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Biodegradation in water: screening tests

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Endpoint:
biodegradation in water: ready biodegradability
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Results derived form a valid read across, with adequate and reliable documentation/justification.
Justification for type of information:
The read across justification is presented in the Biodegradation in water: screening tests Endpoint Summary.
Reason / purpose:
read-across source
Key result
Parameter:
% degradation (O2 consumption)
Value:
80
Sampling time:
28 d
Remarks on result:
other: read-across from Veilex #1
Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The source substance showed 80% biodegradation in an OECD TG 301D test. The 10-day window criterion was met. The target substance is readily biodegradable based on the results of the source substance.
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
16-09-2015 to 15-10-2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
Information used for read across to Veilex#3.
Qualifier:
according to
Guideline:
OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Oxygen conditions:
aerobic
Inoculum or test system:
natural water
Details on inoculum:
River water was sampled from the Rhine near Heveadorp, The Netherlands (10-09-2015). The nearest plant (Arnhem-Zuid) treating domestic wastewater biologically was 3 km upstream. The river water was aerated for 7 days before use to reduce the endogenous respiration (van Ginkel and Stroo, 1992). River water without particles was used as inoculum. The particles were removed by sedimentation after 1 day while moderately aerating.
Duration of test (contact time):
28 d
Initial conc.:
2 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
DEIONISED WATER
Deionized water containing no more than 0.01 mg/L Cu (ISO 17025 certified; non-GLP analysis) was prepared in a water purification system.

TEST BOTTLES
The test was performed in 0.30 L BOD (biological oxygen demand) bottles with glass stoppers.

NUTRIENTS, STOCKS AND ADMINISTRATION
The river water used in the Closed Bottle test was spiked per liter of water with 8.5 mg KH2PO4, 21.75 mg K2HPO4, 33.4 mg Na2HPO4·2H2O, 22.5 mg MgSO4·7H2O, 27.5 mg CaCl2, 0.25 mg FeCl3·6H2O. Ammonium chloride was not added to the river water to prevent nitrification. Accurate administering of the test substance was accomplished by preparing a solid stock of 3.0 mg of the test substance per g of silica gel in a 50-mL serum flask. Only part of the top layer of the silica gel was brought into contact with the test substance. The serum flask was closed with a screw top with aluminum foil and the content was mixed vigorously. Subsequently 0.2 g of silica gel with the test substance was added to the test bottles. The resulting concentration of test substance in the bottles was 2.0 mg/L. Next the bottles were filled with nutrient medium with inoculum and closed. Sodium acetate was added to the bottles using a stock solution of 1.0 g/L.

TEST PROCEDURES
The Closed Bottle test was performed according to the study plan. The study plan was developed from ISO Test Guidelines (1994). Use was made of 10 bottles containing only river water, 10 bottles containing river water and silica gel, 10 bottles containing river water and silica gel with test substance, 6 bottles with river water and sodium acetate. The concentrations of the test substance, and sodium acetate in the bottles were 2.0 and 6.7 mg/L, respectively. Each of the prepared solutions was dispensed into the respective group of BOD bottles so that all bottles were completely filled without air bubbles. The zero time bottles were immediately analyzed for dissolved oxygen using an oxygen electrode. The remaining bottles were closed and incubated in the dark. Two duplicate bottles of all series were withdrawn for analyses of the dissolved oxygen concentration at day 7, 14, 21, and 28.

TEST CONDITIONS
The pH of the media was 8.0 at the start of the test. The pH of the medium at day 28 was 7.8 (test) and 8.0 (controls). Temperatures were within the prescribed temperature range of 22 to 24°C.

CALCULATION OF BIODEGRADATION PERCENTAGES
The biodegradation was calculated as the ratio of the BOD to the theoretical oxygen demand (ThOD). The calculated theoretical oxygen demand (ThOD) of the test substance (mono-constituent) is 2.7 mg/mg. The ThOD of the reference substance sodium acetate is 0.8 mg/mg.
Reference substance:
acetic acid, sodium salt
Test performance:
VALIDITY OF THE TEST
The validity of the test is demonstrated by an endogenous respiration of 1.4 mg/L at day 28 (Table I). Furthermore, the differences of the replicate values at day 28 were less than 20%. The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 89. Finally, the validity of the test is shown with oxygen concentrations remaining at >0.5 mg/L in all bottles over the test period.

