Sulfuric acid, C16-C18 (even numbered) alkyl esters, sodium salts and C16-18 (even numbered) alcohols

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:

December 2017 to January 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

the temperature remained in the required range of 22±2 °C only during the first 16 days of the test, after which it varied between 18.2 and 23.1 °C. This probably did not influence the biodegradation, since the plateau was attained at day 14 already.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Sulfuric acid, C16-C18 (even numbered) alkyl esters, sodium salts and C16-18 (even numbered) alcohols

EC Number:

944-399-8

Cas Number:

not yet assigned

Molecular formula:

see information in structural formula

IUPAC Name:

Sulfuric acid, C16-C18 (even numbered) alkyl esters, sodium salts and C16-18 (even numbered) alcohols

Test material form:

solid

Specific details on test material used for the study:

Identification: Radia 784x
ACH sample no. M1710-10516
Chemical name Reaction mass of Sulfuric acid, C16-18-alkyl esters, neutralized and Alcohols, C16-18

Since the test item consists of large particles/crystals, they were first ground (with a mortar and pestle) to a powder, so as to enhance the homogeneity of the test suspensions.
TOC 80,3%

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

Non-adapted activated sludge from the aeration tank of the ARA Werdhölzli (CH-8048 Zürich), a municipal biological waste water treatment plant. 30 mg/l dry matter in the final mixture. The activated sludge was used after sampling from the treatment plant without adaptation. However, the sludge was pre-conditioned (aerated but not fed) for 7 days, to reduce the amount of O2 consumed by the blank controls. Before the pre-conditioning phase, the sludge was washed twice with tap water, by 2 centrifugation steps. Then the sludge was suspended in test medium at about 2 g/l dry matter. Before the test, this suspension was diluted down to 300 mg/l dry matter, i.e. ten-fold the final concentration, and an appropriate amount of this suspension was added to the flasks

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

25.2 mg/L

Based on:

test mat.

Initial test substance concentration 2

Initial conc.:

20.2 mg/L

Based on:

TOC

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

the following test flasks were set up:
T: Test suspension: containing inoculum, test medium and test item (three replicates)
B: Blank control: containing inoculum and test medium (three replicates)
R: Procedure control: containing inoculum, test medium and sodium benzoate as ready biodegradable reference item (three replicates)
C: Abiotic sterile control: containing test item, ultra-pure water and 0.04 mM HgCl2 as sterilizing agent to prevent microbial decomposition (one replicate)
X: Toxicity control: containing inoculum, test medium, test item and sodium benzoate as ready biodegradable reference item (one replicate)

The test item (in flasks T, X and C) was applied by direct addition in the test flasks, at a concentration of about 20 mg/l with respect to its total organic carbon (TOC) content.
The reference item (in flasks R and X) was applied by direct addition as well, at a concentration of about 20 mg/l with respect to its total organic carbon (TOC) content.
In the toxicity control (flask X), the test and reference item were tested as a mixture, each at a concentration of about 20 mg TOC /l (application by direct addition).
Before the test start, the pH-value was checked and adjusted to 7.4±0.2 with HCl.
Before addition of the sludge, an aliquot of the test mineral medium was taken for measurement of the inorganic carbon (IC) at 0 h (see section 5.3).
The test vessels were stirred thoroughly and aerated with synthetic CO2-free air for a test period of 28 days. The air leaving the individual vessels was passed through gas-absorption bottles filled with KOH. The CO2 trapped in the KOH solutions was determined as inorganic carbon (IC) at frequent time intervals to allow for the assessment of the 10-day window for biodegradation in the vessels.

Biodegradation was determined on days 0, 1, 4, 7, 12, 14, 19, 21, 25 and 28. On day 29, one replicate was used to determine ultimate biodegradation by analysis of DOC.

