Hexanoic acid, 2-ethyl-, C16-18-alkyl esters

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

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

Adequacy of study:

key study

Study period:

06 Sep - 27 Sept 1995

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study tested with acceptable restrictions.

Qualifier:

according to guideline

Guideline:

other: OECD 202, part II (reproduction test) adopted 1984

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
PHYSICO-CHEMICAL PROPERTIES
- Melting point: -53 to -30 °C
- Boiling point: > 300 °C, decomposition probable
- Vapour pressure: < 0.0293 Pa at 20 °C
- Water solubility: < 1 mg/L
- log Pow: log Kow = 6.68 - 8.65

OTHER PROPERTIES
- Results of test for ready biodegradability: 65 - 97% in 28 days: readily biodegradable

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: 1 and 100 mg/L respectively of the test substance were stirred in 1000 mL of test medium for 16 hours to 7 days. After a settling period of 2 hours, the solutions were filtered through a glass fibre filter, which were activated with 1 mL NaOH and washed with deionised water before.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: waterflea
- Source: in-house culture, originally obtained from BGA Berlin, received May 1994
- Age of parental stock : < 24 hours
- Feeding during test:
- Food type: Scenedesmus subspicatus
- Amount: 2.5*10E06 cells/daphnid per day with 0-7 day old animals; 5.0*10E06 cells/daphnid per day with >7 day old animals
- Frequency: daily

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Test temperature:

21.6 - 23.1 °C

pH:

7.7 - 8.2

Dissolved oxygen:

88 - 100%

Nominal and measured concentrations:

Nominal: 1 and 100 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: beaker
- Type: closed with glass plates
- Material: glass; Size: 100 mL; Fill volume: 50 mL
- Renewal rate of test solution: 3 times per week
- No. of organisms per vessel: 1
- No. of vessels per concentration: 10
- No. of vessels per control: 10

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Elendt M4-medium

OTHER TEST CONDITIONS
- Photoperiod: 16 h light/ 8 h dark

EFFECT PARAMETERS MEASURED: The number of neonates was counted daily.

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOELR

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

(WAF)

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

(WAF)

Basis for effect:

reproduction

Details on results:

- Mortality of parent animals: 10% of parent animals died at 100 mg/L. No mortality was observed in the control and and the 1 mg/L treatment.
- No. of offspring produced per day per female: Control: 4.3 neonates/daphnid/d; 1 mg/L: 4.1 neonates/daphnid/day; 100 mg/L: 2.9 neonates/daphnid/day; Significant reduction of the reproduction rate (t-Test, 95%-level)
- Time to first brood release or time to hatch:
control: all 10 daphnids at day 9; 1 mg/L: all 10 daphnids at day 9; 100 mg/L: one daphnid at day 9, 5 daphnids at day 12 and 4 daphnids at day 14
- The effects in the 100 mg/L filtered test solution may be due to a better water soluble by-compound (for example a short chain fatty acid ester) or ,less likely, due to a toxic impurity.

Table 1: Neonate development in control

Control
Daphnid no.	Day 9	Day 12	Day 14	Day 16	Day 19	Day 21	Σoffspring in 21 days per animal	Average offspring/ daphnid/d
1	11	12	21		11	14	69	4.6
2	10	11	21		16	12	70	4.7
3	7	10	19		9	14	59	3.9
4	8	10	19		11	14	62	4.1
5	10	10	21		15	14	70	4.7
6	11	11	18		14	14	68	4.5
7	6	11	19		14	14	64	4.3
8	11	8	19		13	12	63	4.2
9	9	6	17		14	14	60	4.0
10	7	10	17		13	12	59	3.9
Average							64.4	4.3

Table 2: Neonate development at 1 mg/L

Nominal 1 mg/L
Daphnid no.	Day 9	Day 12	Day 14	Day 16	Day 19	Day 21	Σoffspring in 17 days per animal	Average offspring/ daphnid/d
1	10	11	17		11	9	58	3.9
2	10	14	16		12	10	62	4.1
3	7	11	18		12	13	61	4.1
4	7	14	20		9	13	63	4.2
5	8	9	21		11	11	60	4.0
6	10	10	17		11	11	59	3.9
7	11	12	20		10	13	66	4.4
8	9	12	18		9	13	61	4.1
9	10	10	18		13	11	62	4.1
10	5	9	23		11	10	58	3.9
Average							61.0	4.1

Table 3: Neonate development at 100 mg/L

Nominal 100 mg/L
Daphnid no.	Day 9	Day 12	Day 14	Day 16	Day 19	Day 21	Σoffspring in 17 days per animal	Average offspring/ daphnid/d
1		6	15		16	11	48	3.2
2		6		15	23		45	3.0
3		7		14	22		43	2.9
4			13		11	14	38	2.5
5			11	10	21		42	2.8
6			2		11	T/0
7		2	15		23	7	47	3.1
8			13	2	17	9	41	2.7
9		8		11	20		39	2.6
10	5	2	22		14	11	54	3.6
Average							44.1	2.9

Reference 2

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

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

Adequacy of study:

key study

Study period:

06 Oct - 28 Oct 1995

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study tested with acceptable restrictions.

