1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-(C12-18(even numbered) acyl) derivs., hydroxides, inner salts

Long-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

fish early-life stage toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

April 26, 2018 - January 18, 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 210 (Fish, Early-Life Stage Toxicity Test)

Deviations:

yes

Remarks:

Range-finding test: a temporary deviation from the minimum level of temperature occurred. Final test: additional samples were taken on Day 4 and Day 5 from 0.32 mg solid content/L after a failure in the dosing system.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

As the test item is a UVCB, its quantification was based on the response of its main constituent, i.e. C12-alkyl derivatives.

Details on sampling:

- Concentrations: samples for possible analysis were taken from all test concentrations and the control; samples were also taken on Day -1 from the stock and on Day 28 from the mixing flasks.
- Sampling frequency: one day before the start of exposure to check the functioning of the system; at the start, after 7, 14, 21, 28 and 32 days of exposure; samples were also taken on Day 4 and Day 5 from 0.32 mg solid content/L after a failure in the dosing system.
- Sampling volume: 1.5 mL from one replicate of each test group changing systematically amongst replicates.
- Sample storage conditions before analysis: samples not analysed on the day of sampling were stored in a freezer (≤-15°C) until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
Stock solutions of 100 mg solid content/L were prepared in Milli-RO water every 48 hours. All stock solutions were clear and colourless. Exact volumes of these stock solutions were dosed with syringes via a computer-controlled system.

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISM
- Common name: Fathead minnow (Pimephales promelas, Teleostei Cyprinidae) Rafinesque.
- Source: In house culture.

Test type:

flow-through

Water media type:

freshwater

Remarks:

ISO medium

Limit test:

no

Total exposure duration:

32 d

Hardness:

161-196 mg CaCO3

Test temperature:

24.4-25.7°C

pH:

7.4-8.1

Dissolved oxygen:

6.3-8.6 mg/L

Nominal and measured concentrations:

Nominal concentrations: 0.00 (control), 0.010, 0.032, 0.10, 0.32 and 1.0 mg solid content/L.
Measured concentrations (arithemetic means): 0.00 (control), 0.0082, 0.020, 0.072, 0.19 and 1.0 mg solid content/L.

Details on test conditions:

TEST SYSTEM
- Test vessel: stainless steel vessels (~1.7L) with an approximate exposure volume of 1.5L; covered with a Perspex plate (open test system).
- Aeration: no.
- Renewal rate of test solution: continuous (1 litre per hour).
- Introduction of embryos: before cleavage of the blastodisc commenced (approximately 2-4 hours after fertilisation).
- No. of embryos per vessel: 20 embryos.
- No. of vessels per concentration (replicates): 4.
- No. of vessels per control (replicates): 4.

TEST MEDIUM
Adjusted ISO medium, formulated using RO-water (tap-water purified by reverse osmosis).

OTHER TEST CONDITIONS
- Light period: 16h photo-period daily, between 726 and 805 lux.
- Feeding:
o Embryonic phase: no feeding;
o Larvae and juvenile fish: brine shrimp Nauplii 24 or 48-hours old (supplied ad libitum).

EFFECT PARAMETERS MEASURED
- Stage of embryonic development: at the beginning of exposure from at least a representative sample of eggs.
- Hatching and survival: daily, numbers of hatched larvae and dead embryos, larvae and juvenile fish were recorded. Dead embryos, larvae and juvenile fish were removed directly after recording.
- Abnormal appearance/behavior: daily, abnormalities were recorded, e.g. hyperventilating, uncoordinated swimming, atypical quiescence and atypical swimming behaviour.
- Body weight: at the end of the test, all surviving fish were weighed on a replicate basis (blotted dry weight).
- Body length: at the end of the test, individual lengths from the surviving fish were measured.

RANGE-FINDING STUDY
- Test concentrations (nominal): 0.0 (control), 1.0, 10 and 100 mg solid content/L.

