Amides, C16-18, N-[3-(dimethylamino)propyl]

Sediment toxicity

Description of key information

Two endobenthic organisms. the worm Lumbriculus variegatus and the abundant nematode Caenorhabditis elegans, were used to determine a potential sediment toxicity of C20/22 ATQ. These sediment data are used for read-across to the registration substance SAPDMA.

Key value for chemical safety assessment

EC10, LC10 or NOEC for freshwater sediment:

62.5 mg/kg sediment dw

Additional information

No experimental toxicity tests with sediment dwelling organisms are available for C16-18 DMAPA amidoamine. However, studies with the read-across substance C20/22 ATQ are available. A justification for read-across is given below.

The effects of the test item C20/22 ATQ on the oligochaete Lumbriculus variegatus in a water-sediment system were determined according to OECD Guideline 225 (2007). Test duration was 28 days after insertion of the test organisms. The study was performed by spiking the test item into the sediment with five concentrations of 62.5 – 125 – 250 – 500 and 750 mg/kg sediment dry weight. Six replicates per control as well as solvent control and four replicates per test item concentration were set up for the biological investigation. Water quality parameters as temperature, pH-value, O₂-concentration, ammonium and total hardness were determined throughout the study.

The concentrations of C20/22 ATQ were analytically investigated at the test item concentrations of 250, 500 and 750 mg/kg sediment dry weight at day 0 (worm insertion), day 14 and day 28 in the sediment and in the overlying water via LC-MS/MS.

The measured concentrations of C20/22 ATQ (C20 fraction) in the sediment at the beginning of the test were in the range of 95 - 98 % of the applied concentrations. After 14 days, the recoveries of C20/22 ATQ (C20 fraction) in the sediment were at all levels between 91 – 93 % of the applied concentrations. At the end of the study the recoveries were in the range of 83 – 86 %.

The measured concentrations of C20/22 ATQ (C22 fraction) in the sediment at the beginning of the test were in the range of 79 - 86 % of the applied concentrations. After 14 days, the recoveries of C20/22 ATQ (C22 fraction) in the sediment were at all levels between 86 – 88 % of the applied concentrations. At the end of the study the recoveries were in the range of 71 – 79 %.

These recoveries demonstrate that the nominal range of the test item concentrations was met.

After 28 days of exposure C20/22ATQ induced a mortality of 15, 22.5 and 50 % at the test item concentrations of 250, 500 and 750 mg/kg sediment dry weight. The total number of worms and also the reproduction was statistically significantly reduced at 125, 250, 500 and 750 mg/kg sediment dry weight compared to the pooled control. The EC50 for reproduction was determined to be at 169 mg/kg sediment dry weight. Also, the biomass was statistically significantly reduced compared to the pooled control after 28 days of exposure to the test item at 125, 250, 500 and 750 mg/kg sediment dry weight.The EC50 for worm biomass was set at 221 mg/kg sediment dry weight. 

Concluded from the results, the Lowest Observed Effect Concentration (LOEC) was set at 125 mg/kg sediment dry weight. Thus, the No Observed Effect Concentration (NOEC) was determined to be at 62.5 mg/kg sediment dry weight

The effects of the test item C20/22 ATQ on the bacterivorous nematode Caenorhabditis elegans in a sediment system were determined according to the Draft guideline ISO/DIS 10872 (2008). Test duration was 96 hours after insertion of the test organisms. The study was performed by spiking the test item into the sediment with the test item concentrations of 62.5 - 125 - 250 - 500 - 1000 mg test item/kg sediment dry weight (nominal concentration). Four replicates per control, solvent control and each test item concentration were set up.

After four days of exposure C20/22 ATQ did not induce biologically significant mortality of C. elegans at all test item concentrations. Also, adult nematode fertility was not affected at all test item concentrations. However, the reproduction of nematodes was statistically significant reduced at 500 and 1000 mg/kg sediment dry weight. Also, C20/22 ATQ induced a statistically significant reduction of growth at 500 and 1000 mg/kg sediment dry weight.

