Hexadecan- l-ol, ethoxylated

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

- Concentrations:
100 mg/l (WAF) on Days 0, 7, 14 and 20 (fresh media) and on Days 1, 8, 15 and 21 (old media)
Procedural recoveries: duplicate volumes of test medium were spiked with th test item at the required concentration (nominal 0.00546 mg/L) and treated as per the test samples.
- Sample storage conditions before analysis: Duplicate samples were taken on each occasion and stored frozen for further analysis if required.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Nominal amounts of test item (5.0, 16, 50, 160 and 500 mg) were each separately weighed onto a glass slide which was suspended below the surface of 5 liters of test water to give the 1.0, 3.2, 10, 32 and 100 mg/L loading rates respectively. After the addition of the test item, the test water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a dimple at the water surface. The stirring was stopped after 23 hours and the mixtures allowed to stand for 1-Hour. Visual observations made on the WAFs indicated that a significant amount of dispersed test item was present in the water column and hence it was considered justifiable to remove the WAFs by filtering through a glass wool plug (2 to 4 cm in length). A 10 mL and a 2.0 mL pipette, covered at one end with Nescofilm, were submerged into the vessels (sealed end down to mid depth within the vessel). A length of Tygon tubing was attached to the 10 mL pipette and the Nescofilm was removed using the 2.0 mL pipette. A glass wool plug was inserted into the opposite end of the tubing and the WAFs removed by mid depth siphoning (the first approximate 75 to 100 mL discarded) to give the 1.0, 3.2, 10, 32 and 100 mg/L loading rate WAFs. Further filtration through a single sheet of coarse filter paper was conducted in order to remove as much undissolved test item as possible. Microscopic observations of the WAFs were performed after filtering and showed there to be no micro-dispersions of test item present. A dilution was made from the 3.2 mg/L loading rate WAF to give the 0.32 mg/L loading rate WAF.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Age at study initiation (mean and range, SD): <24 h
- Method of breeding: Adult Daphnia were maintained in 150 mL glass vessels containing Elendt M4 medium in a temperature controlled room at approximately 20 °C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and dusk transition periods. Each culture was fed daily with a mixture of algal suspension (Raphidocelis subcapitata (formally known as Pseudokirchneriella subcapitata)) and GEMMA Micro 300 fish food suspension.
- Source: in-house culture
- Feeding during test
- Food type: algal suspension (Raphidocelis subcapitata), GEMMA Micro 300 fish food suspension
- Amount: 5.0 to 20 µL of an algal suspension, 20 µL fish food--> 0.1 to 0.2 mg carbon/daphnid/day,
- Frequency: algae daily, fish food unitl day 3

ACCLIMATION
- Acclimation period: <24 h
- Acclimation conditions (same as test or not): same

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

240 to 264 mg/L as CaCO3

Test temperature:

19-21 °C

pH:

7.1 - 8.2

Dissolved oxygen:

9.7- 8.3 mg/L

Nominal and measured concentrations:

Nominal loading rates (WAF): 0.32, 1.0, 3.2, 10, 32 and 100 mg/L

Chemical analysis of the fresh test preparations on Days 0, 7, 14 and 20 showed measured test concentrations to range from less than the limit of detection (LOD), determined to be 0.000025 mg/L, to 0.0044 mg/L. Measured concentrations below the limit of detection does no infer that no test item was in solution, just that any dissolved test item was at a concentration that could not be quantified.
Analysis of the corresponding aged preparations on Days 1, 8, 15 and 21 showed measured test concentrations of either less than the LOD, or less than the limit of quantification (LOQ), determined to be 0.00010 mg/L, were obtained.
Given that the toxicity cannot be attributed to a single component or a mixture of components, but to the test item as a whole, the results were based on nominal loading rates only.

