethal body burdens instead of LC50 of compounds does
not improve our understanding of the differences in their toxicity to fish.
References APHA, AWWA, WEF, 1998. Standard Methods for the Examination of Water and Wastewater, Washington, DC. Boone, J.S., Chambers, J.E., 1997. Biochemical factors contributing to toxicity differences among chlorpyrifos, parathion, and methyl parathion in mosquitofish (Gambusia affinis). Aquat. Toxicol. 39, 333–343. Deener, J.W., Budde, B.J., Weijers, A., 1999. Variations in the lethal body burdens of organophosphorus compounds in the guppy. Chemosphere 38, 1671–1683. Delistraty, D., 2000. Acute toxicity to rats and trout with a focus on inhalation and aquatic exposures. Ecotoxicol. Environ. Saf. 46, 225–233. Greenfield, R.A., Brown, B.R., Hutchins, J.B., Iandolo, J.J., Jackson, R., Slater, L.N., Bronze, M.S., 2002. Microbiological, biological, and chemical weapons of warfare and terrorism. Am. J. Med. Sci. 323, 326–340. Levin, E.D., Chrysanthis, E., Yacisin, K., Linney, E., 2003. Chlorpyrifos exposure of developing zebrafish: effects on survival and long-term effects on response latency and spatial discrimination. Neurotox. Teratol. 25, 51–57. OECD (Organization for Economic Co-operation and Development), 1993. OECD guidelines for testing of chemicals. OECD, Paris. Turkish Official Gazette (Resmi Gazete), 1991. Su Kirlilig˘i ve Kontrolu¨ Yo¨netmelig˘i Numune Alma ve Analiz Metodları Teblig˘i. Zehirlilik Seyreltme Fakto¨ru¨ (ZSF) Tayini. Tarih: 7.1.1991, Sayı: 20106. U.S. EPA, 1999. LC50 software program, version 1.00. Center for Exposure Assessment Modeling (CEAM) Distribution Center. U.S. EPA, 2001. Fed. Regist. 66, 51665–51666. U.S. EPA, 2002. Fed. Regist. 67, 20118–20121. URL 1: http://www.hclrss.demon.co.uk/chlorpyrifos-methyl. html URL 2: http://www.inchem.org/documents/pds/pds/pest33_24e. htm URL 3: http://www.pesticideinfo.org/Detail_ChemReg.jsp? Rec_Id=PC35494 URL 4: http://www.ipm.msu.edu/CAT00_field/FC08-03-00. htm URL 5: http://www.epa.gov/oppsrrd1/op/chlorpyrifos-methyl/ chlormeth_faq.htm URL 6: http://www.epa.gov/oppsrrd1/op/chlorpyrifos-methyl/ rev_toxicology.pdf URL 7: http://www.epa.gov/pesticides/op/chlorpyrifos-methyl/ overview.htm URL 8: http://epa.gov/tri/chemical/hazard_enviro95.pdf URL 9: http://www.cerc.cr.usgs.gov/data/acute/acute.html
Investigation of acute toxicity of chlorpyrifos-methyl on guppy Poecilia reticulata Mahmut Selvi, Rabia Sarıkaya, Figen Erkoc¸ *, Oner Koc¸ak
1
Department of Biology Education, Gazi University, Teknikokullar, 06500 Ankara, Turkey Received 3 May 2004; received in revised form 19 August 2004; accepted 29 November 2004 Available online 26 January 2005
Abstract Static bioassays were made to determine acute toxicity of chlorpyrifos-methyl, a wide spectrum organophosphorus insecticide and potential toxic pollutant of aquatic ecosystem, Guppy fish (Poecilia reticulata). Bioassays were made at a regulated temperature of 22 ± 1 °C and were repeated three times. Lethal doses of the insecticides were determined using LC50 software programme of U.S. EPA based on FinneyÕs Probit Analysis statistical method. The 96 h LC50 value and 95% confidence limit of chlorpyrifos-methyl for Guppy was estimated as 1.79 (1.47–2.10) mg/l. The fish exposed to chlorpyrifos-methyl exhibited behavioral changes in the form of neurotoxin toxicity: less general activity than control group, loss of equilibrium, erratic swimming and staying motionless at a certain location generally at mid-water level for prolonged periods. The 1 mg/l (lowest) concentration had similar behavior (NOEC) with the control group. Ó 2005 Elsevier Ltd. All rights reserved. Keywords: Chlorpyrifos-methyl; Acute toxicity; Guppy; Poecilia reticulata; Bioassay; FQPA; Behavioral effects
1. Introduction Chlorpyrifos-methyl [CAS 5598-13-0, O,O-dimethyl O-(3,5,6-trichloro-2-pyridinyl) phosphorothioate, formula: C7H7Cl3NO3PS] is a wide spectrum insecticide of the organophosphorus group (URL 1, URL 2). It is poorly soluble in water (4 mg/l), readily soluble in organic solvents such as acetone, benzene and chloroform and stable under normal storage conditions. It is hydrolyzed more rapidly at higher pH and undergoes rapid photodecomposition in UV light. The insecticide is mainly effective against rice stem borer, aphids, cutworms, plant * Corresponding author. Tel.: +90 312 2126470; fax: +90 312 2228483. E-mail address: [email protected] (F. Erkoc¸). 1 Insecticide Test Laboratory, Hacettepe University, Beytepe, Ankara, Turkey.