Thus, the Lowest Observed Effect Concentration (LOEC) concerning all test parameters was set to at 500 mg/kg sediment dry weight. The Overall No Observed Effect Concentration (NOEC) was determined to be 250 mg/kg sediment dry weight. All effect levels given are based on the nominal concentrations of C20/22 ATQ.

1. Read-across hypothesis and justification

This read-across is based on the hypothesis that source and target substances have similar ecotoxicological properties because

·        they are manufactured from similar resp. identical precursors under similar conditions

·        the metabolism pathway leads to comparable products (amine backbone and long chain fatty acids) and non-common products predicted to have no toxicological effects (long chain fatty acids).

·        of their structural similarities: target and source substances are comprised of a hydrophobic (alkyl) and hydrophilic (positively charged ammonium) part; due to theses structural elements they form micelles and have surface active properties.

·        of their similar molecular weight, physicochemical properties and similar ecotoxicological profile in aquatic tests

Therefore, read-across from the existing ecotoxicological studies on the source substances is considered as an appropriate adaptation to the standard information requirements of Annex VII 9.1.1, 9.1.2, 9.2.1, Annex VIII 9.1.3, 9.1.4, Annex IX 9.1.5, 9.1.6, 9.2.1, and 9.4 of the REACH Regulation for the target substance, in accordance with the provisions of Annex XI, 1.5 of the REACH Regulation.

The justification of the proposed read-across approach is elaborated in the next chapters.

2. Justification for read-across

2.1 Substance Identity

Table 1: Substance identities

	Source substances		Target substance
	Stearic acid 3-(dimethylaminopropyl)amide	N,N,N-trimethyl-C20-22-(even numbered)-alkyl-1-aminium chloride; C20/22- alkyltrimethylammonium chloride (C20/22-ATQ)	C16-18 DMAPA amidoamine
Substance type	mono constituent substance	UVCB	UVCB
CAS number	7651-02-7	68607-24-9
EC number	231-609-1	271-756-9
Chain length distribution	< C16: < 1.6% C16: < 7% C18: > 89.8% > C18: < 1.6%	C16: <1% C18: ca. 4% C20: ca. 12% C22: ca. 82% C24: ca. 1%	C14: <= 5 % C16: 25-35 % C18: >= 61 %
DMAPA	<0.002%	N/A	<=0.01%

 

2.1 Substance Identity

Substance descriptions

The target substance C16-18 DMAPA amidoamine is a UVCB substance manufactured from saturated C16-18 fatty acids and N, N-dimethylpropylenediamine (DMAPA). It is composed of C16 and C18 amides of DMAPA, with C18 being the larger part (>/= 61%)

The source substance Stearic acid 3-(dimethylaminopropyl)amide is manufactured from octadecanoic acid and N, N-dimethylpropylenediamine. It is composed of mainly C18 amides (> 89.8%) of DMAPA and small amounts of the C16 amide (<7%).

The source substance C20/22-ATQ is manufactured from behenyl alcohol and dimethylamine, resulting in the corresponding tertiary amine dimethylalkylamine. In step two, the dimethylalkylamine is treated with methylchloride at elevated pressure to form the quaternary ammonium chloride.

2.2 Common breakdown products

The source substance Stearic acid 3-(dimethylaminopropyl)amide is the main component of the UVCB target substance C16-18 DMAPA amidoamine. The only difference is the chain length distribution: the target substance also contains a significant amount of the C16 amide.

This is not considered to be of relevance for metabolism.Both substances are amides which after resorption may be hydrolysed by amidases resulting in free fatty acids and DMAPA. The carboxylic acids then are further degraded by the mitochondrial beta-oxidation process (for details see common text books on biochemistry). The fatty acids enter normal metabolic pathways and are therefore indistinguishable from fatty acids from other sources including diet. The amine compounds are not expected to be further metabolised, but excreted via the urine mainly unchanged. 