Details on test conditions:

TEST SYSTEM
- Test vessel: 150 mL glass vessels
- Type (delete if not applicable): covered with a plastic lid
- Fill volume: 100 mL
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): daily
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10
- No. of vessels per control (replicates): 10

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted Water – Elendt M4 Medium
- Intervals of water quality measurement: at the start and end of the test and before and after each test media renewal.

OTHER TEST CONDITIONS
- Photoperiod:16 h light, 8 h darkness
- Light intensity: 839 to 997 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Daily assessment of mortality, numbers of live and dead "Filial" (F1) Daphnia and number of discarded unhatched eggs


TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 1.0, 10 and 100 mg/L.
- Results used to determine the conditions for the definitive study: No immobilization was observed at the nominal loading rates of 1.0 and 10 mg/L. However, 100% immobilization was observed at 100 mg/L loading rate WAF. Sub lethal effects of exposure were observed in the 100 mg/L loading rate WAF test group prior to death

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

0.32 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

loading rate WAF

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

0.32 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

loading rate WAF

Basis for effect:

other: body length

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

loading rate WAF

Basis for effect:

mortality

Details on results:

- Behavioural abnormalities: no
- Observations on body length and weight: A single adult in the 0.32 mg/L loading rate WAF test group was observed to be either pale, small and pale or small and pale and coated in debris between Days 5 and 18. This observation corresponded with the notably reduced number of neonate produced by this replicate.
- Other biological observations: There was a significant difference in parental mortality between the range-finding and definitive tests at a concentration of 100 mg/L loading rate WAF. It was considered possible that the degradation product(s) formed were more toxic than the parent test item and as such the implementation of a daily media renewal regime reduced the concentration of and time to which the daphnids were exposed to the degradation product(s).
- Mortality of control: no

Reported statistics and error estimates:

ToxRatPro software package Version 3.2.1 (TOXRAT, 2015)

Table 1. Number of live young produced per surviving adult

Nominal Loading Rate (mg/L)		Number of Live Young Produced per Surviving Adult by Day 21	Percentage Reduction in Live Young Produced Compared to the Control (%)
Control	Mean	193	Not applicable
	Standard Deviation	10.0
0.32	Mean	189	2
	Standard Deviation	10.1
1.0	Mean	171	11
	Standard Deviation	12.1
3.2	Mean	175	9
	Standard Deviation	12.3
10	Mean	162	16
	Standard Deviation	15.4
32	Mean	170	12
	Standard Deviation	10.1
100	Mean	138	28
	Standard Deviation	29.1

Table 2: Validity criteria for OECD 211.

Criterion from the guideline	Outcome	Validity criterion fulfilled
The mortality of the parent animals in the control (female Daphnia) does not exceed 20% at the end of the test.	0%	yes
The mean number of living offspring produced per parent animalsurvivingat the end of the test is ≥ 60 in the control.	193	yes

Validity criteria fulfilled:

yes

Remarks:

Please refer to table 2 at "Any other information on results incl. tables"

Additional information

The chronic toxicity of Hexadecan-1-ol, ethoxylated to Daphnia magna was assessed in a study following OECD guideline 211. Daphnids were exposed to test substance concentrations of 0.32, 1.0, 3.2, 10, 32 and 100 mg/L (nominal loading rate, WAF). The test solutions were renewed daily. Fresh and aged test solutions were sampled for analytical monitoring by HPLC/MS-MS. Effects on the survival rate of the parental generaltion were not observed. Based on the reproduction rate a NOELR (21 d) of 0.32 mg/L (nominal, WAF) was recorded.

Studies investigating the acute and chronic toxicity of the category substance Alcohols, C16-18 (even numbered), ethoxylated, < 2.5 EO (CAS No. 68439-49-6) are still ongoing. The outcome of the studies is expected to have an impact on the evaluation of the aquatic toxicity of Hexadecan-1-ol, ethoxylated.

Therefore, the current assessment of the aquatic toxicity of the target substance Hexadecan-1-ol, ethoxylated is considered preliminary and will be updated as soon as the final study reports are available.