Ammonium 2-mercaptopropionate

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

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:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OPPTS 835.3110

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

Source: A mixed population of activated sewage sludge microorganisms was obtained on 7 December 1999 from the aeration stage of the Severn Trent Water Plc sewage treatment plant at Belper Derbyshire, UK, which treats predominantly domestic sewage.
Preparation of inoculum: The sample of activated sewage sludge was maintained on continuous aeration upon receipt. A sample of the activated sewage sludge was washed three times by settlement and resuspension in culture medium to remove any excessive amounts of dissolved organic carbon (DOC) that may have been present. A sub-sample of the washed sewage sludge was then removed and the suspended solids concentration determined.
Culture medium: The culture medium used in this study was that recommended in the OECD Guidelines.

Duration of test (contact time):

28 d

Initial conc.:

10 mg/L

Based on:

TOC

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

The test substance was exposed to activated sewage sludge microorganisms at a concentration of 10 mg C/L with culture medium in sealed culture vessels in the dark at 21 °C for 28 days. The degradation of the test substance was assessed by the determination of carbon dioxide produced. Control solutions with inoculum and the standard material, sodium benzoate, together with a toxicity control were used for validation purposes.

Test substance preparation:
The test substance was prepared by a direct solution in culture medium. 1000 mg of the test substance was dissolved in culture medium and the volume was adjusted to 1 L to give a 1000 mg/L stock solution. Subsequently, an aliquot (147 mL) of this stock solution was dispersed in inoculated culture medium and the volume was adjusted to 3 L to give a final concentration of 49.0 mg/L, equivalent to 10 mg carbon/L.

Standard material control:
For the purposes of the study a standard material, sodium benzoate was used. An initial stock solution of 1000 mg/L was prepared by direct solution in culture medium and a 51.4 mL aliquot added to the test vessel to give a final test concentration of 17 .1 mg/L, equivalent to 10 mg carbon/L.

Preparation of test system: The following test solutions were prepared and inoculated in 5 L glass culture vessels each containing 3 L of solution:
a) A control, in duplicate, consisting of inoculated culture medium.
b) The standard material (sodium benzoate), in duplicate, in inoculated culture medium to give a final concentration of 10 mg carbon/L.
c) The test substance, in duplicate, in inoculated culture medium to give a final concentration of 1 0 mg carbon/L.
d) The test substance plus the standard material in inoculated culture 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 study was carried out in a temperature controlled room at 21 °C, in darkness.
Approximately 24 hours prior to addition of the test and standard materials the vessels were filled with 2400 mL of culture medium and 33.3 mL of inoculum and aerated overnight. On day 0 the test and standard materials were added and the volume in all the vessels adjusted to 3 L by the addition of culture medium.
The culture vessels were sealed and CO2 free air bubbled through the solution at a rate of approximately 40 mL/min and stirred continuously by magnetic stirrer.
The CO2 free air was produced by passing compressed air through a glass column containing self-indicating soda lime (Carbosorb) 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 purified degassed water.

Reference substance:

benzoic acid, sodium salt

Key result

Parameter:

% degradation (CO2 evolution)

Value:

14

Sampling time:

28 d

Details on results:

- The test substance attained 14 % degradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No 301 B.
- The total CO2 evolution in the control vessels on day 28 was 32.51 mg/L (= 97.53 mg/3 L).
- 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 control Replicate 1 and test substance Replicate 2. These increases are considered to be due to CO2 present in solution being driven off by the addition of hydrochloric acid on day 28 and resulted in an increase in the percentage degradation value for the test substance from 14 % on day 28 to 15 % on day 29.
- The toxicity control attained 59 % degradation after 28 days thereby confirming that the test substance was not toxic to the sewage treatment micro-organisms used in the study. The increase in inorganic carbon in the first absorber vessels on day 29 resulted in an increase in the percentage degradation value for the toxicity control from 59 % on day 28 to 62 % on day 29.
- Sodium benzoate attained 74 % degradation after 28 days thereby confirming the suitability of the inoculum and test conditions. The increase in inorganic carbon in the first absorber vessels on day 29 resulted in an increase in the percentage degradation value for the standard substance from 74 % on day 28 to 77 % on day 29.
- Inorganic carbon analysis of the samples from the second absorber vessels on day 29 confirmed that no significant carry-over of CO2 into second absorber vessels occurred.
- Analysis of the test media from the test substance culture vessels on days 0 and 28 for DOC gave percentage degradation values of 24 % and 34 % respectively for replicates R1 and R2 and 64 % for the toxicity control. Sodium benzoate attained 95 % and 100 % degradation respectively for replicates R1 and R2 calculated from the results of the DOC analyses.

