4-methylstyrene

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From November 9, 1979 to November 28, 1979

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

according to guideline

Guideline:

other: Standard Methods for the Examination of Water and Wastewater, 14th edition (1976)

Principles of method if other than guideline:

The method was designed to determine biological oxygen demand (BOD) and chemical oxygen demand (COD). Modifications to the study guideline included the use of a carbon standard and extension of the usual test period from 5 to 20 d. In addition, re-oxygenation was conducted at intervals to maintain a concentration of >2mg/L dissolved oxygen.

GLP compliance:

not specified

Remarks:

At the time this study was conducted GLP was not required for chemical testing.

Oxygen conditions:

aerobic

Remarks:

>2mg/L

Inoculum or test system:

sewage, domestic, non-adapted

Details on inoculum:

no information is provided on the innoculum used.

Duration of test (contact time):

20 d

Initial conc.:

> 2 - < 4 other: uL/300ml

Based on:

test mat.

Parameter followed for biodegradation estimation:

other: BOD

Parameter followed for biodegradation estimation:

other: COD

Details on study design:

The study it was conducted in 300 ml BOD bottles. The test substance was evaluated at two concentrations 2 and 4 uL/300 ml and analysed at 5, 10, 15 and 20 days. The results were compare with results obtained from the glucose and the glucose/glutamic acid control. Re-aeration was conducted as necessary to maintain dissolved oxygen concentrations at levels >2 mg/L.

Key result

Parameter:

% degradation (O2 consumption)

Value:

> 3

Sampling time:

5 d

Key result

Parameter:

% degradation (O2 consumption)

Value:

6

Sampling time:

10 d

Key result

Parameter:

% degradation (O2 consumption)

Value:

13

Sampling time:

15 d

Key result

Parameter:

% degradation (O2 consumption)

Value:

32

Sampling time:

20 d

Details on results:

The BOD showed initial slow by bio-oxidation that increased through time to 32 % after 20 days indicating that the unadapted microbial population in the domestic sewage sample used in the study required time to adapt to degrade the test substance.
The COD expressed as mg oxygen/mg was measured at 2.24 which compared well with a calculated value of 3.1.

Key result

Parameter:

COD

Value:

2.24 g O2/g test mat.

Remarks on result:

other: measured

Results with reference substance:

The glucose/glutamic acid control analysed after five days gave 99.1% of the theoretical oxygen demand indicating that the sewage sample used was performing effectively.

Test substance	COD mg oxygen/mg compound
	Measured	Calculated
purity 97%	2.24	3.1

Test substance	Bio-oxidation (BOD x100)                       ThOD
	Day 5	Day 10	Day 15	Day 20
purity 97%	3.0	6.0	13.0	32.0

The results from an earlier study gave the following data.

Test substance	Bio-oxidation (BOD x100)                       ThOD
	Day 5	Day 10	Day 15	Day 20
purity 97%	3.0	24.0	26.0	34.0

Validity criteria fulfilled:

not specified

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

Under the study conditions, unadapted microorganisms therefore could not degrade the test substance within a 28 d period to a level allowing identification as readily biodegradable. Nevertheless, adapted microorganisms present where continuous exposure to the test substance occurs will readily remove the test substance from sewage systems.

Executive summary:

A study was conducted to determine the Biological Oxygen Demand (BOD) and the Chemical Oxygen Demand (COD) of the test substance according to US Standard Methods for the Examination of Water and Wastewater (1976). The method was modified to include analyses at additional time points after inoculation with domestic sewage sludge (10, 15 and 20 d) in order to determine whether the microorganisms required a time interval to adapt to the new food source (test substance). Exposure to the test substance occurred at a concentration of 2 to 4 µL/300 ml. The control groups confirmed that the inoculum was active and the results indicated that adaptation was necessary in order for domestic sludge microorganisms to utilise the test substance as a substrate. Under the study conditions, 13% of test substance was removed in the first 15 d of the sampling period and 32% was degraded from 15 to 20 d. Under the study conditions, unadapted microorganisms therefore could not degrade the test substance within a 28 d period to a level allowing identification as readily biodegradable. Nevertheless, adapted microorganisms present where continuous exposure to the test substance occurs will readily remove the substance from sewage systems (Gezo, 1980).

Reference 2

Endpoint:

biodegradation in water: screening test, other

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 and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

QSAR prediction from an well known and acknowledged tool. See below under 'Overall remarks, attachments' for applicability domain and 'attached background material section' for methodology.

