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Long-term toxicity to aquatic invertebrates

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Reference
Endpoint:
long-term toxicity to aquatic invertebrates
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
5th September - 26th September 2008
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in accordance with generally accepted scientific principles, possibly with incomplete reporting or methodological deficiencies, which do not affect the quality of the relevant results. Study conducted on shale oil, middle fraction and read across.
Qualifier:
according to guideline
Guideline:
OECD Guideline 211 (Daphnia magna Reproduction Test)
Deviations:
no
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
All WAFs testing loading rates were measured for total organic carbon in new (0 hours) and old (48 or 72 hours) medium three times during the test.
Vehicle:
no
Details on test solutions:
WAFs were prepared by serial dilution of a single stock WAF, because of very low test concentration. Adequate amounts of multi-component substance were added directly to test medium. Stock solution of WAF was prepared in deionised water, 50 mg was added to 500 ml of deionised water and mixed on magnetic stirrer using a stirring rate such that a vortex was formed to give a slight dimple in water surface. The stirring was stopped after 96 hours. After one hour standing the aqueous phase was removed and than filtered trough nylon membrane filters with pore size 0.45 μm and diameter 47 mm first 50 ml discarded.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: freshwater water flea
- Strain/clone: daphnia magna straus
- Justification for species other than prescribed by test guideline: N/A
- Source: DHI Water & Environment, Horsholm, Denmark
- Age of parental stock (mean and range, SD): NDA
- Feeding during test
- Food type: green algae Desmodesmus subspicatus and Brewer's yeast
- Amount: The amount of supplied diet was based on the amount of organic carbon (C) provided to each parent animal of Daphnia magna (0.1 mg C/daphnia/day are sufficient for achieving the required number of offspring to meet the test validity criteria).
- Frequency: 3 times per week

ACCLIMATION
NDA
Test type:
semi-static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
21 d
Post exposure observation period:
NDA
Hardness:
NDA
Test temperature:
19.8 - 23.4
pH:
6.51 - 8.90
Dissolved oxygen:
84.6 - 126.5 %
Salinity:
NDA
Nominal and measured concentrations:
Nominal loading rates, 0, 0.05, 0.10, 0.21, 0.47, 1.03, 2.27 and 5.00 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel: vial
- Aeration: NDA
- Type of flow-through (e.g. peristaltic or proportional diluter):NDA
- Renewal rate of test solution (frequency/flow rate): three times a week
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates):10
- No. of vessels per control (replicates): 10
- No. of vessels per vehicle control (replicates): N/A
- Biomass loading rate: NDA

TEST MEDIUM / WATER PARAMETERS
Standard Elendt M4 and M7 media.

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: 16 hours light / 8 hours dark
- Light intensity: 600 - 800 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortality, time to production of first brood, number and size of broods per animal, number of aborted broods, presence of males or ephippia.

VEHICLE CONTROL PERFORMED: N/A
Reference substance (positive control):
yes
Remarks:
3, 4 dichloraniline
Duration:
21 d
Dose descriptor:
EL50
Effect conc.:
5 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
21 d
Dose descriptor:
NOELR
Effect conc.:
0.1 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Details on results:
- Mortality of parent animals: 2/20 mortalities in control group. 1/10 in 0.05, 0.21 and 0.47 mg/l groups. No other mortalities observed.
- Time to first brood release or time to hatch: 10th day
Reported statistics and error estimates:
Results of the Dunnet one-tailed test providing the NOEC value and TOXEDO results providing the EL50 value are reported in "Any other information on results incl. tables".

OFFSPRING AND REPRODUCTION DATA

Analysis of Dunnet one-tailed test providing "no-observed-effect concentration" (NOEC) values gave following results:

Summary Statistics and ANOVA

Loading rate
mg/l(*)
n mean s.d. cv%
control 18 72.72 5.77 7.9
0.05 9 69.44 13.61 19.6
0.1 10 63.8 9.25 14.5
0.21* 9 59.22 10.47 17.7
0.47* 9 56.67 9.41 16.6
1.03* 10 59.6 7.89 13.2
2.27* 10 55.3 12.44 22.5
5.0* 10 62.9 8.66 13.8

* - the mean for this group is significantly less than the control mean at alpha = 0.05 (1- sided) by a

t– test with Bonferroni adjustment of alpha level

s.d. - standard deviation

n - number of parent animal

cv - coefficient of variation

The No Observed Effect Loading Rate (NOELR) after 21-day exposure was 0.10 mg/l. NOEC is based upon no significant change of reproduction for exposed group compared to control group. The Lowest Observed Effect Loading Rate (LOELR) after 21-day exposure was 0.21 mg/l.