Qualifier:

according to guideline

Guideline:

other: OECD 202, part 2 (1993)

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Eluate of 1 and 100 mg/L at 0, 24, 48 and 72 hours

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: Stock solutions of 1 and 100 mg/L in test medium (M4) were prepared and stirred for 16 h up to 5 d. The suspension was filtered via glass fibre filter, the water phase, supposedly satisfied with test substance to the limit of solubility, was used without further dilution

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: water flea
- Source: in-house culture, originally obtained from BGA Berlin (May 1994)
- Feeding during test: yes
- Food type: Scenedesmus subspicatus
- Amount: 2.5*10E6 cells/daphnid/day for 0-7 day old animals; 5.0*10E6 cells/daphnid/day for >7 day old animals
- Frequency: daily

ACCLIMATION
- 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:

not stated

Test temperature:

20.9 - 22.1 °C

pH:

7.8 - 8.2

Dissolved oxygen:

89 - 100 % (air saturation)

Nominal and measured concentrations:

Nominal: 1 and 100 mg/L
Measured: <0.01 and <0.01 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flask
- Type: closed by watchglass
- Size: 100 mL; Fill volume: 50 mL
- Renewal rate of test solution: every 2 to 3 days
- No. of organisms per vessel: 1
- No. of vessels per concentration: 10
- No. of vessels per control: 10

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: same, M4-Elendt

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light, 8 hours dark
- Light intensity: 300 Lux

EFFECT PARAMETERS MEASURED: The number of offspring and mortality of parental animals were recorded daily.

Reference substance (positive control):

no

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtered test solution

Basis for effect:

mortality

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtered test solution

Basis for effect:

reproduction

Remarks:

number of offspring

Details on results:

- Measured concentration of test substance: < 0.01 mg/L in the 1 mg/L as well as in the 100 mg/L sample at all sampling times.
- Mortality of parent animals: 0% in control and 1 mg/L (nominal) at the end of the test, 90% died on day 19 in 100 mg/L (nominal);
- No. of offspring produced per day per female: 5.8 offspring/daphnid/day in control over 21 days; 6.0 offspring/daphnid/day in 1 mg/L (nominal) over 21 days and 6.7 offspring/daphnid/day in 100 mg/L (nominal) over first 17 days as well as 4.5 offspring/daphnid/day in 100 mg/L (nominal) over full 21 days (23% reduced due to mortality of parent animals);
- other: limit of solubility of test substance <0.01 mg/L.
- The authors state, that the deaths of 90% of parent animals on day 19 in the 100 mg/L (nominal) sample could have resulted from sample contamination. Another course to explain the given results would be an error in eluate preparation. Either way, it is safe to say that a NOEC > 1 mg/L (nominal) is given for reproduction (number of offspring) and mortality (parent animals).

The analytic shows a similarly small concentration of test substance (< 0.01 mg/L) in the 1 mg/L as well as in the 100 mg/L sample at all sampling times.

Table 1: Number of offspring and average offspring per daphnid per day in the control

Control
Daphnid no.	Day 10	Day 12	Day 14	Day 17	Day 19	Day 21	Σoffspring in 21 days per animal	Average offspring/ daphnid/d
1	8	23	0	20	20	13	84	6.5
2	7	0	24	18	21	0	70	5.4
3	6	0	28	19	18	0	71	5.5
4	12	0	21	21	19	11	84	6.5
5	5	13	19	0	12	18	67	5.2
6	5	19	0	21	24	12	81	6.2
7	10	0	21	18	19	15	83	6.4
8	7	0	22	19	25	0	73	5.6
9	6	0	16	18	18	17	75	5.8
10	4	20	0	22	10	13	69	5.3
Average							75.7	5.8

Table 2: Number of offspring and average offspring per daphnid per day in the 1 mg/L concentration

Nominal concentration: 1 mg/L (analytically <0.01 mg/L)
Daphnid no.	Day 10	Day 12	Day 14	Day 17	Day 19	Day 21	Σoffspring in 21 days per animal	Average offspring/ daphnid/d
1	11	19	0	15	19	16	80	6.2
2	0	12	17	10	0	33	72	5.5
3	11	0	21	16	20	0	68	5.2
4	8	17	0	21	29	16	91	7.0
5	7	19	0	21	13	16	76	5.8
6	10	0	23	19	26	0	78	6.0
7	10	0	20	18	21	15	84	6.5
8	9	18	0	18	18	20	83	6.4
9	5	17	0	18	22	18	81	6.2
10	8	0	14	18	29	0	67	6.2
Average							78.0	6.0