Qualifier:

according to guideline

Guideline:

other: REACH guidance on QSARs: Chapter R.6. QSARs and grouping of chemicals

GLP compliance:

no

Key result

Parameter:

half-life in days (QSAR/QSPR)

Value:

ca. 3.897

Remarks on result:

readily biodegradable based on QSAR/QSPR prediction

Results

Chemical	SMILES	Molecular weight	BioHCwin results (Half life - days)	domain evaluation
p-methylstyrene	CC1=CC=C(C=C1)C=C	118.18	3.897	In domain - molecular weight and molecular fragments

BioHCwin
SMILES : Cc1ccc(cc1)C=C
CHEM  :	Domain
MOL FOR: C9 H10
MOL WT : 118.18	ID	70.14	478.94
-------------------------- BioHCwin v1.01 Results ---------------------------
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |  LOG Hydrocarbon FRAGMENT DESCRIPTION    | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------
Frag | 1 | Aromatic-CH3                             | 0.0369 | 0.0369	ID	4
Frag | 4 | Aromatic-H                               | 0.0419 | 0.1678	ID	14
Frag | 3 | -C=CH [alkenyl hydrogen]                | -0.0066 | -0.0199	ID	6
Frag | 1 | Benzene                                  | -0.0838 | -0.0838	ID	1
Const| * | Equation Constant                        |        | 0.4898
============+============================================+=========+=========
RESULT  |       LOG BioHC Half-Life (days)         |        | 0.5907
RESULT  |         BioHC Half-Life (days)           |        |  3.897
============+============================================+=========+=========

Validity criteria fulfilled:

not applicable

Interpretation of results:

readily biodegradable

Conclusions:

Based on the BioHCwin model result, the primary biodegradation half-life for the test substance was determined to be 3.897 days and it was considered to be readily biodegradable.

Executive summary:

The biodegradation potential of the test substance, p-methylstyrene, was estimated using the BioHCwin v1.01 program of EPI Suite v4.11. BioHCwin is a new predictive model within EPI Suite, which determines quantitative primary biodegradation half-lives of individual petroleum hydrocarbons. This model uses a fragment-based approach similar to BioWin. Using SMILES code as the input parameter the primary biodegradation half-life for the test substance was determined to be 3.897 days; indicating that the substance can be considered to be readily biodegradable (US EPA, 2018). The prediction for the substance is considered to be reliable, as it falls within of the applicability domain.

 

Reference 3

Endpoint:

biodegradation in water: screening test, other

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 and falling into its applicability domain, with adequate and reliable documentation / justification

Justification for type of information:

QSAR prediction from an well known and acknowledged tool. See below under 'Overall remarks, attachments' for applicability domain and 'attached background material section' for methodology.

Qualifier:

according to guideline

Guideline:

other: REACH guidance on QSARs: Chapter R.6. QSARs and grouping of chemicals

GLP compliance:

no

Key result

Parameter:

other: Biowin1 (Linear Model Prediction)

Remarks on result:

other: Biodegrades fast

Remarks:

(prediction as per EPISuite BioWin 4.11)

Key result

Parameter:

other: Biowin2 (Non-Linear Model Prediction)

Remarks on result:

other: Biodegrades fast

Remarks:

(prediction as per EPISuite BioWin 4.11)

Key result

Parameter:

other: Biowin3 (Ultimate Biodegradation Timeframe):

Remarks on result:

other: Weeks

Remarks:

(prediction as per EPISuite BioWin 4.11)

Key result

Parameter:

other: Biowin4 (Primary Biodegradation Timeframe):

Remarks on result:

other: Days-Weeks

Remarks:

(prediction as per EPISuite BioWin 4.11)

Key result

Parameter:

other: Biowin5 (MITI Linear Model Prediction)

Remarks on result:

other: Not readily degradable

Remarks:

(prediction as per EPISuite BioWin 4.11)

Key result

Parameter:

other: Biowin6 (MITI Non-Linear Model Prediction):

Remarks on result:

other: Not readily degradable

Remarks:

(prediction as per EPISuite BioWin 4.11)

Key result

Parameter:

other: Biowin7 (Anaerobic Model Prediction):

Remarks on result:

other: Does not biodegrade fast

Remarks:

(prediction as per EPISuite BioWin 4.11)

Key result

Parameter:

other: Ready Biodegradability Prediction

Remarks on result:

other: No

Remarks:

