1,3-Propanediol, 2,2-bis(hydroxymethyl)-, polymer with 2-(chloromethyl)oxirane, 2-propenoate

Toxicity to microorganisms

Administrative data

Endpoint:

activated sludge respiration inhibition testing

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

November 2012-February 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted according to OECD guideline 209 and under GLP.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Pentaerythritol, ethoxylated, esters with acrylic acid

EC Number:

500-111-9

EC Name:

Pentaerythritol, ethoxylated, esters with acrylic acid

Cas Number:

51728-26-8

IUPAC Name:

51728-26-8

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): Pentaerythritol, ethoxylated, esters with acrylic acid
- Substance type: UVCB
- Physical state: clear, yellowisch liquid
- Lot/batch No.: 0008190677
- Expiration date of the lot/batch: 21 February 2013
- Storage condition of test material: room temperature

Sampling and analysis

Analytical monitoring:

not required

Test solutions

Vehicle:

yes

Details on test solutions:

- Test item
The test concentrations (10, 100, 1000 mg/L) were chosen to permit the determination of the EC50. The highest concentration was 1000 mg/L. The test item was not compatible with the test system at a concentration of 1000 mg/L (insoluble, forming a sticky mass of test item on the bottom of the glass vessel). Because of the low solubility, the test item was weighed onto microscope glass cover slips, and weighed before adding into the test vessels. In order to maximise the dispersion of test item into the liquid of the vessel, the mixture was heated to up to a maximum of 50 °C, then cooled to ~20 °C before adding the inoculum.

- Abiotic Control (ATS)
In parallel to the study, abiotic control flasks were tested without inoculum, but with the highest concentration of the test item, under identical test conditions.

- Untreated Control (B)
Five replicates of the untreated control group (deionised water, synthetic sewage and inoculum, without addition of the test item) were tested.

- Reference Control (REF)
In parallel with the test item, the reference item 3,5-Dichlorophenol was tested (at nominal test concentrations of 0.6; 2.2; 7.2; 24.0 and 80.0 mg/L) under otherwise identical test conditions. A stock solution of 3,5-Dichlorophenol was prepared according to the OECD Guideline No. 209 (0.50 g of 3,5-Dichlorophenol dissolved in 500 mL of water). Warm distilled water and ultrasonication was used to help the dissolution of the 0.5 g reference substance and the volume of the solution was made up to 0.5 litre with deionised water, when cooled to room temperature. The pH of the solution was checked and adjusted, with NaOH solution (1 mol/L) to pH 7.5.

Test organisms

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

Species: Activated sludge, microorganisms from a domestic waste water treatment plant.

Source: The activated sludge was supplied from the sewage plant for domestic sewage in Veszprém, Hungary

Conditioning:
The activated sludge used for this study was washed and centrifuged and the supernatant liquid phase was decanted. The solid mat erial was re-suspended in isotonic saline solution and again centrifuged. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to dry weight determined. Based on this ratio, calculated amounts of wet sludge were suspended in isotonic saline solution to yield a concentration equivalent to 3 g per litre (on dry weight basis). The pH of the activated sludge inoculum was determined to be pH 7.65. The activated sludge was used directly after conditioning.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

no

Test conditions

Hardness:

no data

Test temperature:

20 ± 2 °C

pH:

7.5 ± 0.5

Dissolved oxygen:

13.58 mg oxygen/l/g solid

Salinity:

no data

Nominal and measured concentrations:

The test concentrations (10, 100, 1000 mg/L) were chosen to permit the determination of the EC50. The highest concentration was 1000 mg/L. Because of the low solubility, the test item was added by directly weighing of the test item into the bottles.

Details on test conditions:

TEST SYSTEM
- Test vessel:
Type and size: Infusion bottles of approximately 500 ml volume, and BOD bottles with special neck of (~330 ml volume). Identification: Each test flask (controls and test item treated test mixtures) was uniquely identified with at least study code, treatment and replicate codes.

Preparation of the test flasks containing test item for both the biotic and abiotic test mixtures: 16 mL synthetic sewage feed and an adequate amount (see the Table 1) of the test item was placed (directly) into the test flasks and filled up with warm deionised water to 250 mL. Before the start of the incubation the above mixtures were shaken for a one hour period, to help the dispersion of test item into the test mixtures.

At the start of the test, 250 mL activated sludge inoculum with a sludge concentration of 3 g per litre of suspended solids was added, with the exception of the abiotic control flasks. The exact amount of sludge used in the inoculum of the test: 400 g wet sludge (= 25.8 mg solids)/8.6 litres dechlorinated tap water (= 3 g/L solids being in the inoculum). Abiotic controls in 5 parallels, and one untreated blank control was started as the first step of the test. Then at appropriate time intervals of approximately 30 minutes (an arbitrary but convenient interval), the further test groups were started. The second series were 5 different concentrations (no replicates) of the reference item and to the second blank control. The third series, were the 5 replicates of the highest test item concentration and the third blank control. The fourth series was the five replicates of second test item concentration ) and the fourth blank control. Finally the series with the lowest concentration and the fifth blank control.