As no ecotoxicological data are available for the target substance C16-18 DMAPA amidoamine, but only its main constituent Stearic acid 3 -(dimethylaminopropyl)amide, comparison of ecotoxicological data of the second source substance C20/22 ATQ is only possible to the latter. However, as explained above, the small amount of additional C16 is not expected to have any relevant influence on toxicity.

Both, target and source substances are cationic surfactants and are strongly sorbing to solids due to ionic interactions and van der Waals forces. Both substances are also readily and ultimately biodegradable in an OECD 301B CO2 Evolution test. In addition the microbial metabolic pathway is the same for both substances as in the first step the alkyl chain is cleaved from the nitrogen forming the corresponding aldehyde and ammonium compound. The aldehyde is then oxidised to the fatty acid which is subsequently degraded by beta oxidation (Kees van Ginkel, Handbook of Surfactants, Volume F, 1995).

2.3 Common structural elements

Target and source substances are comprised of a hydrophobic (alkyl) and hydrophilic (positively charged ammonium) part. Due to theses structural elements they form micelles and have surface active properties.

2.4 Differences

The slight differences in fatty acid chain length (higher percentage of C16 in the target substance vs. corresponding higher percentage C18 in the source substance) are not considered to be of relevance for ecotoxicity.

C16-18 DMAPA amidoamine as well as Stearic acid 3-(dimethylaminopropyl)amide are protonated to a large degree at environmentally relevant pH. Whereas C20/22 ATQ is a quaternary ammonium chloride. This difference is not considered to be of ecotoxicological relevance.

C16-18 DMAPA amidoamine as well as Stearic acid 3-(dimethylaminopropyl)amide on the one side contain DMAPA as amine-backbone; C20/22 ATQ on the other side is based on dimethylamine as amine-backbone. However, as it is shown in the aquatic toxicity tests with Stearic acid 3-(dimethylaminopropyl)amide and C20/22 ATQ, this has no big influence on ecotoxicity.

3. Physicochemical properties:

Table 2: Physicochemical properties

	Source substances		Target substance
Endpoints	Stearic acid 3-(dimethylaminopropyl) amide	C20/22-ATQ	C16-18 DMAPA amidoamine
Molecular weight [g/mol]	368.64	ca. 400	340.59 - 368.64
Physical state at 20°C / 1013 hPa	Solid (paste)	Solid	Solid (waxy)
Melting point	67.4°C	Decomposition at 220-240°C	41.8°C
Boiling point	412.3°C	Decomposition	320.5°C
Surface tension	37.86 mN/m at 0.22 g/L	47.0 mN/m at 0.01 g/L	26.7 mN/m at 0.027 g/L
Water solubility	10 mg/L at 20°C	10 mg/L at 25°C	3.65 mg/L at 23°C
Log Kow	2.01 at 20°C, pH7	3.29 at 20°C	2.01 at 20°C, pH7; read-across from Stearic acid 3-(dimethylaminopropyl) amide
Vapour pressure	3.4E-08 Pa at 20°C	7E-05 Pa at 20°C	3.4E-08 Pa at 20°C; read-across from Stearic acid 3-(dimethylaminopropyl) amide
Adsorption / desorption	no data (read-across fromC20/22-ATQ)	log Koc = 3 – 5.7 (batch equilibrium method)	log Koc = 7.8-8.0 at 25°C (HPLC method)

As demonstrated in the table above, the source substances have a similar physicochemical profile compared to the target substance. The substances are cationic surfactants and are strongly sorbing to solids due to ionic interactions and van der Waals forces.

4. Comparison of data from ecotoxicological endpoints

5.1 Ecotoxicity data of the target and source substances

Cationic surfactants sorb strongly to negatively charged surfaces like glass or biota. In order to avoid sorption to the glass of the test vessel and on the test organism well characterised river was used as aquatic medium to allow reliable test results.

