1,2-Benzisothiazole, 3-methyl-, 1,1-dioxide

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016-11-16 to 2016-12-14

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)

Version / remarks:

May 30, 2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)

Version / remarks:

July 17, 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

Version / remarks:

January 1998

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

- Lot: 216-106
- Expiry date: 2018-06-16
- Storage conditions: Keep at room temperature

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: The (controlled) activated sludge was supplied by the sewage plant for domestic sewage in Balatonfüred, Hungary, 5 days before the test.
- Preparation of inoculum for exposure: The activated sludge was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in isotonic saline solution and again centrifuged. This procedure was repeated twice. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to its dry weight was determined. Based on this ratio, calculated aliquots of washed sludge suspension, corresponding to 5 g dry material per litre was mixed with test water (mineral medium) and then aerated under test conditions (for 5 days) until use. The pH of the activated sludge inoculum after preparation was 7.29, just before use: 7.00. A pH adjustment of activated sludge inoculum was not performed.
- Pretreatment: Pre-conditioning consisted of aerating (2 L/minute) activated sludge (in mineral medium) for 5 days at the test temperature. The cell count was determined and inoculum was diluted to the necessary cell concentration. After preparation the sludge was filtered through cotton wool.
- Concentration of sludge: The approximate cell count of aerated inoculum fell in the range of 10E8-10E9/L.
- Initial cell/biomass concentration: 10E6 cells/L
- Water filtered: no

Duration of test (contact time):

28 d

Initial conc.:

3 mg/L

Based on:

test mat.

Initial conc.:

5.04 mg/L

Based on:

ThOD

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: mineral medium accroding to OECD guideline
- Additional substrate: no
- Test temperature: 20.0 - 20.2 °C.
- pH: prior study 7.24
- pH adjusted: no
- Aeration of dilution water: The test medium was aerated for 20 minutes and allowed to stand for about 20 hours at the test temperature. The dissolved oxygen concentration was checked and found 9.26 mg/L.
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: Winkler bottles (300 mL, coded) with special neck and glass stoppers.
- Number of culture flasks/concentration: 10 (+ 2 reserve)
- Method used to create aerobic conditions: aerating (2 L/minute)
- Test performed in closed vessels: yes

SAMPLING
- Sampling frequency: days 0, 7, 14, 21 and 28
- Sampling method: oxygen was measured directly, samples for nitrate and nitrite analysis were taken from all vessels
- Sample storage before analysis: For technical reason the nitrate and nitrite analysis of the start (day 0) samples was performed together with the 7-day samples on the 7th day of the test, the analysis of 14-day samples was performed one week later, together with the 21-day samples. The samples were stored in freezer until determination of nitrate and nitrite. The 7-day, 21-day and 28-day samples were analysed directly after oxygen measurements.

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes, only inoculum
- Procedure control: yes, reference item and inoculum
- Toxicity control: yes, test item, reference item and inoculum

STATISTICAL METHODS:
The BOD (mg O2 per mg test item) expected after each period was calculated. The percentage biodegradation of the test item and of the reference item were calculated.

Reference substance:

benzoic acid, sodium salt

Remarks:

Sodium benzoate

Preliminary study:

The test item solubility, behavior, and toxicity were tested in a 14-day preliminary experiment. The test design was the same as described at the main experiment. In the preliminary experiment the test item was investigated at the concentration of 3 mg/L. No toxic effect of the test item was found at this investigated concentration.

Test performance:

Inoculum control:
The oxygen depletion in the inoculum control did not exceed 1.5 mg O2/L after 28 days. It was 1.42 mg O2/L in average.

Oxygen concentration:
The residual oxygen concentration in the test flasks did not drop below 0.5 mg O2/L at any time. (The lowest value was 3.45 mg O2/L, it was measured on the 28th day in the procedure control.)

