Cetrimonium chloride

Toxicity to soil macroorganisms except arthropods

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Study period:

From October 13, 2003 to November 11, 2003

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

KL2 due to RA

Justification for type of information:

Refer to the Quaternary ammonium salts (QAS) category or section 13 of IUCLID for details on the category justification.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

yes

Details on preparation and application of test substrate:

Four groups of ten worms were treated with each concentration of test substance. Doses were applied as aqueous solutions and the same volume for all dose levels was used. The control was treated with water only. Worms were rinsed in dechlorinated water, blotted dry and individually weighed at the beginning of the study. At the end of the study they were cleaned in the same manner and weighed in groups of ten.

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

Test organisms:
- Common name: Earthworm (Eisenia foetida foetida)
- Source: Huntingdon Life Sciences
- Age at test initiation (mean and range, SD): More than 2 months
- Weight at test initiation (mean and range, SD): 300 - 600 mg

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

14 d

Test temperature:

21.1 - 21.7°C

pH:

5.86

Moisture:

Moisture content was 35% of the dry weight at the beginning and 32-34% at the end of the study.

Details on test conditions:

Test system:
- Test container (material, size): One litre glass container covered with perforated plastic to retain moisture.
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 4
- No. of replicates per control: 4

Source and properties of substrate (if soil):
- Artificial OECD 207 soil, Batch 13/03, (Industrial quartz sand, kaolin clay and sphagnum peat) was used for the main test (pH approximately 5.86)

Other test conditions:
- Photoperiod: continuous light
- Light intensity: 684 lux

Effect parameters measured: Mortality, behavioural and pathological signs

Vehicle control: yes

NB. On Days 7 and 14 the medium was emptied from the containers and worms were removed and counted. On Day 7, the medium was returned to the containers and surviving worms placed on the soil surface.

Nominal and measured concentrations:

Nominal concentrations: 0, 953, 1715, 3086, 5556 or 10000 mg a.i./kg dw of soil

Reference substance (positive control):

yes

Remarks:

Chloroacetamide

Key result

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

7 160 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 95% confidence limits 5560 - 7590 mg a.i./kg dw of soil

Key result

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

7 070 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

mortality

Remarks on result:

other: 95% confidence limits 5560 - 7400 mg a.i./kg dw of soil

Key result

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

953 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

other: body weight

Details on results:

- Mortality at end of exposure period: After 7d, all worms at 10000 and 2 worms at 5556 mg a.i./kg dw of soil were dead. By Day 14, one additional worm died at 5556 mg a.i./kg dw of soil.
- Changes in body weigth of live adults (% of initial weight) at end of exposure period: Group mean body weights were affected by treatment with test substance at 1,715 mg a.i./kg dw of soil and above.
- Morphological abnormalities: No
- Behavioural abnormalities: No
- Other biological observations: At 5556 mg a.i./kg soil dw several live worms were visible at the sides of the containers on Days 1 to 3 then again on Days 8 and 9. At the highest dose level, live worms were visible at the sides and on the surface of the soil in some containers during Days 1 to 6.

Results with reference substance (positive control):

The LC50 values of chloroacetamide to the earthworm, determined in a separate GLP compliant study, were 41 (95% confidence limits 37-43) and 39 (95% confidence limits 35-41) mg a.i./kg dw of soil at Day 7 and 14, respectively. The results obtained in the positive control test were considered acceptable.

Reported statistics and error estimates:

The LC50 values were calculated using a logistic model (Berkson, 1944). 95% confidence limits were estimated by the likelihood ration method (Williams, 1986).

Table 1. Mortality rate (%)

Group No.	Treatment	Initial	Mortality		Total worms
	(mg a.i./kg dw of soil	No. worms	Day 7	Day 14	as %
1	0	40	0	0	0
2	953	40	0	0	0
3	1715	40	0	0	0
4	3086	40	0	0	0
5	5556	40	2	3	7.5
6	10000	40	40	40	100

 

Table 2. Mean bodyweights (mg)

Group	Treatment (mg a.i./kg dw of soil	Day 0	Day 14	% decrease
1	0	388	334	14
2	953	389	340	13
3	1715	390	323	17
4	3086	386	304	21
5	5556	389	250	36
6	10000	392	n/d	n/d

 

n/d: no data (as all worms in this group had died)

For further details refere to the attachment under 'attached background materials'.

