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Toxicity to microorganisms

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Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1992
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Qualifier:
according to
Guideline:
other: Method B-21
Principles of method if other than guideline:
Not available
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None
Analytical monitoring:
not specified
Details on sampling:
Not available
Vehicle:
not specified
Details on test solutions:
Not available
Test organisms (species):
other:
Details on inoculum:
Not available
Test type:
not specified
Water media type:
not specified
Limit test:
no
Remarks on exposure duration:
No data
Post exposure observation period:
Not available
Hardness:
Not available
Test temperature:
22.2 °C
pH:
6.8
Dissolved oxygen:
Not available
Salinity:
Not available
Nominal and measured concentrations:
Not available
Details on test conditions:
Not available
Reference substance (positive control):
not specified
Duration:
3 h
Dose descriptor:
IC50
Effect conc.:
> 320 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
not specified
Remarks on result:
other: non toxic
Details on results:
No data
Results with reference substance (positive control):
3,5-Dichlorophenol IC50 (3 h) = 11.0 mg/L
Reported statistics and error estimates:
No data

None

Validity criteria fulfilled:
not specified
Conclusions:
IC50 : >320 mg/L
Executive summary:

IC50 : >320 mg/L

Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1984
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Qualifier:
no guideline followed
Principles of method if other than guideline:
Not available
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None
Analytical monitoring:
not specified
Details on sampling:
Not available
Vehicle:
not specified
Details on test solutions:
Not available
Test organisms (species):
other:
Details on inoculum:
Not available
Test type:
other: Suspension test
Water media type:
not specified
Limit test:
no
Remarks on exposure duration:
No data
Post exposure observation period:
Not available
Hardness:
Not available
Test temperature:
Not available
pH:
Not available
Dissolved oxygen:
Not available
Salinity:
Not available
Nominal and measured concentrations:
Not available
Details on test conditions:
Not available
Reference substance (positive control):
not specified
Dose descriptor:
other: BST
Effect conc.:
> 300 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
not specified
Remarks on result:
other: non toxic
Details on results:
No data
Results with reference substance (positive control):
No data
Reported statistics and error estimates:
No data

None

Validity criteria fulfilled:
not specified
Conclusions:
Toxicity to activated sludge: > 300 mg/L
Executive summary:

Toxicity to activated sludge: > 300 mg/L

Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1992
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
no
Principles of method if other than guideline:
Guidelines followed
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None
Analytical monitoring:
not specified
Details on sampling:
Not available
Vehicle:
not specified
Details on test solutions:
Not available
Test organisms (species):
other:
Details on inoculum:
Not available
Test type:
not specified
Water media type:
not specified
Limit test:
no
Remarks on exposure duration:
No data
Post exposure observation period:
Not available
Hardness:
Not available
Test temperature:
Not available
pH:
Not available
Dissolved oxygen:
Not available
Salinity:
Not available
Nominal and measured concentrations:
Not available
Details on test conditions:
Not available
Reference substance (positive control):
not specified
Duration:
3 h
Dose descriptor:
IC50
Effect conc.:
> 320 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
not specified
Remarks on result:
other: non toxic
Details on results:
No data
Results with reference substance (positive control):
Not available
Reported statistics and error estimates:
No data

None

Validity criteria fulfilled:
not specified
Conclusions:
IC50 : > 320 mg/L
Executive summary:

IC50 : > 320 mg/L

Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1975
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Qualifier:
no guideline followed
Principles of method if other than guideline:
Not available
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None
Analytical monitoring:
not specified
Details on sampling:
No data
Vehicle:
not specified
Details on test solutions:
No data
Test organisms (species):
activated sludge
Details on inoculum:
Bacteria:
Bacteria used have to be isolated from sewage sludge. The original German document provided as an annex to this paper lists a number of gram positive and gram negative bacteria that need to be screened.