and leaf hoppers, mole crickets and some moths; stored grain pests. Present main use is on stored grain. Mode of action is cholinesterase inhibition (Boone and Chambers, 1997; Levin et al., 2003). Maximum residue limits of the insecticide have been recommended by the joint FAO/WHO Meeting on Pesticide Residues (URL 2). Chlorpyrifos-methyl was initially registered in 1985 but is being evaluated with other organophosphate pesticides under FQPA Tolerance Reassessment Progress and Interim Risk Management Decision (TRED) by the United States Environmental Protection Agency (U.S. EPA, 2001, 2002). The significant data gaps including developmental neurotoxicity studies are required for all organophosphate pesticides under FQPA to evaluate their safety to children, and chemical-specific occupational exposure studies. Registrants of chlorpyrifos-methyl (Dow Agrosciences) have requested voluntary cancellation of their products rather than develop the additional
0045-6535/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.chemosphere.2004.11.093
94
M. Selvi et al. / Chemosphere 60 (2005) 93–96
data requested by EPA to complete the toxicology data base. EPA will issue its final tolerance reassessment decision for chlorpyrifos-methyl as soon as the cumulative risk assessment for the insecticide is completed. Present international regulatory status is: ‘‘Not Listed’’ under UNEP Persistent Organic Pollutant, UNEP Prior Informed Consent Chemical, WHO Obsolete Pesticide lists and ‘‘Unlikely to be Hazardous’’ under WHO Acute Hazard Ranking where the parent chemical chlorpyrifos is ‘‘II, Moderately Hazardous’’ (URL 3). While EPA and Dow are discussing the phase-out process and alternatives to chlorpyrifos-methyl, for now, it can still be used on stored barley, oats, rice, sorghum and wheat (URL 4; URL 5). Hazard characterization and human health risk assessment including dietary risk can be found in the EPA web sites (URL 6 and URL 7). Recently in a review on the microbiological, biological, and chemical weapons of warfare and terrorism, Greenfield et al. (2002) estimated lethal dose of the parent compound chlorpyrifos liquidas approximately 15 000 000 lg/person, drawing attention to health risks of this group of agricultural insecticides. It is toxic to non-target organisms such as shrimps, crabs, fish (96 h LC50 for rainbow trout: 12.6 ppb) and Daphnia (48 h LC50 for daphnia: 1.11 ppb) (URL 8). Although most of the information on possible ecological risk of chlorpyrifos exist in the open literature, there is very little published on the aquatic toxicity of chlorpyrifos-methyl. The Columbia Environmental Research Center Acute Toxicity Database (URL 9) reports 96 h chlorpyrifos-methyl toxicity to various standard fish test species in static test system as: Rainbow trout (1.40 g) technical grade 301 lg/l at 13 °C Brook trout (1.15 g) technical grade 200 lg/l at 12 °C Brook trout (1.15 g) 61% emulsifiable concentrate 100 lg/l at 12 °C Fathead minnow (0.90 g) technical grade 678 lg/l at 18 °C. The widespread use of these pesticides consequently leads to the exposure of manufacturing workers, field applicators, the ecosystem and finally the public to the possible toxic effects of these pesticides. The aim of this study is to provide fish acute toxicity data for chlorpyrifos-methyl to contribute to the significant ecological risk data gap. Therefore, static bioassays were made to determine LC50 value of chlorpyrifos-methyl to Guppy (Poecilia reticulata).
