Dimethyldioctylammonium chloride

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2001

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

Standard laboratory study to OECD Guideline 209 and GLP. An initial range-finding study was undertaken prior to the definitive test. The results are reported as nominal concentrations.

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Test material was supplied by Lonza as the commercial product Bardac 22, which contains 50% didecyldimethyl ammonium chloride active ingredient (a.i.). The results are reported as a.i.
Purity: 50.8%
Description: Extremely pale yellow, slightly viscous liquid
Batch: DEGE030167
Date received: 16 February 2001
Storage: Room temperature in darkness
Sponsor identification: LZ1043

Analytical monitoring:

no

Details on sampling:

No information reported

Vehicle:

no

Details on test solutions:

A stock solution was prepared using 500 mg of test material and dissolved in water to 1000 ml to give a concentration of 500 mg/L. Aliquots of the stock solution were each separately dispersed with synthetic sewage (16 ml), activated sludge (200 ml) and water to give a final volume of 500 ml. The reference material 3,5-dichlorophenol was used as a positive control. Two stock solutions were prepared of 50 and 160 mg/l. Aliquots of each were dispersed with activated sludge, synthetic sludge and water to give final concentrations of 10 mg/l (50 mg/l stock), and 3.2 and 32 mg/l (160 mg/l stock), respectively.

An initial range-finding study exposed sewage sludge microorganisms to 1, 10, 100 and 1000 mg/l nominal test material. Based on the result of the range-finding test, the following nominal test concentrations were assigned to the definitive study: 0, 10, 18, 32, 56 and 100 mg/l. The study included two control groups - a negative control (water) and a positive control 3,5-dichlorophenol.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

A mixed population of activated sewage sludge microorganisms was obtained on 22 February 2001 from the aeration stage of the Severn Trent Water Plc sewage treatment plant at Belper, Derbyshire, UK, which treats predominantly domestic sewage. The activated sludge was maintained on continuous aeration in the laboratory at a temperature of 21 degrees C and was used on the day of collection. The pH of the sample was 7.1 and the suspended solids equal to 3.9 mg/l prior to use.

Synthetic sewage was added to each vessel to act as a respiratory substrate with the following constituents dissolved in 1 litre of water:

16 g Peptone
11 g Meat extract
3 g Urea
0.7 g NaCl
0.4 g CaCl2.2H2O
0.2 g MgSO4.7H2O
2.8 g K2HPO4

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Remarks on exposure duration:

Respiration inhibition measurements were taken at 30 minutes and 3 hours contact time.

Post exposure observation period:

No information reported

Hardness:

Laboratory tap water dechlorinated was passed through an activated carbon filter and partly softened giving water a total hardness of 100 mg/l as CaCO3

Test temperature:

21 degrees C

pH:

pH in the two negative control replicates were 7.6 and 7.7

pH in the test concentration series ranged from 7.7 to 8.0

pH in the positive controls was 7.8, 7.9 and 8.0 (3.2, 10 and 32 mg/l, respectively)

Dissolved oxygen:

In some instances the initial and final dissolved oxygen concentrations were below those recommneded in the test guidelines (6.5 mg O2/l and 2.5 mg O2/l, respectively), but the laboratory considered this to have had no adverse effect on the results of the study given that in all cases the oxygen consumption rate was measured over the linear portion of the exygen consumption trace.

Salinity:

No information provided

Conductivity:

No information provided

Nominal and measured concentrations:

Nominal concentrations of the test material were prepared at 0, 18, 32, 56 and 100 mg/l.

Details on test conditions:

At time "0" 16 ml of synthetic sewage was diluted to 300 ml with water and 200 ml of inoculum added to a 500 ml conical flask. The mixture was aerated with compressed air via narrow bore glass tubes at a rate of approximately 0.5 to 1 litre per minute. Thereafter, at 15 minutes intervals the procedure was repeated with appropriate amounts of reference material being added. Flasks were inverted several times to ensure homogeneity of the solution.

As each vessel reached 30 minutes contact time an aliquot was removed from the conical flask and poured into the measuring vessel (250 ml darkened glass Biological Oxygen Demand bottle) and the rate of respiration measured using a Yellow Springs dissolved oxygen meter fitted with a BOD probe. The contents of the measuring vesle were stirred continuously by magnetic stirrer. The rate of respiration for each flask was measured over the linear portion of the oxygen consumption trace for an approximate 10 minute period (between approximately 8.9 mg O2/l and 2.4 mg O2/l). After the measure,emt, the contents of the BOD bottle were returned to the test vessel. This procedure was rpeated after 3 hours contact time.