TOXICITY
Inhibition of the endogenous respiration of the inoculum by the test substance tested was not detected. Therefore, no inhibition of the biodegradation due to the initial concentration of the test compound is expected.
Key result
Parameter:
% degradation (O2 consumption)
Value:
80
Sampling time:
28 d
Details on results:
Over 60% biodegradation was achieved in a period of 10 days immediately following the attainment of 10% biodegradation. The test item therefore fulfilled the time window criterion for ready biodegradable compounds. Hence the test substance should be classified as readily biodegradable.
Results with reference substance:
The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 89.

Tabel 1: Dissolved oxygen concentrations (mg/L) in the closed bottles.

Time (days)

Oxygen concentration (mg/L)

Ocs

Ot

Oc

Oa

0

8.7

8.7

8.7

8.7

 

8.7

8.7

8.7

8.7

Mean

8.7

8.7

8.7

8.7

7

8.0

6.7

7.9

3.5

 

7.9

6.8

8.0

3.9

Mean

8.0

6.8

8.0

3.7

14

7.7

4.3

7.6

2.7

 

7.7

3.8

7.6

2.9

Mean

7.7

4.1

7.6

2.8

21

7.4

3.4

7.4

-

 

7.4

3.3

7.6

-

Mean

7.4

3.4

7.5

-

28

7.3

2.8

7.3

-

 

7.2

3.2

7.3

-

Mean

7.3

3.0

7.3

-

Ocs: River water with nutrients and silica gel.

Ot: River water with nutrients, test material (2.0 mg/L) and silica gel.

Oc: River water with nutrients.

Oa: River water with nutrients and sodium acetate (6.7 mg/L).

Table 2: Oxygen consumption (mg/L) and the percentages biodegradation of the test substance and the reference substance

Time (days)

Oxygen consumption (mg/L)

Biodegradation (%)

Test substance

Acetate

Test substance

Acetate

0

0.0

0.0

0

0

7

1.2

4.3

22

80

14

3.6

4.8

67

89

21

4.0

-

74

-

28

4.3

-

80

-

Validity criteria fulfilled:
yes
Remarks:
see 'Test performance'.
Interpretation of results:
readily biodegradable
Conclusions:
The substance showed 80% biodegradation in an OECD TG 301D test. The 10-day window criterion was met.
Executive summary:

In order to assess the biotic degradation, a ready biodegradability test was performed which allows the biodegradability to be measured in an aerobic aqueous medium. The ready biodegradability was determined in the Closed Bottle test (OECD TG 301D) under GLP. The test substance is non-inhibitory to the inoculum based on absence of inhibition of the inoculum. Veilex 1 is biodegraded by 80% at day 28 in the Closed Bottle test. Over 60% biodegradation was achieved in a period of 10 days immediately following the attainment of 10% biodegradation. Hence, this substance should be classified as readily biodegradable. The test is valid as shown by an endogenous respiration of 1.4 mg/L, by the total mineralization of the reference compound, sodium acetate (89% within 14 days) and the oxygen concentrations remained at >0.5 mg/L in all bottles over the test period.

Description of key information

The biodegradability of Veilex #3 is assessed based on read-across from close structural analogue Veilex #1 (CAS# 63449-88-7) and is considered readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable
Type of water:
freshwater

Additional information

In this Endpoint summary the experimental information of Veilex #1 is presented. The read across justification is presented thereafter.

In order to assess the biotic degradation, a ready biodegradability test was performed which allows the biodegradability to be measured in an aerobic aqueous medium. The ready biodegradability was determined in the Closed Bottle test (OECD TG 301D) under GLP. The test substance is non-inhibitory to the inoculum based on absence of inhibition of the inoculum. Veilex 1 is biodegraded by 80% at day 28 in the Closed Bottle test. Over 60% biodegradation was achieved in a period of 10 days immediately following the attainment of 10% biodegradation. Hence, this substance should be classified as readily biodegradable. The test is valid as shown by an endogenous respiration of 1.4 mg/L, by the total mineralization of the reference compound, sodium acetate (89% within 14 days) and the oxygen concentrations remained at >0.5 mg/L in all bottles over the test period.

Biodegradation of Veilex #3 (CAS #1193-81-3) based on read across from Veilex #1 (CAS #63449-88-7).

