4,4'-(4-methylpentane-2,2-diyl)bis((heptyloxy)benzene)

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11 April to 19 April 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

1-(heptyloxy)-4-{2-[4-(heptyloxy)phenyl]-4-methylpentan-2-yl}benzene

EC Number:

830-582-9

Cas Number:

1951440-04-2

Molecular formula:

C32H50O2

IUPAC Name:

1-(heptyloxy)-4-{2-[4-(heptyloxy)phenyl]-4-methylpentan-2-yl}benzene

Test material form:

liquid: viscous

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: AS455433
- Expiration date of the lot/batch: 01 November 2020
- CAS number 1951440-04-2
-Purity: 98.5%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature protected from the light
- Stability under test conditions: not stable
- Solubility and stability of the test substance in the solvent/vehicle: Preliminary data indictaed the test substance to have a water solubility of <0.023mg/l using a slow stirring flask method.

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0.1mg/l
- Sampling method: 2.0ml was taken from the approximate centre of the test vessels
- Sample storage conditions before analysis: Samples were stored in a freezer (≤-15°C) until analysis at the analytical laboratory of the Test Facility.

Test solutions

Vehicle:

yes

Remarks:

tetrahydrofuran (THF)

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Preparation of test solutions started with dissolving test item in tetrahydrofuran (THF) to obtain a stock solution with a nominal concentration of 1.0 mg/mL. No other treatment than vigorous shaking was needed to completely dissolve the test item. The stock solution was then directly spiked into the test vessels (100 µl).
- Controls: Controls were spiked with 100µl of THF without test substance.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): tetrahydrofuran (THF)
- Concentration of vehicle in test medium (stock solution and final test solution(s) or suspension(s) including control(s)): 0.10 mg/l THF in test media
- Evidence of undissolved material (e.g. precipitate, surface film, etc.): No evidence of undissolved test item was observed in the stock or test solution.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Water flea
- Strain/clone: not stated
- Age at study initiation (mean and range, SD): at least third generation, obtained by a cyclical parthenogenesis under specified breeding conditions.
- Method of breeding: clinical parthenogenesis
- Source: not stated
- Feeding during test : not stated
- Food type: not stated

ACCLIMATION /BREEDING
- Acclimation period: 4 weeks stated as maximum age of cultures
- Acclimation conditions (same as test or not): not stated
- Type and amount of food: daily, a suspension of fresh water algae
- Feeding frequency: daily
- Health during acclimation (any mortality observed): not stated
-Test medium: various salts and vitamins were added to the test medium during the breeding.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test conditions

Hardness:

180 mg/l

Test temperature:

19 to 20 °C

pH:

7.8 to 8.3

Dissolved oxygen:

9.2 to 9.5 mg/l

Salinity:

not stated

Conductivity:

not stated

Nominal and measured concentrations:

Nominal concentration: 0.1mg/l
Measured concentration: At time 0 the concentration was 0.073mg/l, and after 48 hours it had decreased to 0.026mg/l. The average exposure concentration was calculated to be 0.043mg/l which was used for expression of effects.

Details on test conditions:

TEST SYSTEM
- Test vessel: 60ml, all glass
- Type (delete if not applicable): not stated
- Material, size, headspace, fill volume: 10ml
- Volume of solution: 50ml
- Aeration: no aeration of the test solutions was applied
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per vehicle control (replicates): 4 (only vehicle control)
- Biomass loading rate: not stated

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water purified by Reverse Osmosis (RO-water, GEON Waterbehandeling, Berkel-Enschot, The Netherlands):
- Culture medium different from test medium:
CaCl2.2H2O 211.5 mg/L
MgSO4.7H2O 88.8 mg/L
NaHCO3 46.7 mg/L
KCl 4.2 mg/L

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: a daily photoperiod of 16 hours
- Light intensity: not stated

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : immobility at 24 and 48 hours

VEHICLE CONTROL PERFORMED: yes

RANGE-FINDING STUDY : not performed. Based on the very low water solubility of the test item, no acute toxic effects were expected. Therefore, testing started with a limit test.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Results and discussion

Effect concentrations

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 0.043 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