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

Under the study conditions, the test substance attained 14% degradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict conditions of OECD Guideline 301B.

Executive summary:

A study was conducted to determine the ready biodegradability of the test substance according to OECD Guideline 301B (CO2 evolution test), EU Method C.4 - C, in compliance with GLP. This study was performed in order to evaluate aerobic elimination and degradation potential of the test substance throughout a 28 day period in a test for ready biodegradability, using a test substance concentration of nominally 10 mg C/L with culture medium in sealed culture vessels in the dark at 21 °C. The degradation of the test substance was assessed by the determination of carbon dioxide produced. Control solutions with inoculum and the standard material, sodium benzoate, together with a toxicity control were used for validation purposes. Under the study conditions, the test substance attained 14% degradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict conditions of OECD Guideline 301B (Mead, 2000).

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Remarks:

publication

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Deviations:

yes

Remarks:

see 'Principles of method if other than guideline'

Principles of method if other than guideline:

- Test substance concentration used: approx. 30 mg ThOD/L (50-100 mg ThOD/L according to guideline)
- Bacterial density of: 80 mL STP effluent/L final solution (10+E8 CFU/mL according to guideline)

GLP compliance:

yes

Remarks:

(not applicable)

Inoculum or test system:

activated sludge (adaptation not specified)

Details on inoculum:

The inoculum for each experiment was freshly taken from the Lüneburg municipal STP.

Duration of test (contact time):

28 d

Initial conc.:

30 mg/L

Based on:

ThOD

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

The test was run for 28 d at 20±1 °C in the dark with gentle stirring. Oxygen consumption was recorded daily using an OxiTop control OC110 system (WTW, Weilheim, Germany), measuring the pressure decrease in the headspace (about 1/3 of bottle volume), while CO2 produced was removed by adsorption/absorption to NaOH pellets/conc. NaOH solution. The temperature was monitored daily; the pH was measured at days 0 and 28, was adjusted to 6.5-8 if necessary at day 0, and was in this range at day 28.

Key result

Parameter:

% degradation (O2 consumption)

Value:

71.2

St. dev.:

6.6

Sampling time:

28 d

Details on results:

Results from two replicates:
66.6 % and 75.9 % biodegradation

The theoretical oxygen demand ThOD was calculated under the assumption that sulfur is oxidized to SO42-.

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

Under the study conditions, the degree of biodegradation reached 71.2% after 28 days, therefore the test substance was considered to be readily biodegradable.

Executive summary:

A study was conducted to evaluate the ready biodegradability of the tests substance according to a modified OECD Guideline 301F (Manometric Respirometry). The test was run for 28 d at 20±1 °C in the dark with gentle stirring. Oxygen consumption was recorded daily using an OxiTop control OC110 system, measuring the pressure decrease in the headspace (about 1/3 of bottle volume), while CO2 produced was removed by adsorption/absorption to NaOH pellets/conc. NaOH solution. The theoretical oxygen demand ThOD was calculated under the assumption that sulfur is oxidized to SO42-. Under the study conditions, the degree of biodegradation reached 71.2% after 28 days, therefore the test substance was considered to be readily biodegradable (Rücker, 2017).