-Measurement of pH, Dissolved Oxygen and Water Temperature
The pH and the oxygen concentrations were determined at the start and at the end of the incubation period in all treatments. The temperature was measured in the laboratory (it is the place of the flasks during the incubation period) with a min/max thermometer during the incubation period. The temperature of the test mixtures was recorded during the oxygen measurement in all BOD bottles.

-Measurement of Respiration Rate
For the measurement of the respiration rate a well-mixed sample of each treatment was poured into a BOD flask after about 3 hours incubation time, and was not further aerated. The oxygen concentration was measured for about fifteen minutes, with an O2 electrode and was recorded in every half minute. The oxygen consumption (in mg O2/L/minute) was determined from the most linear part of the respiration curve.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Results and discussion

Effect concentrations

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

In comparison to the inoculum controls, the respiration rate of the activated sludge was inhibited between 0% and 61.31% in the examined Pentaerythritol, ethoxylated, esters with acrylic acid range of 10 – 1000 mg/L. The average specific respiration rate of the untreated (blank) controls (B1-B5) was 13.58 mg O2/L/h/g solid. The coefficient of variation of oxygen uptake rate in above untreated replicates was 4.72 %.

The calculated EC50 is in the range of 400 – 500 mg/L.

Results with reference substance (positive control):

The following nominal concentrations of the positive reference control 3,5-dichlorophenol were tested on the same activated sludge and under identical conditions as the test item: 0.6, 2.2, 7.2, 24.0 and 80.0 mg/L.

In comparison to the controls, the respiration rate of the activated sludge was inhibited by 42.23 % at the lowest nominal concentration of 0.6 mg/L.
At the nominal concentrations of 2.2, 7.2, 24.0 and 80.0 mg/L, the respiration rate was inhibited by 46.09 %, 45.94 %, 89.07 % and 95.56 %, respectively. The 3-hour EC50 of 3,5-dichlorophenol was calculated to be 2.23 mg/L with 95% confidence limits of 1.56 to 3.19 mg/L.

Any other information on results incl. tables

Table.    Influence of test item on oxygen consumption of activated sludge

 

Test group		Conc. of test item in test mix. (mg/L)	Total oxygen consumption rate (RT)           (mg O2/L/h)	Specific respiration rate (RS)         (mg O2/L/h/g solid)	Inhib. (%)	pH values^#		Temperature (C°)		Variation coefficient of O2consumption rate (%)
ID	Name					start	end	Lab.*	Flasks **
ATS1	Abiotic	1000	0.09	–	–	7.55	7.42	19.0/20.8	17.6	–
ATS2		1000	0.09	–	–	7.54	7.39		17.5	–
ATS3		1000	0.04	–	–	7.53	7.37		17.4	–
ATS4		1000.	0.06	–	–	7.53	7.38		17.4	–
ATS5		1000	0.04	–	–	7.50	7.37		17.6	–
B1	Blank	0.00	20.02	13.35	–	7.52	7.75	19.0/20.8	17.3	4.72
B2		0.00	19.54	13.03	–	7.55	7.56		17.0
B3		0.00	21.85	14.57	–	7.53	7.78		17.8
B4		0.00	20.77	13.84	–	7.53	7.90		18.0
B5		0.00	19.64	13.09	–	7.53	7.79		17.8
REF1	Ref. item	80.00	0.93	–	95.56	7.60	7.96	19.0/20.8	17.1	–
REF2		24.00	2.20	–	89.07	7.59	7.93		18.0	–
REF3		7.20	10.63	–	45.94	7.58	7.95		17.6	–
REF4		2.20	10.60	–	46.09	7.62	7.98		16.9	–
REF5		0.60	11.35	–	42.23	7.64	7.92		16.9	–
TS1	Test item	1000	8.52	–	61.31	7.43	7.74	19.0/20.8	***17.6	–
TS2		100	20.96	–	-0.63	7.51	7.71		***17.6	–
TS3		10	20.62	–	-4.64	7.59	7.83		***18.2	–

#:      Individual values of the BOD flasks, except TS1-3, where the average values are reported

*:      Maximum/minimum temperature (°C) measured in the Laboratory, during the preparation and treating period

**:    Average temperature (°C) in the individual BOD flasks, during the measuring period

 

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Pentaerythritol, ethoxylated, esters with acrylic acid was found to have a EC50 of >100 mg/l. The calculated EC50 is in the range of 400 – 500 mg/L. At 100 mg/l, Pentaerythritol, ethoxylated, esters with acrylic acid had no effect on Respiration Rate in this test system, so no further testing is required.

Executive summary:

A laboratory test was carried out with Pentaerythritol, ethoxylated, esters with acrylic acid to evaluatethe effect of the test item on microorganisms by measuring the respiration rate.The test concentrations (10, 100 and 1000 mg/l) were chosen to permit the determination of theapproximate EC₅₀. The reference control results showed that in this study, the inoculum and methodology used provided a good measure of inhibition of activated sludge respiration rate. Pentaerythritol, ethoxylated, esters with acrylic acid was found to have a EC₅₀ of >100 mg/L. The calculated EC₅₀ is in the range of 400 – 500 mg/L. At 100 mg/L, Pentaerythritol, ethoxylated, esters with acrylic acid had no effect on the respiration rate in this test system, so no further testing is required.