N-phenylprop-2-enamide

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: screening tests

Type of information:

(Q)SAR

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

results derived from a valid (Q)SAR model, but not (completely) falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

1. SOFTWARE : VEGA v1.1.4

2. MODEL (incl. version number) : Ready Biodegradability Model (IRFMN) v 1.0.9

3. SMILES OR OTHER IDENTIFIERS USED AS INPUT FOR THE MODEL : C=CC(=O)Nc1ccccc1

4. SCIENTIFIC VALIDITY OF THE (Q)SAR MODEL
[Explain how the model fulfils the OECD principles for (Q)SAR model validation. Consider attaching the QMRF or providing a link]
See attached QMRF

5. APPLICABILITY DOMAIN
[Explain how the substance falls within the applicability domain of the model]
See the attached QPRF for a detailed list of related substances in the training dataset.
The QSAR indicates that the substance fall possibly outside of the applicability domain. However, looking at the structural analogues in the QSAR training set, al but one of the 6 substances are readily biodegradable (experimental value) and correctly predicted to be (possibly) ready biodegradable. Because there is a clear similarity with these substances and the majority of these substances are correctly predicted, the QSAR prediction is considered reliable.

6. ADEQUACY OF THE RESULT
[Explain how the prediction fits the purpose of classification and labelling and/or risk assessment]

Qualifier:

according to guideline

Guideline:

other: Guidance on information requirements and chemical safety assessment Chapter R.6: QSARs and grouping of chemicals

GLP compliance:

not specified

Remarks on result:

readily biodegradable based on QSAR/QSPR prediction

Remarks:

Possible readily biodegradable

Validity criteria fulfilled:

not applicable

Interpretation of results:

readily biodegradable

Conclusions:

According to the Ready Biodegradability Model (IRFMN) v1.0.9, the substance is possible readily biodegradable.

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Deviations:

yes

Remarks:

Short deviation (4h) of guideline temperature range was deemed to have negligible effects on study results

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

yes

Remarks:

Short deviation (4h) of guideline temperature range was deemed to have negligible effects on study results

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): municipal sewage treatment plant 'Waterschap de Maaskant', 's Hertogenbosch, the Netherlands
- Laboratory culture: no
- Method of cultivation: not applicable
- Storage conditions: The sludge was kept under continuous aeration until further treatment.
- Storage length: not reported
- Preparation of inoculum for exposure: Before use, the sludge was allowed to settle (39-60 minutes) and the liquid was decanted for use as inoculum at the amount of 10 ml/L of mineral medium.
- Pretreatment: not applicable
- Concentration of sludge: concentration suspended solids: 4.4 (first test) and 3.7 g/L (second test)
- Initial cell/biomass concentration:
- Water filtered: yes
- Type and size of filter used, if any: tap-water was purified by reverse osmosis (milli-RO) and subsequently passed over activated carbon and ion-exchange cartridges (Milli-Q)

Duration of test (contact time):

28 d

Initial conc.:

12 mg/L

Based on:

TOC

Remarks:

based on the molecular formula

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: 1 litre mineral medium contains: 10 mL of solution (A), 1 mL of solutions (B) to (D) and Milli-RO water.
A) 8.50 g KH2PO4
21.75 g K2HPO4
67.20 g Na2HPO4.12H2O
0.50 g NH4Cl
dissolved in Milli-RO water and made up to 1 litre, pH 7.4 ± 0.2
B)22.50 g MgSO4.7H2O dissolved in Milli-RO water and made up to 1 litre.
C)36.40 g CaCl2.2H2O dissolved in Milli-RO water and made up to 1 litre.
D)0.25 g FeCl3.6H2O dissolved in Milli-RO water and made up to 1 litre.
- Additional substrate: /
- Solubilising agent (type and concentration if used): /
- Test temperature:
first test: 22.2 - 23.9 °C, short period (4h) exceeded 24°C due to technical malfunction
second test: 21.5 - 23.0 °C
- pH: 7.4-7.9
- pH adjusted: no
- CEC (meq/100 g): /
- Aeration of dilution water: A mixture of oxygen (ca. 21%) and nitrogen (ca. 79%) was passed through a bottle, containing 0.5 - 1 litre 0.0125 M Ba(OH)2 solution to trap CO2 which might be present in small amounts. The synthetic air was sparged through the scrubbing solutions at a rate of approximately 1-2 bubbles per second (ca. 30-100 mL/min).
- Suspended solids concentration: 4.4 (first test) and 3.7 g/L (second test)
- Continuous darkness: yes
- Other: /