Validity criteria fulfilled:

yes

Conclusions:

Based on the results of the read across study, the 14d LC50 and NOEC values for earthworm (Eisenia foetida) were 7070 and 953 mg a.i./kg soil dw (nominal) respectively.

Executive summary:

A study was conducted to determine the toxicity to soil macroorganisms of the read across substance C12-16 ADBAC (49.5% active) according to OECD Guideline 207, in compliance with GLP. Six groups of forty earthworm (Eisenia foetida) were allocated to an artificial soil containing 0, 953, 1715, 3086, 5556 or 10000 mg a.i./kg soil dw (nominal concentrations). No analytical dose verification was performed. Mortality was recorded on Days 7 and 14. Worms were weighed at the beginning and end of the study. After 7 days, all worms at 10000 and 2 worms at 5556 mg a.i./kg soil dw were dead. By Day 14, one additional worm died at 5556 mg a.i./kg soil dw. A treatment-related reduction in body weight was observed. Group mean body weights were affected by treatment with read across substance at 1715 mg a.i./kg soil dw and above. Under the study conditions, the 7 and 14 d LC50 values were 7160 and 7070 mg a.i./kg soil dw, respectively and the NOEC was 953 mg a.i./kg soil dw (nominal) (Rodgers, 2004). Based on the results of the read across study, similar EC50 and NOEC values is expected for the test substance.

Reference 2

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Study period:

From December 02, 1998 to February 26, 1999

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Justification for type of information:

Refer to the Quaternary ammonium salts (QAS) category or section 13 of IUCLID for details on the category justification.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Deviations:

yes

Remarks:

analyses were not performed because the substance was readily biodegradable and has a halflife of 40d in soil. It can not be ascertained whether or not the test substance was stable during the test.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

- Species/strain: Eisenia fetida fetida
- Source of the initial stock: Aquatic Research Organisms INC, Hampton, USA
- Culturing techniques: They were kept in covered plastic vessels (volume 60 L) containing potting compost.
- Age/weight: The weight of the adult earthworms was 0.30 to 0.60 g.
- Pre-treatment: The worms had been acclimatised to artificial soil one d prior to the test.

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

14 d

Test temperature:

18-21°C

pH:

5.56 - 6.06

Moisture:

32.9-35.8%

Details on test conditions:

- Adjustment of pH: No
- Light intensity / photoperiod: Continuous light
- Relevant degradation products: Not determined

Details on test system:
- Artificial soil test substrate: 10% peat (air dried and finely ground), 20% kaolin (kaolinite content 84-87%), 70% quartz sand (sand with particle
size of 100-400 µm) and 0.5% calcium carbonate. PH was 5.73 and moisture content was 20.7%.
- Test mixture: A total weight of 750 g of test substrate was put in each vessel.
- Size, volume and material of test container: Glass dishes of 1.0 l.
- Amount of artificial soil (kg)/ container: 0.750 kg w wt/container
- Nominal levels of test concentrations: 100, 180, 320, 580 and 1000 mg/kg dry weight artificial soil, based product with 51.7% in water
- Active ingredient levels of test concentrations: 52, 93, 165, 300 and 517 mg/kg dry weight artificial soil
- Number of replicates/concentration: 4
- Number of earthworms/test concentration: 40
- Number of earthworms/container: 10
- Light source: 400 to 800 lux
- Test performed in closed vessels due to significant volatility of test substrate: Yes, with perforated plastic film, but not because of volatilisation
but to prevent the test substrate from drying.

The test method was static. The test substance was applied once at test start. The individual live weights of the worms were reported after 14d of exposure. Other effects (pathological symptoms, changes in behaviour of the worms) of the test substance were reported after 7 and 14d of exposure.