Table 3: Number of offspring and average offspring per daphnid per day in the 100 mg/L concentration up to day 17*

Nominal concentration: 100 mg/L (analytically <0.01 mg/L)
Daphnid no.	Day 10	Day 12	Day 14	Day 17	Day 19	Day 21	Σoffspring in 17 days per animal	Average offspring in/ daphnid/d
1	10	19	0	21	T		50	6.3
2	13	25	0	25	T		63	7.9
3	5	21	26	0	0	T/9	52	6.5
4	8	20	0	23	6	T	51	6.4
5	7	26	0	25	T		58	7.3
6	9	0	24	16	T		49	6.1
7	6	20	27	11	0	12	64	8.0
8	8	23	0	23	T/8		54	6.8
9	3	15	23	0	9	T	41	5.1
10	10	5	23	17	T		55	6.9
Average							53.7	6.7

* On day 19 parent animals started dying, so this values were excluded (T = animal found dead)

Description of key information

No adverse effects up to the limit of water solubility (< 0.15 mg/L) for Daphnia magna (OECD 211)

Additional information

Since no studies investigating the long-term toxicity of Hexanoic acid, 2-ethyl-, C16-18 alkyl esters (CAS No. 90411-68-0) to aquatic invertebrates are available for this endpoint, in accordance to Regulation (EC) No. 1907/2006 Annex XI, 1.5 a read-across to the structurally related source substances C8-16-Fatty acid-2-ethylhexyl ester (CAS No. 135800-37-2) and 2-Ethylhexyloleat (CAS No. 26399-02-0) are conducted.

A read across from C8-16-Fatty acid-2-ethylhexyl ester and 2-Ethylhexyloleat to Hexanoic acid, 2-ethyl-, C16-18 alkyl esters to cover the long-term toxicity to aquatic invertebrates endpoint is acceptable as firstly all substances have common functional groups being fatty acid esters. The fact that the target substance is an ester of 2-ethylhexanoic acid and fatty alcohols with carbon chain lengths of C16 - C18 while the source substances are esters of 2-ethylhexanol and fatty acids with a carbon chain length of C8-C16 and C18, is not considered to invalid the read-across approach. This is assumed due to the similar physico-chemical properties and the common breakdown products of the substances, which are expected to be rapidly metabolised in the body via enzymatic hydrolysis to the starting materials (e.g., see Barron et al. 1999 for metabolism of esters in fish), which then fed into body’s natural catabolic processes, such as ß-oxidation and citric acid cycle (e.g., Rahn et al., 1973; Henderson, 1996). 

All substances are measured or calculated to be insoluble in water and the main components of the substances have similar molecular weights of 368.6 - 396.7 g/mole (CAS No. 90411-68-0), 256.43 – 312.54 g/mole (CAS No. 135800-37-2) and 394.69 g/mole (CAS No. 26399-02-0). The volatility of all substances is low (< 0.01 Pa at 20 °C). Hence, the behaviour, the fate, the bioaccumulation and thus the toxicity of the substances in the aquatic environment are considered to be identical.

The read-across study with C8-16-Fatty acid-2-ethylhexyl ester (CAS No. 135800-37-2) was conducted under GLP according to OECD guideline 202, part II using Daphnia magna as test organism (Wierich, 1998). Test concentrations of 1 and 100 mg/L were applied as filtered test solutions. No effects on mortality and reproduction were observed in the 1 mg/L treatment. A significant reduction of reproduction was observed in the 100 mg/L treatment. The authors supposed that the effects in the 100 mg/L treatment was due to a better water soluble by-compound or, less likely, due to a toxic impurity. Hence, the 21d-NOEC is determined to be ≥ 1 mg/L.

The second read-across study with 2-Ethylhexyloleat (CAS No. 26399-02-0) was also performed under GLP according to OECD guideline 202, part II using Daphnia magna as test organism (Kuech, 1995). Test concentrations of 1 and 100 mg/L were applied as filtered test solutions. Test concentrations were measured to be < 0.01 mg/L in both treatments. No effects on mortality and reproduction were observed in the 1 mg/L treatment. 90% mortality (on day 19) and 23% reduction of reproduction (day 21) was observed in the 100 mg/L treatment. The authors stated that the deaths in the higher concentration could be attributed to sample contamination. However, as the 21d-NOEC is determined to be ≥ 1 mg/L, the substance is considered not to cause toxic effects on the reproduction of aquatic invertebrates.

Due to structural and property similarities to the analogue substances C8-16-Fatty acid-2-ethylhexyl ester (CAS No. 135800-37-2) and 2-Ethylhexyloleat (CAS No. 26399-02-0), as explained above, the results are also applicable to read across to Hexanoic acid, 2-ethyl-, C16-18 alkyl esters (CAS No. 90411-68-0). 

A detailed reference list is provided in the technical dossier (see IUCLID, section 13) and within the CSR.