(prediction as per EPISuite BioWin v 4.11)

Results

Chemical	SMILES	Molecular weight	Biowin results	domain evaluation
p -methylstyrene	Cc1cccc(c1)C=C	118.18	Biowin1 (Linear Model Prediction): Biodegrades Fast Biowin2 (Non-Linear Model Prediction): Biodegrades Fast Biowin3 (Ultimate Biodegradation Timeframe): Weeks Biowin4 (Primary Biodegradation Timeframe):Days-Weeks Biowin5 (MITI Linear Model Prediction): Not Readily Degradable Biowin6 (MITI Non-Linear Model Prediction): Not Readily Degradable Biowin7 (Anaerobic Model Prediction): Does Not Biodegrade Fast Ready Biodegradability Prediction: NO	In domain - molecular weight and molecular fragments

BIOWIN
SMILES : Cc1cccc(c1)C=C
CHEM  :
MOL FOR: C9 H10		Training set
MOL WT : 118.18	ID	30.02	697.65
--------------------------- BIOWIN v4.10 Results ----------------------------
Biowin1 (Linear Model Prediction)   : Biodegrades Fast
Biowin2 (Non-Linear Model Prediction): Biodegrades Fast
Biowin3 (Ultimate Biodegradation Timeframe): Weeks
Biowin4 (Primary Biodegradation Timeframe): Days-Weeks
Biowin5 (MITI Linear Model Prediction)   : Not Readily Degradable
Biowin6 (MITI Non-Linear Model Prediction): Not Readily Degradable
Biowin7 (Anaerobic Model Prediction): Does Not Biodegrade Fast
Ready Biodegradability Prediction: NO
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |      Biowin1 FRAGMENT DESCRIPTION        | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------		Training set
Frag | 1 | Alkyl substituent on aromatic ring       | 0.0547 | 0.0547	ID	3
MolWt| * | Molecular Weight Parameter               |        | -0.0563
Const| * | Equation Constant                        |        | 0.7475
============+============================================+=========+=========
RESULT  |   Biowin1 (Linear Biodeg Probability)    |        | 0.7460
============+============================================+=========+=========
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |      Biowin2 FRAGMENT DESCRIPTION        | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------		Training set
Frag | 1 | Alkyl substituent on aromatic ring       | 0.5771 | 0.5771	ID	3
MolWt| * | Molecular Weight Parameter               |        | -1.6782
============+============================================+=========+=========
RESULT  | Biowin2 (Non-Linear Biodeg Probability)  |        | 0.8708
============+============================================+=========+=========
A Probability Greater Than or Equal to 0.5 indicates --> Biodegrades Fast
A Probability Less Than 0.5 indicates --> Does NOT Biodegrade Fast
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |      Biowin3 FRAGMENT DESCRIPTION        | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------		Training set
Frag | 1 | Alkyl substituent on aromatic ring       | -0.0749 | -0.0749	ID	3
MolWt| * | Molecular Weight Parameter               |        | -0.2612
Const| * | Equation Constant                        |        | 3.1992
============+============================================+=========+=========
RESULT  | Biowin3 (Survey Model - Ultimate Biodeg) |        | 2.8632
============+============================================+=========+=========
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |      Biowin4 FRAGMENT DESCRIPTION        | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------		Training set
Frag | 1 | Alkyl substituent on aromatic ring       | -0.0685 | -0.0685	ID	3
MolWt| * | Molecular Weight Parameter               |        | -0.1705
Const| * | Equation Constant                        |        | 3.8477
============+============================================+=========+=========
RESULT  |  Biowin4 (Survey Model - Primary Biodeg) |        | 3.6087
============+============================================+=========+=========
Result Classification:  5.00 -> hours    4.00 -> days   3.00 -> weeks
(Primary & Ultimate)   2.00 -> months   1.00 -> longer
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |      Biowin5 FRAGMENT DESCRIPTION        | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------		Training set
Frag | 1 | Aromatic-CH3                             | 0.0415 | 0.0415	ID	3
Frag | 4 | Aromatic-H                               | 0.0082 | 0.0329	ID	15
Frag | 3 | -C=CH [alkenyl hydrogen]                | 0.0062 | 0.0186	ID	9
MolWt| * | Molecular Weight Parameter               |        | -0.3516
Const| * | Equation Constant                        |        | 0.7121
============+============================================+=========+=========
RESULT  | Biowin5 (MITI Linear Biodeg Probability) |        | 0.4535
============+============================================+=========+=========
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |      Biowin6 FRAGMENT DESCRIPTION        | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------		Training set
Frag | 1 | Aromatic-CH3                             | 0.3072 | 0.3072	ID	3
Frag | 4 | Aromatic-H                               | 0.1201 | 0.4806	ID	15
Frag | 3 | -C=CH [alkenyl hydrogen]                | 0.0285 | 0.0855	ID	9
MolWt| * | Molecular Weight Parameter               |        | -3.4117
============+============================================+=========+=========
RESULT  |Biowin6 (MITI Non-Linear Biodeg Probability)|        | 0.4968
============+============================================+=========+=========
A Probability Greater Than or Equal to 0.5 indicates --> Readily Degradable
A Probability Less Than 0.5 indicates --> NOT Readily Degradable
------+-----+--------------------------------------------+---------+---------
TYPE | NUM |      Biowin7 FRAGMENT DESCRIPTION        | COEFF | VALUE
------+-----+--------------------------------------------+---------+---------		Training set
Frag | 1 | Alkyl substituent on aromatic ring       | -0.1145 | -0.1145	ID	3
Frag | 1 | Aromatic-CH3                             | -0.2573 | -0.2573	ID	3
Frag | 4 | Aromatic-H                               | -0.0954 | -0.3817	ID	13
Frag | 3 | -C=CH [alkenyl hydrogen]                | -0.0735 | -0.2206	ID	11
Const| * | Equation Constant                        |        | 0.8361
============+============================================+=========+=========
RESULT  |  Biowin7 (Anaerobic Linear Biodeg Prob)  |        | -0.1380
============+============================================+=========+=========
A Probability Greater Than or Equal to 0.5 indicates --> Biodegrades Fast
A Probability Less Than 0.5 indicates --> Does NOT Biodegrade Fast
Ready Biodegradability Prediction: (YES or NO)
----------------------------------------------
Criteria for the YES or NO prediction: If the Biowin3 (ultimate survey
model) result is "weeks" or faster (i.e. "days", "days to weeks", or
"weeks" AND the Biowin5 (MITI linear model) probability is >= 0.5, then
the prediction is YES (readily biodegradable). If this condition is not
satisfied, the prediction is NO (not readily biodegradable). This method
is based on application of Bayesian analysis to ready biodegradation data
(see Help). Biowin5 and 6 also predict ready biodegradability, but for
degradation in the OECD301C test only; using data from the Chemicals
Evaluation and Research Institute Japan (CERIJ) database.