Bacterial pre-cultures:
Once per week a pre-culture is prepared taking one inoculation loop of bacteria from an agar plate (BHI-agar) to be cultivated in Nutrient-Broth medium. Each day one inoculation loop from this pre-culture is used to inoculate 5 ml of Nutrient-Broth which is then cultured at 22 °C.

Active sewage sludge or activated sludge:
This is taken from the aerated compartment of an Husmann apparatus and has to be prepared freshly on a weekly basis.
Test type:
other: suspension test
Water media type:
not specified
Limit test:
no
Total exposure duration:
24 h
Remarks on exposure duration:
No data
Post exposure observation period:
No data
Hardness:
No data
Test temperature:
25 ± 2 °C
pH:
No data
Dissolved oxygen:
No data
Salinity:
No data
Nominal and measured concentrations:
No data
Details on test conditions:
Suspension Test:
Test tubes are inoculated with one drop of the bacteria sewage sludge suspension and incubated for 12-24 hours at 25 ± 2 °C in an incubator

TEST CONCENTRATIONS
Tested are 1, 5, 10, 20, 50, 100, 200 and 300 ppm of a test substance. In special cases 1, 3.2, 10, 32, 100, 320 and 1000 ppm are used (as indicated in the short report). A dilution scheme is attached to the original German version of the method description. 0.2 ml of the dilution is mixed with 15 ml of liquid nutrient agar (tempered to 45 °C) and plated on sterile petri dishes (agar incorporation test). Solvent and growth controls have to be used in parallel. In case of the suspension test 0.1 ml of dilutions are mixed with 4.5 ml of Nutrient broth in a test tube.
Reference substance (positive control):
not specified
Duration:
24 h
Dose descriptor:
other: BST
Effect conc.:
> 300 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
not specified
Details on results:
No data
Results with reference substance (positive control):
No data
Reported statistics and error estimates:
No data

No data

Validity criteria fulfilled:
not specified
Conclusions:
BST >300 mg/l
Executive summary:

BST >300 mg/l

Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1982
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Qualifier:
no guideline followed
Principles of method if other than guideline:
Not available
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None
Analytical monitoring:
not specified
Details on sampling:
No data
Vehicle:
not specified
Details on test solutions:
No data
Test organisms (species):
activated sludge
Details on inoculum:
Bacteria:
Bacteria used have to be isolated from sewage sludge. The original German document provided as an annex to this paper lists a number of gram positive and gram negative bacteria that need to be screened.

Bacterial pre-cultures:
Once per week a pre-culture is prepared taking one inoculation loop of bacteria from an agar plate (BHI-agar) to be cultivated in Nutrient-Broth medium. Each day one inoculation loop from this pre-culture is used to inoculate 5 ml of Nutrient-Broth which is then cultured at 22 °C.

Active sewage sludge or activated sludge:
This is taken from the aerated compartment of an Husmann apparatus and has to be prepared freshly on a weekly basis.
Test type:
other: suspension test
Water media type:
not specified
Limit test:
no
Total exposure duration:
24 h
Remarks on exposure duration:
No data
Post exposure observation period:
No data
Hardness:
No data
Test temperature:
25 ± 2 °C
pH:
No data
Dissolved oxygen:
No data
Salinity:
No data
Nominal and measured concentrations:
No data
Details on test conditions:
Suspension Test:
Test tubes are inoculated with one drop of the bacteria sewage sludge suspension and incubated for 12-24 hours at 25 ± 2 °C in an incubator

TEST CONCENTRATIONS
Tested are 1, 5, 10, 20, 50, 100, 200 and 300 ppm of a test substance. In special cases 1, 3.2, 10, 32, 100, 320 and 1000 ppm are used (as indicated in the short report). A dilution scheme is attached to the original German version of the method description. 0.2 ml of the dilution is mixed with 15 ml of liquid nutrient agar (tempered to 45 °C) and plated on sterile petri dishes (agar incorporation test). Solvent and growth controls have to be used in parallel. In case of the suspension test 0.1 ml of dilutions are mixed with 4.5 ml of Nutrient broth in a test tube.
Reference substance (positive control):
not specified
Duration:
24 h
Dose descriptor:
other: BST
Effect conc.:
> 300 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
not specified
Details on results:
No data
Results with reference substance (positive control):
No data
Reported statistics and error estimates:
No data

