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Ecotoxicological information

Sediment toxicity

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Endpoint:
sediment toxicity: long-term
Type of information:
experimental study
Adequacy of study:
key study
Study period:
08/02/08-21/03/08
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study conducted on read-across material
Remarks:
GLP and conducted to current guideline, but read across to an analogous substance prevents use of highest reliability score. The read-across from MnCl2 to Mn(NO3)2 is justified on the following basis: both substances are very soluble in water hence bioavailable and both will release Mn2+ ions. Therefore, from an ecotoxicity standpoint, the chloride or nitrate anions are not considered to have any influence on the effective toxicity of Mn2+ or any toxicity in their own right, so the anions can be disregarded. Therefore any effect will be related to the Mn2+ cation, and the data from MnCl2 ecotoxicity tests is regarded as a suitable surrogate for read-across
Justification for type of information:
See the read-across report attached in Section 13.
Cross-reference
Reason / purpose:
other: Read-across target
Reference
Endpoint:
sediment toxicity: long-term
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study conducted on read-across material
Justification for type of information:
See the read-across report attached in Section 13.
Reason / purpose:
read-across source
Duration:
42 d
Dose descriptor:
NOEC
Effect conc.:
285.9 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Dissolved
Basis for effect:
other: Survival
Duration:
42 d
Dose descriptor:
LOEC
Effect conc.:
627.2 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Dissolved
Basis for effect:
other: Survival
Duration:
42 d
Dose descriptor:
EC10
Effect conc.:
188.5 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Dissolved
Basis for effect:
other: Survival
Duration:
28 d
Dose descriptor:
EC10
Effect conc.:
96.4 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
other: Dissolved
Basis for effect:
growth rate

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report Date:
2009

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 218 (Sediment-Water Chironomid Toxicity Test Using Spiked Sediment)
Principles of method if other than guideline:
This study was conducted according to the protocol, ''42- day Chronic Toxicity of Manganese to the freshwater amphipod crustacean, Hyalella aztecaThis protocol was designed to comply with methods described in the following references:
Environmental Canada(1997). Biological Test Methods: Test for Growth and Survival in Sediment Using the Freshwater Amphipod Hyalella azteca. Report EPS 1/RM/33.
U.S. Environmental Protection Agency(2000). Methods for measuring the toxicity and bioaccumulation of sediment associated contaminants with freshwater invertebrates. EPA 600/R-99/064,Duluth,MN.
GLP compliance:
yes (incl. certificate)
Remarks:
U.S. EPA GLP standard (40 CFR 792)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Manganese(II) chloride, anhydrous
- Physical state: Solid
- Analytical purity:≥ 97%
- Impurities (identity and concentrations): Iron - 0.01% , Nickel - 0.002%
- Composition of test material, percentage of components: MnCl2- 99%, H20- 1.0%, Iron - 0.01%, Nickel -0.002%.
- Lot/batch No: (Lot # J25R029)
- Storage condition of test material: Stored at room temperature
- Other:The manufacturer's material safety data sheet reported the solubility of MnCl2 in water as 723 grams per litre at 25°C.

Sampling and analysis

Analytical monitoring:
yes
Details on sampling:
- Concentrations: were determined at test initiation and then determined twice a week
- Sampling method: Analytical samples from each treatment were collected for total recoverable and dissolved Mn analysis at test initiation. For the remainder of the test, samples for dissolved Mn analysis were collected from alternating replicates within each treatment twice a week.
- Sample storage conditions before analysis: Samples were preserved with nitric acid ( trade metal grade) to pH < 2 prior to analysis.

Test substrate

Vehicle:
yes
Details on sediment and application:
PREPARATION OF SPIKED SEDIMENT
Artificially- formulated sediment was used as the sediment type in the toxicity test. Components and characteristics of the formulated sediments are provided in Table 1. Preparation instructions for the formulated sediments are detailed in OECD Guideline 218 (OECD 2004).

