1-[2-(allyloxy)ethyl-2-(2,4-dichlorophenyl)-1H-imidazolium hydrogen sulphate

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Data from the related substance imazalil base is used to cover this endpoint. The justification for read across is attached in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Key result

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

0.009 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

imazalil sulfate (recalculated from imazalil concentration)

Basis for effect:

reproduction

Reference 2

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27 August to 17 September 2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.20 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: ZR023979G3L431
- Expiration date of the lot/batch: 29-5-2011
- Purity test date: 06-09-2005

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: in original container, at room temperature (+10°C to +30°C) in the dark
- Stability under test conditions: The analytically determined mean test item concentrations in the analysed test media varied in the range from 90% to 114% of the nominal value.

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0.008, 0.025, 0.08, 0.25, 0.8 mg/L

- Sampling method:
* one sample from the freshly prepared stock solution
* duplicate samples from the freshly prepared test media of all test concentrations and the control at the first treatment (day 0) and at a treatment period in the second and third week (days 7 and 16)
* duplicate samples from the aged test media of all test concentrations and the control at day 2 and 9 (48h old) and at day 19 (72h old, because of the weekend)

The concentrations of the test item were measured in the duplicate test medium samples from the test concentrations.
From the control samples, only one of the duplicate samples was analysed from each sampling date.

- Sample storage conditions before analysis:
According to the study plan, samples were to be stored in a freezer (< -10°C) if the analysis directly after sampling is not possible. However, due to an error, the vials for (containing the prepared samples) were stored in a refrigerator (2-6°C) in the dark until measurement, whereas the sapre samples were kept frozen. No effect on the study is presumed, since the fortified samples were treated in the same way and there was no loss of imazalil during storage in the refrigerator.
Secondly, due to technical problems, the freezer temperature reached -10°C on some days (instead of the < -10°C) indicated in the study plan. However, only a slight deviation is presumed as an effect on the study.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution was prepared by dissolving 10 mg test item into 500 mL culture medium by intense stirring for 20 minutes and ultrasonic treatment for 10 minutes. Adequate volumes of this stock solution were dilute with culture medium to prepare the test media. Test concentrations were prepared just before introduction of the Daphnia (= start of the test).
- Controls: yes

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Strain/clone: clone 5
- Source: in-house laboratory culture
- Feeding during test: yes
- Food type: green algae (Desmodesmus subspicatus)
- Amount: The amount of food was based on the concentration of total organic carbon (TOC) in the algal food suspension. The fed daily amounts of algal TOC/Daphnia were as follows:
* days 0-4: 0.10 mg
* days 5-20: 0.15 mg
- Frequency: daily

ACCLIMATION
No acclimation necessary since breeding was performed in the same kind of test water.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

250 mg/L CaCO3

Test temperature:

20 to 21°c i the freshly prepared media
19 to 21°C in the aged media

pH:

7.9 to 8.2 in the freshly prepared media
7.6 to 8.3 in the aged media

Dissolved oxygen:

8.7 to 9.2 mg/L in the freshly prepared media
8.0 to 9.3 mg/L in the aged media

Salinity:

not applicable

Conductivity:

not applicable

Nominal and measured concentrations:

Nominal concentrations: 0.8, 0.25, 0.08, 0.025, 0.008 mg/L
Mean measured concentrations: 0.705, 0.229, 0.0716, 0.0278, * mg/L

* below LOQ

Details on test conditions:

TEST SYSTEM
- Test vessel: glass beaker
- Type (delete if not applicable): open / closed
- Material, size, headspace, fill volume: glass beakers of 100 mL volume with 80 mL test medium
- Renewal rate of test solution (frequency/flow rate): The test media of all test concentrations and of the control were renewed on days 2, 5, 7, 9, 12, 14, 16 and 19. At these dates, the test animals were carefully transferred by glass tubes from the old test vessels into the freshly prepared test media of the corresponding concentration.
- No. of organisms per vessel: 1 Daphnia per replicate in 80 mL test medium
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4
- Intervals of water quality measurement: Measurements were taken in all test concentrations and in the control at the start and end of each exposure period.