Summary data analysed using software TOXEDO (5) providing EL50 values, gave following results:

Loading rate
mg/l
Females at the end Offspring at termination Offspring per female Inhibition in percent
Control 18 1309 72.72 -
0.05 9 625 69.44 4.5
0.1 10 638 63.8 12.3
0.21 9 533 59.22 18.6
0.47 9 510 56.67 22.1
1.03 10 596 59.6 18
2.27 10 553 55.3 24
5 10 629 62.9 13.5

There was not observed reproduction toxicity more than 50% in daphnids exposed to test substance WAF of all loading rates after 21 days. EL50 was greater than 5 mg/l. Time of first brood production was the 10th day of test duration for all WAFs and control.

Complex data, for each concentration of test substance, for reproduction toxicity with Daphnia magna are given in APPENDIX 1.

Chemical analysis

The test preparations were analysed for Total Organic Carbon (TOC). Analysis of the new medium (at 0 hours) test preparations showed measured test concentration (corrected for control) to range from 0.013 to 1.424 mg/l. Analysis of the old medium (after 48 or 72 hours) test preparations showed measured test concentration (corrected for control) to range from 0.055 to 1.719 mg/l with the one exception., 4.196 mg/l at loading rate 0.10 mg/l. There was no significant decline in measured test concentrations over 48 and 72 hours period in the highest test concentration.

Chemical analysis of TOC showed that measured concentrations in old medium for ranged from 83% to 97% of nominal concentration of WAF at loading rate 5 mg/l. In the WAFs with the lower loading rates, the TOC concentration in new medium was

approximately in the same range as in the control vessel and in old medium the TOC concentration was mostly higher than in control (this consider due to feeding of animals). Thus, the measurements of TOC originated form the test items was not possible due to low

test concentration and feeding of animals.

Given that toxicity cannot be attributed to a single component or mixture of components but to the test material as a whole the results were based on loading rates only.

Table 1: Daphnia Reproduction Data

Loading rate (mg/L)

Females alive at end of test

Offspring at termination

Offpsring per female

Inhibition in percent

Control

18

1309

72.72

-

0.05

9

625

69.44

4.5

0.10

10

638

63.80

12.3

0.21

9

533

59.22

18.6

0.47

9

510

56.67

22.1

1.03

10

596

59.60

18.0

2.27

10

553

55.30

24.0

5.00

10

629

62.90

13.5

Validity criteria fulfilled:
yes
Remarks:
The mortality of the parent animals (female) did not exceed 20%, at the end of the test it was 10% and the mean number of live offspring produced per parent animal surviving at the end of the test was 72.72.
Conclusions:
The EL50 (Effective Loading Rate) of the test substance, Shale oil, in the test of reproduction to Daphnia magna with 21-day exposure was greater than 5 mg/l. The No Observed Effect Loading Rate (NOELR) after 21-day exposure was the concentration of WAF at loading rate 0.10 mg/l.
Executive summary:

In an 21-day Daphnia magna reproduction toxicity test according to OECD 211 and performed to GLP standard, daphnia were exposed to Shale oil middle fraction in indivivdual vessels at nominal loading rates of 0.05, 0.10, 0.21, 0.47, 1.03, 2.27 and 5.00 mg/L.

The EL50 (Effective Loading Rate) of the test substance, Shale oil, in the test of reproduction to Daphnia magna with 21-day exposure was greater than 5 mg/l. The No Observed Effect Loading Rate (NOELR) after 21-day exposure was the concentration of WAF at loading rate 0.10 mg/l.

Based on the rationale for read-across, it is considered acceptable to use this study to address the same endpoint for the heavy fraction of shale oil.

Description of key information

The EL50 (Effective Loading Rate) of the test substance, Shale oil, in the test of reproduction to Daphnia magna with 21-day exposure was greater than 5 mg/l. 
The No Observed Effect Loading Rate (NOELR) after 21-day exposure was the concentration of WAF at loading rate 0.10 mg/l.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
0.1 mg/L

Additional information

Similar values for short-term toxicity to daphnia were observed between the middle fraction and heavy fraction of shale oil. This, combined with analytical data which showed the two fractions to be compositionally similar, support the validity of a read-across approach to address the long-term toxicity to aquatic invertebrates endpoint. Hence, the study conducted on the middle fraction of shale oil, is considered robust enough to be the key study for the heavy fraction.