2. Materials and methods Male, adult specimens of Guppy were obtained from a local breeder in Ankara and brought to the laboratory within 30 min in plastic bags with sufficient air. The plas-
tic bags were placed into the maintenance aquarium for about 30–35 min for acclimatization. Then the bags were cut open and the fish were allowed to swim into the aquarium water. Test chambers were glass aquaria of about 25 l capacity. Temperature was regulated at 22 ± 1 °C by using heaters. At the time of dosing air was turned off; it was on at all times otherwise. The water was continuously aerated for several days before putting the fish in, to remove chlorine. Test chambers were filled with 20 l of tap water. Characteristics of this aquarium water were as follows; temperature 22 ± 1 °C, dissolved oxygen 6.3–6.6 mg/l and conductivity 0.189–0.207 mS, French hardness 21– 24 FS°, total ammonia 0.017–0.021 mg/l, nitrite 0.008– 0.009 mg/l, nitrate 0.11–0.16 mg/l. Technical grade (92%) chlorpyrifos-methyl was obtained from the Insecticide Testing Laboratory of Hacettepe University, Ankara (source: Shenzen Co. Ltd., China) and was stored at +4 °C until stock solution preparation. Stock solution was prepared by bringing chlorpyrifos-methyl to room temperature then weighing a certain amount and diluting it in acetone to give the stock material. Dosing solutions were prepared from this stock by diluting with acetone to give the dosing concentrations of 1, 1.5, 2, 2.5, 3 mg/l. Groups of experimental animals, each consisting of 10 individuals, were selected at random and placed into aerated aquaria. After 48 h of adaptation, the different concentrations of chlorpyrifos-methyl in acetone were added to the experimental aquaria. During the last 24 h of adaptation, and throughout the duration of the experiment, animals were not fed. Mortality was assessed at 24, 48, 72 and 96 h after the start of the tests. Dead individuals were removed immediately. Behavioral changes were followed closely. The dosing volume never exceeded 0.2 ml. Control group received acetone at the maximum acetone volume used in the dilution of the dosing concentrations. Each bioassay was repeated three times. The bioassay system was as described in standardized methods (OECD, 1993; APHA, 1998) and the national regulation (Turkish Official Gazette, 1991). The selected species is also as recommended in these references. LC50 and 95% confidence limits were calculated by a computer program (U.S. EPA, 1999).
3. Results The calculated 96 h acute LC50 value (95% confidence limits) of technical chlorpyrifos-methyl, dissolved in acetone, using a static bioassay system to adult, male Guppy Poecilia reticulata was 1.79 mg/l (1.47–2.10). Control mortality was zero. Results are in Table 1 and Fig. 1.
M. Selvi et al. / Chemosphere 60 (2005) 93–96
95
Table 1 Acute 96 h toxicity of technical chlorpyrifos-methyl in adult male Guppy (Poecilia reticulata) Point
Concentration (mg/l)
95% Confidence limits
Slope ± SE
Intercept ± SE
LC LC LC LC LC LC LC LC LC
0.76 0.98 1.12 1.22 1.79 2.62 2.86 3.27 4.20
0.33–1.05 0.52–1.26 0.67–1.39 0.78–1.48 1.47–2.10 2.21–3.72 2.38–4.36 2.63–5.54 3.16–8.78
6.28 ± 1.51
3.41 ± 0.47
1.00 5.00 10.00 15.00 50.00 85.00 90.00 95.00 99.00
Note: Control group (theoretical spontaneous response rate) = 0.0000.
Observations of behavioral response of Guppy were conducted at 1–8, and every 12 h during the acute toxicity tests. The control group showed normal behavior during the test period. The changes in behavioral response started 1 h after dosing. The 1 mg/l (lowest) concentration had similar behavior with the control group. Observed behavioral changes/effects were typical of neurotoxin toxicity: less general activity than control group, loss of equilibrium, erratic swimming and staying motionless at a certain location generally at mid-water level for prolonged periods. Fish exposed to 1.50 mg/l
showed less general activity with occasional loss of equilibrium. Loss of equilibrium become more frequent in the 2 mg/l concentration. The 2.50 mg/l concentration group stayed motionless close to the water surface and later fell to the aquarium bottom in an uncontrolled manner. The highest concentration of 3 mg/l showed all responses at high intensities: the loss of equilibrium, hanging vertically in water, rapid gill movement, erratic swimming, sudden swimming motion in a spiral fashion, after long periods of motionlessness lying down on the aquarium bottom and suddenly starting to move.
Fig. 1. Plot of adjusted probits and predicted regression line for chlorpyrifos-methyl to Guppies (P. reticulata).
