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Ecotoxicological information

Long-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report date:
2015

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 211 (Daphnia magna Reproduction Test)
Deviations:
no
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
2-butoxyethyl benzoate
EC Number:
226-685-8
EC Name:
2-butoxyethyl benzoate
Cas Number:
5451-76-3
Molecular formula:
C13H18O3
IUPAC Name:
2-butoxyethyl benzoate
Test material form:
other: liquid
Details on test material:
2-Butoxyethyl benzoate, Lot # 201303443-19; Purity 99.2 %
Specific details on test material used for the study:
Test Material Name: 2-Butoxyethyl benzoate
Chemical Name: 2-Butoxyethanol benzoate
Supplier, City, State (Lot, Reference Number): The Dow Chemical Company, Midland, Michigan (Lot # 201303443-19).
Purity/Characterization (Method of Analysis and Reference): The purity of the test material was determined to be 99.2% area (corrected for water) by gas chromatography with identification by nuclear magnetic resonance and gas chromatography mass spectrometry (Gobbi, 2014).

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
The bulk test solutions were sampled at test initiation and on days 1, 7, and 14 of the
study for analytical verification of 2-butoxyethyl benzoate concentrations. Spent test
solutions (10 replicates per test level, when applicable) were pooled by test level and
analyzed on days 1, 7, 14, and 21. The samples were analyzed for 2-butoxyethyl
benzoate by high-performance liquid chromatography with photodiode array detection
(HPLC/DAD). To assess analytical method precision and solution homogeneity, three
additional samples were collected on day 0, 1, 7, and 14 from the lowest and highest
concentration bulk treatment solutions with surviving organisms.

Test solutions

Vehicle:
no
Details on test solutions:
Bulk test solutions were prepared daily (excluding day 21) at nominal (target)
concentrations of 0 (water control), 0.75, 1.5, 3.0, 6.0, and 12.0 mg 2-butoxyethyl
benzoate/L. The 12.0 mg/L bulk test solution, which also served as the stock solution for
preparing the remaining bulk test solutions, was prepared by injecting (subsurface) 24 μL
of 2-butoxyethyl benzoate into a 2 L volumetric flask filled to volume with lab dilution
water (LDW). A stopper was placed on the flask, and the solution was shaken vigorously
to ensure a homogenous solution. The remaining bulk test solutions were prepared as
dilutions of the 12.0 mg 2-butoxyethyl benzoate/L solution as follows:
Nominal Test Solution
Concentration
(mg 2-butoxyethyl
benzoate/L)
Stock Solution
Concentration
(mg 2-butoxyethyl
benzoate /L)
Stock Solution
Transfer Volume
(L)
Final Volume
(L)
0.75 12.0 0.0625 1
1.5 12.0 0.125 1
3.0 12.0 0.250 1
6.0 12.0 0.500 1
After transfer of the stock solution aliquots, the test solutions were brought to volume in
1 L volumetric flasks with lab dilution water (LDW). All test solutions were noted as
clear and colorless. Analytical results of the bulk test solutions indicate that the test
material was soluble at the concentrations tested. Test solutions were shaken vigorously
prior to analytical sampling and distribution into test vessels. The laboratory control
vessels were filled with LDW and capped prior to preparation of the bulk stock solutions.

Test organisms

Test organisms (species):
Daphnia magna

Study design

Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
21 d

Test conditions

Hardness:
44-70 mg/L CaCO3)
pH:
7.2-8.0
Dissolved oxygen:
5.8-9.3
Conductivity:
160-180 us/cm
Nominal and measured concentrations:
The study was conducted with daphnids (one individual per replicate with ten replicates per
treatment level) exposed to nominal concentrations of 0 (water control), 0.75, 1.5, 3.0, 6.0,
and 12 mg 2-butoxyethyl benzoate/L.

