Ecotoxicity of glyphosate and aterbane ® br surfactant on guaru ( Phalloceros caudimaculatus )

Aquatic macrophytes are important components of aquatic ecosystems, but these plants have become a problem due to their occurrence in different regions. Some studies aimed to demonstrate the effectiveness of herbicides to control these macrophytes; however, few studies report the possible ecotoxicological effects. The objective of this study was to estimate the acute toxicity (LC (I)50;96h) and assess water quality variables for glyphosate in the Rodeo formulation, Aterbane BR surfactant and mixtures of glyphosate + 0.5% and 1.0% of surfactant, for the guaru fish (Phallocerus caudimaculatus). The guaru was exposed to increasing concentrations of glyphosate and a mixture of glyphosate + 0.5 and 1.0% of surfactant. The mixture of glyphosate and glyphosate + 0.5 and 1.0% of surfactant showed (LC (I)50;96h) > 975.0 mg L. For the surfactant, the rate was 5.81 mg L. The glyphosate and mixtures of glyphosate + 0.5% and 1.0% of surfactant caused a decrease in pH and dissolved oxygen and increased the electrical conductivity of water. Glyphosate in the Rodeo formulation and the mixtures with surfactant Aterbane BR can be classified as practically nontoxic, whereas surfactant Aterbane BR can be considered as moderately toxic to guaru.


Introduction
Aquatic ecosystems are formed by a great variety of aquatic macrophytes.However, certain human activities can cause alterations in water bodies and promote the development of dense and extensive monospecific populations, which may cause negative impacts on the environment, interfering with the multiples uses of water (CHAMBERS et al., 2008).Among the possible actions to control these macrophytes is the use of chemical control products with herbicidal activity, which has provided satisfactory control performance, cost-effective, low environmental impacts and with a history of use in other countries (MARCONDES et al., 2003).
Currently, to use these products in the aquatic environment it is necessary to perform ecotoxicological studies for fish and other non-target organisms, to assess the possible negative impacts.Therefore, the choice of test organisms must follow certain criteria: representation of an ecological taxonomic group, easy availability to conduct a test, known information of the biology of the species (RAND; PETROCELLI, 1985) and, whenever possible, belonging to native species (ARAUCO et al., 2005 The general objective of a toxicity test is the characterization of a biological answer to an environmental sample, substance or a chemical product (CAIRNS Jr. et al., 1998).The toxicity of gliphosate in the Roundup ® formulation was assessed for piaçu (Leporinus macrocephalus) (ALBINATI et al., 2007), curimbatá (Prochilodus lineatus) (LANGIANO; MARTINEZ, 2008), and fingerlings of jundiá (Rhamdia quelen) (KREUTZ et al., 2008).
One of the problems of the use of herbicides to control macrophytes is the effect of these variables on water quality.The herbicide groups act differently and specifically in water, varying according to their molecular composition.
Furthermore, the surfactants used in the composition of commercial formulations of herbicides, for improved efficacy in control of aquatic macrophytes (DIAMOND; DURKIN, 1997), can be more toxic to fish, algae, bacteria, protozoa and crustaceans than the herbicide itself (TSUI; CHU, 2003).
Therefore, the herbicide Rodeo  , with the active ingredient glyphosate, has been used to control floating aquatic macrophytes for its broad spectrum of action, application in post-emergence, systemic action, ease of application and excellent cost/benefit relationship.
The objective of this study was to determine the acute toxicity (LC (I)50;96h) and assess the variables of water quality during the test of glyphosate in the Rodeo ® formulation, of Aterbane ® BR surfactant and the mixtures of glyphosate + 0.5 and 1.0% of Aterbane ® BR surfactant for the guaru fish (Phallocerus caudimaculatus).The assessment of the mixtures was performed to simulate the real toxicity that may occur during the application of the herbicide on an aquatic environment.

Material and methods
This work was performed at São Paulo State University, Unesp, in the Faculty of Agriculture and Veterinary Sciences, Center for Environmental Studies and Research in Weed Science, Nepeam.
The products used in this work were: herbicide Rodeo ® , with active ingredient glyphosate on the concentration of 480 g L -1 , belonging to toxicity class IV, considered low toxic, and to environmental class III, dangerous product, and Aterbane ® BR surfactant (mix alkyl polyglycol ether nonionic surfactant), in the concentration of 466 g L -1 , belonging to toxicity class IV, considered low toxic.
For the beginning of the experiments, the fish were previously acclimatized in a living-bioassay, with controlled temperature to 26 ± 2°C and photoperiod of 12 hours of light, in a box with a capacity of 250 L, with aeration and continuous water flow, for ten days.
In this period, the animals were fed once a day, ad libitum, with commercial feed.
For sensory control of the organisms, acute toxicity tests were performed periodically using a reference substance with 96h of duration.The reference substance used was potassium chloride (Pro-analysis), with a 99.9% level of purity (ABNT, 2004).The weight of organisms used ranged from 0.70 to 1.50 g.Three experiments were conducted with six concentrations (0. 0, 0.5, 1.0, 1.5, 2.0 and 2.5 g L -1 ), three replicas and five fishes per replication.To the guaru, the lethal concentration of 50% (LC (I)50;96h), average estimate of potassium chloride, was 1.67 ± 0.06 g L -1 , with upper limit of 1.94 ± 0.03 g L -1 and lower limit of 1.45 ± 0.11 g L -1 .
The water used in the tests was from a semi artesian well and the variables of water quality were maintained in: pH 7.4 ± 0.2, dissolved oxygen between 6 and 8 mg L -1 , electric conductivity around 170 μS cm -1 , hardness between 50 and 60 mg L -1 in CaCO 3 and water temperature between 25 and 26ºC, according to ABNT (2004).
To estimate the lethal concentration 50% (LC (I)50;96h) of glyphosate, of Aterbane ® BR and from the mixture of glyphosate + 0.5 and 1.0% of Aterbane ® BR, experiments were performed in a completely randomized design.
Assessment of the mortality was performed daily with the removal of dead fish from the experimental units.The variables of water quality, pH, dissolved oxygen and electrical conductivity were measured at 24, 48, 72 and 96 hours.The values of LC 50;96h were calculated using the Trimmed Spearman-Karber method (HAMILTON et al., 1977).