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

1 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: hatchability

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

0.097 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: post-hatch survival

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

0.12 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: larval growth (length)

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

0.075 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: larval growth (weight)

Duration:

32 d

Dose descriptor:

NOEC

Effect conc.:

0.072 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: hatchability

Duration:

32 d

Dose descriptor:

NOEC

Effect conc.:

0.072 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: post-hatch survival

Duration:

32 d

Dose descriptor:

NOEC

Effect conc.:

0.008 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: larval growth (length)

Duration:

32 d

Dose descriptor:

NOEC

Effect conc.:

< 0.008 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: larval growth (weight)

Details on results:

EMBRYONIC SURVIVAL AND HATCHING
The overall survival of embryos at the end of hatching was 98% for the control, which complies with the requirements of the guideline (i.e. at least 70% of the embryos in the control should hatch). Hatching success was 98-100% in concentrations up to and including 0.072 mg solid content/L and not statistically different from the control. Hatching success at the two highest concentrations of 0.19 and 1.0 mg solid content/L was 93 and 90%,
respectively, but proved to be statistically different from the control. The test item caused earlier onset of hatching. First larvae were observed on day three of exposure and were mainly related to the two highest concentrations tested (i.e. 25 and 31%, of the introduced embryos, respectively, had hatched). On day four the percentage of hatching in all five test concentrations was 9, 29, 49, 71 and 86%, while hatching in the control did not start until day 5. Hatching was complete for the control and all test concentrations on day five of exposure.

LARVAL SURVIVAL AND DEVELOPMENT
The mean post-hatch larval survival was 96% in the control. The validity criterion for posthatch survival of at least 75% was met. Mean survival rates in the test concentrations ranged from 0 to ≥96% and were clearly dose-related. Statistical analyses indicated that larval survival was not statistically different from the control at test concentrations up to and including 0.072 mg solid content/L.

EFFECTS ON LARVAL GROWTH
A statistically significant reduction of length was observed at concentrations of 0.020 mg solid content/L and higher, while reduction of weight was statistically significant at all concentrations tested.


See below (Any other information on results incl. tables + Attachments) for further information on results.

Reported statistics and error estimates:

Calculation of ECx-values was based on probit analysis using linear maximum likelihood regression with the percentages of post-hatch mortality and the percentages of reduction of growth (both weight and length) versus the logarithms of the corresponding average exposure concentrations of the test item. All analyses were performed with ToxRat Professional 3.2.1 (ToxRat Solutions® GmbH, Germany).

Post-hatch survival at the end of exposure (day 32)

Concentration (mg solid content/L)	Total hatched	Survived	Dead	% Post-hatch mortality
Control 0.0082 0.020 0.072 0.19 1.0	78 78 80 78 74 72	75 76 77 75 40 0	3 2 3 3 34 72	3.8 2.6 3.8 3.8 45.9* 100.0*

* Effect was statistically significant (p ≤0.05)

Mean body length (mm) and percentage of reduction at the end of exposure (day 32)

Concentration (mg solid content/L)	Mean	Std. dev.	n		% Reduction
Control 0.0082 0.020 0.072 0.19	25.38 25.08 24.80 24.83 17.57	0.386 0.369 0.258 0.222 0.350	4 4 4 4 4		1.2 2.3* 2.2* 30.7*

* Effect was statistically significant (p ≤0.05)

Mean body weight (mg wet weight) and percentage of reduction at the end of exposure (day 32)

Concentration (mg solid content/L)	Mean	Std. dev.	n		% Reduction
Control 0.0082 0.020 0.072 0.19	158.250 147.250 143.500 144.500 55.00	12.0934 8.1803 3.8730 6.2450 3.1623	4 4 4 4 4		7.0 9.3* 8.7* 65.2*

* Effect was statistically significant (p ≤0.05)

Effect parameters (mg solid content/L)

Parameter		NOEC	LOEC	EC10	EC50
Embryonic survival	Value (1)	0.072	0.19	1.0	>1.0
Post-hatch survival	Value (2) lower 95%-cl upper 95%-cl	0.072	0.19	0.097 0.075 0.12	0.20 0.17 0.24
Growth (length)	Value (2) lower 95%-cl upper 95%-cl	0.0082	0.020	0.12 0.11 0.12	0.26 0.25 0.28
Growth (weight)	Value (2) lower 95%-cl upper 95%-cl	<0.0082	0.0082	0.075 0.068 0.081	0.15 0.15 0.16

(1) For day 5 of exposure / (2) For day 32 of exposure / cl – confidence limit

Validity criteria fulfilled:

yes

Conclusions:

The results led to the following conclusions:
- the 32d-NOEC and EC10 for hatching were 0.072 and 1.0 mg solid content/L, respectively;
- the 32d-NOEC and EC10 for post-hatch survival were 0.072 and 0.097 mg solid content/L, respectively;
- the 32d-NOEC and EC10 for larval growth (length) were 0.0082 and 0.12 mg solid content/L, respectively;
- the 32d-NOEC and EC10 for larval growth (weight) were <0.0082 and 0.075 mg solid content/L, respectively.

