Alcohols, C8-10, ethoxylated, sulfates, ammonium salts

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

7th Jun - 14th Oct 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

2019

Qualifier:

according to guideline

Guideline:

other: OECD Series on Testing and Assessment No. 23 Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures

Version / remarks:

2019

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

LC-MS/MS

Details on sampling:

- Concentrations: all treatment groups including the control
- Sampling time: fresh media at 0 h and 48 h; old media at 48 h and 96 h
- Sampling method: per treatment group triplicate 5 mL samples of the medium were transferred to 20 mL glass scintillation vials containing 5 mL of methanol
- Sample storage conditions before analysis: One replicate was analyzed on the day of receipt, the other two replicates were frozen as 'back-up' samples (<-10°C)

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: application via stock solution (ca. 3571 mg of the test item dissolved in 1 L treated mains water, resulting in a 1200 mg AI/L stock solution); An aliquot of 700 mL stock solution was diluted with mains water (7 L) resulting in a concentration of 120 mg AI/L (nominal).
- Eluate: treated mains water
- Controls: treated mains water only
- Evidence of undissolved material: no; test media and control were checked for the tyndall effect with a turbidity meter (Hach 2100 N); no significant differences observed between treatment and control group.

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISM
- Common name: fathead minnow
- Source: in-house breeding system
- Age at study initiation: same age and originated from the same source and population
- Length of a sub-sample of ten fish at the end of the acclimatisation period: 2.8 - 3.1 cm (mean: 3.0 cm)
- Weight of a sub-sample of ten fish at the end of the acclimatisation period: 0.3009 – 0.5406 g (mean: 0.4257 g)

- Method of breeding:
- Tanks: holding tanks appropriate to their size, under continuous water renewal (flow-through) conditions
- Light: artificial light (with a 16-hour light:8 hour dark photo-period with a ca 30 minute dawn/dusk period)
- Temperature: temperature-controlled room
- Food: fed ad libitum using freshly hatched brine shrimp (Artemia salinis) nauplii and also with Tetramin® flake food, which was added to the holding tank in quantities dictated by the size of the fish. The food was not considered to contain contaminants likely to affect the outcome of the study. Uneaten food and debris was siphoned or cleaned from the tanks as required.
- Water source and treatment: laboratory treated (activated carbon filter) mains supply (quality checked)

ACCLIMATION
- Acclimation period: for at least 9 days prior to testing
- Acclimation conditions: same as test
- Type and amount of food during acclimation: same as under breeding conditions
- Feeding frequency during acclimation: same as during normal breeding, except that the fish were not fed for at least 24 hours before the start of the definitive test
- Health during acclimation (any mortality observed): 0% mortality in the seven days prior to the start of the definitive test

FEEDING DURING TEST
- Food type: no feeding during the test

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

< 18 mg/L CaCO3

Test temperature:

22.6 – 24.2°C

pH:

6.69 – 7.85

Dissolved oxygen:

≥ 80.3%, with the exception of a single reading. At 48 hours, the dissolved oxygen in the old media
for the 120 mg AI/L was measured to be 55.3% ASV. Aearation was adjusted and was measured to be 80.3% ASV thereafter. This was considered not to affect the integrity of the test given that no mortality or adverse effects were observed.

Conductivity:

194 µS/cn

Nominal and measured concentrations:

Nominal: 0 (control), 120 mg a.i./L (equivalent to 357.1 mg test substance/L)
Analysis of the 120 mg a.i.L test concentration at 0 hours showed recoveries of between 127 – 157% of nominal for all the targeted constituents.
Measured: 0 (control), 96 mg a.i./L (TWA)

Details on test conditions:

TEST SYSTEM
- Test vessel: 10 L constructed glass aquaria, each fitted with a clear plastic lid, containing 7 L of media
- Aeration: yes
- Renewal rate of test solution: every 48 h (semi static)
- No. of organisms per vessel: 7 fish per replicate, randomly selected
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.4257 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: same as used for breeding
- Analysis of metals, pesticides: yes, 30.03.2022
- Chlorine (free/total (Cl2)): < 0.03 mg/L / < 0.06 mg/L
- Alkalinity: < 18 mg/L (as CaCO3)
- Ca/mg ratio: 8.1
- Culture medium different from test medium: no
- Water quality determinations (temperature, dissolved oxygen, pH): start and in 24 h intervals
- Light intensity: at test start and end (test start: 612 – 763 Lux; test end: 616 - 812 Lux)

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: same as breeding
- Light intensity: 616 to 812 Lux

EFFECT PARAMETERS MEASURED:
- Fish observation (behaviour, survival, appearance): ca 3 and 5 hours after addition and then at
24-hour intervals (24, 48, 72 and 96 hours) throughout the duration of the test


TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 0 (control), 80 and 120 mg AI/L (n=1); remaining test solution was prepared from the stock (120 mg AI/L) using a 2:3 dilution
- Test vessels: 3 L constructed glass aquaria with perspex clear lid
- Results used to determine the conditions for the definitive study: yes
- Chemical analysis of the test medium: yes, every 24 h

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 96 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: Concentration based on eight representative constituents

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 120 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Details on results:

- Other abnormalities: no
- Observations on body length and weight: sub-sample of ten fish taken at random from all surviving fish after 96 h: Mean 3.0 cm (2.8 - 3.1 cm); wet weight mean: 0.4257 g (0.3009 - 0.5406 g)
- Mortality of control: 0%
- Other adverse effects control: no
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: no
- Effect concentrations exceeding solubility of substance in test medium: no
- Test medium occurrence: colorless solutions with foam at the surface of the 120 mg AI/L due to the aeration of the tank at fresh media. The old media appeared as colorless solution for the control and as homogenous hazy dispersion for the 120 mg AI/L.