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M. Selvi et al. / Chemosphere 60 (2005) 93–96
4. Discussion The results show that chlorpyrifos-methyl is less toxic to Guppy than to most other species. A range of 100– 678 lg/l have been reported for other standard test species (URL 9). It is interesting to note that only a few studies on the acute toxicity of one of the most aquatic toxic organophosphates, namely chlorpyrifos-methyl, to fish exist in the open literature. Chlorpyrifos-methyl is a highly toxic insecticide widely used in agriculture. Here special attention is drawn to itsÕ heavy use in grain storage and other aquatic uses which necessitate in-depth sub-chronic and chronic toxicity tests to fish species and to non-target species to be undertaken. In addition, potential risk from chlorpyrifos-methyl metabolites should be investigated to get a more complete picture in terms of toxicity. The low toxicity of chlorpyrifos-methyl to mammals may be misleading at this point in terms of ecotoxicology and lead to extrapolation problems to aquatic species. Delistraty (2000) in the study of examining relationships among physicochemical properties and acute toxicity endpoints of 231 chemicals in rats and trout concluded that; trout aquatic LC50 was predicted from rat inhalation LC50 with moderate success. Deener et al. (1999) reported log Kow (log of octanol–water partition coefficient) of chlorpyrifos-methyl as 4.37 ± 0.01 and lethal body burdens in Guppy of 9.9 ± 5.6 for low exposure and 7.8 ± 3.6 lmol/g for high exposure levels. Chlorpyrifos-methyl was in the highest lethal body burden group among the 13 organophosphorus chemicals tested. They concluded that this situation may be indicative of less formation of toxic metabolite. Therefore such data are useful in ecological risk assessment but there are limitations and uncertainties. 5. Conclusion Further work with toxicity testing methods directly on fish will be very useful in assessing possible ecological risk assessment of these pesticides. To overcome discrepancies and potential synergistic effects from the components of chlorpyrifos-methyl formulations, toxicity tests with formulations must be included together with active ingredient tests. Using only the active ingredient in the tests may be misleading. Special attention is drawn to the registration status of chlorpyrifos-methyl where availability of missing data is of significance. Using the lethal body burdens instead of LC50 of compounds does
not improve our understanding of the differences in their toxicity to fish.
References APHA, AWWA, WEF, 1998. Standard Methods for the Examination of Water and Wastewater, Washington, DC. Boone, J.S., Chambers, J.E., 1997. Biochemical factors contributing to toxicity differences among chlorpyrifos, parathion, and methyl parathion in mosquitofish (Gambusia affinis). Aquat. Toxicol. 39, 333–343. Deener, J.W., Budde, B.J., Weijers, A., 1999. Variations in the lethal body burdens of organophosphorus compounds in the guppy. Chemosphere 38, 1671–1683. Delistraty, D., 2000. Acute toxicity to rats and trout with a focus on inhalation and aquatic exposures. Ecotoxicol. Environ. Saf. 46, 225–233. Greenfield, R.A., Brown, B.R., Hutchins, J.B., Iandolo, J.J., Jackson, R., Slater, L.N., Bronze, M.S., 2002. Microbiological, biological, and chemical weapons of warfare and terrorism. Am. J. Med. Sci. 323, 326–340. Levin, E.D., Chrysanthis, E., Yacisin, K., Linney, E., 2003. Chlorpyrifos exposure of developing zebrafish: effects on survival and long-term effects on response latency and spatial discrimination. Neurotox. Teratol. 25, 51–57. OECD (Organization for Economic Co-operation and Development), 1993. OECD guidelines for testing of chemicals. OECD, Paris. Turkish Official Gazette (Resmi Gazete), 1991. Su Kirlilig˘i ve Kontrolu¨ Yo¨netmelig˘i Numune Alma ve Analiz Metodları Teblig˘i. Zehirlilik Seyreltme Fakto¨ru¨ (ZSF) Tayini. Tarih: 7.1.1991, Sayı: 20106. U.S. EPA, 1999. LC50 software program, version 1.00. Center for Exposure Assessment Modeling (CEAM) Distribution Center. U.S. EPA, 2001. Fed. Regist. 66, 51665–51666. U.S. EPA, 2002. Fed. Regist. 67, 20118–20121. URL 1: http://www.hclrss.demon.co.uk/chlorpyrifos-methyl. html URL 2: http://www.inchem.org/documents/pds/pds/pest33_24e. htm URL 3: http://www.pesticideinfo.org/Detail_ChemReg.jsp? Rec_Id=PC35494 URL 4: http://www.ipm.msu.edu/CAT00_field/FC08-03-00. htm URL 5: http://www.epa.gov/oppsrrd1/op/chlorpyrifos-methyl/ chlormeth_faq.htm URL 6: http://www.epa.gov/oppsrrd1/op/chlorpyrifos-methyl/ rev_toxicology.pdf URL 7: http://www.epa.gov/pesticides/op/chlorpyrifos-methyl/ overview.htm URL 8: http://epa.gov/tri/chemical/hazard_enviro95.pdf URL 9: http://www.cerc.cr.usgs.gov/data/acute/acute.html