- Other biological observations: 70 to 60% of the daphnids exposed to the limit concentration were entrapped at the surface of the solution after 24 and 48 hours. As the test item is highly hydrophobic and poorly soluble in aqueous media, it was hypothesized that daphnids became buoyant due to adsorption of test item to their body’s outer surface. However, microscopic observation revealed that no visible test item was attached to the daphnids. According to the definition of ‘immobility’ as described in Annex 1 of the OECD Test Guideline 202 (2004), all daphnids exposed to the limit concentration were mobile after being re-immersed in the solution. Therefore, the study was concluded, and based on the result of the limit test, EC50-values for immobility were considered to be beyond the maximum concentration tested.
- Mortality of control: None observed.
- Other adverse effects control: None observed.
- Immobilisation of control: None observed
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No precipitation observed.
- Water solubility: the 48 EC50 was thought to be above the water solubility limit.

Results with reference substance (positive control):

- Results with reference substance valid: the sensitivity of this batch of Daphnia magna was in agreement with ISO International Standard 6341, October 2012 and the historical data collected at Charles River Den Bosch.
- Relevant effect levels: 0.10 to 1.8mg/l
- Limit test: no
- Dose-response test: yes
- ECx: The 24h-EC50 was 0.92 mg/L with a 95% confidence interval between 0.78 and 1.1 mg/L. The 48h-EC50 was 0.55 mg/L with a 95% confidence interval between 0.47 and 0.64 mg/L.

Any other information on results incl. tables

Table 1. Number of Introduced Daphnids and Incidence of Immobility in the Limit Test

 

Time (h)	Replicate	4,4’-(4-methylpentane-2,2-diyl)bis((heptyloxy)benzene) Average exposure concentration(mg/L)
	Control		0.043
0	A	5	5
	B	5	5
	C	5	5
	D	5	5
	Total introduced	20	20
24	A	0	0 (4)
	B	0	0 (4)
	C	0	0 (4)
	D	0	0 (2)
	Total immobilised	0	0
	Effect %	0	0
48	A	0	0 (4)#
	B	0	0 (3)
	C	0	0 (3)
	D	0	0 (3)
	Total immobilised	0	0
	Effect %	0	0

( ) - Number of daphnids observed trapped at the surface of the test solutions. These organisms were re- immersed into the respective solutions before recording of mobility,#Microscopic observation revealed that no visible test item was attached to the entrapped daphnids in this replicate.

Table 2. Limit Test: Test Samples Concentrations

Time ofsampling[hours]	Date ofsampling	Date of analysis1	Concentration [mg/L]		Relative to nominal [%]	Relative to initial [%]
			Nominal	Analyzed
0	16 Apr 2019	24 Apr 2019	0	n.d.	n.a.
			0.10	0.0732	73
48	18 Apr 2019	24 Apr 2019	0	n.d.	n.a.	n.a.
			0.10	0.0262	26	35

1         Samples were stored in the freezer (≤ -15°C) until the day of analysis.

2         Estimated value, calculated by extrapolation of the calibrationcurve.

n.d. Not detected.

n.a. Not applicable.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The 48h-EC50 for immobility of Daphnia magna exposed to 4,4’-(4-methylpentane-2,2-diyl)bis((heptyloxy)benzene) was beyond the maximum concentration tested, i.e. exceeded an average exposure concentration of 0.043 mg/L which was above the water solubility limit.

Executive summary:

Assessment of the acute toxicity of 4,4’-(4-methylpentane-2,2-diyl)bis((heptyloxy)benzene) to Daphnia magna was carried out according to the OECD 202 Guidelines and in accordance with GLP. Due to the low solubility of the substance a stock solution was prepared with tetrahydrofuran (THF) and this was used to spike the test medium. A limit test was performed due to the low solubility of the substance with daphnids exposed to nominal concentrations of 0.1mg/l. Analytical confirmation of the test concentrations at time 0 and time 48 hours indicated a decreased in exposure concentration with an average exposure concentration calculated at 0.043mg/l.

No immobility was observed in the control and at the limit concentration throughout the test, although there were issues with the daphnids becoming trapped in the surface layer in the exposure vessels depsite no visible signs of the test material in the test solutions or on the surface of the daphnids. The 48 hour EC50 for immobility was calculated at >0.043mg/l, which was above the water solubility limit.