Hexadecyltrimethylammonium methyl sulphate

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:

From 04 April, 2012 to 26 April, 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Test substance was added to the incubation vessels using a stock solution of approximately 1.0 g/L.The stock solution of quaternary ammonium compounds, C12-14 (evennumbered)-alkylethyldimethyl, ethyl sulphates was prepared by dissolving 0.78 g C12-14 alkyl ethyl dimethyl ethyl sulphates QAS in 780 mL deionized water.

pH stock solution C12-14 alkyl ethyl dimethyl ethyl sulphates QAS =6.6

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

Unadapted secondary activated sludge was obtained (24-04-2012) from the WWTP Nieuwgraaf in Duiven. The WWTP Duiven is an activated sludge plant treating predominantly domestic wastewater. Prior to use the activated sludge was homogenized with a syringe. The dry weight of the homogenized activated sludge was determined and subsequently concentrated by settlement to the required dry weight concentration. The dry weight of the activated sludge in the incubation vessels was 1.5 g/L.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Test temperature:

Temperature varied from 19.8 to 20.6°C.

pH:

The pH of the reaction mixtures after the incubation period ranged from 7.2 to 7.4.

Dissolved oxygen:

Aerobic during test

Nominal and measured concentrations:

Test substance:  10 mg/L, 20 mg/L, 40 mg/L, 80 mg/L and 160 mg/L (nominal)
Reference compound 3,5-dichlorophenol: 2.5 mg/L, 5.0 mg/L,10.0 mg/L, 20.0 mg/L and 40.0 mg/L

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Key result

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

ca. 9 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

ca. 30 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: with 95% confidence limits of 13 and 72 mg/L

Details on results:

Test conditions
The pH of the reaction mixtures after the incubation period ranged from 7.2 to 7.4. Temperature varied from 19.8 to 20.6°C. These conditions allow
respiration of the activated sludge used.

Validity of the test
The validity of the test is demonstrated by three criteria. First, the coefficient of variation of the replicates of the control oxygen uptake rates was 10%, which fulfils the maximum prescribed variation of <30%. Secondly, the prescribed average oxygen uptake rate of > 20 mg O2/g dry weight/hour for the control measurements was achieved (30 mg O2/g dry weight/hour). The third criterion was met as shown by the EC50 of the reference compound of 15 mg/L, which is within the prescribed range of 2 to 25 mg/L.

Activated sludge respiration inhibition test
The inhibitory effect of test substance at a particular concentration is expressed as a percentage of the two controls. From the results EC values were calculated. The EC50 of test substance for activated sludge after 3 hours contact time is 30 mg/L with 95% confidence limits of 13 and 72 mg/L. The EC10, EC20 and EC80 are 9, 14 and 66 mg/L, respectively.

Results with reference substance (positive control):

The EC50 of the reference compound was 15 mg/L at a contact time of 3 h, respectively. These EC50 values is within the prescribed range of 2 to 25 mg/L.

Results

Table I Respiration rates of the activated sludge, inhibition percentages, pH and temperature at various concentrations of 3,5-dichlorophenol after 3 hours contact time

Concentration (mg/L)	Activity (mgO2/g/min)	Inhibition (%)	pH at start test	pHatend test	Temperature at end test (°C)
Control	0.5222	-	7.0	7.2	19.8
Control	0.5222	-	7.1	7.2	20.0
Control*	0.4556	-	7.1	7.2	20.2
Control*	0.4556	-	7.1	7.3	20.4
2.5	0.5000	4	7.1	7.3	20.1
5.0*	0.3778	17	7.1	7.2	20.0
10.0	0.3444	34	7.1	7.3	20.0
20.0	0.1500	71	7.0	7.3	20.2
40.0	0.1143	78	7.0	7.3	20.4

Concentration stock solution 3,5-dichlorophenol = 999 mg/L pH stock solution 3,5-dichlorophenol = 7.1

* = corresponding control and reference concentration

Table II Respiration rates of the activated sludge, inhibition percentages, pH and temperature at various concentrations of test substance after 3 hours contact time.