Executive summary:

The chronic toxicity of the test item (= substance EC 939-455-3 in aqueous commercial product) to the freshwater fish species Pimephales promelas was investigated in a GLP-compliant study performed in accordance with OECD Guideline 210 (Fish, Early-Life Stage Toxicity Test). Lethal and sub-lethal effects were assessed and compared with control values to determine the various effect concentrations. The results led to the following conclusions:

- the 32d-NOEC and EC10 for hatching were 0.072 and 1.0 mg solid content/L, respectively;

- the 32d-NOEC and EC10 for post-hatch survival were 0.072 and 0.097 mg solid content/L, respectively;

- the 32d-NOEC and EC10 for larval growth (length) were 0.0082 and 0.12 mg solid content/L, respectively;

- the 32d-NOEC and EC10 for larval growth (weight) were <0.0082 and 0.075 mg solid content/L, respectively.

The reported effect concentrations are based on measured concentrations.

Reference 2

Endpoint:

fish early-life stage toxicity

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
C8-C18 AAPHS and C12-18 AAPHS have the same functional groups, and general composition. The main variable resides in the alkyl chain distribution.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Source chemical = C8-18 cocamidopropyl hydroxysultaine (EC 939-457-4).
Target chemical = C12-18 cocamidopropyl hydroxysultaine (EC 939-455-3).

3. ANALOGUE APPROACH JUSTIFICATION
Cf. attached Read-Across Justification Document (§13 Assessment reports).

4. DATA MATRIX
Cf. attached Read-Across Justification Document (§13 Assessment reports).

Reason / purpose for cross-reference:

read-across source

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

1 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: hatchability

Remarks on result:

other: Pimephales promelas (freshwater sp.)

Remarks:

read-across from EC 939-455-3

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

0.097 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: post-hatch survival

Remarks on result:

other: Pimephales promelas (freshwater sp.)

Remarks:

read-across from EC 939-455-3

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

0.12 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: larval growth (length)

Remarks on result:

other: Pimephales promelas (freshwater sp.)

Remarks:

read-across from EC 939-455-3

Key result

Duration:

32 d

Dose descriptor:

EC10

Effect conc.:

0.075 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

other: solid content

Basis for effect:

other: larval growth (weight)

Remarks on result:

other: Pimephales promelas (freshwater sp.)

Remarks:

read-across from EC 939-455-3

Conclusions:

By analogy with the source substance EC 939-455-3, a 32d-EC10 (reproduction) in the range 0.075-1 mg solid content/L on the freshwater fish species Pimephales promelas was reported for the target substance 1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-(C12-18(even numbered) acyl) derivs., hydroxides, inner salts (EC 939-457-4).

Description of key information

As the chronic toxicity to fish of the substance EC 939-457-4 has not been investigated experimentally, a read-across approach was followed to fill in the endpoint. The chronic toxicity to the freshwater fish species Pimephales promelas of a structural analogue substance (= source substance EC 939-455-3) was investigated in a GLP-compliant study (Bouwman, 2019) performed in accordance with OECD Guideline 210 (Fish, Early-Life Stage Toxicity Test). Lethal and sub-lethal effects were assessed and compared with control values to determine the various effect concentrations. The results led to the following conclusions for the substance:

- the 32d-NOEC and EC10 for hatching were 0.072 and 1.0 mg solid content/L, respectively;

- the 32d-NOEC and EC10 for post-hatch survival were 0.072 and 0.097 mg solid content/L, respectively;

- the 32d-NOEC and EC10 for larval growth (length) were 0.0082 and 0.12 mg solid content/L, respectively;

- the 32d-NOEC and EC10 for larval growth (weight) were <0.0082 and 0.075 mg solid content/L, respectively.

The reported effect concentrations are based on measured concentrations. By analogy, the same NOEC and EC10 values were reported for the target substance 1-Propanaminium, N-(3-aminopropyl)-2-hydroxy-N,N-dimethyl-3-sulfo-, N-(C12-18(even numbered) acyl) derivs., hydroxides, inner salts (EC 939-457-4).

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

0.075 mg/L

Additional information

The chronic toxicity to fish of the structural analogue substance (= source substance EC 939-455-3) was investigated in a GLP-compliant study (Bouwman, 2019) performed in accordance with standard methods, with minor deviations. The study is considered as reliable (Klimisch 1) and was selected as key study for the endpoint of the target substance EC 939-457-4.