Validity criteria fulfilled:

not applicable

Interpretation of results:

not readily biodegradable

Conclusions:

Based on the BIOWIN v4.11 model estimation, the test substance is predicted to be not readily biodegradable. The individual submodules (i.e., BioWin 2 and 3 or BioWin 3 and 6) however suggest that the substance do not meet the persistence criteria as per Annex XIII criteria as set out in the REACH Technical Guidance Document Chapter R.11 (Table R.11—4).

Executive summary:

The biodegradation potential of the test substance, p-methylstyrene, was estimated using the BioWin v4.11 program of EPI Suite v4.11. BIOWIN contains seven separate models.This version (v4.11) designates the models as Biowin1 (linear probability model), Biowin2 (nonlinear probability model), Biowin3 (expert survey ultimate biodegradation model), Biowin4 (expert survey primary biodegradation model), Biowin5 (MITI linear model), Biowin6 (MITI nonlinear model), Biowin7 (anaerobic biodegradation model). Using SMILES or CAS number as the input parameter, the biodegradability estimates are based upon fragment constants that were developed using multiple linear or non-linear regression analyses, depending upon the model. For p-methylstyrene, the Biowin 1 and Biowin 2 models both predicted 'biodegrades fast, Biowin3 and Biowin 4 predicted 'weeks' and 'days-weeks' respectively, Biowin 5 and Biowin 6 both predicted 'not readily biodegradable' (with probability of biodegradation as 0.36 and 0.387 respectively) and Biowin 7 predicted 'does not biodegrade fast'. As the criteria for overall 'yes' ready biodegradability prediction is based on the 'weeks' or faster and biodegradation probability of >=0.5 predictions by Biowin 3 and Biowin 5 respectively, the overall ready biodegradability prediction for the substance was 'No' (US EPA, 2018). The individual submodules (i.e., BioWin 2 and 3 or BioWin 3 and 6), however suggest that the substance do not meet the persistence criteria as per Annex XIII criteria as set out in the REACH Technical Guidance Document Chapter R.11 (Table R.11—4). Overall, the prediction for the substance is considered to be reliable, as it falls within of the applicability domain.

