Bromoacetic acid

Toxicity to microorganisms

Administrative data

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From August 2000 to January 2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study (OECD, EPA, etc)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Bromoacetic acid

EC Number:

201-175-8

EC Name:

Bromoacetic acid

Cas Number:

79-08-3

Molecular formula:

C2H3BrO2

IUPAC Name:

2-bromoacetic acid

Details on test material:

- Name of test material (as cited in study report): Monobromoacetic acid
- Substance type: Light brownish liquid
- Physical state: Liquid (80% solution)
- Analytical purity: 80%
- Purity test date: 80%
- Lot/batch No.: 00-2391
- Expiration date of the lot/batch: 11 August 2001
- Storage condition of test material: room temperature and no protection from the light

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
PHYSICO-CHEMICAL PROPERTIES are detailed in the section 4 of IUCLID.

Sampling and analysis

Analytical monitoring:

no

Details on sampling:

No analytical monitoring

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: A stock solution of the test substance was prepared by adding 1.0003 g of the test substance to 500 ml ultrapure water.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): No vehicle used.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): No

Test organisms

Test organisms (species):

other: Activated suldge taken from an oxidation ditch used to treat domestic sewage

Details on inoculum:

- Source: Approximately 20 litres of activated sludge was taken from an oxidation ditch situated at the district of Hazerswoude Dorp, the Netherlands on 25 September 2000.
- Preparation of inoculum for exposure: The sample of activated sludge was centrifuged and the supernatant discarded. The activated sludge was washed three times by centrifugation (10 min at 2000 rpm = max. 1218 g) and resuspension in tap water twice and twice in dilution water.
The dry weight of the sludge suspension was determined by drying on a Mettler AMl00 balance with a LPI6-M infrared drier. The mixed liquor suspended solids
(MLSS) content was found to be 5.3 g/l. The suspension was aerated vigorously, and kept at approximately 20°C in the dark.
50 ml of synthetic sewage feed was added daily per litre of sludge suspension. The dry weight was checked prior to use in the test, and a mixed liquor suspended solids level of 3.9 g/l was found.

Study design

Test type:

not specified

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Remarks on exposure duration:

20°C

Test conditions

Hardness:

Not reported

Test temperature:

20°C

pH:

Neutral: 7.0-7.6

Dissolved oxygen:

Not reported

Salinity:

Not applicable

Nominal and measured concentrations:

The final nominal concentrations of the substance tested were: 0, 1.0, 3.2, 10, 32, 100, 320, 1000 mg/l and and 320 and 1000 mg/l with pH
neutralisation.

Details on test conditions:

TEST SYSTEM
- Material, size, headspace, fill volume: one litre glass beakers :
* Control mixtures of 16 ml synthetic sewage feed and 284 mI dilution water + 200 ml of sludge suspension.
* Test mixtures were prepared by combining 16 ml synthetic sewage feed with the appropriate volumes of the test substance stock solutions and dilution water to a volume of 300 ml + 200 ml of sludge suspension.
- Aeration: Vigorously aerated
- No. of vessels per concentration (replicates): 1 replicate
- No. of vessels per control (replicates): 1 replicate
- Biomass loading rate: 3.9 g(d.w.)/l

TEST MEDIUM / WATER PARAMETERS
* Dilution water. (Prescription according to draft NEN No 6634:
- StockA2
KH2P04 2.125 g
K2HP04 5.44 g
Na2HP04·7H20 8.35 g
NH4CI 0.42 g
ultrapure water 250 ml
(pH 7.2)
- Stock B
MgS04·7H20 2.25 g
ultrapure water 100 ml
- Stock C
CaCl2·2H20 3.63 g
ultrapure water 100 ml
- Stock D
FeCl3·6H20 0.025 g
ultrapure water 100 ml

Dilution water preparation by adding 1 ml of stocks 1 to 4 to each litre of ultrapure water and store at 20°C in the dark with aeration.

* Synthetic sewage feed (The following amounts of substances to be dissolved in two litres of ultrapure water, according to DECO Guideline No. 209)
peptone 32 g
meat extract 22 g
urea 6 g
NaCI 1.4 g
CaCl2·2H20 0.8 g
MgS04·7H20 0.4 g
K2HP04 5.6 g

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
After three hours of incubation, a sample of each mixture was poured into a BOD bottle (volume about 295 mI) and stirred vigorously on a magnetic stirrer. An
oxygen electrode (connected to a WTW OXI 2000 meter) was inserted into the sample and the oxygen concentration in the sample was continuously measured.
The oxygen concentration was recorded every minute during a period of about 10 minutes.

