Reaction products of N-Phenyl-1-Naphthylamine with Nonene, branched

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

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test (Carbon and Ammonium Oxidation))

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Test substance No.: 20/0205-4
Batch identification: SP66MF (L1+L2)
CAS No.: 63451-49-0
Content: 99.92 g/100 g
100 g/100 g (test item) – 0.03 g/100 g (water) – 0.054
g/100 g (PANA)
Identity: confirmed
Homogeneity: No
Storage stability: Expiry date: 14 Jan 2024
Date of production: 15 Jan 2021
Physical state/appearance: liquid, high viscous / brown to red
Water solubility: At 20.0 °C ± 0.5 °C 0.7 μ/L ± 0.5 μg/L for tests with 0.1 g/L; 0.7
μg/L ± 0.3 μg/L for tests with 1.0 g/L
Storage conditions: Ambient (room temperature)

Details on sampling:

Test system: municipal Activated sludge
Age: 1 day
Origin: aeration tank of the wastewater treatment plant of Lambsheim, Germany
Collection of the test system: 30 Aug 2021
Arrival in the test facility: 30 Aug 2021
After arrival of the activated sludge suspension in the test facility the suspension was sieved with a fine woven mesh (mesh size about 1 mm). This suspension was pre-aerated over night at room temperature. At the next day the sludge suspension was washed once with drinking water and the suspension was adjusted to 3 g/L Dw.

Details on test solutions:

TEST SUBSTANCE PREPARATIONS
Application of the test substance: Direct addition by initial weight into the test flasks. The test substance was weighed in the required amounts for the test concentrations directly to the test vessels.
3.4.1. Reference substance preparations
1000.4 mg of the reference substance was added to about 800 mL of deionized water and stirred at room temperature until the reference substance was completely dissolved. The pH value of the stock solution was measured and adjusted to 7.2 with 1 M sodium hydroxide solution. Following this the stock solution was made up to 1L with deionized water. The stock solution appeared colorless-clear and no undissolved reference substance was visible.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

Inoculum from aeration tank of the wastewater treatment plant of Lambsheim, Germany.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 d

Test temperature:

20±2 °C

Nominal and measured concentrations:

Test concentrations: 1000, 500, 250, 125, 62.5 mg/L as nominal concentration
based on test substance without correction of purity and blank controls.

Details on test conditions:

Test replicates: 6 replicates for the control,
3 replicates for each test substance concentration,
2 replicates for each reference substance concentration.

Test conditions
Incubation time: 3 hours
Test temperature: 20.1-21.7°C
Test vessels: Glas-beakers (nominal volume 1L)
Test volume: 500 mL
Synthetic medium: 16 mL/test vessel of 100-fold concentrated OECD medium
Oxygen consumption of the test system:
9.8 mg O2/g*h, deviation from the default value 20 mg O2/g*h
Oxygen concentration during aeration: > 2 mg/L
Oxygen concentration immediately before measurement:
> 7 mg/L
Sludge concentration in the test vessel: 1.5 g/L Dw

The test substance was added in the required amounts according to the test concentrations directly to the test vessels with 234 mL deionized water. Aliquots of the reference substance stock solution were dosed to the test vessels and made up with deionized water to a volume of 234 mL. 16 mL synthetic medium were dosed to each test vessel with test substance and reference substance afterwards. To prepare the blank control assays 234 mL of deionized water and 16 mL synthetic medium were mixed.
The pH-values in all test vessels were checked. An adjustment was not necessary.
After addition of 250 mL of inoculum suspension (3 g/L DW) the incubation was started by aeration of the test vessels with pressured air. The vessels for the blank control assays were prepared to the same procedure without addition of test- or reference substance.
After 3 hours incubation at 20  2°C the mixtures in the test vessels were placed subsequently for oxygen measuring. The temperature was measured for seven times in a separate vessel filled with aerated deionized water during incubation phase. The content of oxygen at the start of the measurements was > 7 mg/L. The oxygen uptake was measured for a period of about 8 to 10 minutes.
The total oxygen consumption of was measured in the sequence BC1-3 and RS1, RS2-5, RS6 and TS1-3, TS4-7, TS8-11 and TS12-15. The oxygen consumption of the blank control BC4-6 were measured at last. No abiotic control was tested.

Reference substance (positive control):

yes

Remarks:

3,5 dichloropheno

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

Key result

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth inhibition

Details on results:

Validity criteria:
The coefficient of variation of the six replicates of blank control was 10.5 % O2 consumption.
The EC50 of 3,5-dichlorophenol was 5.5 mg/L
The mean oxygen uptake of the blank controls was 9.8 mg/g*h
The results in this study were consistent with the validity criteria with one exception. The mean
oxygen uptake in the blank controls was lower than 20 mg/g*h (10 mg/g*h). The reference
substance shows an EC50 in the specified range (usual range of EC50 of the last 20 values
(within the values in this study) in the laboratory was 4.7-10.5 mg/L). The used activated sludge was collected from a municipal waste water treatment plant. The last 20 collections showed a maximum oxygen consumption of 18.7 mg/g*h and minimum oxygen consumption of 9.8 mg/g*h (within the values in this study). For this the study is classified as valid.

Results with reference substance (positive control):

The EC50 of 3,5-dichlorophenol was 5.5 mg/L

Validity criteria fulfilled:

yes

Conclusions:

In an OECD 209 test the test item was not acute toxic toward microorganism (activated sludge). The EC10 and EC50 were> 1000 mg/L nominal.

Description of key information

In an OECD 209 test the test item was not acute toxic toward microorganism (activated sludge). The EC10 and EC50 were> 1000 mg/L nominal.

Key value for chemical safety assessment

EC50 for microorganisms:

1 000 mg/L

EC10 or NOEC for microorganisms:

1 000 mg/L