2-Octen-1-ylsuccinic anhydride, mixture of cis and trans

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

activated sludge respiration inhibition testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

April 28-28, 1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

A category approach will be used for the hazard assessment of several endpoints. The hypothesis for the category of C8-12 Alkenyl Succinic Anhydrides is that data can be read-across among members of the category because their properties and behaviours are similar, based on common functional groups and similar breakdown products, and based on a constant pattern in changing of the potency of properties of the various carbon chain lengths. These features are in accordance with Annex XI, Section 1.5, of Regulation EC No. 1907/2006.  Common functional groups are: a dihydro-2,5-Furandione (cyclic anhydride) ring, carbon chain of length 8 to 12 carbons, with or without branching alkyl groups, and a single carbon-carbon double bond in the carbon chain, location unspecified. There are no additional functional groups which would contribute incremental or different toxicity.
 
For each category member, the breakdown product is the dioic acid of the corresponding anhydride; these dioic acids also have common functional groups. A constant pattern may also be displayed in acute toxicity, dermal irritancy and biodegradation, with the lowest carbon chain length (C8) displaying the highest activity. Irritation, toxicity and degradation potential diminish with increasing carbon chain length. Read-across among the category members is substantiated by the common behaviour in physico-chemical and toxicity behaviours, as provided in the Chemical Category Report Format (CCRF) attached to the IUCLID file. It is adequate to fulfil the information requirements of Annex IX, to be the basis for classification and labelling decisions, and for risk assessment.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

activated sludge, domestic

Details on inoculum:

The test system was a mixture of the microbial inoculum, the synthetic sewage, and water. Activated sludge from a domestic wastewater treatment plant (Mentor, OH) was used as the microbial inoculum. The sludge was washed thoroughly and resuspended in water at the specific mixed liquor suspended solid level (4 g/L ± 10%) before used in the test. The synthetic sewage feed was added to provide the nutrient for the microbial respiration. The activity of the sludge was checked both as blank control and with a reference substance 3,5-dichlorophenol, which is a known inhibitor.

The inoculum used was the return sludge collected from a local waste water treatment plant (Mentor, Ohio). On return to the laboratory the sludge was centrifuged and the supernatant decanted. The sludge was then washed in water three times by re-suspension and centrifugation. A small amount of the washed sludge was dried and weighed. From this result one calculates the amount of wet sludge which must be suspended in water in order to obtain an activated sludge with a mixed liquor suspended solids level of 4 g/L (± 10%). This level gives a concentration of 1.6 g/L in the test medium with the test procedure described below.
If the sludge could not be used on the day of collection, 50 mL synthetic sewage was added to each liter of the activated sludge prepared above; this was then aerated overnight at room temperature. It was then kept aerated for use during the day. Before use the pH was checked to be within 6.0-8.0 (using pH paper).
If the same batch of sludge was required to be used on subsequent days (maximum four days), a further 25-50 mL of synthetic sewage feed was added to each liter of the sludge at the end of each working day.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Hardness:

.

Test temperature:

Not monitored.

pH:

Before use, the pH of the activated sludge was checked to be within 6.0-8.0 (using pH paper). pH of the reference solution was in the range of 7 to 8.

Nominal and measured concentrations:

The targeted test concentrations for the test substance were 1, 10, 100, 1,000 and 10,000 ppm (mg/L).

Reference substance (positive control):

yes

Remarks:

3,5 dichlorophenol - reference substance obtained from Aldrich Chemical Company (Milwaukee, WI), Lot#00410KZ. Three concentrations of this substance were used in the test: 10, 20, and 40 ppm.

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

800 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

The test substance OS#18230, up to the concentration of 100 ppm, did not show significant toxicity toward the microbial population. At the concentration of 1,000 and 10,000 ppm, however, the test substance did cause substantial inhibition (62% and 85%) of the microbial respiration. The EC50 was estimated to be at 800 ppm and the NOEC estimated to be at 100ppm. Therefore, the test substance is non-inhibitory at concentrations lower than 100ppm. OS#18230 does inhibit microbial respiration at higher concentrations with an estimated EC50 of 800 ppm.

Results with reference substance (positive control):

The EC50 (3 hours) of the reference substance, 3,5-dichlorphenol, is in the accepted range of 5 to 30 mg/L.

Reported statistics and error estimates:

Statistics:
1) The respiration rate is calculated from the recorder trace as mg O2/L.hr. The portion of the respiration curve over which the respiration rate is measured should be linear.
2) The following equation is used to calculate the inhibitory effect of a test/reference substance at a particular concentration.