Results and discussion
During the 96 hours of exposition to glyphosate, on the formulation Rodeo ® , and to the mixtures of 0.5 and 1.0% of the surfactant, the lethal concentration estimated for guaru (LC (I)50;96h) was > 975 mg L -1 (Table 1).
Table 1.Acute toxicity of the glyphosate and of the surfactant Aterbane ® BR and their mixtures to guaru.
The toxicity of glyphosate, in the Rodeo ® formulation, and the mixture of glyphosate + 0.5% of surfactant Aterbane ® BR were similar to the formulation Rodeo ® with the surfactant X-77 to Salmo gairdneri and Oncorhynchus tshawytscha with (LC 50;96h) of 1070 and 1440 mg L -1 (MITCHELL et al., 1987); to Hybognathus amarus and to Pimephales promelas with (LC 50;96h) higher than 1000 mg L -1 (BEYERS, 1995).The Glyphosate is less toxic than Diquat for tilapia (Oreochromis niloticus) with LC (I) (50;96h) of 37.28 mg L -1 , with lower limits of 33.12 mg L -1 and upper limits of 41.44 mg L -1 , other herbicide that could be used to control macrophytes (HENARES et al. 2008).
The addition of the surfactant in the formulation did not alter the toxicity of glyphosate.The recommended agricultural dose was used, which is 0.5% of surfactant and one concentration above 1.0%.Higher concentrations were not tested, since they are not normally used, but such concentrations could come forward with toxicity.
During tests of acute toxicity of glyphosate in the Rodeo ® formulation, no change was observed in the pH of water used in acute toxicity tests (Table 2).
For the electrical conductivity of water during the tests of acute toxicity of glyphosate, considerable variations were observed when compared to the control treatment.However, there were no considerable variations among the treatments (Table 2).The presence of glyphosate in the formulation Rodeo ® formulation in water altered the pattern of electrical conductivity of water during the acute toxicity tests, providing more ions dissolved in water.
During the tests of acute toxicity of the surfactant ® for guaru (P.caudimaculatus) the pH, dissolved oxygen and electrical conductivity of the water did not differ between treatments (Table 3), remaining within the limits established by Ibama (1987) and Usepa (2002).
The values of pH of the water tests of acute toxicity of glyphosate + 0.5% Aterbane ® BR decreased between the treatments over time of exposure and response to treatment with 0.0 mg L -1 (Table 4).
During tests of acute toxicity of the mixture of glyphosate + 0.5% of Aterbane ® BR was marked decrease in levels of dissolved oxygen in water from 48 hours of exposure in all treatments compared to treatment with 0.0 mg L -1 (Table 3).These values are below the limit of 4 mg L -1 established by Ibama (1987) and the Usepa (2002) for acute toxicity test with fish.
In acute toxicity test with glyphosate + 0.5% surfactant, the values of electrical conductivity of water increased in relation to treatment with 0.0 mg L -1 (Table 4).
In tests of acute toxicity of glyphosate + 1.0% Aterbane ® BR surfactant, pH, dissolved oxygen and electrical conductivity of water were similar to the test with a mixture of glyphosate + 0.5% surfactant (Table 5).Glyphosate in the Rodeo ® formulation and mixtures of glyphosate + 0.5, 1.0% of Aterbane ® BR exhibited similar pattern of change during the acute toxicity tests for guaru, indicating that the addition of surfactant Aterbane ® BR does not contribute to change in water quality.These two formulations have caused decrease in pH and dissolved oxygen and increase the electrical conductivity of water during the acute toxicity tests for the species studied.

Conclusion
Glyphosate in the Rodeo ® formulation and mix with surfactant Aterbane ® BR can be considered practically non-toxic to guaru, while the surfactant Aterbane ® BR as moderately toxic.Glyphosate and mix with surfactant changed electrical conductivity and dissolved oxygen from the water.

Table 2 .
Average ± standard desviation of pH, dissolved oxygen (mg L -1 ) and electrical conductivity (μS cm -1 ) of water during the test of acute toxicity of glyphosate.

Table 3 .
Average ± standard deviation of pH, dissolved oxygen (mg L -1 ) and electrical conductivity (μS cm -1 ) of water during the test of acute toxicity of the surfactant.Tabela 4. Average ± standard deviation of pH, dissolved oxygen (mg L -1 ) and electrical conductivity (μS cm -1 ) of water during the test of acute toxicity of glyphosate + 0.5% Aterbane ® BR.