Quaternary ammonium compounds, C12-14-alkyltrimethyl, Me sulfates

Endpoint summary

Description of key information

Based on the results of the 90-day and 28 -day studies with the read across substance, Coco TMAC and C16 -TMAC in rats respectively, the NOAEL of 40.3 mg a.i./kg bw/day has been considered further for the hazard assessment of the test substance, Quaternary ammonium compounds, C12-14 alkyltrimethyl, Me sulfates. Overall, considering that the available studies revealed NOAELs based on reduction in body weight and body weight gain (consistent with decreased food consumption), it can be concluded that all effects could be attributed to local gastrointestinal irritaton/corrosion and consequent reduced food intake without observing any primary systemic effect. Therefore, the derivation of a DNEL for systemic effects was deemed inappropriate.

With regard to the dermal route, due to deficiencies in the available 28 -day dermal toxicity study with read across substance, C16 TMAC (reduced number of test animals per group and limited histopathology), the 90-day oral study with read across substance can be considered for the systemic hazard assessment of the test substance, Quaternary ammonium compounds, C12-14 alkyltrimethyl, Me sulfates.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Reference 1

Endpoint:

sub-chronic toxicity: oral

Type of information:

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

Adequacy of study:

key study

Study period:

From 29 October, 2001 to 18 June, 2002

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

KL2 due to RA study

Justification for type of information:

Refer to section 13 for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)

Deviations:

no

GLP compliance:

yes

Species:

rat

Strain:

other: Sprague-Dawley, Crl:CD® (SD) IGS BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Charles River (UK) limited, Margate, Kent
- Age at study initiation: 6 wk
- Weight at study initiation: Males: 141-183 g, females: 132-161 g
- Acclimation period: 14 d

ENVIRONMENTAL CONDITIONS
- Temperature: 21±2 °C
- Humidity: 55±15%
- Photoperiod (hrs dark / hrs light): 12 h/12 h

Route of administration:

oral: feed

Vehicle:

other: mixed with diet

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Mean dietary admixture concentrations were within acceptable limits for the purpose of the study (Gas chromatography).

Duration of treatment / exposure:

90 days

Frequency of treatment:

Daily in feed

Remarks:

Doses / Concentrations:
0, 100, 500 and 2,000 ppm (i.e., equivalent to 22, 113 and 273 mg/kg bw/day after correction for 35.5% purity); the highest dose of 2,000 ppm was reduced to 1,000 ppm from Day 29 onwards due to deterioration in health of treated animals at 2,000 ppm.
Basis:
nominal in diet

No. of animals per sex per dose:

10 males and 10 females per dose

Control animals:

yes, plain diet

Details on study design:

- Post-exposure recovery period in satellite groups: None

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once daily

BODY WEIGHT: Yes
- Time schedule for examinations: Day 0 (the day before start of treatment) and weekly thereafter

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Weekly throughout the study

FOOD EFFICIENCY: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: daily for each cage group

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Before start of treatment and before termination of treatment (during week 12)
- Dose groups that were examined: All

HAEMATOLOGY: Yes
- Time schedule for collection of blood: End of treatment (Day 90)

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: End of treatment (Day 90)

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Day 0 (the day before start of treatment) and weekly thereafter
- Dose groups that were examined: All
- Battery of functions tested: Sensory activity

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, all animals
HISTOPATHOLOGY: Yes, Macroscopic lesions, Adrenals, Aorta, Bone and bone marrow (femur including stifle joint), Bone and bone marrow (sternum), Brain (including cerebrum, cerebellum and pons), Cecum, Colon, Duodenum, Epididymides, Eyes, Gross lesions, Heart, ileum, Jejunum, Kidneys, Liver, Lungs (with bronchi), Lymph nodes (cervical and mesenteric), Mammary gland, Muscle (skeletal), Oesophagus, Ovaries, Pancreas, Pituitary gland, Prostate, Rectum, Salivary glands (submaxillary), Sciatic nerve, Seminal vesicles, Skin (hind limb), Spinal cord (cervical), Spleen, Stomach, Testes, Thymus, Thyroid/parathyroid, Tongue, Trachea, Urinary bladder, Uterus.