TEST SYSTEM
- Culturing apparatus: /
- Number of culture flasks/concentration:
Test suspension: containing test item and inoculum (2 bottles).
Inoculum blank: containing only inoculum (2 bottles)
Positive control: containing reference item and inoculum (1 bottle).
Toxicity control: containing test item, reference item and inoculum (1 bottle).
- Method used to create aerobic conditions: A mixture of oxygen (ca. 21%) and nitrogen (ca. 79%) was passed through a bottle, containing 0.5 - 1 litre 0.0125 M Ba(OH)2 solution to trap CO2 which might be present in small amounts. The synthetic air was sparged through the scrubbing solutions at a rate of approximately 1-2 bubbles per second (ca. 30-100 mL/min).
- Method used to create anaerobic conditions: /
- Measuring equipment: The CO2 produced in each test bottle reacted with the barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2 produced was determined by titrating the remaining Ba(OH)2 with 0.05 M standardized HCl (1:20 dilution from 1 M HCl (Titrisol® ampoule), Merck, Darmstadt, Germany).
- Test performed in closed vessels due to significant volatility of test substance: /
- Test performed in open system: /
- Details of trap for CO2 and volatile organics if used: At the start of the test (day 0), test and reference item were added to the bottles containing the microbial organisms and mineral components. The volumes of suspensions were made up to 2 litres with Milli-RO water, resulting in the mineral medium described before.
Three CO2-absorbers (bottles filled with 100 mL 0.0125 M Ba(OH)2) were connected in series to the exit air line of each test bottle.
- Other: /

SAMPLING
- Sampling frequency: Titrations were made every second or third day during the first 10 days, and thereafter at least every fifth day until day 28, for the inoculum blank and test item.
- Sampling method: Each time the CO2-absorber nearest to the test bottle was removed for titration; each of the remaining two absorbers were moved one position in the direction of the test bottle. A new CO2-absorber was placed at the far end of the series. Phenolphthalein (1% solution in ethanol, Merck) was used as pH-indicator. On the penultimate day, the pH of all test media was measured and 1 mL of concentrated HCl (37%, Merck) was added to the test bottles. The bottles were aerated overnight to drive off CO2 present and the final titration was made.
- Sterility check if applicable: /
- Sample storage before analysis: /
- Other: /

CONTROL AND BLANK SYSTEM
- Inoculum blank: containing only inoculum (2 bottles)
- Abiotic sterile control: /
- Toxicity control: containing test item, reference item and inoculum (1 bottle).
- Other: Positive control: containing reference item and inoculum (1 bottle).

STATISTICAL METHODS:

Test performance:

In the toxicity control for both tests, 25% degradation did not occur within 14 days. The test substance thus might have an inhibiting effect on bacteria.

Parameter:

% degradation (CO2 evolution)

Remarks:

test 1

Value:

>= 4 - <= 33

Sampling time:

19 d

Parameter:

% degradation (CO2 evolution)

Remarks:

test 1

Value:

>= 55 - <= 80

Sampling time:

28 d

Parameter:

% degradation (CO2 evolution)

Remarks:

test 2

Value:

>= 32 - <= 34

Sampling time:

19 d

Parameter:

% degradation (CO2 evolution)

Remarks:

test 2

Value:

>= 79 - <= 95

Sampling time:

28 d

Details on results:

Because the difference between the two bottles was too high (>20% difference), the test was repeated.

In the second test, the relative degradation values revealed no significant degradation of CH03543 until day 14. After this lag phase, more than 60% biodegradation was reached in both test bottles. However, 60% degradation was reached within a 10-day window in bottle B but not in bottle A. The difference of duplicate values for %-degradation was more than 20 on days 27 and 29.
The biodegradation curves of the second test are included as illustration.