Table 4: Ecotoxicological profiles for the source substances Stearic acid 3-(dimethylaminopropyl)amide and C20/22-ATQ, and the target substance C16-18 DMAPA amidoamine

	Source substances		Target substance
Endpoints	Stearic acid 3-(dimethylaminopropyl) amide	C20/22-ATQ	C16-18 DMAPA amidoamine
Short-term toxicity to fish	96 h LC50 >0.1 - <1 mg/L(nominal)	96 h LC50 = 3.5 mg/L (meas., geom.. mean)	No data; read-across
Long-term toxicity to fish	9 d NOEC(behaviour) = 0.1 mg/L (nominal)	9 d NOEC = 0.24 mg/L (meas., geom.. mean)	No data; read-across
Short-term toxicity to aquatic invertebrates	48 h EC50 = 381 µg/L (nominal)	48 h EC50 = 1.39 mg/L (nominal)	No data; read-across
Long-term toxicity to aquatic invertebrates	21 d NOEC(mortality) = 200 µg/L (nominal); 21 d EC10 (mortality) =200 µg/L (nominal)	21 d NOEC = 128 µg/L	No data; read-across
Toxicity to aquatic algae and cyanobacteria	72 h EC50 = 140 µg/L (nominal); 72 h EC10 = 71 µg/L (nominal)	72 h EC50 = 3.48 mg/L (meas., geom. mean); 72 h EC10 = 0.78 mg/L (meas., geom. mean)	No data; read-across
Toxicity to microorganisms	3 h EC50 >100 - < 1000 mg/L; 3 h NOEC = 100 mg/L	3 h EC50 >100 - < 1000 mg/L; 3 h NOEC = 100 mg/L	No data; read-across
Biodegradation in water; screening	readily biodegradable	readily biodegradable	readily biodegradable
Toxicity to soil macroorganisms except arthropods (earthworm)	No data	54 d NOEC (reproduction, mortality,body weight) = 250mg/kg soil dw   14 d NOEC(mortality, biomass, development) = 1000 mg/kg soil dw	No data; read-across
Toxicity to terrestrial arthropods (Collembola)	No data	28 d NOEC (mortality, reproduction) = 500 mg/kg soil dw;   28 d LD50 > 1000 mg/kg soil dw	No data; read-across
Toxicity to soil microorganisms	No data	28 d EC50 = 76 mg/kg soil dw; 28 d EC10 = 15 mg/kg soil dw; 28 d NOEC = 10 mg/kg soil dw	No data; read-across

 

No experimental ecotoxicity data are available for the target substance C16-18 DMAPA amidoamine. However, as demonstrated above, Stearic acid 3-(dimethylaminopropyl)amide is the main constituent of the target substance. Thus, aquatic toxicity data of the source substance Stearic acid 3-(dimethylaminopropyl)amide are considered to be relevant also for the target substance C16-18 DMAPA amidoamine.

Terrestrial ecotoxicity data are available for the source substance C20/22-ATQ. Based on the close similarity of Stearic acid 3-(dimethylaminopropyl)amide and the target substance C16-18 DMAPA amidoamine, also the read-across from C20/22-ATQ is justified:

The substances are readily and ultimately biodegradable. In addition the microbial metabolic pathway is the same for both substances as in the first step the alkyl chain is cleaved from the nitrogen forming the corresponding aldehyde and ammonium compound. The aldehyde is then oxidised to the fatty acid which is subsequently degraded by beta oxidation (Kees van Ginkel, Handbook of Surfactants, Volume F, 1995).

In the following table the river water test results for Stearic acid 3-(dimethylaminopropyl)amide and C20/22 ATQ are given.