The resulting mean measured test concentrations were 0.578, 1.00,
1.90, 3.55, and 10.6 mg/L. No 2-butoxyethyl benzoate was detected in the water control
above the lowest level quantitated (LLQ) of 0.250 mg/L.
Details on test conditions:
Dissolved oxygen levels ranged from 5.8-9.3 mg/L (67-104% air
saturation), temperatures ranged from 20-21°C, and pH ranged from 7.2-8.0. Light
intensity ranged from 1119-1472 lux.
Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
3.55 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
reproduction
Key result
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
3.55 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth
Key result
Duration:
21 d
Dose descriptor:
LOEC
Effect conc.:
10.6 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
The chronic toxicity values for Daphnia magna exposed to 2-butoxyethyl benzoate over a
21-day static-renewal (daily) exposure period were statistically and/or empirically
determined using mean measured test concentrations. The resulting endpoint values are
presented below:
 The LOEC and MATC for reproduction, length and weight (at study termination)
were > 3.55 mg/L.
 The LOEC and MATC for survival (at study termination) were 10.6 and
6.13 mg/L, respectively.
 The NOEC for survival, reproduction, and length and weight (at study
termination) was 3.55 mg/L.
Reported statistics and error estimates:
Test endpoints consisted of the determination of the NOEC, LOEC, and MATC values
for survival, reproduction (total number of offspring) and growth as both length and dry
weight. In addition, the 21-day EC50 value for survival was determined. Replicates were
excluded from statistical analysis if an accidental/inadvertent mortality occurred or if
there was evidence that observed mortalities did not follow a concentration-response
pattern (OECD 2012). A Cochran Armitage trend test with a lack of fit test was used to
determine if the mortality data were consistent with a monotone dose-response
relationship. Since the Cochran Armitage lack of fit test was significant, the data were
determined to be non-monotonic. Therefore, the NOEC for mortality was determined
using a Fisher Exact test with a Bonferroni-Holm correction applied to the p-values.
Due to mortality, weight, length and offspring data for the 10.6 concentration level were
not available for analysis. A Spearman rank correlation was used to determine if the
offspring and growth data were consistent with a monotone dose-response. The
correlation coefficients were below 0.5 for all parameters so the data were judged as
being not monotonic. The raw data were tested for normality using the Shapiro-Wilk’s
test at a Type I error rate of 0.01. If the data were not normally distributed, the logarithmic, inverse, and square root transformations were tested sequentially to search
for a normalizing transformation. Next, the data and the transformed variables were
tested for homogeneity of variance using Levene’s test at a Type I error rate of 0.05.
The offspring and length data were both normal and homogeneous, so a parametric
analysis was conducted using a Dunnett’s test to compare each treated group with the
control. A one-tailed Dunnett’s test, looking for a significant decrease from the control
group, was conducted at a Type I error rate of 0.05.

Any other information on results incl. tables

The LOEC and MATC for reproduction, length and weight (at study termination)

were > 3.55 mg/L.

 The LOEC and MATC for survival (at study termination) were 10.6 and

6.13 mg/L, respectively.

 The NOEC for survival, reproduction, and length and weight (at study

termination) was 3.55 mg/L.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The chronic toxicity values for Daphnia magna exposed to 2-butoxyethyl benzoate over a
21-day static-renewal (daily) exposure period were statistically and/or empirically
determined using mean measured test concentrations. The resulting endpoint values are
presented below:
 The LOEC and MATC for reproduction, length and weight (at study termination)
were > 3.55 mg/L.
 The LOEC and MATC for survival (at study termination) were 10.6 and
6.13 mg/L, respectively.
 The NOEC for survival, reproduction, and length and weight (at study
termination) was 3.55 mg/L.
Executive summary:

The purpose of this study was to assess the chronic toxicity of 2-butoxyethyl benzoate to the freshwater daphnid, Daphnia magna. A 21-day static-renewal (daily) exposure was conducted and endpoints included adult daphnid survival, reproduction (total young produced per surviving adult female), and growth (length and weight of surviving adults).

The study was conducted with daphnids (one individual per replicate with ten replicates per treatment level) exposed to nominal concentrations of 0 (water control), 0.75, 1.5, 3.0, 6.0, and 12 mg 2-butoxyethyl benzoate/L. Test solutions were renewed daily throughout the 21- day exposure. Daily observations were made and the number of surviving daphnids recorded. Reproduction was evaluated by counting and removing neonates during renewals. Lengths and dry weights of all surviving adult daphnids were measured and recorded following termination of exposure.

Freshly-prepared bulk test solutions were sampled at test initiation and on days 1, 7, 14, and 19 of the study for analytical verification of 2-butoxyethyl benzoate concentrations. Spent test solutions (10 replicates per dose level when applicable) were pooled and analyzed on days 1, 7, 14, and at the end of exposure (day 21). The samples were analyzed for 2-butoxyethyl benzoate by high-performance liquid chromatography with photodiode array detection (HPLC/DAD). The resulting mean measured test concentrations were 0.578, 1.00, 1.90, 3.55, and 10.6 mg/L. No 2-butoxyethyl benzoate was detected in the water control above the lowest level quantitated (LLQ) of 0.250 mg/L.

The chronic toxicity values for Daphnia magna exposed to 2-butoxyethyl benzoate over a 21-day static-renewal (daily) exposure period were statistically and/or empirically determined using mean measured test concentrations. The resulting endpoint values are presented below:

- The LOEC and MATC for reproduction, length and weight (at study termination) were > 3.55 mg/L.

- The LOEC and MATC for survival (at study termination) were 10.6 and 6.13 mg/L, respectively.

- The NOEC for survival, reproduction, and length and weight (at study termination) was 3.55 mg/L.