 

Introduction and hypothesis for the analogue approach

Veilex #3 consist of a cyclohexylethyl backbone with an alcohol as functional group. For this substance there are no experimental biodegradation data available. Biodegradation information may be generated by other means, i.e. applying alternative methods such as QSARs, grouping and read-across. For assessing the ready biodegradability of Veilex #3, the analogue approach is selected because for a closely related analogue, Veilex #1, ready biodegradation information is available which can be used for read across.

Hypothesis: Veilex #3 has the same biodegradation potential as Veilex #1. 

Available information: For Veilex #1 a ready biodegradation study was performed according to OECD TG 301D (Rel. 1). In this study Veilex #1 was biodegraded by 80% at day 28 in the Closed Bottle test. Over 60% biodegradation was achieved in a period of 10 days immediately following the attainment of 10% biodegradation. Hence, this substance is considered readily biodegradable. The data is reliable without restrictions (Klimisch 1).

Target chemical and source chemical(s)

Chemical structures of Veilex #3 and Veilex #1 are shown in the data matrix, including their physico-chemical properties.

Purity / Impurities

Veilex #3 is a mono-constituent substance with purities close to 100%, which does not contain any impurities that will affect the read across for biodegradation.

Analogue approach justification

According to Annex XI 1.5 read across can be used to replace testing when the similarity can be based on a common backbone and a common functional group. When using read across the result derived should be applicable for C&L and/or risk assessment and it should be presented with adequate and reliable documentation. The analogue Veilex #1 is selected from IFFs portfolio.

Structural similarities and differences: Veilex #3 and Veilex #1 share the same cyclohexylethyl backbone. The difference is that in Veilex #3 the functional group consists of an alcohol while in Veilex #1 it is a butyl ester.

Bioavailability: Veilex#3 has a considerably lower log Kow and higher water solubility compared Veilex#4, this means that Veilex #3 will be much more bioavailable and mor susceptible to biodegradation.

Biodegradable fragments:Veilex #3 as a whole is an integral part of Veilex #1 and has therefore the same fragments. The ester of Veilex #1 will be rapidly degraded by carboxylesterases and Veilex #3 will be formed as well as butyric acid. The ready biodegradability is further supported by BIOWIN.

Only BIOWIN modules 5 (‘MITI-linear’) and 6 (‘MITI-non-linear’) are used because these two models are based on ready biodegradability tests, which is the endpoint that needs to be predicted. From these BIOWIN predictions it can be seen that both Veilex #3 and Veilex #1 are readily biodegradable with values of respectively 0.5527 and 0.6260, respectively for BIOWIN 5. The values 0.6902 and 0.7499, respectively are found for BIOWIN 6. All these are above the cut-off value of 0.5. The straight alkyl butyric acid is the reason for a slightly higher biodegradation potential of Veilex#1.

Uncertainty of the prediction:There are no remaining uncertainties other than those already addressed in the previous sections.

Conclusions for environmental fate

For Veilex#3 no biodegradation information is available. The analogue Veilex#1 was found for read across. When using read across the result derived should be applicable for C&L and/or risk assessment and be presented with adequate and reliable documentation. This documentation is presented in the current document. For Veilex #1 a well conducted ready biodegradability test is available which can be used for read across to Veilex #3. Veilex #1 is readily biodegradable and therefore Veilex #3 will be assessed as readily biodegradable as well.

Final conclusion:Veilex #3 is considered to be readily biodegradable.

Data matrix for read across to Veilex #3 from Veilex #1

Common name

Veilex #3

Veilex #1

 

Target

Source

Chemical name

1-cyclohexylethanol

1-cyclohexylethyl butyrate

Chemical structures

CAS no

1193-81-3

63449-88-7

Einecs

214-780-7

264-158-4

REACH registration

For Intermediated use and for 2018

Registered

Empirical formula

C8H16O

C12H22O2

Physico-chemical data

 

 

Molecular weight

128.21

198.30

Physical state

liquid

liquid

Melting point,oC

< -20

< -20

Boiling point,oC

195.8

246.5

Vapour pressure, Pa

36.8 (at 24°C)

6.5 (at 24 °C)

Water solubility, mg/L

5163.2 (at 24°C)

23.8 (at 24 °C)

Log Kow

1.6

5.2

Fate and behaviour

 

 

Biodegradation

Derived from read across:

Readily biodegradable

 

Readily biodegradable (OECD TG 301D)