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Type of water:

freshwater

Additional information

Study 1:

A study was conducted to determine the ready biodegradability of the test substance according to OECD Guideline 301B (CO2 evolution test), EU Method C.4 - C, in compliance with GLP. This study was performed in order to evaluate aerobic elimination and degradation potential of the test substance throughout a 28 day period in a test for ready biodegradability, using a test substance concentration of nominally 10 mg C/L with culture medium in sealed culture vessels in the dark at 21°C. The degradation of the test substance was assessed by the determination of carbon dioxide produced. Control solutions with inoculum and the standard material, sodium benzoate, together with a toxicity control were used for validation purposes. Under the study conditions, the test substance attained 14% degradation after 28 days and therefore could not be considered to be readily biodegradable under the strict conditions of OECD Guideline 301B (Mead, 2000).

 

Study 2:

A study was conducted to evaluate the ready biodegradability of the tests substance according to a modified OECD Guideline 301F (Manometric Respirometry). The test was run for 28 d at 20±1 °C in the dark with gentle stirring. Oxygen consumption was recorded daily using an OxiTop control OC110 system, measuring the pressure decrease in the headspace (about 1/3 of bottle volume), while CO2 produced was removed by adsorption/absorption to NaOH pellets/conc. NaOH solution. The theoretical oxygen demand ThOD was calculated under the assumption that sulfur is oxidized to SO42-. Under the study conditions, the degree of biodegradation reached 71.2% after 28 days, therefore the test substance was considered to be readily biodegradable (Rücker, 2017).

​

Overall assessment:

It is assumed that structurally similar compounds should have comparable behaviors when tested according to the same OECD 301 protocol. However, published biodegradation data for substances containing both an acid or ester and a mercaptan functional group are in many cases contradictive (Rücker et al., 2017).

Results of repeated biodegradation experiments for the same substance in the same test using inoculum from the same source may differ considerably. These findings are also supported by the relevant guidelines: “Realising that ready biodegradability tests may sometime fail because of the stringent test conditions, positive test results should generally supersede negative test results” (ECHA 2017, page 208; OECD 2006, page 3), and “When contradictory results in ready biodegradability tests are obtained the positive results could be considered valid irrespective of negative results, when the scientific quality of the former is good and the positive test results are well documented, …” (ECHA 2017, page 230).

Based on the above mentioned information, wa eight of evidence approach was taken to assess the biodegradability of the test substance. The following studies were taken into consideration (I) a study conducted according to OECD Guideline 301 B (Mead 2000) and (2) published data by Rücker et al., 2017 which provides results from an OECD study 301 F.

In the study by Rücker et al., 24 substances containing divalent sulfur (i. e., mercaptocarboxylic acids, their esters, disulfides, sulfides and mercaptans) were tested in two standardised biodegradation tests, OECD 301 D (Closed Bottle Test, CBT) and OECD 301 F (Manometric Respirometry Test, MRT) in the same laboratory.

For most of the test substances strong differences between CBT (OECD 301 D) and MRT (OECD 301 F) results were observed with MRT test being more effective. For our substance of interest, MRT test biodegradation of 71.2 % on Day 28 and a biodegradation of 12.4 % with CBT test were achieved. Based on the literature and experimental biodegradation information presented in the article, mercaptocarboxylic acids and their esters as a class are found to be either readily biodegradable or at least biodegradable to a significant extent.

Based on the weight of evidence approach, the test substance is concluded to be readily biodegradable.

 

References:

ECHA, 2017: Guidance on information requirements and chemical safety assessments, Chapter R.7b: Endpoint specific guidance, June 2017,

OECD, 2006: Organisation for Economic Co-operation and Development, OECD Guidelines for the testing of chemicals, Revised introduction to the OECD Guidelines for testing of chemicals, Section 3, adopted 23 March 2006,

Rücker Ch., Mahmoud W. M. M., Schwartz D., Kümmerer K. (2017). Biodegradation tests of mercaptocarboxylic acids, their esters, related divalent sulfur compounds and mercaptans. Manuscript (24.11.2017).