(1-2Rs/(Rc1 + Rc2))* 100 = per cent inhibition
Where :
Rs = oxygen consumption rate at tested concentration of substance
Rc1 = oxygen consumption rate, Control 1
Rc2 = oxygen consumption rate, Control 2

3) Estimate the range of concentrations where EC50 value for the reference substance (3,5-dicholorphenol) resides in.
4) Determine the NOEC of the test substance as the highest test concentration which observed inhibition less than or equal to 15%.
5) Determine the EC50 of the test substance by plotting the “percent inhibition” employing a linear Y axis versus the concentration of the test substance on a logarithmic X axis. Visually fit a straight line through the apparent slope of the data points were concentration-effect was observed. (Since only two or three data points were involved in the line fitting, no statistical method was used.) The EC50 was the concentration of the test candidate that resulted in a 50%
inhibition of respiration.

Validity criteria fulfilled:

yes

Remarks:

Respiration rates of the two controls were within 15% of each other and the EC50 of the reference substance, 3,5 - dichlorophenol, was between 10 and 20 ppm. Therefore, both criteria for validity check were met.

Conclusions:

The test substance is non-inhibitory at concentrations lower than 100ppm (NOEC estimated at 100 mg/L). OS#18230 does inhibit microbial respiration at higher concentrations with an estimated EC50 of 800 ppm. Data can be read-across among members of the C8-12 Alkenyl Succinic Anhydrides Category, based on common functional groups, similar break-down products and potency patterns among carbon-chain length. This is adequate to fulfil the information requirements, to be the basis for classification and labelling decisions, and for risk assessment.

Executive summary:

In the study entitled “Activated Sludge, Respiration Inhibition Screening Test on Test Substance OS-18230” (Kung – Ricerca, 1995) and based on OECD Guideline Method 209, activated sludge obtained from a domestic sewage treatment plant is used as the microbial source to test the ecotoxicity of OS#18230 (CAS RN 26544-38 -7). The effects of the test substance on micro-organisms was determined by measuring the microbial respiration rate under defined conditions in the presence of different concentrations of the test substance (1, 10, 100, 1,000 and 10,000 mg/L).

The test substance was determined to be non-inhibitory to microbial respiration at concentrations lower than 100 ppm (NOEC estimated at 100 mg/L). However, OS#18230 does inhibit microbial respiration at higher concentrations with an estimated EC₅₀ of 800 ppm.

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

April 28-28, 1995

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

activated sludge, domestic

Details on inoculum:

The test system was a mixture of the microbial inoculum, the synthetic sewage, and water. Activated sludge from a domestic wastewater treatment plant (Mentor, OH) was used as the microbial inoculum. The sludge was washed thoroughly and resuspended in water at the specific mixed liquor suspended solid level (4 g/L ± 10%) before used in the test. The synthetic sewage feed was added to provide the nutrient for the microbial respiration. The activity of the sludge was checked both as blank control and with a reference substance 3,5-dichlorophenol, which is a known inhibitor.

The inoculum used was the return sludge collected from a local waste water treatment plant (Mentor, Ohio). On return to the laboratory the sludge was centrifuged and the supernatant decanted. The sludge was then washed in water three times by re-suspension and centrifugation. A small amount of the washed sludge was dried and weighed. From this result one calculates the amount of wet sludge which must be suspended in water in order to obtain an activated sludge with a mixed liquor suspended solids level of 4 g/L (± 10%). This level gives a concentration of 1.6 g/L in the test medium with the test procedure described below.
If the sludge could not be used on the day of collection, 50 mL synthetic sewage was added to each liter of the activated sludge prepared above; this was then aerated overnight at room temperature. It was then kept aerated for use during the day. Before use the pH was checked to be within 6.0-8.0 (using pH paper).
If the same batch of sludge was required to be used on subsequent days (maximum four days), a further 25-50 mL of synthetic sewage feed was added to each liter of the sludge at the end of each working day.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Hardness:

.

Test temperature:

Not monitored.

pH:

Before use, the pH of the activated sludge was checked to be within 6.0-8.0 (using pH paper). pH of the reference solution was in the range of 7 to 8.

Nominal and measured concentrations:

The targeted test concentrations for the test substance were 1, 10, 100, 1,000 and 10,000 ppm (mg/L).

Reference substance (positive control):

yes

Remarks:

3,5 dichlorophenol - reference substance obtained from Aldrich Chemical Company (Milwaukee, WI), Lot#00410KZ. Three concentrations of this substance were used in the test: 10, 20, and 40 ppm.

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

800 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

The test substance OS#18230, up to the concentration of 100 ppm, did not show significant toxicity toward the microbial population. At the concentration of 1,000 and 10,000 ppm, however, the test substance did cause substantial inhibition (62% and 85%) of the microbial respiration. The EC50 was estimated to be at 800 ppm and the NOEC estimated to be at 100ppm. Therefore, the test substance is non-inhibitory at concentrations lower than 100ppm. OS#18230 does inhibit microbial respiration at higher concentrations with an estimated EC50 of 800 ppm.

Results with reference substance (positive control):

The EC50 (3 hours) of the reference substance, 3,5-dichlorphenol, is in the accepted range of 5 to 30 mg/L.

Reported statistics and error estimates:

Statistics:
1) The respiration rate is calculated from the recorder trace as mg O2/L.hr. The portion of the respiration curve over which the respiration rate is measured should be linear.
2) The following equation is used to calculate the inhibitory effect of a test/reference substance at a particular concentration.