Test organisms

Test organisms (species):
Hyalella azteca
Details on test organisms:
TEST ORGANISM
- Common name: terrestrial amphipods
- Source: Aquatic Biosystems (Fort Collins, CO, USA)
- Age of parental stock (mean and range, SD): Arrived at the testing laboratory as 5 to 7 day old organisms
- Age of animals at beginning of exposure: 7-9 days old
- Feeding during test
The organism were held in-house for 2 days as per PERL standard operating procedure(SOP) # 5102, with appropriate feeding s and water renewals to maintain health in the organisms prior to test initiation.


ACCLIMATION
- Acclimation period: 2 days
- Acclimation conditions (same as test or not):The organism were held in-house for 2 days as per PERL standard operating procedure (# 5102, with appropriate feeding and water renewals to maintain health in the organisms prior to test initiation.
- Health during acclimation (any mortality observed): Visual observation, the organism appeared healthy at test initiation.

Study design

Study type:
laboratory study
Test type:
flow-through
Water media type:
freshwater
Type of sediment:
artificial sediment
Exposure duration
Duration:
42 d
Exposure phase:
total exposure duration
Post exposure observation period:
Not reported

Test conditions

Hardness:
104±5 mg/L as CaCO3
Test temperature:
23±2°c
pH:
7.3-8.1
Dissolved oxygen:
Dissolved oxygen concentration was maintained above 2.5 mg/L.
Salinity:
Not applicable
Ammonia:
<1.0 mg/L
Nominal and measured concentrations:
Nominal Concentration - 250, 500, 1000, 2000, 4000, and 8000 µg Mn/L
Average Measured Dissolved Mn Concentration - <62.5, 136.0, 285.9, 627.2, 1402.6, 3351.1, and 6663.6 µg/L
Details on test conditions:
TEST SYSTEM
- Test container (material, size): 1L glass jars with a mesh covered drainage hole.
- Sediment volume: Containers was filled with 100-mL of sterilised sand. A piece of nitex screen was placed in each chamber above the surface of the sand.
- Overlying water volume: The overlying volume of water in each chamber was approximately 500-600 mL.

EXPOSURE REGIME
- No. of organisms per container (treatment):10
- No. of replicates per treatment group:12
- No. of replicates per control / vehicle control:12
- Feeding regime: Daily. Flow to each test chamber was diverted for 2-3 hours to allow food to settle and become available to the organisms. If excess food or bacterial growth begins to form on the sediment surface, feeding may be suspended for one or more days.
- Type and preparation of food: Each chamber was fed YTC (Yeast, Trout Chow, Cereal Leaves).
- Amount of food: Each chamber was fed 1.0 mL of an 1800 mg/L stock.


RENEWAL OF OVERLYING WATER
- Details on volume additions: A Mariotte bottle was used to continuously deliver an appropriate volume of Mn stock solution to the diluter. A flow-splitting chamber was used for each test concentration( treatment) to promote mixing of the test solution and to allocate equally the test solution among the twelve replicate test chambers.
- Flow-rate: The stock solution was delivered to the dilutor at a rate of 5.0 mL/mixture. The flow rate to each test chamber was approximately 10 mL/min providing approximately 24 volume exchanges per replicate per day.


OVERLYING WATER CHARACTERISTCS
- Type of water : The dilution and control water used for this study was laboratory-blended water ( well water blended with reverse osmosis-treated water with a targeted hardness of 80- 120 mg/L as CaCO3).
- Alkalinity:129±5 mg/L as CaCO3
- Conductivity: 339±22 µS/cm
- Total organic carbon:<0.50 mg/L (DOC)
- Unionized ammonia:<1.0 mg/L
- Residual chlorine:37.3 mg/L


CHARACTERIZATION OF ARTIFICIAL SEDIMENT
Components and Characteristics of the formulated sediment are provided in Table 1. Preparation instruction for the formulated sediment is as per OECD guideline 218 (OECD 2004).


