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Endpoint:
activated sludge respiration inhibition testing
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
2002-07-01 to: 2002-07-12
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Tungstic acid (WO3•H2O) is the hydrated form of tungsten trioxide (WO3). Based on Annex V, hydrates and water free forms (anhydrous) of compounds can be regarded as the same substance for registration purposes. Therefore, tungstic acid will be considered equivalent to WO3. For more details on the rationale, refer to the attached description of the read-across approach.
Qualifier:
according to guideline
Guideline:
other: Commission Directive 88/302/EEC
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
no data
Analytical monitoring:
no
Details on sampling:
no data
Vehicle:
no
Details on test solutions:
The test substance was added to about 130 mL of deionized water and stirred overnight before testing (equilibration phase)
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
no data
Test type:
not specified
Water media type:
not specified
Limit test:
no
Total exposure duration:
3 h
Remarks on exposure duration:
permanent aeration
Post exposure observation period:
no data
Hardness:
no data
Test temperature:
20 +/- 2 ⁰C
pH:
pH of the suspension before application : 6.1
pH of the various test substance concentratoins: 7.2 - 7.4
Dissolved oxygen:
no data
Salinity:
N/A
Nominal and measured concentrations:
Nominal test concentrations (mg/L): 1000, 1800, 3200, 5600, 10000
Details on test conditions:
TEST SYSTEM
- Test vessel: aeration tank of waste water treatment plant treating predominately domestic sewage (Wupper area water authority)
- Test concentration of the activated sludge : 480 mg/L suspended solids
Reference substance (positive control):
yes
Remarks:
3,5-Dichlorophenol (purity of 97%)
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
> 10 000 mg/L
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
- Tungsten trioxide showed 3.1 % respiration inhibition of activated sludge at a test substance concentration of 10,000 mg/L.
Results with reference substance (positive control):
- Results with reference substance valid? Yes
- Relevant effect levels: EC50 = 6.1 mg/L
Reported statistics and error estimates:
- The EC values were determined by statistical methods (probit analysis)

Test concentration: 1000 mg/L - Inhibition: 0.0% Test Concentration: 1800 mg/L - Inhibition: 15.4% Test Concentration: 3200 mg/L - Inhibition: 7.7% Test Concentration: 5600 mg/L - Inhibition: 7.7% Test Concentration: 10000 mg/L - Inhibition: 3.1% Physical-Chemical oxygen consumption was 0 mg/L-h for all test concentrations. The Respiratory rate-phsical-chemical oxygen consumption ranged from 22.0 mg/L-h to 27.0 mg/L-h

Conclusions:
Tungsten trioxide showed 3.1% respiration inhibition of activated sludge at the test substance concentration of 10,000 mg/L. The EC50 > 10,000 mg/L
Executive summary:

No microorganisms toxicity data are available for tungstic acid (target substance). However, micoorganisms toxicity data are available for tungsten trioxide (source substance), which were used for read-across. Tungstic acid (WO3•H2O) is the hydrated form of tungsten trioxide (WO3). Based on Annex V, hydrates and water free forms (anhydrous) of compounds can be regarded as the same substance for registration purposes. Therefore, tungstic acid will be considered equivalent to WO3. For more details on the rationale, refer to the attached description of the read-across approach.

Endpoint:
activated sludge respiration inhibition testing
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
key study
Study period:
2008-09-18 to: 2009-03-13
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
Well documented, scientifically sound study that was conducted in accordance to GLP and OECD Guideline 209 with no deviation to the protocol. The reliability of this study for the substance tested is a K1, but in application of read-across to a different substance ECHA’s guidance specifies that the score can be a maximum of K2. Due to lower transformation/dissolution results for tungsten metal (the target substance) than sodium tungstate (the source substance), the resulting toxicity potential would also be expected to be lower, so read-across is appropriate. In addition, read-across is justified because the classification and labelling is the less severe for the target substance and the PBT/vPvB profile is the same. Finally, the dose descriptor for the target substance is expected to be sufficiently lower than the source chemical, and read-across to the source chemical is adequately protective. For more details refer to the attached description of the read across approach.
Justification for type of information:
REPORTING FORMAT FOR THE CATEGORY APPROACH
1. HYPOTHESIS FOR THE CATEGORY APPROACH: The hypothesis is that properties are likely to be similar or follow a similar pattern because of the presence of a common metal ion, in this case tungstate.
2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES):
Source: Sodium Tungstate
Target: Tungstic acid
3. CATEGORY APPROACH JUSTIFICATION: See Annex 3 in CSR
4. DATA MATRIX: See Annex 3 in CSR
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
according to guideline
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
no data
Analytical monitoring:
no
Details on sampling:
- Concentrations: A stock solution of the test substance was prepared in reagent water at a concentration of 2.00 mg total product/mL. Stock solution volumes of 2.5, 8.0, 25, 80, and 250 mL were added to the 10, 32, 100, 320, and 1,000 mg total product/L test flasks, respectively, for the range finding test. Since there was no appreciable inhibition (eg >50%) up to a nominal of 1,000 mg/L, no further testing was required.

