3',5'-dichloro-4'-ethyl-2'-hydroxypalmitanilide

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:

from 1989, august 22nd to 1989, september 20th

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study performed according to the Test Guidelines described in the EEC Directive 84/449/EEC.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Report date:

1989

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.5 (Degradation: Biochemical Oxygen Demand)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

3',5'-dichloro-4'-ethyl-2'-hydroxypalmitanilide

EC Number:

406-200-8

EC Name:

3',5'-dichloro-4'-ethyl-2'-hydroxypalmitanilide

Cas Number:

117827-06-2

Molecular formula:

C24H39Cl2NO2

IUPAC Name:

N-(3,5-dichloro-4-ethyl-2-hydroxyphenyl)hexadecanamide

Details on test material:

Identification UC—136
RCC NOTOX Substance 1763
Description White powder
Purity 98.5%
Instructions for test article storage : At room temperature in the dark in a tightly sealed container
Stability of test article : Stable for one year maximal under storage conditions
Safety precautions : Gloves, goggles and face mask will be sufficient to ensure personnel health and safety

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, adapted

Details on inoculum:

Inoculum
The source of test organisms was activated
sludge freshly obtained from a municipal
sewage treatment plant, Waterschap de Aa,
Schijndel, The Netherlands.
Treatment
The activated sludge was aerated for 4 hours.
It was then left to settle for about 1 hour.
The supernatant was decanted to provide a
sufficient large volume for a 1% inoculum for
each CO2 test flask.
Temperature
20 ± 2°C
Reason for selection
This test has been found suitable by the OECD
Expert Group Degradation! Accumulation for
determining the ready biodegradability of
organic chemicals under aerobic conditions.
It has been tested in the OECD Laboratory
Intercomparison Test Programme (1978—1980).
Because of the stringency of this test method
a result of less than 60% yield of CO2
(within 28 days) does not necessary mean that
the test substance is not biodegradable under
environmental conditions, but indicates that
more work will be necessary to establish
biodegradability.

Duration of test (contact time):

28 d

Initial test substance concentration

Initial test substance concentration 1

Initial conc.:

10 mg/L

Based on:

test mat.

Initial test substance concentration 2

Initial conc.:

20 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

Total carbon content
The theoretical carbon content was calculated from the bruto formula.
Pre—incubation test medium
30 ml of the inoculum was added to the test medium present in each of the 3 litre test
bottles. This mixture was aerated with CO2 free air for 24 hours.
Preparations
After the aeration period, three CO2 absorber bottles were filled with 80 ml 0.025 N
Ba(OH)2 and connected in series to the exit air line of each test bottle.
Test material was added to two bottles, nominal concentrations 10 and 20 mg/l,
Q Trehespceocntitvroelly.substance was added to one bottle at a nominal concentration of 20 mg/l.
The fourth bottle contained only inoculum blank.
Subsequently, the four bottles were made up to 3 litre with Miili—Q water.
Principle
The CO2 produced in each test bottle reacted with the barium hydroxide in the gas
scrubbing bottle and was precipitated out as barium carbonate; the amount of CO2 produced
was determined by titrating the remaining Ba(OH)2 with 0.05 N standardized HCL.
Periodically the CO2 absorber nearest to the test bottle was removed for titration. The
remaining two absorbers were each moved one position in the direction of the test bottle,
and a new absorber filled with 80 ml of 0.025 N Ba(OH)2 (normalised the same day) was
placed at the far end of the series.
Measurements
Titrations were made when necessary (before significant BaCO3 precipitate was evident in
the second trap), approximately every other day for the first 10 days, and thereafter
every fifth day until the 28th day. Phenolphthalein was used as indicator.
On the 26th day, the pH of the four test bottles was measured, and then 1 ml of
concentrated HCl was added to drive off inorganic carbonate. The bottles were aerated
overnight, and samples were removed from each bottle to enable the possibility for
dissolved organic carbon (DOC) analysis. The final titration was made on day 28.

Reference substance

Reference substance:

acetic acid, sodium salt

Results and discussion

% Degradation

% Degradation 1

Parameter:

% degradation (CO2 evolution)

Value:

6.4

Sampling time:

9 d

Remarks on result:

other: 10 mg/l low

% Degradation 2

Parameter:

% degradation (CO2 evolution)

Value:

10.5

Sampling time:

28 d

Remarks on result:

other: 10 mg/l low

% Degradation 3

Parameter:

% degradation (CO2 evolution)

Value:

1.3

Sampling time:

9 d

Remarks on result:

other: 20 mg/l high

% Degradation 4

Parameter:

% degradation (CO2 evolution)

Value:

4.1

Sampling time:

28 d

Remarks on result:

other: 20 mg/l high

Details on results:

THEORETICAL CO2—PRODUCTION
The theoretical CO2—production of UC—136 (TCO2) (brutoformula:(C24H19C12NO2
MW=424) was calculated to be 2.491 mg C02/mg test substance.
The low concentration was 30.3 mg UC—136 in 3 litre test medium and the
high concentration was 60.5 mg UC—136 in 3 litre test medium. Hence, the
theoretical C02—production following complete degradation was 75.5 mg per
3 litre for the low and 150.7 mg per 3 litre for the high concentration.
The positive control contained 60.3 mg sodium acetate (TCO2= 1.073 mg
C02/mg) resulting in a theoretical C02—production of 64.7 mg per 3 litre.
BIODEGRADATION
The relative degradation values calculated from the measurements performed
during the test period revealed no significant degradation of UC—136 at
the high concentration (<10 %) and hardly at the low i.e. 11% .
The control substance degraded with more than 60% within 16 days and 80 %
biodegradation was reached at the end of the test period.
CO2 evolution in the blank reached a total value of 21.7 mg.
MONITORING OF TEMPERATURE AND pH
The temperature of the climate room varied between 18.5 °C and 21.5°C.
The pH values of the different test media were: a. blank: 5.9; b. positive
control: 5.7;c. UC—136 low: 5.9; d. UC—136 high: 4.2.

BOD5 / COD results

Results with reference substance:

%deg 40.9 CO2 evolution 9d
%deg 79.5 CO2 evolution 28d

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

valid study

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

Hence, UC—136 appears to be not readily biodegradable under the conditions
used in the present test.