OTHER TEST CONDITIONS
- Photoperiod: 16- hours and 8-hours dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Observations of live and dead organisms was recorded daily from test initiation until termination. Dead organism was recorded and immediately removed. At test day 28 organisms from four of the replicates was used for survival and growth measurements. The surviving organisms from the remaining eight replicates were replaced into water only chambers (without sand) and returned to the flow- through exposure system. On day 35, offspring was counted and removed from each chamber. Endpoint measured include survival (Day 28, 35, and 42), growth (as dry weight on Day 28 and 42), and reproduction (number of young/ femaleproduced from day 28 to Day 42). For dry weight measurement, organisms from each chamber was placed into a pre-weighed aluminium pan and dried for at least 12 hours at 105°C to determine dry weight

VEHICLE CONTROL PERFORMED: Yes


TEST CONCENTRATIONS
- Spacing factor for test concentrations: Six test treatments was tested using a 0.5 dilution scheme.
- Range finding study
- Test concentrations: 320 µg Mn/L
- Results used to determine the conditions for the definitive study: These concentration were selected based on findings from a previous chronic life-cycle H. azteca Mn test where concentration up to 320 µg Mn/L resulted in no effect on exposed organism.
Reference substance (positive control):
no

Results and discussion

Effect concentrationsopen allclose all
Duration:
42 d
Dose descriptor:
NOEC
Effect conc.:
285.9 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
dissolved
Basis for effect:
other: Survival
Duration:
42 d
Dose descriptor:
LOEC
Effect conc.:
627.2 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
dissolved
Basis for effect:
other: Survival
Duration:
42 d
Dose descriptor:
EC10
Effect conc.:
188.5 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
dissolved
Basis for effect:
other: Survival
Remarks on result:
other: 95% CL
Duration:
28 d
Dose descriptor:
EC10
Effect conc.:
96.4 µg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
dissolved
Basis for effect:
growth rate
Remarks on result:
other: 95% CL
Details on results:
Not reported
Results with reference substance (positive control):
Not reported
Reported statistics and error estimates:
All statistical analyses were performed using the mean dissolved Mn concentrations. Differences in survival (at Days 28, 35, and 42), growth (at day 28 and 42), and reproduction (at day 35 and 42) were evaluated using a statistical computer package (comprehensive Environmental Toxicity Information System [CETIS], McKinleyville, CA, USA, and Toxicity Relationship Analysis Program [TRAP], Duluth, MN, USA). If the data met the assumptions of normality and homogeneity, the NOEC and LOEC were estimated using an analysis of variance to compare (p≤0.05) organism performance in the experimental treatments with that observed in the control. The effective concentration required to reduce the survival, growth, or reproduction by 10% relative to control performance (EC10 and EC20) were estimated using a piecewise linear regression analysis. The median-lethal concentration (LC50) was also estimated using a piecewise linear regression analysis. Exposure concentration were log-transformed before determination of EC10 and EC20 and LC50 values.

Any other information on results incl. tables

  Table 6. Survival Data1

 

Average Measured Dissolved Mn (mg/L)

% Survival (±SD)

Day 28

Day 35

Day 42

<62.5

91±10

81±15

51±32

136.0

83±22

60±38

56±30

285.9

91±11

79±27

48±42

627.2

72±24*

48±26*

13±15*

1402.6

58±35*

37±34*

20±23*

3351.1

23±23*

8±11*

0±0*

6663.6

0±0*

-

-

1Survival is reported as the average survival within a treatment (combination of all replicates within a treatment).

* Significant less than control (p≤0.05).

 SD = Standard Deviation

 Table 7. Organism Growth Data (Biomass)1

 

Average Measured Dissolved Mn Concentration (µg/L)

Average Dry weight per original organism in mg (±SD)

Day 28

Day 42

<62.5

0.18±0.01

0.08±0.07

136.0

0.19±0.01

0.15±0.09

285.9

0.12±0.04

0.12±0.08

627.2

0.10±0.05*

0.04±0.04

1402.6

0.16±0.052,*

0.07±0.03

3351.1

0.03±0.01*

-

6663.6

-

-

1Biomass (average dry weight per original organism) is reported as a combination of all replicates within a treatment

2Day 28 average includes ‘outliers’ replicate.