Concentration (mg/L)	Activity (mgO2/g/min)	Inhibition (%)	pH at start test	pHatend test	Temperature end test (°C)
Control	0.6133	-	7.1	7.3	20.2
Control	0.5333	-	7.1	7.3	20.6
Control*	0.5278	-	7.1	7.4	20.5
Control*	0.4889	-	7.2	7.3	20.3
5*	0.5048	1	7.1	7.3	20.4
10	0.4714	18	7.1	7.3	20.2
20	0.3444	40	7.2	7.3	20.0
40	0.2519	56	7.2	7.4	20.3
80	0.0778	86	7.2	7.4	20.1
160	0.0238	96	7.2	7.4	20.3

Concentration stock solution test substance = 1 g/L

pH stock solution of test substance = 6.6

* = corresponding control and reference concentration

Validity criteria fulfilled:

yes

Conclusions:

Under the study conditions, the EC50 of test substance in sludge after 3 h of contact time was determined to be 30 mg a.i./L (nominal).

Executive summary:

A study was conducted to determine the toxicity to microorganisms of the test substance, C12-14 ADMAES (96% active), in an activated sludge respiration inhibition testaccording to OECD 209 Guideline, in compliance with GLP. The toxicity of the test substance to activated sludge was determined at a contact time of 3 h, using various concentrations of the test substance. The inhibitory effect of the substance at a particular concentration is expressed as a percentage of the two controls. No analytical determination was performed for the test substance. The EC50 of test substance in sludge after 3 h of contact time was determined to be 30 mg/L (nominal) (equiavalent to 28.8 mg a.i./L) (95% confidence limits: 13 and 72 mg/L). The 3 h EC10, EC20, EC80 were determined to be 9, 14 and 66 mg/L (nominal) (equiavalent to 8.64, 13.44 and 63.36 mg a.i./L), respectively. The test substance was therefore considered harmful to activated sludge. The test was valid as shown by the EC50 of the reference compound 3,5 -dichlorophenol (15 mg/L), the coefficient of variation of the replicates of the control oxygen uptake rates (10%) and the average oxygen uptake rate for the control replicates (30 mg O2/g dry weight/hour). Under the study conditions, the EC50 of test substance in sludge after 3 h of contact time was determined to be 30 mg a.i./L (nominal) (Geerts, 2012).

Description of key information

Based on the results of the read across study, the 3 h EC50 and EC10 values for toxicity in micro-organisms is considered to be 30 and 9 mg a.i./kg; these values have been considered further for hazard/risk assessment.

Key value for chemical safety assessment

EC50 for microorganisms:

30 mg/L

EC10 or NOEC for microorganisms:

9 mg/L

Additional information

A study was conducted to determine the toxicity to microorganisms of the read across substance, C12-14 ADMAES (96% active), in an activated sludge respiration inhibition testaccording to OECD 209 Guideline, in compliance with GLP. The toxicity of the read across substance to activated sludge was determined at a contact time of 3 h, using various concentrations of the read across substance. The inhibitory effect of the substance at a particular concentration is expressed as a percentage of the two controls. No analytical determination was performed for the read across substance. The EC50 of read across substance in sludge after 3 h of contact time was determined to be 30 mg/L (nominal) (equivalent to 28.8 mg a.i./L) (95% confidence limits: 13 and 72 mg/L). The 3 h EC10, EC20, EC80 were determined to be 9, 14 and 66 mg/L (nominal) (equivalent to 8.64, 13.44 and 63.36 mg a.i./L), respectively. The read across substance was therefore considered harmful to activated sludge. The test was valid as shown by the EC50 of the reference compound 3,5 -dichlorophenol (15 mg/L), the coefficient of variation of the replicates of the control oxygen uptake rates (10%) and the average oxygen uptake rate for the control replicates (30 mg O2/g dry weight/hour). Under the study conditions, the EC50 of read across substance in sludge after 3 h of contact time was determined to be 30 mg a.i./L (nominal) (Geerts, 2012). Based on the results of the read across study, similar effect levels can be considered for the test substance.