(1-2Rs/(Rc1 + Rc2))* 100 = per cent inhibition
Where :
Rs = oxygen consumption rate at tested concentration of substance
Rc1 = oxygen consumption rate, Control 1
Rc2 = oxygen consumption rate, Control 2

3) Estimate the range of concentrations where EC50 value for the reference substance (3,5-dicholorphenol) resides in.
4) Determine the NOEC of the test substance as the highest test concentration which observed inhibition less than or equal to 15%.
5) Determine the EC50 of the test substance by plotting the “percent inhibition” employing a linear Y axis versus the concentration of the test substance on a logarithmic X axis. Visually fit a straight line through the apparent slope of the data points were concentration-effect was observed. (Since only two or three data points were involved in the line fitting, no statistical method was used.) The EC50 was the concentration of the test candidate that resulted in a 50%
inhibition of respiration.

Validity criteria fulfilled:

yes

Remarks:

Respiration rates of the two controls were within 15% of each other and the EC50 of the reference substance, 3,5 - dichlorophenol, was between 10 and 20 ppm. Therefore, both criteria for validity check were met.

Conclusions:

The test substance is non-inhibitory at concentrations lower than 100ppm (NOEC estimated at 100 mg/L). OS#18230 does inhibit microbial respiration at higher concentrations with an estimated EC50 of 800 ppm. Data can be read-across among members of the C8-12 Alkenyl Succinic Anhydrides Category, based on common functional groups, similar break-down products and potency patterns among carbon-chain length. This is adequate to fulfil the information requirements, to be the basis for classification and labelling decisions, and for risk assessment.

Executive summary:

In the study entitled “Activated Sludge, Respiration Inhibition Screening Test on Test Substance OS-18230” (Kung – Ricerca, 1995) and based on OECD Guideline Method 209, activated sludge obtained from a domestic sewage treatment plant is used as the microbial source to test the ecotoxicity of OS#18230 (CAS RN 26544-38 -7). The effects of the test substance on micro-organisms was determined by measuring the microbial respiration rate under defined conditions in the presence of different concentrations of the test substance (1, 10, 100, 1,000 and 10,000 mg/L).

The test substance was determined to be non-inhibitory to microbial respiration at concentrations lower than 100 ppm (NOEC estimated at 100 mg/L). However, OS#18230 does inhibit microbial respiration at higher concentrations with an estimated EC₅₀ of 800 ppm.

Description of key information

The toxicity of the substance to micro-organisms was evaluated in an activated sludge respiration inhibition test conducted on tetrapropenyl succinic anhydride in accordance with OECD Guideline 209, by read-across based on a grouping of substances (category approach).

Key value for chemical safety assessment

EC50 for microorganisms:

800 mg/L

EC10 or NOEC for microorganisms:

100 mg/L

Additional information

The study “Activated Sludge, Respiration Inhibition Screening Test on Test Substance OS-18230” (Kung, Ricerca, 1995), in which tetrapropenyl succinic anhydride was evaluated based on OECD Guideline Method 209, was identified as the key study,by read-across based on a grouping of substances (category approach). The activated sludge obtained from a domestic sewage treatment plant was used as the microbial source to test the toxicity of the substance. The effects of the test substance on micro-organisms was determined by measuring the microbial respiration rate under defined conditions in the presence of different concentrations of the test substance (1, 10, 100, 1,000 and 10,00 mg/L). The test substance was determined to be non-inhibitory to microbial respiration at concentrations lower than 100 ppm (NOEC estimated at 100 mg/L). However, the test substance was shown to inhibit microbial respiration at higher concentrations with an estimated EC₅₀of 800 ppm (mg/L); this EC₅₀ value will be used to represent the substance, by read-across based on a grouping of substances (category approach).

A category approach is used for the hazard assessment of several endpoints. The hypothesis for the category of C8-12 Alkenyl Succinic Anhydrides is that data can be read-across among members of the category, because the properties and behaviours of category members are similar, based on common functional groups, similar breakdown products, and demonstration of a constant pattern associating the potency of properties with the various carbon chain lengths. Functional groups include a dihydro-2,5-furandione cyclic anhydride ring, a carbon chain of length 8 -12 carbons, and a single carbon-carbon double-bond within the carbon chain. The primary functional group associated with toxicity is the succinic anhydride moiety, which is quickly hydrolysed to form a butanedioic acid. A constant pattern may also be displayed in acute toxicity, dermal irritancy and biodegradation, with the lowest carbon chain length (C8) displaying the highest activity. Irritation, toxicity and degradation potential diminish with increasing carbon chain length. Read-across among the category members is substantiated by the common behaviour in physico-chemical and toxicity behaviours, as provided in the Chemical Category Report Format (CCRF) attached to the IUCLID file. It is adequate to fulfil the information requirements of Annex IX, to be the basis for classification and labelling decisions, and for risk assessment.