Fatty acids, C18-unsatd., reaction products with acrylic acid and polyethylenepolyamines

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:

03 June to 09 June 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted in accordance with international guidelines and in accordance with GLP. All relevant validity criteria were met.

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes

Remarks:

GLP compliance statement

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
n/a

Analytical monitoring:

no

Details on sampling:

n/a

Vehicle:

no

Details on test solutions:

2000 mg of test item dissolved in water and volume adjusted to 1 litre to provide a 2000 mg/L stock solution. Dilutions made to produce 200 & 20 mg/L stocks.
A 250ml aliquot of 20mg/L stock was dispersed in 16mL synthetic sewage, 200mL activated sludge and water to a final volume of 500mL to give a 10m/L test solution. Test solutions of 32, 100, 320 and 1000 mg/L were prepared similarly.
Test solution flasks inverted severla times to ensure homogeneity.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

A mixed population of activated sewage sludge obtained on 9 June 2010 from the aeration stage of Severn Trent Water Plc sewage treatment plant, Loughborough, UK, which treats predominantly domestic sewage.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

After a 3 hour incubation period the concentration of dissolved oxygen (mg/l) was measured every minute in each vessel for 10 minutes.

Hardness:

Water partially softened giving a total hardness of 140mg/L as CaCO3.

Test temperature:

Room temperature controlled at 21+/-1 deg C

pH:

pH at 0 hours = 7.1 - 7.5
pH at 3 hours = 7.4 - 8.3

Dissolved oxygen:

The concentration of dissolved oxygen (mg/l) was measured in each vessel after 3 hour incubation.
Results shown in summary Table 1.

Salinity:

n/a

Nominal and measured concentrations:

Nominal test concentrations of 0 (control), 10, 32, 100, 320, 1000 mg/L were used.

Details on test conditions:

At time zero 16mL of synthetic sewage was diluted to 300mL with water and 200mL of inoculum were added in a 500mL conical flask (Control 1). The mixture was aerated at arate of 0.5-1 L/min. At 15 minute intervals the procedure was repeated with the reference substance being added and finally a second control prepared.
The test solutions were prepared as described in the "Details on test solution" section.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol (3,5-DCP)

Duration:

3 h

Dose descriptor:

other: EC20

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Duration:

3 h

Dose descriptor:

other: EC80

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Key result

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Details on results:

It was considered unnecessary and unrealistic to test at concentrations in excess of 1000mg/L.

Results with reference substance (positive control):

The EC50 for the reference substance 3,5-DCP was 6.4 mg/L, satisfying the OECD validity condition that the EC50 for this substance be in the range 5 to 30 mg/L. The second validity condition, given in OECD 209, that the two control flasks had respiration rates within 15% of each other was also satisfied in this test, with a variation of +/- 2%.
The 95% CI for the positive reference was 4.8 - 8.5 mg/L. The lower end of this range was slightly below the value given in the validation criteria, however this was considered to have no effect on the validity of the study.

Summary Results Table

	PR-4758 - Technical Grade		3,5-dichlorophenol
	ECx (3hours) (mg/L)	95% CI (mg/L)	ECx (3 hours) (mg/L)	95% CI (mg/L)
EC20	>1000	-	1.7	-
EC50	>1000	-	6.4	4.8 - 8.5
EC80	>1000	-	24	-
NOEC	1000	-	1.4	-

 

Table 1 – Oxygen Consumption Rates

Nominal Concentration (mg/L)		Initial O2 Reading (mg O2/L)	Measurement Period (minutes)	Final O2 Reading (mg O2/L)	O2 Consumption Rates (mg O2/L/min)	% Inhibition
Control	R1	6.3	9	2.2	0.46	-
	R2	6.6	9	2.3	0.48	-
Test Item	10	6.2	9	1.8	0.49	[4]
	32	5.3	8	1.7	0.45	4
	100	5.1	6	2.2	0.48	[2]
	320	4.7	5	2.5	0.44	6
	1000	3.9	4	2.2	0.43	9
3,5-DCP	3.2	7	10	3.8	0.32	32
	10	7.9	10	6.2	0.17	64
	32	8.3	9	7.7	0.07	85

 [] denotes an increase in respiration rate in comparison to the control

Table 2 - pH Values

Nominal Concentration (mg/L)		pH
		0 hours	3 hours
Control	R1	7.3	7.9
	R2	7.5	7.8
Test Item	10	7.4	7.8
	32	7.4	7.7
	100	7.4	7.6
	320	7.3	7.6
	1000	7.1	7.4
3,5-DCP	3.2	7.3	8.0
	10	7.4	8.1
	32	7.4	8.3

 

Validity criteria fulfilled:

yes

Conclusions:

The effect of the test item on the respiration of activated sewage sludge micro-organisms gave a 3 hour EC50 >1000mg/L, and the NOEC was 100mg/L.

Executive summary:

This report describes a study to determine the effect of the test substance on activated sludge micro-organisms by measuring the respiration rate, under defined conditions, in the presence of different concentrations of the test substance. The method is based on that described in OECD test guideline 209 in which activated sludge, freshly obtained from a sewage treatment plant receiving predominantly domestic sewage, is used as the microbial source. The purpose of the study is to assess the possible effect of the test substance on aerobic microbial treatment plants and to indicate suitable non-inhibitory concentrations of the test substance for biodegradability tests.   

Two controls, without test substance, were included in the study to determine the degree of experimental variation. The sensitivity of the activated sludge was estimated using three concentrations of the reference substance 3,5-dichlorophenol (3,5-DCP).

The EC50 for the reference substance 3,5-DCP was 6.4 mg/L, satisfying the OECD validity condition that the EC50 for this substance be in the range 5 to 30 mg/Ll. The second validity condition, given in OECD 209, that the two control flasks had respiration rates within 15% of each other was also satisfied in this test.

At all test concentrations 10 to 1000 mg/L of the test substance the inhibition rate ranged from 9% to -4%. The EC50 was >1000 mgL, and the NOEC = 1000 mg/L.

Description of key information

Sewage microorganism EC50 = >1000 mg/L; OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test); N Clarke (2010)

Key value for chemical safety assessment

EC50 for microorganisms:

1 000 mg/L

EC10 or NOEC for microorganisms:

1 000 mg/L

Additional information

One key study is available. The study was conducted to OECD 209 guidance, in accordance with GLP standards. Validity criteria were met.