Reference substance

Reference substance:

benzoic acid, sodium salt

Results and discussion

Test performance:

The results of a first test was repeated by mistake, as it was though - in a first judgement of the Study Director - that results are invalid according to the criteria of the OECD guideline (difference of extremes on day 28).
The results of the second test were valid according to the criteria of the OECD guideline.

% Degradation

% Degradation 1

Key result

Parameter:

% degradation (CO2 evolution)

Value:

61

Sampling time:

14 d

% Degradation 2

Parameter:

% degradation (CO2 evolution)

Value:

25

Sampling time:

28 d

Details on results:

Two tests were performed. The results of the first test were considered invalid in the first place (and as stated in the study report), since the results did not to meet the criteria of study plan and guideline OECD 301B on day 28 (difference of extremes 68.8%). Hence, it was decided to repeat the test. Validity of the first test was confirmed in retrospective, since variability of extreme at the end of the 10-day window was < 20%.Both results are hence valid and discussed and reported in the study report. Following OECD guideline 301 § 24, the repeat test is "considered that more work was necesswary to establish biodegradability". Hence the results of the repeat test were taken to establish the key result on ready biodegradation.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

The substance is readily biodegradable.

Executive summary:

In a valid GLP study according to OECD 301B the test item was aerobically exposed at a concentration of 25.2 mg/L (20.2 mg/L TOC) to standard OECD 301 mineral medium and 30 mg dry mass/L activated, non-adapted sludge. The test item was Oleon 784x, reaction mass of Sulfuric acid, C16-18-alkyl esters, neutralized and alcohols, C16-18.The inoculum was obtained from a domestic STP, i.e., the municipal sewage treatment plant „Weidhölzli“, Zürich, Switzerland. Exposure was for 28 days in the dark at pH 7.3 to 7.5, under stirring conditions and at 22 ± 2 °C. Eleven closed bottle glass vessels with a total volume of 2.3 L each were used and constantly aerated with CO2 free air. Each test vessel had received a total volume of 2 L inoculum and mineral medium. Five test groups were incubated: Test item (20.2 mg TOC/L, three replicates), blank control (three replicates), reference substance (20 mg TOC/L sodium benzoate), a toxicity control (20.2 mg/L TOC test substance plus 20 mg/L TOC of reference substance, one replicate) and a abiotic control, (20.2 mg TOC/L) sterilized with 0.04 mM HgCL2. Each test vessel had received inoculum and mineral medium. CO2 as produced by the inoculum was absorbed by 125 mL of 0.13 M KOH, which had been exposed in gas-washing bottles connected serially to the test vessels. Trapped carbon dioxide was determined as inorganic carbon (IC) by means of an appropriate IC analyser. For that purpose, duplicate samples were used. Biodegradation was determined on days 0, 1, 4, 7, 12, 14, 19, 21, 25 and 28. On day 29, one replicate was used to determine ultimate biodegradation by analysis of DOC. Two tests were performed. The results of the first test were considered invalid in the first place (as stated in the study report), since the results did not to meet the criteria of study plan and guideline OECD 301B on day 28 (difference of extremes 68.8%). Hence, it was decided to repeat the test. Validity of the first test was confirmed in retrospective, since variability of extremes at the end of the 10 -day window was 15.6%, i.e. < 20%. Both results are hence valid and discussed and reported as such in the study report. Following OECD guideline 301 § 24, the repeat test is "considered that more work was necessary to establish biodegradability". Hence the results of the repeat test were taken to establish the key result on ready biodegradation and reported as follows. The results of the reference substance show that 83% of the reference substance was degraded within 14 days. This result validates suitability of inoculum and testing procedures and conditions. The abiotic (sterile) control did not show more than 0.4% biodegradation. Hence the results reported here are due to microbial biodegradation. Test items results show that 61% of the test substance were degraded within 14 days (end of the 10 - day window). The 10 -day window was hence met and the substance is readily biodegradable. The substance is furthermore not toxic to STP microorganisms, since 56% of CO2were evolved by the toxicity control within 14 days of incubation.