The study was conducted under normal laboratory lighting and in a temperature controlled room at 21 degrees C.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

11 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

5 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Details on results:

The range-finding test indicated significant inhibition of respiration was observed at 100 and 1000 mg/l, however there were no inhibitory effects at 1.0 and 10 mg/l. The concentration gradient for the definitive study ranged from 0 to 100 mg/l. The validation criteria for the control respiration rates and reference material EC50 values were satisfied. No undissolved test material in the test vessel solution was observed during the test.

Oxygen consumption rates and percentage inhibition values for the control, test and reference concentrations are as follows:

Bardac 22 product EC50 (30 minutes) = 29 mg/l
Test substance EC50 (30 minutes) = 14.5 mg a.i./l
Variation in respiration rates of controls 1 and 2 after 30 minutes = +/- 5%

Bardac 22 product EC50 (3 hours) = 22 mg/l
Test substance EC50 (3 hours) = 11 mg a.i./l
Variation in respiration rates of controls 1 and 2 after 3 hours = +/- 7%

Bardac 22 product NOEC (3 hours) = 10 mg/l
Test substance NOEC (3 hours) = 5 mg a.i./l

Results with reference substance (positive control):

The results with the positive control are as follows:

3,5-dichlorophenol EC50 (30 minutes) = 11 mg/l
Variation in respiration rates of controls 1 and 2 after 30 minutes = +/- 5%

3,5-dichlorophenol EC50 (3 hours) = 62 mg/l
Variation in respiration rates of controls 1 and 2 after 3 hours = +/- 7%

Reported statistics and error estimates:

No reported statistical analysis.

Validity criteria fulfilled:

yes

Conclusions:

In a 3-hour study of the assessment of the inhibitory effect on the respiration of activated sewage sludge of the test substance (DDAC or didecyldimethyl ammonium chloride) resulted in an EC50 of 11 mg a.i./l and a NOEC of 5.0 mg a.i./l as a nominal concentration.

Executive summary:

An assessment of the inhibitory effect on the respiration of activated sewage sludge of the test substance DDAC resulted in a 3-hour EC50 of 11 mg a.i./l and a NOEC of 5.0 mg a.i./l. The study is in accordance with the OECD Test Guideline 209 and was undertaken by a commercial laboratory using sewage sludge from a treatment plant receiving primarily domestic sewage. A range-finding test preceded the definitive test. No adverse effects were observed in the control groups of microorganisms.

Description of key information

Two studies are available for toxicity to microorganisms, for the read-across substances didecyldimethyl ammonium chloride DDAC (Mead, 2001) and N,N-Didecyl-N,N-dimethylammonium carbonate (DDACarbonate) (Safepharm Laboratories, 2004).

An assessment of the inhibitory effect on the respiration of activated sewage sludge of the test substance DDAC resulted in a 3-hour EC50 of 11 mg a.i./L and a NOEC of 5 mg a.i./L. The study is in accordance with the OECD Test Guideline 209 and was undertaken by a commercial laboratory using sewage sludge from a treatment plant receiving primarily domestic sewage. A range-finding test preceded the definitive test. No adverse effects were observed in the control groups of microorganisms. 

An assessment was also made on the inhibitory effect of DDACarbonate to domestic sewage sludge, according to the OECD Guideline 209, to GLP. The effect of the test material on respiration of activated sludge micro-organisms gave a 3-hour EC50 of 64 mg/L (51 mg/L as 100 % active ingredient). The No Observed Effect Concentration (NOEC) after 3 hours exposure was 10 mg/L (8 mg/L as 100 % active ingredient).

It is recommended that the microorganism study for DDAC (Mead, 2001) is used for the PNEC development as this is the most sensitive study, with an EC50 of 11 mg a.i./L and a NOEC of 5 mg a.i./L.

Key value for chemical safety assessment

EC50 for microorganisms:

11 mg/L

EC10 or NOEC for microorganisms:

5 mg/L

Additional information

The EC50 and NOEC values are expressed as the active ingredient, which is 50% of the commercial product.