No data

Validity criteria fulfilled:
not specified
Conclusions:
BST >300 mg/l
Executive summary:

BST >300 mg/l

Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1983
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Qualifier:
according to
Guideline:
other: Method B-10
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None
Analytical monitoring:
not specified
Details on sampling:
No data
Vehicle:
not specified
Details on test solutions:
No data
Test organisms (species):
activated sludge
Details on inoculum:
Bacteria:
Bacteria used have to be isolated from sewage sludge. The original German document provided as an annex to this paper lists a number of gram positive and gram negative bacteria that need to be screened.

Bacterial pre-cultures:
Once per week a pre-culture is prepared taking one inoculation loop of bacteria from an agar plate (BHI-agar) to be cultivated in Nutrient-Broth medium. Each day one inoculation loop from this pre-culture is used to inoculate 5 ml of Nutrient-Broth which is then cultured at 22 °C.

Active sewage sludge or activated sludge:
This is taken from the aerated compartment of an Husmann apparatus and has to be prepared freshly on a weekly basis.
Test type:
other: suspension test
Water media type:
not specified
Limit test:
no
Total exposure duration:
24 h
Remarks on exposure duration:
No data
Post exposure observation period:
No data
Hardness:
No data
Test temperature:
25 ± 2 °C
pH:
No data
Dissolved oxygen:
No data
Salinity:
No data
Nominal and measured concentrations:
No data
Details on test conditions:
Suspension Test:
Test tubes are inoculated with one drop of the bacteria sewage sludge suspension and incubated for 12-24 hours at 25 ± 2 °C in an incubator

TEST CONCENTRATIONS
Tested are 1, 5, 10, 20, 50, 100, 200 and 300 ppm of a test substance. In special cases 1, 3.2, 10, 32, 100, 320 and 1000 ppm are used (as indicated in the short report). A dilution scheme is attached to the original German version of the method description. 0.2 ml of the dilution is mixed with 15 ml of liquid nutrient agar (tempered to 45 °C) and plated on sterile petri dishes (agar incorporation test). Solvent and growth controls have to be used in parallel. In case of the suspension test 0.1 ml of dilutions are mixed with 4.5 ml of Nutrient broth in a test tube.
Reference substance (positive control):
not specified
Duration:
24 h
Dose descriptor:
other: BST
Effect conc.:
> 300 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
not specified
Details on results:
No data
Results with reference substance (positive control):
No data
Reported statistics and error estimates:
No data

No data

Validity criteria fulfilled:
not specified
Conclusions:
BST >300 mg/l
Executive summary:

BST >300 mg/l

Endpoint:
toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1984
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Qualifier:
according to
Guideline:
other: Method B-10
Principles of method if other than guideline:
Not available
GLP compliance:
no
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
None
Analytical monitoring:
not specified
Details on sampling:
No data
Vehicle:
not specified
Details on test solutions:
No data
Test organisms (species):
activated sludge
Details on inoculum:
Bacteria:
Bacteria used have to be isolated from sewage sludge. The original German document provided as an annex to this paper lists a number of gram positive and gram negative bacteria that need to be screened.

Bacterial pre-cultures:
Once per week a pre-culture is prepared taking one inoculation loop of bacteria from an agar plate (BHI-agar) to be cultivated in Nutrient-Broth medium. Each day one inoculation loop from this pre-culture is used to inoculate 5 ml of Nutrient-Broth which is then cultured at 22 °C.