Reported statistics and error estimates:

The highest concentration where no significant mortality was observed i.e. the no observed effect concentration (NOEC) was also reported. No effects were observed throughout the test therefore the results were determined empirically.

Sublethal observations / clinical signs:

Preliminary test:

In general, all constituents appeared to be stable at the 120 and 80 mg a.i./L test concertation over a 24-hour and 48-hour period.
The results of the biology indicated that the LC50 for the test substance would be >120 mg a.i./L (no mortality observed in all treatment groups and controls after 96 h).

 

Test concentrations definitive test:

Analysis of the 120 mg a.i./L test concentration at 0 hours showed recoveries between 127 – 157% of nominal for all the targeted constituents. Analysis of the corresponding old media sample at 48 hours showed recoveries between 0.25 – 154% of nominal for all the targeted constituents. Analysis of fresh media at 48 hours showed recoveries between 106 – 191% of nominal for all the targeted constituents. Analysis of the corresponding old media sample at 96 hours showed recoveries between 0.15 – 107% of nominal for all the targeted constituents.
More pronounced losses in concentration of the constituents were observed over the second dosing period. This was considered to be possibly due to bacterial build-up due to the presence of the fish. Given that measured concentrations of the targeted constituents were generally
outside the 80 to 120% range of nominal value throughout the duration of the test, the results were based on time-weighted average exposure concentrations. The time-weighted average was calculated to be 96 mg/L as test substance.

 

Table 1: Analysis of test media per constituent during the definitive test

		% of nominal corrected per constituent
Nominal concentration (mg AI/L)		0 hour (Old)	48- hour	48-hour (Fresh)	96- hour
Control (for all constituents)		*-	*-	*-	*-
Ammonium octyl sulfate
120		129	31	106	0.34
1-Octanol, ethoxylated (1 EO), sulfated, ammonium salt
120		131	104	114	34
1-Octanol, ethoxylated (2 EO), sulfated, ammonium salt
120		127	136	185	100
1-Octanol, ethoxylated (3 EO), sulfated, ammonium salt
120		137	142	191	107
Ammonium decyl sulfate
120		132	0.25	127	0.22
1-Decanol, ethoxylated (1 EO), sulfated, ammonium salt
120		131	44	106	0.15
1-Decanol, ethoxylated (2 EO), sulfated, ammonium salt
120		149 (**)	145	111	51
1-Decanol, ethoxylated (3 EO), sulfated, ammonium salt
120		157 (***)	154	114	82

*<LOQ (Limit of quantification (LOQ) validated to 0.02 mg/L as test substance; Individual LOQ see details on analytical methods)

** "B" and "C" analysed to confirm original results. "A" sample was 9.47 mg/L, "B" was 11.0 mg/L and "C" samples was 11.2 mg/L. The mean of the "B" and "C" samples reported as the difference between the mean of the repeats and original samples was >15%

*** "B" and "C" analysed to confirm original results. "A" sample was 8.51 mg/L, "B" was 10.2 mg/L and "C" samples was 10.4 mg/L. The mean of the "B" and "C" samples reported as the difference between the mean of the repeats and original samples was >15%

 

Table 2: Mortality of fish recorded during the definitive test

		Cumulative Mortalities Recorded
Nominal concentration (mg AI/L)	Number of Pimephales promelas exposed	24 h	48 h	72 h	96 h
Control	7	0	0	0	0
120	7	0	0	0	0

 

Table 3: Validity criteria for OECD 203 (2019)

Criterion from the guideline	Outcome	Validity criterion fulfilled
The mortality in the control(s) should not exceed 10% (or one fish if less than ten are used) at the end of the test	0%	yes
The dissolved oxygen concentration must have been at least 60 per cent of the air saturation value throughout the test	≥ 80.3%*
There must be evidence that the concentration of the substance being tested has been satisfactorily maintained, and preferably it should be at least 80% of the nominal concentration throughout the test. If the deviation from the nominal concentration is greater than 20 per cent, results should be based on the measured concentration.	deviation from nominal concentration > 20% therefore effect values were based on TWA	yes

*with the exception of a single reading. At 48 hours, the dissolved oxygen in the old media
for the 120 mg AI/L was measured to be 55.3% ASV. Aearation was adjusted and was measured to be 80.3% ASV again. This was considered not to affect the integrity of the test given that no mortality or adverse effects were observed.

Validity criteria fulfilled:

yes

Remarks:

Please refer to table 3 at "Any other infrmation on results incl. tables"

Description of key information

EC50 (96 h) > 120 mg a.i./L (P. promelas, OECD 203)

Additional information

The assessment is based on the data currently available. New studies, based on the category review and the final decisions issued for some of the category substances, which are also relevant for this assessment, are currently being conducted. The hazard assessment with respect to aquatic toxicity will be updated once all ongoing studies have been finalised.

The acute toxicity of Alcohols, C8-10, ethoxylated, sulfates, ammonium salts to fish was investigated in a semi-static limit test following OECD guideline 203 (GLP). Seven fish of the species Pimephales promelas were exposed to a nominal test substance concentration of 120 mg a.i./L. The test medium was renewed every 48 h during the 96 h test period. The test concentrations was analytically verified by measuring the concentration of eight representative constituents by LC-MS/MS. Samples for analysis were taken from fresh and aged medium. The time weighted mean measured concentrations, based on the eight representative constituents of the UVCB substance was 96 mg a.i./L. No mortality or toxic symptoms were observed. The determined EC50 (96 h) was > 120 mg a.i./L nominal (EC50 (96 h) > 96 mg a.i./L, TWA).