Tetramethylcyclotetrasiloxane substituted dipropylether bisphenol A

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13-July-2020 to 08-February-2021

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2021

Report date:

2021

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1992

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

Version / remarks:

Paragraph (m)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

No. 440/2008

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Reaction mass of (Tetramethylcyclotetrasiloxane propylether) prop-1-enylether bisphenol A and Di(tetramethylcyclotetrasiloxane propylether) bisphenol A

Cas Number:

203874-34-4

Molecular formula:

(OSiHxCH3)4-CH2-CH2 -CH2- O-C6H4-C(CH3)2-C6H4-0-CH2-CH2-CH2-(SiHxCH3O)4

IUPAC Name:

Reaction mass of (Tetramethylcyclotetrasiloxane propylether) prop-1-enylether bisphenol A and Di(tetramethylcyclotetrasiloxane propylether) bisphenol A

Test material form:

liquid

Details on test material:

Appearance: Pale yellow liquid

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

- Source of inoculum: A mixed population of activated sewage sludge micro-organisms was obtained from the aeration stage of a sewage treatment plant at UK, which treats predominantly domestic sewage.
- Preparation of inoculum for exposure: The activated sewage sludge sample was washed twice by settlement and re-suspension in mineral medium to remove any excessive amounts of Dissolved Organic Carbon (DOC) that may have been present.
- Storage conditions: The washed sample was maintained on continuous aeration in the laboratory at a temperature of approximately 21ºC.
- Storage length: The washed sample used on the day of collection.
- Filtration of the sample: Determination of the suspended solids level of the activated sewage sludge was carried out by filtering a sample (100 mL) of the washed activated sewage sludge by suction through a pre-weighed Whatman GF/A filter paper using a Buchner funnel.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial conc.:

10 mg/L

Based on:

DOC

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

Preliminary Solubility Work
No information was available on the water solubility of the test item. Preliminary solubility/dispersibility work was performed in order to determine the most suitable method of preparation.

Test Item Preparation
From the preliminary solubility work and following the recommendations of the International Standards Organisation (ISO 10634, 1995) and Handley et al (2002) it was concluded that the best testable dispersions were found to be obtained when using the high shear mixing method of preparation and the silica gel with high shear mixing of preparation. As there was no significant difference between the appearance of these dispersions it was considered appropriate to use high shear mixing without the use of silica gel for the test.
The test item was dispersed directly in mineral medium.
An amount of test item (66.6 mg) was dispersed directly in approximately 400 mL of mineral medium, in duplicate, with the aid of high shear mixing (approximately 7500 rpm, 15 minutes). They were then allowed to cool to a temperature of approximately 22°C prior to dispersal in inoculated mineral medium. The volume was adjusted to 3 L to give a final concentration of 22.2 mg/L, equivalent to 10 mg carbon/L.

Reference Item Preparation
A reference item, sodium benzoate (C6H5COONa), was used to prepare the procedure control vessels. An initial stock solution was prepared by dissolving a nominal amount of the reference item (1000 mg) directly in mineral medium and the volume adjusted to 1 L to give a 1000 mg/L stock solution. An aliquot (51.4 mL) of this stock solution was added to the test vessel containing inoculated mineral medium prior to the volume being adjusted to 3 L to give a final test concentration of 17.1 mg/L, equivalent to 10 mg carbon/L. The volumetric flask containing the reference item was inverted several times to ensure homogeneity of the solution.

Toxicity Control
A toxicity control, containing the test item and sodium benzoate, was prepared in order to assess any toxic effect of the test item on the sewage sludge micro-organisms used in the test.
An amount of test item (66.6 mg) was dispersed in approximately 400 mL of mineral medium with the aid of high shear mixing (approximately 7500 rpm, 15 minutes) prior to allowing to cool to a temperature of approximately 23°C prior to dispersal in inoculated mineral medium. An aliquot (51.4 mL) of the sodium benzoate stock solution was also added to the test vessel and the volume adjusted to 3 L to give a final concentration of 22.2 mg test item/L plus 17.1 mg sodium benzoate/L, equivalent to a total of 20 mg carbon/L.