Active sewage sludge or activated sludge:
This is taken from the aerated compartment of an Husmann apparatus and has to be prepared freshly on a weekly basis.
Test type:
other: suspension test
Water media type:
not specified
Limit test:
no
Total exposure duration:
24 h
Remarks on exposure duration:
No data
Post exposure observation period:
No data
Hardness:
No data
Test temperature:
25 ± 2 °C
pH:
No data
Dissolved oxygen:
No data
Salinity:
No data
Nominal and measured concentrations:
No data
Details on test conditions:
Suspension Test:
Test tubes are inoculated with one drop of the bacteria sewage sludge suspension and incubated for 12-24 hours at 25 ± 2 °C in an incubator

TEST CONCENTRATIONS
Tested are 1, 5, 10, 20, 50, 100, 200 and 300 ppm of a test substance. In special cases 1, 3.2, 10, 32, 100, 320 and 1000 ppm are used (as indicated in the short report). A dilution scheme is attached to the original German version of the method description. 0.2 ml of the dilution is mixed with 15 ml of liquid nutrient agar (tempered to 45 °C) and plated on sterile petri dishes (agar incorporation test). Solvent and growth controls have to be used in parallel. In case of the suspension test 0.1 ml of dilutions are mixed with 4.5 ml of Nutrient broth in a test tube.
Reference substance (positive control):
not specified
Duration:
24 h
Dose descriptor:
other: BST
Effect conc.:
> 300 mg/L
Nominal / measured:
not specified
Conc. based on:
not specified
Basis for effect:
not specified
Details on results:
No data
Results with reference substance (positive control):
No data
Reported statistics and error estimates:
No data

No data

Validity criteria fulfilled:
not specified
Conclusions:
BST >300 mg/l
Executive summary:

BST >300 mg/l

Endpoint:
activated sludge respiration inhibition testing
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given, acceptable for assessment
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that the source and the target substance have very similar physicochemical and (eco)toxicological properties because their chemical structures are nearly identical. An analogue approach has thus been employed. The target substance is the meta-isomer of the dye Reactive Blue 049, where the sulphonate group is bound at the meta-position of the aminobenzene moiety. The source chemical is the reaction mass of both the meta-isomer and the para-isomer of Reactive Blue 049.
The presence of sulphonate groups make both dyes highly water soluble and therefore less critical for human health and environmental issues. Based on their chemical similarity, similar properties are expected in both humans and the environment.

2. SOURCE AND TARGET CHEMICAL(S)
Source: Reactive Blue 49 meta/para (CAS# 72214-18-7 / EC# 276-481-8)
Target: Reactive Blue 49 meta (CAS# 72927-99-2 / EC# 277-040-2)

3. ANALOGUE APPROACH JUSTIFICATION
see attachment under 4.2 Melting point / freezing point
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Reference substance (positive control):
yes
Duration:
3 h
Dose descriptor:
IC50
Effect conc.:
> 300 mg/L
Conclusions:
The weight-of-evidence approach used for the source substance describes seven internal Huntsman studies. Only basic data is given. The reported BST is the lowest concentration which clearly inhibits bacterial growth, based on visible opacity of the test after 24 hours.
solution. In five studies, the BST is reported to be >300 mg/L. In the two other studies, the 3h-IC50 is reported to be >320 mg/L.
The structurally related target substance will show the same behaviour and therefore it can be anticipated that the IC50 will be >300 mg/L

Description of key information

IC50 > 300 mg/L

Key value for chemical safety assessment

EC50 for microorganisms:
300 mg/L

Additional information

The weight-of-evidence approach used for the source substance describes seven internal Huntsman studies. Only basic data is given. The reported BST is the lowest concentration which clearly inhibits bacterial growth, based on visible opacity of the test

solution after 24 hours. In five studies, the BST is reported to be >300 mg/L. In the two other studies, the 3h-IC50 is reported to be >320 mg/L.

The structurally related target substance will show the same behaviour and therefore it can be anticipated that the IC50 will be >300 mg/L.