Nominal and measured concentrations:

Nominal: 100, 180, 320, 580 and 1000 mg/kg dw of soil

Reference substance (positive control):

yes

Remarks:

2-Chloracetamide (99.5%)

Key result

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: weight

Remarks on result:

other: corresponding to >517 mg a.i./kg soil dw

Key result

Duration:

14 d

Dose descriptor:

LC0

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: weight

Remarks on result:

other: corresponding to >517 mg a.i./kg soil dw

Details on results:

LC0 >1000 mg/kg dw
LC50 >1000 mg/kg dw
LC100 >1000 mg/kg dw
No significant mortality and effects were seen in any of the tested concentrations after 14d of exposure. There was no statistically significant difference at the test concentrations 100 - 1000 mglkg compared to the control group. The earthworms in the concentrations 100 to 1000 mg/kg dw showed normal behaviaur and no pathological symptoms. The highest tested concentration without mortality and any other effects was 1000 mg/kg dw.

Results with reference substance (positive control):

Results of the reference test with 2-Chloracetamide showed that the method is sensitive and valid.

For result tables, kindly refer to the attached background material section of the IUCLID.

Validity criteria fulfilled:

yes

Conclusions:

Under the study conditions, the 14d NOEC in earthworm (Eisenia foetida) was determined at 1000 mg/kg dw (or 517 mg/kg dw based on a.i.) (nominal).

Executive summary:

A study was conducted to determine the toxicity to soil macroorganisms of the read across substance, C12 -16 ADBAC (51.7% active) according to OECD Guideline 207, in compliance with GLP. Earthworms (Eisenia foetida) were exposed to a single dose of the read across substance at nominal concentrations of 100, 180, 320, 580 or 1,000 mg/kg dw of artificial soil. No analytical dose verification was performed. The individual live weights of the worms were reported after 14 d of exposure. Other effects (pathological symptoms, behaviour of the worms) were reported after 7 and 14 d of exposure. Results of the reference test with 2 -chloracetamide show that the method was sensitive and valid. The substance did not cause a change in behaviour, weight and mortality of the earthworm at any of the tested concentrations after 14 d of exposure. This was probably due to adsorption onto soil. The highest tested concentration without mortality and any other effects was 1000 mg/kg dw. Under the study conditions, the 14 d NOEC in earthworm was 1000 mg/kg dw (or 517 mg a.i./kg dw) and the 14 d LC0 was > 1000 mg/kg dw (or > 517 mg a.i./kg dw) (Noack, 1999). Based on the results of the read across study, similar NOEC value is expected for the test substance.

Description of key information

Based on the above information and in line with the biocides assessment report on the read across substance C12 -16 ADBAC, the 14 d LC50 of 7070 has been selected to express the acute toxicity of the test substance. Further, based on the chronic toxicity study with the read across substance, the 28-d NOEC of 125 mg/kg bw/day has been considered further for hazard/risk assessment.    

Key value for chemical safety assessment

Short-term EC50 or LC50 for soil macroorganisms:

7 070 mg/kg soil dw

Long-term EC10, LC10 or NOEC for soil macroorganisms:

125 mg/kg soil dw

Additional information

Short-term toxicity study:  

Study 1.  A study was conducted to determine the toxicity to soil macroorganisms of the read across substance C12-16 ADBAC (49.5% active) according to OECD Guideline 207, in compliance with GLP. Six groups of forty earthworms (Eisenia foetida) were allocated to an artificial soil containing 0, 953, 1715, 3086, 5556 or 10000 mg a.i./kg soil dw (nominal concentrations). No analytical dose verification was performed. Mortality was recorded on Days 7 and 14. Worms were weighed at the beginning and end of the study. After 7 days, all worms at 10000 and 2 worms at 5556 mg a.i./kg soil dw were dead. By Day 14, one additional worm died at 5556 mg a.i./kg soil dw. A treatment-related reduction in body weight was observed. Group mean body weights were affected by treatment with read across substance at 1715 mg a.i./kg soil dw and above. Under the study conditions, the 7 and 14 d LC50 values were 7160 and 7070 mg a.i./kg soil dw, respectively and the NOEC was 953 mg a.i./kg soil dw (nominal) (Rodgers, 2004).