		Stearic acid 3-(dimethylaminopropyl) amide	C20/22 ATQ	Ratio highest:lowest value
Chronic fish OECD 212 River water test	NOEC repro (9d)	0.1 mg/L	0.24 mg/L	2.4
Chronic daphnia OECD 211River water test	NOEC repro (21d)	0.2 mg/L	0.13 mg/L	1.5
Algae OECD 201River water	ErC10 (72h)	0.07 mg/L	0.93 mg/L	13.2
Algae OECD 201Reconstituted water	ErC10 (72h)	0.2 mg/L	0.93 mg/L(RW)	4.7

 

The ecotoxicity ratios for Chronic fish and Chronic daphnia is 2.4 and 1.5 respectively. These ratios are well within the boundaries on variability / uncertainty accepted e.g. for reference substances. The ratio for ErC10 (72h) for Stearic acid 3-(dimethylaminopropyl)amide and C20/22 ATQ is 12.9 and much higher than expected. But from the tertiary alkyl dimethylamines (see above) it is known that the algae ecotoxicity in river water is sometimes higher than in reconstituted water. When comparing the Stearic acid 3-(dimethylaminopropyl)amide algae ErC10 (72h) in reconstituted water with the algae value for C20/22 ATQ river water the ratio for the endpoint drops from 12.9 to 4.5. The value 4.5 is well within the range for variability / uncertainty acceptable for algae test (3 standard deviations). The PNEC freshwater and marine for Stearic acid 3-(dimethylaminopropyl) amide is derived from the ErC10 (72h) river water algae test of 0.07 mg/L whereas the PNEC freshwater and marine for C20/22 ATQ is derived from the NOEC repro (21d) daphnia of 0.13 mg/L. The ratio between these two NOEC is 0.13/0.07=1.9 and well acceptable with respect to uncertainty / variability of test results.

Based on these data as well as on similar physicochemical properties and on the similar toxicological profile the read-across approach for long-term toxicity as well as terrestrial toxicity is considered to be appropriate. The available data are comparable for source and target substance, supporting the validity of the grouping approach.

5.3 Quality of the experimental data of the analogues:

Aquatic toxicity:

The source substance Stearic acid 3-(dimethylaminopropyl)amide has been tested in reliable (RL1) GLP-compliant studies according to OECD TG 201 and 202, and a reliable (RL2) study according to OECD TG 203 for short-term toxicity as well as reliable (RL1) GLP-compliant studies according to OECD TG 211 and 212 for long-term toxicity.

The source substance C20/22 ATQ has been tested in reliable (RL1) GLP-compliant studies according to OECD TG 201, 202 and 203 for short-term toxicity as well as reliable (RL1) GLP-compliant studies according to OECD TG 211 and 212 for long-term toxicity.

Sediment toxicity:

The source substance C20/22 ATQ has been tested in reliable (RL1) GLP-compliant studies according to OECD TG 225 and Draft ISO/DIS 10872.

The available data from the source chemical are sufficiently reliable to justify the read-across approach.

5.4 Classification and labelling

Concerning environmental effects, the source substance Stearic acid 3-(dimethylaminopropyl)amide is classified as Aquatic Acute 1 (M-factor = 1) and Aquatic Chronic 2. The source substance C20/22 ATQ is classified as Aquatic Acute 1 (M-factor = 1).

Based on read-across, the target substance C16-18 DMAPA amidoamine is classified as Aquatic Acute 1 (M-factor = 1) and Aquatic Chronic 2.

The substances are neither PBT nor vP/vB substances.

6. Conclusion

The structural and physicochemical similarities between the source and the target substances and the similarities in their breakdown products presented above support the read-across hypothesis. Adequate and reliable scientific information indicates that the source and target substances and their subsequent degradation products have similar toxicity profiles.

As demonstrated, Stearic acid 3-(dimethylaminopropyl)amide is the main constituent of the target substance C16-18 DMAPA amidoamine.

Based on close the relationship the results from aquatic, sediment and terrestrial toxicity data obtained with the source substances Stearic acid 3-(dimethylaminopropyl)amide and C20/22 ATQ are also relevant for the target substance C16-18 DMAPA amidoamine.