Validity criteria fulfilled:

no

Remarks:

Another repeat of the test was not considered to lead to different results since this variety was substance related due to low water solubility and inhibition of microbial activity at the concentration applied.

Interpretation of results:

other: criterion for ready biodegradability not met, but very close to ready biodegradability

Conclusions:

An OECD 301 (modified Sturm test) test was performed to determine the ready biodegradabilty of CH03543. Because of the high difference between the two replicates, the test was repeated.
Both tests were similar in that degradation only started after >=14 days and that degradation rate was variable. Although the validity criterion for the difference of duplicate values for %-degradation was not met, another repeat of the test was not considered to lead to different results since this variety was test substance related due to low water solubility and inhibition of microbial activity at the concentration applied. Therefore the study was finalised.

After a lag phase of 14-19 days, CH03543 was degradated significantly (80, 56, 67 and 95%) during the test period of 28 days. More than 60% degradation was reached within 6-12 days. In the strict sense of the guidelines, the criterion for ready biodegradability was not met. However, CH03543 should be considerd as being very close to ready biodegradable.

Reference 3

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

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

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH
See attached Read-across document.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Deviations:

yes

Remarks:

Short deviation (4h) of guideline temperature range was deemed to have negligible effects on study results

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

yes

Remarks:

Short deviation (4h) of guideline temperature range was deemed to have negligible effects on study results

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): municipal sewage treatment plant 'Waterschap de Maaskant', 's Hertogenbosch, the Netherlands
- Laboratory culture: no
- Method of cultivation: not applicable
- Storage conditions: The sludge was kept under continuous aeration until further treatment.
- Storage length: not reported
- Preparation of inoculum for exposure: Before use, the sludge was allowed to settle (39-60 minutes) and the liquid was decanted for use as inoculum at the amount of 10 ml/L of mineral medium.
- Pretreatment: not applicable
- Concentration of sludge: concentration suspended solids: 4.4 (first test) and 3.7 g/L (second test)
- Initial cell/biomass concentration:
- Water filtered: yes
- Type and size of filter used, if any: tap-water was purified by reverse osmosis (milli-RO) and subsequently passed over activated carbon and ion-exchange cartridges (Milli-Q)

Duration of test (contact time):

28 d

Initial conc.:

12 mg/L

Based on:

TOC

Remarks:

based on the molecular formula

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: 1 litre mineral medium contains: 10 mL of solution (A), 1 mL of solutions (B) to (D) and Milli-RO water.
A) 8.50 g KH2PO4
21.75 g K2HPO4
67.20 g Na2HPO4.12H2O
0.50 g NH4Cl
dissolved in Milli-RO water and made up to 1 litre, pH 7.4 ± 0.2
B)22.50 g MgSO4.7H2O dissolved in Milli-RO water and made up to 1 litre.
C)36.40 g CaCl2.2H2O dissolved in Milli-RO water and made up to 1 litre.
D)0.25 g FeCl3.6H2O dissolved in Milli-RO water and made up to 1 litre.
- Additional substrate: /
- Solubilising agent (type and concentration if used): /
- Test temperature:
first test: 22.2 - 23.9 °C, short period (4h) exceeded 24°C due to technical malfunction
second test: 21.5 - 23.0 °C
- pH: 7.4-7.9
- pH adjusted: no
- CEC (meq/100 g): /
- Aeration of dilution water: A mixture of oxygen (ca. 21%) and nitrogen (ca. 79%) was passed through a bottle, containing 0.5 - 1 litre 0.0125 M Ba(OH)2 solution to trap CO2 which might be present in small amounts. The synthetic air was sparged through the scrubbing solutions at a rate of approximately 1-2 bubbles per second (ca. 30-100 mL/min).
- Suspended solids concentration: 4.4 (first test) and 3.7 g/L (second test)
- Continuous darkness: yes
- Other: /