TEST CONCENTRATIONS
- Test concentrations: 0, 1.0, 3.2, 10, 32, 100, 320, 1000 mg/l

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol - 3 concentrations: 5, 12, 30 mg/l

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Effect concentrations 2

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

320 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

The results were obtained with the test substance: "80 % solution of monobromoacetic acid".
The results are generally based on nominal concentrations of the test substance, and numbers in parenthesis refer to nominal concentrations of the active ingredient.
Respiration inhibition was found at a test substance concentrations of 32 and 1000 mg/l (25.6 and 800 mg.a.i/l) because no inhibition was found at the test substance concentrations 100 and 320 mg/l (80 and 256 mg.a.i./l) the NOEC was assessed to be 320 mg/l (256 mg.a.i./l). At the highest test substance concentration tested (800 mg.a.i./1) less than 50% inhibition was observed (36.1 %).
Therefore the EC50-value was assessed to be >1000 mg/l (>800 mg.a.i./l).
In the additional test with pH neutralisation, no effect was found at 1000 mg/l (800 mg.a.i./l). The observed effect can thus be attributed to the acidity rather than the toxicity of the test substance.

- VALIDITY CRITERIA:
The respiration rates of the two controls during the test with monobromoacetic acid were found to be 24.91 and 29.89 mg O2/g/h, respectively. The difference between the two values amounted to 16.0 % being not within the validity criterion (15%) of the OECO Guideline. This is a small deviation from the criteria of 15% and did not influence the outcome of this test.
The EC50 of 3,5 -dichlorophenol after a contact time of 3 hours, 15.1 mg/l was also in the accepted range of the Guideline (range 5 to 30 mg/l).

Results with reference substance (positive control):

The EC50 of 3,5-dichlorophenol, was found to be 15.1 mg/l, which is within the range prescribed by DEeD Guideline No. 209

Reported statistics and error estimates:

the respiration rate (in mg o2 per g dry weight active sludge per hour) was calculated by linear regressionn of the oxygen concentration meter readings in the linear part of the oxygen depletion curve in accordance with the OECD Guideline No 209.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

According to the validity criteria was met, the NOEC of monobromoacetic acid was assessed to be 320 mg/l.
The EC50-value was assessed to be >1000 mg/l, because the highest concentration of 1000 mg/l resulted in less than 50 % inhibtion.

Executive summary:

The effect of monobromoacetic acid on activated sludge from an oxidation ditch, treating domestic sewage only, was detennined in a respiration inhibition test according to the OECD Guideline No. 209: 'Activated Sludge. Respiration Inhibition Test' and the OECD principles of Good Laboratory Practice.

Monobromoacetic acid is a light brownish liquid that is soluble in water. The nominal concentrations of 0, 1.0,3.2,10,32, 100.320 and 1000 mg/l ,were tested without pH adjustment and 320 and 1000 mg/l were tested with pH neutralisation.

Mixtures of activated sludge (3.9 g(d.w.)/l, synthetic sewage feed and test substance were aerated for 3h at 20 ± 2°C. The decrease in the oxygen concentration in the mixtures was measured during a 10 minute period, and the respiration rate for each test substance concentration was calculated.

The concentrations given in this report are nominal concentrations of the test substance monobromoacetic acid.

A control test with the reference substance 3,5-dichlorophenol yielded an EC50 value of 15.1 mg/l, which is in the range prescribed by the Guideline. The validity criteria of the Guideline were fulfilled.

A respiration inhibition of 36.1 % was found at a test substance concentration of 1000 mg/l. A slightly reduced respiration was observed at 32 mg/l and no inhibtion was observed at the test substance concentrations 100 and 320 mg/l.

The NOEC of monobromoacetic acid was thus assessed to be 320 mg/l. The EC50-value was assessed to be >1000 mg/l, because the highest concentration of 1000 mg/l resulted in less than 50 % inhibtion. In the additional test with pH neutralisation, no effect was found at 1000 mg/l. The observed effect can thus be attributed to the acidity rather than the toxicity of the test substance.