Study 2. A study was conducted to determine the toxicity to soil macroorganisms of the read across substance, C12 -16 ADBAC (51.7% active) according to OECD Guideline 207, in compliance with GLP. Earthworms (Eisenia foetida) were exposed to a single dose of the read across substance at nominal concentrations of 100, 180, 320, 580 or 1,000 mg/kg dw of artificial soil. No analytical dose verification was performed. The individual live weights of the worms were reported after 14 d of exposure. Other effects (pathological symptoms, behaviour of the worms) were reported after 7 and 14 d of exposure. Results of the reference test with 2 -chloracetamide show that the method was sensitive and valid. The substance did not cause a change in behaviour, weight and mortality of the earthworm at any of the tested concentrations after 14 d of exposure. This was probably due to adsorption onto soil. The highest tested concentration without mortality and any other effects was 1000 mg/kg dw. Under the study conditions, the 14 d NOEC in earthworm was 1000 mg/kg dw (or 517 mg a.i./kg dw) and the 14 d LC0 was > 1000 mg/kg dw (or > 517 mg a.i./kg dw) (Noack, 1999).

 Based on the above two studies, the same effect levels were concluded in the biocide assessment report on C12-16 ADBAC by RMS Italy. They further stated that: “The findings of the two tests, although different in absolute values, are not in contrast. Since the second test provides a “higher than” value corresponding to a complete lack of lethal or sublethal effects, the 14d LC50 = 7070 mg/kg dry soil (US ISC) is selected to express the acute toxicity of Alkyl (C12-16) dimethylbenzyl ammonium chloride to soil dwelling invertebrates.” 

 

Long-term toxicity study:  

A study was conducted to determine the effects of read across substance (50% active in water) on mortality, biomass and the reproductive potential of the earthworm speciesEisenia fetida(Annelida, Lumbricidae), according to the OECD TG 222, in compliance with GLP. The study was conducted under static conditions over 8 weeks with the read across substance concentrations 125, 250, 500, 1000, 2000 mg//kg solid dry weight (SDW) corresponding to 62.5, 125, 250, 500, 1000 mg a.i./kg SDW. Each application rate was mixed into artificial soil containing 5% peat. A control including untreated artificial soil was tested under the same conditions as the read across substance treatments. A total of 80 test organisms were divided equally into 8 control replicates adnd another total of 40 test organisms were divided equally into 4 replicates for each read across substance treatment (i.e., 10 earthworms per replicate). They had an individual body weight between 0.36 and 0.55 g at the experimental starting. Each concentration level and control were analysed via LC-MS/MS analysis on Day 0, Day 28 and Day 55 using pooled samples of all replicates. The measured concentrations of the pooled samples of replicates were within the range of 83 to 101 % of the nominal values on Day 0, demonstrating the right preparation of the tested concentrations. After 28 days of exposure in soil, no read across substance-related earthworm mortalities (<10%), pathological symptoms or changes in the behaviour of adult earthworms were observed in the control or all read across substance concentrations. There were no statistically significant differences in earthworm body weights in all read across substance concentrations compared to the control. After an additional 4 weeks, the reproduction rate (average number of juveniles produced) was 83 juveniles in the control and ranged from 18 to 74 juveniles in the read across substance treatment rates. There were no statistically significant differences in earthworm reproduction in the treatment rates 125 and 250 mg read across substance/kg SDW compared to the control. However, at the read across substance concentrations, 500 to 2000 mg read across substance/kg SDW the earthworm reproduction was statistically significantly reduced. All validity criteria recommended by the test guidelines were fulfilled. Under the study conditions, the LOEC (mortality, biomass), NOEC (mortality, biomass), LOEC (reproduction), NOEC (reproduction) and EC50 (reproduction) values for read across substance were reported to be >2000, ≥2000, 500, 250 and 589 mg read across substance/kg SDW, respectively (equivalent to >1000, ≥1000, 250, 125 and 295 mg a.i./kg SDW, respectively). Based on the results of the read across study, similar effect levels can be considered for the test substance.

Therefore, based on the above information and in line with the biocides assessment report on the read across substance C12 -16 ADBAC, the 14 d LC50 of 7070 has been selected to express the acute toxicity of the test substance. Further, based on the chronic toxicity study with the read across substance, the 28-d NOEC of 125 mg/kg bw/day has been considered further for hazard/risk assessment.