*Significantly less than control (p≤0.05).

SD = Standard Deviation.

 

Table 8. Organism Growth Data1

 

Average Measured Dissolved Mn Concentration (µg/L)

Average Dry weight per original organism in mg (±SD)

Day 28

Day 42

<62.5

0.22±0.03

0.15±0.05

136.0

0.20±0.01

0.21±0.08

285.9

0.14±0.05*

0.18±0.03

627.2

**

**

1402.6

**

**

3351.1

**

**

6663.6

**

**

1Dry weight is reported as the average weight per organism (combination of all replicates within a treatment).

*Significantly less than control (p≤0.05).

** Data was not used in statistical analysis (hypothesis testing) because a survival effect was observed at this concentration.

SD = Standard Deviation.

 

Table 8. Organism Reproduction Data1

 

Average Measured Dissolved Mn Concentration (µg/L)

Average Reproduction (±SD) (young/female)

Day 35

Day 42

<62.5

0.75±1.1

1.55±2.57

136.0

0.12±0.24*

0.12±0.24

285.9

0.0±0.0*

0.28±0.45

627.2

**

**

1402.6

**

**

3351.1

**

**

6663.6

**

**

1 Reproduction is reported as the average number of young per female (combination of all replicates within a treatment)

*Significantly less than control (p≤0.05).

** Data was not used in statistical analysis (hypothesis testing) because a survival effect was observed at this concentration.

SD = Standard Deviation

 

Table 10. Summary of Survival Statistics (µg Mn/L)1

 

Endpoint

Survival

28 day

35 day

42 day

NOEC

285.9

285.9

285.9

LOEC

627.2

627.2

627.2

EC10 (95% Cl)

568.6 (349.9-923.9)

382.8(104.7-1399.5)

188.47(27.47-1293.2)

EC20 (95% Cl)

752.6(494.0-1146.4)

512.6(170.1-1544.3)

255.9(51.9-1260.5)

1NOEC = No Observed Effect Concentration. LOEC = Lowest Observed Effect Concentration. EC10= Effective Concentration for 10% reduction in survival relative to control performance. EC20 = Effective Concentration for 20% reduction in survival relative to control performance. CI – Confidence interval.

 

Table 11. Summary of Growth Statistically (µg Mn/L)1

 

Endpoint

Mean Dry Biomass

Dry Weight

28 day2

42 day

28 day

42 day

NOEC

285.9

1402.6

136.0

285.9

LOEC

627.2

>1402.6

285.9

>285.9

EC10( 95% CI) all replicates

1349.8

(138.2-13184.3)

N/A

1720.5 9N/A)

N/A

EC10( 95% CI)

Without outlier replicate

1063.1

(549.0-2058.5)

N/A

96.4(12.9-720.7)

N/A

EC20(95% CI)

all replicates

1610.2(256.0-10126.5)

N/A

1949.0(N/A)

N/A

EC20(95% CI)

Without outlier replicate

1248.6(706.4-2206.9)

N/A

203.4(39.5-1046.9)

N/A

1NOEC = No Observed Effect Concentration. LOEC = Lowest Observed Effect Concentration. EC10= Effective Concentration for 10% reduction in survival relative to control performance. EC20 = Effective Concentration for 20% reduction in growth relative to control performance. CI– Confidence interval. N/A = Non-applicable/determinable.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Remarks:
Control survival was greater than 80% at the end of the test
Conclusions:
The lowest NOEC (survival) was 0.29 mg Mn/L. The read-across from MnCl2 to Mn(NO3)2 is justified on the following basis: both substances are very soluble in water hence bioavailable and both will release Mn2+ ions. Therefore, from an ecotoxicity standpoint, the chloride or nitrate anions are not considered to have any influence on the effective toxicity of Mn2+ or any toxicity in their own right, so the anions can be disregarded. Therefore any effect will be related to the Mn2+ cation, and the data from MnCl2 ecotoxicity tests is regarded as a suitable surrogate for read-across