Parallels:
The difference of duplicate values for the degradation at the plateau, at the end of the test or at the end of the 10-d window was not greater than 20 %.
The highest difference (10.6 %) between the duplicate values from the 14th to the 28th day of the test (this period was taken into consideration as biodegradation plateau) or at the end of the test (test item, procedure control, and toxicity control groups) for degradation was calculated in the test item group, it was observed on the 21st day.

Reference item:
The percentage degradation of the reference item reached the level for ready biodegradability (> 60 %) by exposure day 14. (The percentage degradation of the reference item was 85.5 % on the 14th day.)

Key result

Parameter:

% degradation (O2 consumption)

Value:

11.1

Sampling time:

28 d

Details on results:

Because of the N-containing test item, the oxidised nitrogen (nitrate and nitrite) concentrations were determined following each oxygen measurement with photometric method using nitrite and nitrate cell tests. The LOQ (Limit Of Quantification) of the measurements was 0.03 mg NO2/L and 0.4 mg NO3/L, respectively. The measured quantities of nitrite and nitrate in the inoculum control, test item and toxicity control samples were below the LOQ throughout the study and any correction of the oxygen consumption values was considered as not necessary.
Under the test conditions the percentage biodegradation of the test item reached a mean of 11.1 % after 28 days based on its theoretical oxygen demand (ThODNH4). In the toxicity control containing both, the test item and the reference item, a mean of 37.7 % biodegradation was noted within 14 days and a mean of 40.8 % biodegradation was determined after 28 days of incubation.

Results with reference substance:

The reference item sodium benzoate was sufficiently degraded to a mean of 85.5 % after 14 days, and to a mean of 86.1% after 28 days of incubation, based on its ThODNH4.

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

At the end of the 28-day exposure period, the mean biodegradation of the test item amounted to 11.1 %.

Executive summary:

The test item was investigated for its ready biodegradability in a closed bottle test over 28 days according to OECD 301 D. The concentration of 3.0 mg/L was chosen based on the preliminary test results and based on the theoretical oxygen demand of 1.68 mg O2 per mg test item. Because of the N-containing test item, the oxidised nitrogen (nitrate and nitrite) concentrations were determined. The measured quantities of nitrite and nitrate in the inoculum control, test item and toxicity control samples were below the LOQ throughout the study and any correction of the oxygen consumption values was considered as not necessary. All validity criteria of the study were met. Under the test conditions the percentage biodegradation of the test item reached a mean of 11.1 % after 28 days based on its theoretical oxygen demand (ThODNH4). The pass level for ready biodegradability is the removal of 60 % ThODNH4 in a 10-day window. Minimal biodegradation of the test item occurred in this study; however the value remained far below the pass level; therefore the test item was considered to be not ready biodegradable. According to the test guidelines the test item can be assumed as not inhibitory at the applied concentration level on the activated sludge microorganisms because the degradation in the toxicity control group was higher than 25 % within 14 days.

Description of key information

At the end of the 28-day exposure period, the mean biodegradation of the test item amounted to 11.1 % indicating that the test substance is not readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Additional information

The test item was investigated for its ready biodegradability in a closed bottle test over 28 days according to OECD 301 D. The concentration of 3.0 mg/L was chosen based on the preliminary test results and based on the theoretical oxygen demand of 1.68 mg O2 per mg test item. Because of the N-containing test item, the oxidised nitrogen (nitrate and nitrite) concentrations were determined. The measured quantities of nitrite and nitrate in the inoculum control, test item and toxicity control samples were below the LOQ throughout the study and any correction of the oxygen consumption values was considered as not necessary. All validity criteria of the study were met. Under the test conditions the percentage biodegradation of the test item reached a mean of 11.1 % after 28 days based on its theoretical oxygen demand (ThODNH4). The pass level for ready biodegradability is the removal of 60 % ThODNH4 in a 10-day window. Minimal biodegradation of the test item occurred in this study; however the value remained far below the pass level; therefore the test item was considered to be not ready biodegradable. According to the test guidelines the test item can be assumed as not inhibitory at the applied concentration level on the activated sludge microorganisms because the degradation in the toxicity control group was higher than 25 % within 14 days.