- A stock solution of the reference substance was prepared in reagent water at a concentration of 0.500 mg/mL and was used to add reference substance to the appropriate flasks. Stock solution volumes of 3.2, 10, and 32 mL were added to the 3.2, 10, and 32 mg/L test flasks, respectively.
Vehicle:
no
Details on test solutions:
no data
Test organisms (species):
activated sludge, domestic
Details on inoculum:
-Source of inoculum: Activated sludge, Columbia Wastewater Treatment Plant, Columbia, Missouri
- Preparation of inoculum for exposure: Repeated (11 times) centrifugation of 250 mL aliquots to concentrate the inoculum, followed by 3 rinses in ABC well water, and final suspension in a carboy with stirring and aeration with compressed air. The inoculum was sustained with a synthetic feed solution overnight. Suspended solids concentration was determined in triplicate, and the solution diluted with well water to a final suspended solids concentration between 3.6 and 4.4 g/L.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
30 min
Post exposure observation period:
None
Hardness:
n/a
Test temperature:
19.8-20.2 °C
pH:
8.03- microbial inoculum
8.23-8.35-test flasks, reference, and controls
8.23- abiotic control contact flask (containing no micorbial inoculum)
Dissolved oxygen:
Tested parameter- see Results
Salinity:
n/a
Nominal and measured concentrations:
Nominal concentrations:
Test substance- 0 (control), 10, 32, 100, 320, and 1000 mg test substance/L
Reference substance- 3.2, 10 and 32 mg/L
Details on test conditions:
TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: Glass 1 L, 200 mL inoculum added to flask for 30 min exposure, 500 mL final volume
- Aeration: 500-1000 mL/min
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 2 control, 1 abiotic control, 3 concentrations (1 each) reference substance

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ABC Labs well water

OTHER TEST CONDITIONS
- Adjustment of pH: None
- Light intensity: Ambient

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Oxygen consumption mg/L/hr, one reading per concentration.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3.2

Reference substance (positive control):
yes
Remarks:
3,5 Dichlorophenol
Duration:
30 min
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Details on results:
Test Substance:
The test substance showed 12.4, 31.1, 11.3, 14.8, and 17.9% inhibition at concentrations of 10, 32, 100, 320, and 1,000 mg/L, respectively. Thus, the estimated 30-minute-EC50 value for sodium tungstate was estimated to be >1,000 mg (total product)/L and no further testing was required.

Controls:
- Respiration rates for the control flasks were 81.0 and 71.7 mg O2L-1hr-1 during the 30-minute exposure. These respiration rates were within 15% of each other. The respiration rate of the abiotic control was 0.0 mg O2L-1hr-1, indicating that rate of oxygen uptake by sodium tungstate dihydrate was effectively 0% of the mean respiration rate of the controls, 76.4 mg O2L-1hr-1.
Results with reference substance (positive control):
- Results with reference substance valid? Yes. The 30 minute EC50 based on respiration rate for 3,5 dichlorophenol was 14.4 mg/L.
- Relevant effect levels: The percent inhibitions for the three reference toxicant substance flasks were 22.6, 34.3, and 83.8% at 3,5 dichlorophenol nominal concentrations of 3.2, 10, and 32 mg/L, respectively
- Other: 30 min EC50 was 14.4 mg/L- well within the 5-30 mg/L acceptance range required by the method.
Reported statistics and error estimates:
no data

The ASR system provided the oxygen consumption/depletion curve as a plot of the dissolved oxygen concentration (mg O2/L) versus time (seconds). The analyst selected the appropriate start and end times between which the oxygen depletion curve was linear, and the ASR system automatically calculated the oxygen consumption rate (respiration rate) and expressed these rates as mg O2L-1hr-1.

Conclusions:
The 30 minute EC50 value for sodium tungstate dihydrate to activated sludge is > 1000 mg/ L.
Executive summary:

No microorganisms toxicity data of sufficient quality are available for tungstic acid (target substance). However, microorganisms toxicity data are available for sodium tungstate (source substance), which will be used for read-across. Due to lower water solubility and lower toxicity for the target substance compared to the source substance, the resulting read-across from the source substance to the target substance is appropriate as a conservative estimate of potential toxicity for this endpoint. In addition, read-across is appropriate because the classification and labelling is more protective for the source substance than the target substance, the PBT/vPvB profile is the same, and the dose descriptors are, or are expected to be, lower for the source substance. For more details, refer to the attached description of the read-across approach.

Description of key information

No toxicity data on microorganisms are available for tungstic acid; therefore, data for WO3 and sodium tungstate were evaluated for this endpoint. For WO3, a 3-h respiration inhibition test with activated sludge (predominantly domestic) is available, which determined an EC50 of >10,000 mgWO3/L (ca. >7900 mg W/L), based on nominal concentrations. In a 30-min respiration inhibition test with domestic activated sludge, an EC50 of > 1000 mg/L was reported for sodium tungstate (> 586 mg W/L). The value for sodium tungstate will be used in the derivation of the PNECSTP.

Key value for chemical safety assessment

EC50 for microorganisms:
586 mg/L

Additional information

No toxicity data on sewage microorganisms are available for tungstic acid (target substance). However, data are available for WO3 and sodium tungstate are available, which were evaluated for derivation of the PNEC. Tungstic acid (WO3•H2O) is the hydrated form of tungsten trioxide (WO3). Based on Annex V, hydrates and water free forms (anhydrous) of compounds can be regarded as the same substance for registration purposes. Therefore, tungstic acid will be considered equivalent to WO3. For more details on the rationale, refer to the attached description of the read-across approach. For more details on the read-across rationale to WO3 and sodium tungstate, refer to the read-across category approach document (Annex 3).