TEST SYSTEM
- Culturing apparatus: /
- Number of culture flasks/concentration:
Test suspension: containing test item and inoculum (2 bottles).
Inoculum blank: containing only inoculum (2 bottles)
Positive control: containing reference item and inoculum (1 bottle).
Toxicity control: containing test item, reference item and inoculum (1 bottle).
- Method used to create aerobic conditions: A mixture of oxygen (ca. 21%) and nitrogen (ca. 79%) was passed through a bottle, containing 0.5 - 1 litre 0.0125 M Ba(OH)2 solution to trap CO2 which might be present in small amounts. The synthetic air was sparged through the scrubbing solutions at a rate of approximately 1-2 bubbles per second (ca. 30-100 mL/min).
- Method used to create anaerobic conditions: /
- Measuring equipment: The CO2 produced in each test bottle reacted with the barium hydroxide in the gas scrubbing bottle and precipitated out as barium carbonate. The amount of CO2 produced was determined by titrating the remaining Ba(OH)2 with 0.05 M standardized HCl (1:20 dilution from 1 M HCl (Titrisol® ampoule), Merck, Darmstadt, Germany).
- Test performed in closed vessels due to significant volatility of test substance: /
- Test performed in open system: /
- Details of trap for CO2 and volatile organics if used: At the start of the test (day 0), test and reference item were added to the bottles containing the microbial organisms and mineral components. The volumes of suspensions were made up to 2 litres with Milli-RO water, resulting in the mineral medium described before.
Three CO2-absorbers (bottles filled with 100 mL 0.0125 M Ba(OH)2) were connected in series to the exit air line of each test bottle.
- Other: /

SAMPLING
- Sampling frequency: Titrations were made every second or third day during the first 10 days, and thereafter at least every fifth day until day 28, for the inoculum blank and test item.
- Sampling method: Each time the CO2-absorber nearest to the test bottle was removed for titration; each of the remaining two absorbers were moved one position in the direction of the test bottle. A new CO2-absorber was placed at the far end of the series. Phenolphthalein (1% solution in ethanol, Merck) was used as pH-indicator. On the penultimate day, the pH of all test media was measured and 1 mL of concentrated HCl (37%, Merck) was added to the test bottles. The bottles were aerated overnight to drive off CO2 present and the final titration was made.
- Sterility check if applicable: /
- Sample storage before analysis: /
- Other: /

CONTROL AND BLANK SYSTEM
- Inoculum blank: containing only inoculum (2 bottles)
- Abiotic sterile control: /
- Toxicity control: containing test item, reference item and inoculum (1 bottle).
- Other: Positive control: containing reference item and inoculum (1 bottle).

STATISTICAL METHODS:

Test performance:

In the toxicity control for both tests, 25% degradation did not occur within 14 days. The test substance thus might have an inhibiting effect on bacteria.

Parameter:

% degradation (CO2 evolution)

Remarks:

test 1

Value:

>= 4 - <= 33

Sampling time:

19 d

Parameter:

% degradation (CO2 evolution)

Remarks:

test 1

Value:

>= 55 - <= 80

Sampling time:

28 d

Parameter:

% degradation (CO2 evolution)

Remarks:

test 2

Value:

>= 32 - <= 34

Sampling time:

19 d

Parameter:

% degradation (CO2 evolution)

Remarks:

test 2

Value:

>= 79 - <= 95

Sampling time:

28 d

Details on results:

Because the difference between the two bottles was too high (>20% difference), the test was repeated.

In the second test, the relative degradation values revealed no significant degradation of CH03543 until day 14. After this lag phase, more than 60% biodegradation was reached in both test bottles. However, 60% degradation was reached within a 10-day window in bottle B but not in bottle A. The difference of duplicate values for %-degradation was more than 20 on days 27 and 29.
The biodegradation curves of the second test are included as illustration.

Validity criteria fulfilled:

no

Remarks:

Another repeat of the test was not considered to lead to different results since this variety was substance related due to low water solubility and inhibition of microbial activity at the concentration applied.

Interpretation of results:

other: criterion for ready biodegradability not met, but very close to ready biodegradability

Conclusions:

A structural analogue (CH03543) was found to be almost ready biodegradable. Because of the structural and physicochemical similarities, these results can also be applied to CH03859.

Description of key information

Based on the read-across from a structural analogue, the substance is almost ready biodegradable. To further support this conclusion, a QSAR result (Ready Biodegradability Model (IRFMN) v 1.0.9) is also provided.

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable but failing 10-day window

Type of water:

freshwater