OTHER TEST CONDITIONS
- Photoperiod: 16h light, 8h dark
- Light intensity: 110 - 430 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
The mortality of the test animals and the number of young were recorded each working day. Dead animals and offspring were removed at the same times.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Range finding study: yes
- Test concentrations: not indicated
- Results used to determine the conditions for the definitive study: yes

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

0.25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

0.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Key result

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

0.025 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

0.08 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Key result

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

0.153 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Details on results:

- Other biological observations: With exception of the reported mortality and the reduced reproduction rates, no particular signs of intoxication were observed at the test animals during the tests.
- Mortality of control: 90% surviving on day 21
- Abnormal responses: none indicated
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none indicated
- Effect concentrations exceeding solubility of substance in test medium: no

Reported statistics and error estimates:

The NOEC and the LOEC for the reproduction rate were evaluated by Dunnett's Multiple t-test after analysis of variance (ANOVA). The EC50 (21d) of the reproduction rate was determined by Probit analysis.
The NOEC and LOEC for the mortality rate were evaluated by Multiple sequentially rejective U-test after Bonferroni-Holm.
The software used to perform the statistical analysis was ToxRat Professional, Version 2.09, ToxRat(R) Solutions GmbH, 2005.

Validity criteria fulfilled:

yes

Conclusions:

The influence of Imazalil on the survival and reproduction rate of Daphnia magna were determined during an exposure period of 21 days in a semi-static test according to the OECD guideline 211. The 21-day NOEC was determined to be 0.08 mg test item/L for the reproduction rate and 0.25 mg test item/L for the survival of the adult test animals, respectively. The EC50 for the reproduction rate was 0.153 mg test item/L. The results of the test can be considered reliable without restriction.

Reference 3

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: OECD 202: Daphnia sp., Acute Immobilisation Test and Reproduction Test

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: batch D7303
- Expiration date of the lot/batch: not indicated
- Purity test date: not indicated

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0.01, 0.03, 0.1, 0.3, 1, 3 mg imazalil / l
- Sampling method: At the start of the test and before and after each renewal, 20 ml samples of test medium were sampled from each beaker
- Sample storage conditions before analysis: samples were frozen for subsequent determination of the concentration of test substance

Vehicle:

no

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Source: stock culture held at own laboratory
- Age of parental stock (mean and range, SD): On the day preceding test initiation, daphnids aged about 3 weeks were sieved through a stainless steel sieve of 20 mesh in order to separate the offspring from the parents. Consequently, the offspring produced by the adults within the day preceding the test is guaranteed younger than 24h.
- Feeding during test: yes
- Food type: chlorella suspension (10E8 cells/ml) and tetrametrin suspenion (chemical oxygen demand 11.09 g O2/l)
- Amount and frequency:
* day 0 to 6: 0.64 ml Chlorella suspension per test vessel, 0.36 ml Tetrametrin suspension per test vessel
* day 7 to 9: 1.28 ml Chlorella suspension per test vessel, 0.36 ml Tetrametrin suspension per test vessel
* day 10 to 21: 1.92 ml Chlorella suspension per test vessel, 0.36 ml Tetrametrin suspension per test vessel

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

256.275 mg/l CaCO3 (mean value)

Test temperature:

19.33°C (mean value)

pH:

7.7 (mean value)

Dissolved oxygen:

102 %O2 (mean value)

Salinity:

not applicable

Conductivity:

not applicable

Nominal and measured concentrations:

Nominal concentrations: 0.01, 0.03, 0.1, 0.3, 1, 3 mg Imazalil /L
Mean measured concentration (day 0, start): 0.07, 0.023, 0.080, 0.262, 0.763, 2.481 mg imazalil /L
Mean measured concentration (day 3, after renewal): 0.007, 0.024, 0.075, 0.255, 0.741, 2.444 mg imazalil /L
Mean measured concentration (day 5, after renewal): 0.006, 0.025, 0.076, 0.233, -, 2.451 mg imazalil /L
Mean measured concentration (day 7, after renewal): 0.008, 0.025, 0.082, 0.254, -, - mg imazalil /L
Mean measured concentration (day 10, after renewal): 0.009, 0.033, 0.079, 0.243, -, - mg imazalil /L
Mean measured concentration (day 12, after renewal): 0.008, 0.025, 0.091, 0.260, -, - mg imazalil /L
Mean measured concentration (day 14, after renewal): 0.007, 0.024, 0.078, 0.236, -, - mg imazalil /L
Mean measured concentration (day 17, after renewal): 0.009, 0.025, 0.081, 0.242, -, - mg imazalil /L
Mean measured concentration (day 19, after renewal): 0.005, 0.029, 0.078, 0.264, -, - mg imazalil /L
Mean measured concentration (day 21, mean end): 0.004, 0.025, 0.069, 0.253, -, - mg imazalil /L

Details on test conditions:

TEST SYSTEM
- Test vessel: beakers
- Material, size, headspace, fill volume: all glass beakers of 600 ml filled with 400 ml test medium
- Renewal rate of test solution (frequency/flow rate): renewal took place each Monday, Wednesday and Friday during the 21 days test period
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Dilution water was prepared in accordance with the ISO norm: ISO - 6341, 1982, to which micro-nutrients were added
* NaHCO3 12.95 g/l
* KCl 1.15 g/l
* CaCl2.2H2O 58.8 g/l
* MgSO4.7H2O 24.65 g/l
* K2-EDTA.2H2O 1.086 g/l
* FeCl3.6H2O 1.5 g/l
* MnSO4.H2O 0.310 g/l
* Na2MoO4.2H2O 0.126 g/l
* ZnSO4.7H2O 0.044 g/l
* SeO2 0.014 g/l
* Vitamine B12 0.010 g/l
- Intervals of water quality measurement: at the start of the test and before and after each renewal

OTHER TEST CONDITIONS
- Photoperiod: 16h light - 8h dark
- Light intensity: fluorescent bulb (Philips 58W/33)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
The parent daphnids were inspected daily for formation of eggs, appearance of the first brood, number of layings, morbidity, behavior and mortality.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2
- Justification for the selection of test substance concentrations: The OECD guideline 202 proposes a starting concentration equal to the EC50-24h value. Therefore, 3 mg Imazalil per litre dilution water was chosen as the highest concentration to be tested. Five following concentrations descended with a factor 3.2 to obtain a concentration range in which at least one concentration equals 1/100 of the highest.

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

> 0.03 - < 0.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

0.3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Key result

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

< 0.007 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

> 0.763 - < 2.481 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

reproduction

Details on results:

- Behavioural and physical abnormalities: The behaviour and physical appearance of daphnids exposed to imazalil concentrations of 0.01, 0.03 and 0.1 mg/l was not different from the control group. Imazalil at a concentration of 0.3 mg/l did not have a distinct effect. After 6 days of exposure, a few daphnids were pale and adhered to organic matter. However, after eleven days of exposure these effects disappeared. Almost all daphnids survived but they were not capable of forming viable young daphnids. The following dose groups (1.0 and 3.0 mg/l) exerted a rather severe effect. Daphnids were small, pale and adhered to organic matter. In addition they moulted very difficult. Their general condition was very bad and they died within 6 yo 7 days of exposure.
- Other biological observations: Reproduction started at the same day as the control for all test concentrations where reproduction was possible. A delay of 1 day before the appearance of the fourth brood was observed for a test substance concentration of 0.1 mg Imazalil per litre. Due to this delay, at the day 21 the fifth brood was not complete, resulting in a lower average number of young daphnids per female.
- Mortality of control: 1/40
- Other adverse effects control: none indicated
- Abnormal responses: none indicated
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: None indicated. Deviations from the nominal concentrations can be explained by factors such as the accuracy of the analytical method and removal of test substance from the media with the young daphnids.
- Effect concentrations exceeding solubility of substance in test medium: no

Based on the RAC document (2013) on Imazalil, the measured concentrations were between 70 -87% of the nominal concentrations at the start of the study. These values fall below 80% of the nominal concentrations at the start of the study. therefore the results should be based on the measured concentrations. The RAC considers the value of < 0.007 mg/L based on mean measured concentration as the most appropriate toxicity value. However as no discrete NOEC could be determined (< 0.007 mg/L) the LOEC <= 0.007 mg/L is more appropriate.

Validity criteria fulfilled:

yes

Conclusions:

The influence of Imazalil on the reproduction rate of Daphnia magna were determined during an exposure period of 21 days in a semi-static test according to the OECD guideline 202. The NOEC after 21 days of exposure with respect to mortality was 0.03 < NOEC < 0.10 mg imazalil per litre. The EC50-value after 21 days of exposure with respect to mortality was 0.3 mg imazalil per litre. The NOEC after 21 days of exposure with respect to reproduction was < 0.01 mg imazalil per litre. The EC50-value after 21 days of exposure with respect to reproduction was between 0.1 and 0.3 mg imazalil per litre. The NOEC after 21 days of exposure with respect to behaviour was 0.1 < NOEC < 1 mg imazalil per litre. The results of the test can be considered reliable without restriction.

Description of key information

No reliable study was available for imazalil sulfate and read across was performed with imazalil base. For imazalil base, two long-term toxicity studies are available forDaphnia magna, performed according to OECD guideline 211. The study of Kuhl et al. (2008) reported a 21d NOEC of 0.025 mg imazalil / L which corresponds to 0.033 mg imazalil sulphate/L. The study of Weytjens et al. (1989) reported a 21d LOEC of 0.007 mg imazalil / L which corresponds to 0.0093 mg imazalil sulphate/L.

Based on the RAC document on Imazalil (2014), as there is an unbounded NOEC in the study of Weytjens et al (1989), the LOEC of 0.007 mg /L should be taken forward in classification which corresponds to 0.0093 mg imazalil sulphate /L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

0.009 mg/L