Preparation of Test System
The following were prepared and inoculated in 5 L test culture vessels each containing 3 L of preparation:
a) An inoculated control, in duplicate, consisting of inoculated mineral medium.
b) The procedure control containing the reference item (sodium benzoate), in duplicate, in inoculated mineral medium to give a final concentration of 10 mg carbon/L.
c) The test item, in duplicate, in inoculated mineral medium to give a final concentration of 10 mg carbon/L.
d) The test item plus the reference item in inoculated mineral medium to give a final concentration of 20 mg carbon/L to act as a toxicity control (one vessel only).
Each test vessel was inoculated with the prepared inoculum at a final concentration of 30 mg suspended solids (ss)/L. The test was carried out in a temperature controlled room at temperatures of between approximately 21 and 24 °C for 28 days, in darkness. The temperature was recorded daily from a vessel incubated alongside the test containing 3 L of reverse osmosis water.
Approximately 24 hours prior to addition of the test and reference items the vessels were filled with 2400 mL of mineral medium and 39.1 mL of inoculum and aerated overnight. On Day 0 the test and reference items were added and the pH of all vessels measured using a
Hach HQ30d Flexi handheld meter prior to the volume in all of the vessels being adjusted to 3 L by the addition of mineral medium, which had been purged overnight with CO2-free air. The inoculum control vessels were prepared in a similar manner without the addition of test item or reference item.
In order to confirm that the sodium benzoate solution was prepared correctly, a diluted 100 mg/L stock solution (in reverse osmosis water) was sampled for TOC analysis.
The test vessels were sealed and CO2-free air bubbled through the solution at a rate of 30 to 100 mL/minute per vessel and stirred continuously by magnetic stirrer.
The CO2-free air was produced by passing compressed air through a moisture trap prior to being passed through a glass column containing self-indicating soda lime granules.
The CO2 produced by degradation was collected in two 500 mL Dreschel bottles containing 350 mL of 0.05 M NaOH. The CO2 absorbing solutions were prepared using deionized reverse osmosis water.

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

% Degradation

Key result

Parameter:

% degradation (CO2 evolution)

Value:

5

Sampling time:

28 d

Details on results:

The IC values for the test item, procedure control, toxicity control and inoculum control vessels at each analysis occasion are given in Table 1 below under "Any other information...". Percentage biodegradation values of the test and reference items and the toxicity control are given in Table 2 and the biodegradation curves are presented in Figure 1, Figure 2 and Figure 3. TC and IC values in the culture vessels on Day 0 are given in Table 3. The pH values of the test preparations on Days 0 and 28 are given in Table 4. Observations made on the contents of the test vessels are given in Table 5.

The total CO2 evolution in the inoculum control vessels on Day 28 was 35.72 mg/L and therefore satisfied the validation criterion given in the OECD Test Guidelines.
The IC content of the test item suspension in the mineral medium at the start of the test, before dosing (see Table 3 below), was below 5% of the TC content and hence satisfied the validation criterion given in the OECD Test Guidelines.
The difference between the values for CO2 production at the end of the test for the replicate vessels was <20% and hence satisfied the validation criterion given in the OECD Test Guidelines.
Acidification of the test vessels on Day 28 followed by the final analyses on Day 29 was conducted according to the methods specified in the Test Guidelines. This acidification effectively kills the micro-organisms present and drives off any dissolved CO2 present in the test vessels. Therefore any additional CO2 detected in the Day 29 samples originated from dissolved CO2 that was present in the test vessels on Day 28 and hence the biodegradation value calculated from the Day 29 analyses is taken as being the final biodegradation value for the test item.
The results of the inorganic carbon analysis of samples from the first absorber vessels on Day 29 showed an increase in all replicate vessels with the exception of inoculum control Replicate 1 and test item Replicate 2.
The IC analysis of the samples from the second absorber vessels on Day 29 confirmed that no significant carry-over of CO2 into the second absorber vessels occurred.
The test item attained 5% biodegradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No. 301B.
The toxicity control attained 34% biodegradation after 14 days and 45% biodegradation after 28 days thereby confirming that the test item did not exhibit an inhibitory effect on the sewage treatment micro-organisms used in the test.

BOD5 / COD results

Results with reference substance:

Sodium benzoate attained 76% biodegradation after 14 days with greater than 60% degradation being attained in a 10-day window. These results confirmed the suitability of the inoculum and test conditions and satisfied the validation criterion given in the OECD Test Guidelines. After 28 days 82% biodegradation was attained.
Total Organic Carbon Confirmation
Total organic carbon of the diluted sodium benzoate stock solution confirmed that it had been prepared correctly.