Description of key information

In accordance with the restricted definition of readily biodegradable in screening tests used in the context of the REACH directive the substance must be considered to be not readily biodegradable as preconditioning of the microorganisms is required as per the experimental study with the test substance. However, the individual submodule prediction from BioWin as well as half-life prediction from BioHCwin model of EPI Suite QSAR program, indicate that the substance is biodegradable and will not be persistent in the environment.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Type of water:

freshwater

Additional information

Experimental study

A study was conducted to determine the Biological Oxygen Demand (BOD) and the Chemical Oxygen Demand (COD) of the test substance, p -methylstyrene according to US Standard Methods for the Examination of Water and Wastewater (1976). The method was modified to include analyses at additional time points after inoculation with domestic sewage sludge (10, 15 and 20 d) in order to determine whether the microorganisms required a time interval to adapt to the new food source (test substance). Exposure to the test substance occurred at a concentration of 2 to 4 µL/300 ml. The control groups confirmed that the inoculum was active and the results indicated that adaptation was necessary in order for domestic sludge microorganisms to utilise the test substance as a substrate. Under the study conditions, 13% of test substance was removed in the first 15 d of the sampling period and 32% was degraded from 15 to 20 d. Under the study conditions, unadapted microorganisms therefore could not degrade the test substance within a 28 d period to a level allowing identification as readily biodegradable. Nevertheless, adapted microorganisms present where continuous exposure to the test substance occurs will readily remove the substance from sewage systems (Gezo, 1980).

QSAR model predictions (BioWin and BioHCwin):

The biodegradation potential of the test substance, p-methylstyrene, was estimated using the BioWin v4.11 program of EPI Suite v4.11. BIOWIN contains seven separate models.This version (v4.11) designates the models as Biowin1 (linear probability model), Biowin2 (nonlinear probability model), Biowin3 (expert survey ultimate biodegradation model), Biowin4 (expert survey primary biodegradation model), Biowin5 (MITI linear model), Biowin6 (MITI nonlinear model), Biowin7 (anaerobic biodegradation model). Using SMILES or CAS number as the input parameter, the biodegradability estimates are based upon fragment constants that were developed using multiple linear or non-linear regression analyses, depending upon the model. For p -methylstyrene, the Biowin 1 and Biowin 2 models both predicted 'biodegrades fast, Biowin3 and Biowin 4 predicted 'weeks' and 'days-weeks' respectively, Biowin 5 and Biowin 6 both predicted 'not readily biodegradable' (with probability of biodegradation as 0.36 and 0.387 respectively) and Biowin 7 predicted 'does not biodegrade fast'. As the criteria for overall 'yes' ready biodegradability prediction is based on the 'weeks' or faster and biodegradation probability of >=0.5 predictions by Biowin 3 and Biowin 5 respectively, the overall ready biodegradability prediction for the substance was 'No' (US EPA, 2018). The individual submodules (i.e., BioWin 2 and 3 or BioWin 3 and 6) however suggest that the substance do not meet the persistence criteria as per Annex XIII criteria as set out in the REACH Technical Guidance Document Chapter R.11 (Table R.11—4). The prediction for the substance is considered to be reliable, as it falls within of the applicability domain.

The biodegradation potential of the test substance, p-methylstyrene, was also estimated using the BioHCwin v1.01 program of EPI Suite v4.11.BioHCwin is a new predictive model within EPI Suite, which determines quantitative primary biodegradation half-lives of individual petroleum hydrocarbons. This model uses a fragment-based approach similar to BioWin.Using SMILES code as the input parameter the primary biodegradation half-life for the test substance was determined to be 3.897 d; indicating that the substance can be classed as readily biodegradable (US EPA, 2018). The prediction for the substance is considered to be reliable, as it falls within of the applicability domain.

Overall,in accordance with the restricted definition of readily biodegradable in screening tests used in the context of the REACH directive, the substance must be considered to be not readily biodegradable as preconditioning of the microorganisms is required as per the experimental study with the test substance. However, the individual submodule prediction from BioWin as well as half-life prediction from BioHCwin model of EPI Suite QSAR program, indicate that the substance is biodegradable and will not be persistent in the environment.