Registration Dossier

Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 27 March to 18 April 2006
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report Date:
2006

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Deviations:
no
Qualifier:
according to
Guideline:
other: JMAFF 2-7-2-1, Agrochemicaltest Guidelines 12 nohsan No. 8147. 24 november 2000, revised 26 June 2001. Daphnia immobilization.
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Product code: G30027
Other identification: Atrazine
Batch number: S03-2797
Content of active 96.2% wt/wt
ingredient:
Common name of ai: Atrazine
Appearance: White powder
Source: Syngenta Crop Protection Inc., Analytical and Product Chemistry Department, Greensboro, North Carolina, USA.
Reanalysis date: September 2006
Storage: <30°C
Stability: The test item was assumed to be stable for the period of use in the study, pending concurrent batch re-analysis.
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
not applicable

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
Quadruplicate samples were taken for analysis from the test concentration and the control at the start and end of the test.
At the start of the test, samples were taken from the excess test solutions after filling the test vessels. At the end of the test samples were taken from below the surface of the test solutions.
Sample volumes were 10 ml for the exposure concentration and 30ml for the control.
Samples were analyzed according to procedures documented in Syngenta analytical method ECOL06-00l.

Sub-samples were diluted 50:50 with acetonitrile prior to analysis. Analysis for atrazine was performed by high performance liquid chromatography with UV detection by diode array (HPLC-LTV).
Samples received at the 0 hour time point were processed on the day of sampling. Samples received at the 48 hour time point were stored for a maximum of 5 days at ca 7°C. Unprocessed fortified Elendt M7 medium samples were stored with the samples and processed alongside them.

Test solutions

Vehicle:
not specified
Details on test solutions:
A saturated solution of atrazine in Elendt M7 medium was prepared by adding 199.5 mg of the test item to 2 litres of M7 medium. This was then stirred for 15.5 hours, before being filtered through a 0.45 pm nitrocellulose membrane filter, the filtrate being used as the test solution. The appearance of this solution was clear and colourless.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Daphnia magna Straus 1820, Clone 5
Characteristics that make this test organism suitable for aquatic toxicity testing are its ease of culturing, its sensitivity to a variety of chemicals and the extensive database that exists for this common freshwater invertebrate species.
The test was started using first instar Daphnia (E24 hours old) obtained from in-house cultures. The Daphnia were cultured in the same medium and similar environment conditions to those used in the test. At test initiation, the animals were not from the first brood.
Daphnia magna for the start of the cultures were received on 15 September 2005 from Huntingdon Life Sciences (HLS), Huntingdon Research Centre, Woolley, Alconbury, Huntingdon, Cambridgeshire, PE28 4HS, UK.
The quality of the Daphnia in-house cultures are monitored at least once a year using potassium dichromate as a positive control. The most recent result of the positive control in July 2005 gave a 24 hour EC50 of 1.11 mg/L (Syngenta study number 2033102-A). This result was within the range of 0.6 - 2.1 mg/L as specified in the OECD 202

Study design

Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
48 h
Post exposure observation period:
no post exposure observation period

Test conditions

Hardness:
269.2 mg CaCo3/L
Test temperature:
18 - 22 °C
pH:
6.0 - 9.0
Dissolved oxygen:
ca 60% air saturation
Salinity:
The dilution water used in the test was Elendt M7 medium (M7-medium, OECD guideline 211, 1998). The following analytical grade salts and additives were added to deionised water to obtain the following nominal concentrations:
Macro Nutrients
CaCl x 2H20 293.8 mg/L
MgSO4 x 7H20 123.3 mgI
KCl 5.8 mg/L
NaHCO3 64.8 mg/L
Na2SiO3 x 9H20 10.0 nig/L
NaNO3 0.27 mgI
KH2PO4 0.14 mg/L
K2HPO4 0.18 mg/L

Trace Elements
H3BO3 715.0 µg/L
FeSO4 x 7H20 249.0 µg!L
MnCl2 x 24H20 90.0 µg /L
LiCl 77.0 µg /L
RbCl 18.0 µg /L
SrCI2 x 6H20 38.0 tg/L
NaBr 3.1 µg /L
NaMoO4 x 2H2O 16.0 µg /L
CuCl2 x 2H2O 4.0 µg /L
ZnCl2 13.0 µg /L
CoCl2 x 6H2O 10.0 µg /L
KI 3.3 µg /L
Na2SeO3 2.2 µg /L
NH4VO3 0.6 µg /L
Na2EDTA x 2H2O 625 µg /L

Vitamins
Thiamine HCl 75.0 µg /L
Cyanocobalamine (B12) 1.0 µg /L
Biotine 0.75 µg /L

Nominal and measured concentrations:
The test solution was prepared at the limit of aqueous solubility, as 100% saturated solution. the mean measured concentration of the 100% saturated solution was 29 mg/L at the start and end of the test. the limit of quantification (LOQ) in this study was 2.0 mg/L.
Details on test conditions:
The test consisted of two treatment groups: a dilution water control and one concentration. Four replicates test vassel were prepared for the test concentration. Four replicate test vessels were prepared for the test concentration and for the control. Five Daphnia were added to each test vessel, without conscious bias, as soon as possible after the preparation of the test solution.
The study was carried out in a temperature-controlled room. Throughout the exposure period room temperature was monitored by conditions: means of a data logger in a vessel containing deionised water.
Temperature, pH and dissolved oxygen were also recorded in the test solutions at the start and the end of the study. Hardness of the dilution water was measured prior to use in the test.
Required lighting regime: 16 hours light:8 hours dark photoperiod with 30 minute dawn and dusk transition periods (light intensity 500-1000 Lux)
Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Duration:
24 h
Dose descriptor:
LC50
Effect conc.:
> 29 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: n/a
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
> 29 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: n/a
Duration:
24 h
Dose descriptor:
NOEC
Effect conc.:
29 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: n/a
Duration:
48 h
Dose descriptor:
NOEC
Effect conc.:
29 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: n/a
Details on results:
As no toxic effects were observed the 48 hours EC50 is estimated to be more than 29 mg/L.
the 40 hours NOEC based on visual inspection of the data was 29 mg/L.
The pH of the test solution was 7.72 at 0 hours and 7.50 at 48 hours, while the concentration of dissolved oxygen ranged from 94 to 95% of the air saturation value.
The pH of the control was 7.72 at 0 hours and 7.47 at 48 hours, while the concentration of dissolved oxygen ranged from 98 to 101% of the air saturation value.
The temperature of the test solution at the start and the end of the exposure period was 20.8°C and 2 1.1°C respectively. The temperature of the control at the start and the end of the exposure period was 20.1°C and 21.2°C respectively. Temperature measured by means of a data logger, in a vessel containing deionised water, ranged from 20.2 to 20.8°C throughout the exposure period.
The light intensity was measured at 420 Lux. This is slightly below the minimum value given in the Study Plan.
Results with reference substance (positive control):
not applicable
Reported statistics and error estimates:
The median effect concentration (EC50) could not be determined at 24 or 48 hours from the data.
The NOEC was estimated based on visual observation of the data. No formal test of significance was appropriate for this study design.
All validity criteria were met. Control immobility at 48 hours was 0% (required = 10%). The dissolved oxygen concentration ranged from 94 to 101% saturation (required = 60%).

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
Control immobility must not exceed 10%. Dissolvedoxygen concentrations must be maintained at ca 60% saturation throughout the study.
Conclusions:
The 48 hour EC50 of atrazine was estimated to be more than 29 mg/L based on the mean measured concentration of the test material
The 48 hour NOEC was 29 mg/L based on visual inspection of the data.