Cyclohexanol, 4-[[(2-amino-3,5-dibromophenyl)methyl]amino]-, hydrobromide (1:1), trans-

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:

1999-01-27 to 1999-02-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted under GLP conditions in accordance with the official Guideline (OECD 301 B, 1992).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1999

Report date:

1999

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

(July, 1992)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

(December, 1992)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Mucosolvan HBr

IUPAC Name:

Mucosolvan HBr

Test material form:

other: solid

Details on test material:

- Name of test material (as cited in study report): Mucosolvan HBr
- Physical state: Beige solid
- Storage condition of test material: at room temperature in the dark
- Stability under storage conditions: stable
- Other: Stability in water: not indicated

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

sewage, predominantly domestic (adaptation not specified)

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): activated sludge freshly obtained from a municipal sewage treatment plant 'Waterschap de Maaskant' in 's-Hertogenbosch, the Netherlands
- Treatment: The sludge was kept under continuous aeration until further treatment. The concentration of suspended solids was 3.7 g/L in the concentrated sludge (information obtained from the municipal sewage treatment plant). Before use, the sludge was allowed to settle for at least 30 minutes and the liquid decanted for use as inoculum at the amount of 10 mL/L of mineral medium.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

71 other: mg/ 2 litres

Based on:

test mat.

Initial test substance concentration 2

Initial conc.:

12 mg/L

Based on:

other: TOC

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: Milli-Q water (Tap-water purified by reverse osmosis and subsequently passed over activated carbon and ion-exchange cartridges (Millipore Corp., Bedford, USA)) was used to prepare the media. Salts of analytical grade were added to Milli-Q water to give stock solutions. Stock solutions were diluted in Milli-Q water.
- Test temperature: 20.5 to 22°C
- pH: 7.4 to 7.8
- pH adjusted: no
- Aeration of dilution water: Mineral components, Milli-Q water and inoculum were added to each bottle. This mixture was aerated with CO2-free air overnight to purge the system of CO2.

TEST SYSTEM
- Culturing apparatus: 2 litre all-glass brown coloured bottles
- Number of culture flasks/concentration: 2
- Method used to create aerobic conditions: A mixture of oxygen (21%) and nitrogen (79%) was led through a bottle, containing 0.5-1 litre 0.0125 M Ba(OH)2 solution to trap CO2 which might be present in small amounts. The CO2-free air was sparged through the scrubbing solutions at a constant rate.
- Measuring equipment: Three CO2-absorbers (bottles filled with 100 ml 0.0125 Ba(OH)2) were connected in series to the exit air line of each test bottle.

SAMPLING
- Sampling frequency: Titrations were made every second or third day during the first 10 days, and thereafter at least every fifth day until the 28th day.
- Sampling method: Each time the CO2-absorber nearest to the test bottle was removed for titration; each of the remaining two absorbers was moved one position in the direction of the test bottle. A new CO2-absorber was placed at the far end of the series. Phenolphthalein was used as pH-indicator. On the 28th day, the pH of the test suspensions was measured and 1 mL of concentrated HCl was added to each bottle. The bottles were aerated overnight to drive off CO2 present in the test suspension. The final titration was made on day 29.

CONTROL AND BLANK SYSTEM
- Inoculum blank: containing only inoculum (2 bottles)
- Positive control: containing reference substance (ca. 40 ml/L sodium acetate; TOC = 12 mg/L) and inoculum (1 bottle)
- Toxicity control: containing test substance, reference substance and inoculum (1 bottle)

Reference substance

Reference substance:

acetic acid, sodium salt

Results and discussion

% Degradation

Parameter:

% degradation (CO2 evolution)

Value:

>= 0.8 - <= 2.9

Sampling time:

29 d

Details on results:

The relative degradation values calculated from the measurements performed during the test period revealed no significant degradation of Mocosolvan HBr.
The difference of duplicate values for %-degradation of Mucosolvan HBr was always less than 20.
In the toxicity control more than 25% degradation occurred within 14 days (based on ThCO2). Therefore, the test substance was assumed to be not inhibitory.

BOD5 / COD results

Results with reference substance:

The positive control substance was degraded at least 60% within 7 days.

Any other information on results incl. tables

Theoretical CO₂production

The Theoretical CO₂production (ThCO₂) of Mucosolvan HBr (MW=459.0) was calculated to be 1.25 mg CO₂/mg.

The concentration was 70.9 mg Mucosolvan HBr in 2 litres test medium. Hence, the theoretical CO₂production following complete degradation was 88.6 mg per 2 litres.

The positive control contained 80.2 mg sodium acetate (ThCO₂= 1.07 mg CO₂/mg) resulting in a theoretical CO₂ production following complete degradation of 85.8 mg 2 litres.

The toxicity control contained 80.2 mg sodium acetate and 70.9 mg Mucosolvan HBr in 2 litres of test medium. Hence, the theoretical CO2 production following complete degradation of Mucosolvan HBr plus sodium acetate was 174.4 mg per 2 litres.

Biodegradation

The relative degradation values calculated from the measurements performed during the test period revealed no significant degradation of Mucosolvan HBr.

In the toxicity control more than 25% degradation occurred within 14 days (based on ThCO₂). Therefore, the test substance was assumed to be not inhibitory.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

other: not readily biodegradable

Conclusions:

Mucosolvan HBr was not readily biodegradable under the conditions of the modified Sturm test presently performed.