Any other information on results incl. tables

Table 1 Inorganic Carbon Values on Each Analysis Occasion

Day	Inorganic Carbon (mg IC)
	Inoculum Control				Procedure Control				Test Item				Toxicity Control
	R1		R2		R1		R2		R1		R2		R1
	Abs 1	Abs 2	Abs 1	Abs 2	Abs 1	Abs 2	Abs 1	Abs 2	Abs 1	Abs 2	Abs 1	Abs 2	Abs 1	Abs 2
0	1.40	1.75	1.75	1.75	1.75	1.75	1.75	1.75	1.75	1.75	1.75	1.75	1.40	1.75
6	15.34	-	14.19	-	30.68	-	34.02	-	13.84	-	15.22	-	34.37	-
8	17.43	-	16.17	-	33.71	-	36.92	-	15.14	-	16.63	-	36.58	-
10	19.26	-	18.70	-	35.11	-	41.38	-	17.44	-	19.84	-	41.72	-
14	22.55	-	23.23	-	43.18	-	48.17	-	20.40	-	23.35	-	43.29	-
21	26.03	-	29.18	-	46.64	-	48.22	-	23.89	-	26.03	-	51.26	-
28	29.68	-	28.79	-	45.47	-	49.84	-	29.23	-	29.68	-	52.31	-
29	27.95	2.44	29.39	2.44	46.65	8.47/3.48*	57.34	4.18	31.06	2.44	29.39	2.44	55.55	2.78

R = Replicate
Abs = CO2 absorber vessel
-= No value determined
*Results from analysis of frozen sample due to original result deemed to be erroneous

Table 2             Percentage Biodegradation Values

 

Day	Biodegradation (%)
	Procedure Control	Test Item	Toxicity Control
0	0	0	0
6	59	0	33
8	62	0	33
10	64	0	38
14	76	0	34
21	66	0	39
28	61	1	38
29*	82	5	45

* Day 29 values corrected to include any carry-over of CO2 detected in Absorber 2

 

Table 3  Total and Inorganic Carbon Values in the Culture Vessels on Day 0

 

Test vessel	Total Carbon* (mg/L)	Inorganic Carbon* (mg/L)	IC Content (% of TC)
Test Item 10 mg C/L R1	10.27**	0.02	0
Test Item 10 mg C/L R2	10.19**	0.16	2

R = Replicate

* Corrected for control values

** TC value given is the sum of the TC value obtained from analysis and the nominal TC contribution of the test item

 

Table 4  pH Values of the Test Preparations on Days 0 and 28

 

Test Vessel	pH
	Day 0 Pre-Adjustment	Day 28
Inoculum Control R1	7.6	7.5
Inoculum Control R2	7.6	7.5
Procedure Control R1	7.6	7.6
Procedure Control R2	7.6	7.6
Test Item R1	7.6	7.5
Test Item R2	7.6	7.5
Toxicity Control	7.6	7.6

 

 

Table 5  Observations on the Test Preparations Throughout the Test Period

 

Test Vessel		Observations on Test Preparations
		Day 0	Day 7	Day 14	Day 21	Day 28
Inoculum Control	R1	Light brown dispersion	Light brown dispersion	Light brown dispersion	Light brown dispersion	Light brown dispersion
	R2	Light brown dispersion	Light brown dispersion	Light brown dispersion	Light brown dispersion	Light brown dispersion
Procedure Control	R1	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible
	R2	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible	Light brown dispersion, no undissolved reference item visible
Test Item	R1	Light brown cloudy dispersion with oily specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface
	R2	Light brown cloudy dispersion with oily specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface	Light brown cloudy dispersion with specks of test item visible floating at the surface
Toxicity Control		Light brown cloudy dispersion with oily specks of test item visible floating at the surface, no undissolved reference item visible	Light brown cloudy dispersion with specks of test item visible floating at the surface, no undissolved reference item visible	Light brown cloudy dispersion with specks of test item visible floating at the surface, no undissolved reference item visible	Light brown cloudy dispersion with specks of test item visible floating at the surface, no undissolved reference item visible	Light brown cloudy dispersion with specks of test item visible floating at the surface, no undissolved reference item visible

R: Replicate

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

According to the results of a CO2 Evolution Test, performed according to OECD Guideline No. 301B and GLP principles, the test item attained 5% biodegradation after 28 days and is therefore considered to be not readily biodegradable.

Executive summary:

A CO2 Evolution Test was performed according to A OECD Guideline No. 301B and GLP principles. Carbon dioxide (CO2) evolution from the test item was followed by means of Inorganic Carbon (IC) analysis over a period of 28 days when exposed to sewage sludge micro-organisms under defined conditions. Control solutions with inoculum and the reference item, sodium benzoate, together with a toxicity control were used for validation purposes. Sodium benzoate attained 76% biodegradation after 14 days with greater than 60% degradation being attained in a 10-day window. These results confirmed the suitability of the inoculum and test conditions.

The test item attained 5% biodegradation after 28 days and is therefore considered to be not readily biodegradable.