Statistics:

Data were processed to give group mean values and standard deviation where appropriate. Haematological, blood chemical, organ weight, weekly body weight gain and quantitative functional performance and sensory reactivity data were assessed for control and test substance treatment groups for dose response relationship by linear regression analysis, followed by one way analysis of variance (ANOVA) incorporating Levene's test for homogeneity of variance. Where variances were shown to be homogenous, pairwise comparisons were conducted using Dunnett's test. Where Levene's test showed unequal variances, the data were analysed using non-parametric methods: Kruskal-Wallis ANOVA and Mann-Whitney 'U' test.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Food efficiency:

effects observed, treatment-related

Water consumption and compound intake (if drinking water study):

no effects observed

Ophthalmological findings:

no effects observed

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Behaviour (functional findings):

effects observed, treatment-related

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Details on results:

CLINICAL SIGNS AND MORTALITY: No mortality occurred during the study period.
Clinical signs - High dose animals developed clinical signs of toxicity from Day 7 onwards. These included hunched posture, pilo-erection, tiptoe gait, diarrhoea and red/brown staining of external body surface. Due to these effects, the dose level was reduced to 1000 ppm from Day 29 onwards. Clinical signs persisted following the reduction in dose level and included two incidents of pallor of extremities together with generalised fur loss. No clinically observable signs of toxicity were detected at the mid and the high doses.

Behavioural assessment: Detailed open-field observations conducted during the study confirmed the clinical signs of hunched posture, pilo-erection and tiptoe gait detected in high dose animals. No such effects were detected at the mid or low dose levels.

Functional performance test: No treatment-related changes were detected.

Sensory Reactivity Assessments: High dose females showed an increase in startle reflex parameters compared with controls. No such effects were detected for high dose males or for animals of either sex treated with the lower doses.

BODY WEIGHT AND WEIGHT GAIN: Reduced body weight gain was detected for high dose animals of either sex during the first five weeks of the study compared with controls. Mid dose males were similarly affected but this was confined to week 1 and 2 only. Body weight gain recovered following reduction in the dose level and was comparable with controls thereafter but terminal bodyweights for high dose animals and mid dose males remained lower than controls. No adverse effect on bodyweight gain was detected for 500 ppm females or for animals of either sex treated with the low dose.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Reduced food intake was observed in the high and mid dose animals throughout the study period compared with controls.

FOOD EFFICIENCY: Food efficiency was reduced over the first three weeks of the study but this was confined to the high dose group only. No adverse effect on dietary intake or food efficiency was detected at the low dose.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No appreciable intergroup differences were detected. High dose animals showed a reduced water intake on Day 6 of the study which recovered thereafter.

OPHTHALMOSCOPIC EXAMINATION: No treatment-related effects.

HAEMATOLOGY: No treatment-related effects.

CLINICAL CHEMISTRY: No treatment-related effects.

ORGAN WEIGHTS: No toxicologically important organ weight changes were detected. The reductions in absolute weight (including heart, kidneys, liver and thymus weight at the high dose and heart weight at the mid dose) and increases in relative weight (including high dose brain epididymides, kidneys, spleen, testes and ovaries weight), were all considered to be a result of reduced bodyweight development rather than test substance toxicity.

GROSS PATHOLOGY: No treatment-related macroscopic abnormalities.

HISTOPATHOLOGY: NON-NEOPLASTIC: Treatment-related changes were observed in the spleen and kidneys. Lower severities of pigment accumulation were observed in the spleen of high dose male rats but not for the females (p <0.05). A higher incidence of pigment accumulation was observed in the kidneys of the high and mid dose male rats. In both tissues the pigment reacted positively to Perl's staining technique and was considered to be haemosiderin.
Haemosiderin is normally observed in kidneys of aged rats, where it accumulates in tubular epithelium. There are no indications of bleeding (black faeces, increase spleen weight, accelerated erythropoiesis).

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 40 mg/kg bw/day (actual dose received)

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: Clinical signs of toxicity, reduced body weight gain, reduced food efficiency and occurence of haemosiderine in kidneys of high dose animals.

Remarks on result:

other: reduction of body weight and food consumption can be considered secondary compared to the corrosive properties of the test substance; Haemosiderin is normally observed in kidneys of aged rats, where it accumulates in tubular epithelium.

Key result

Critical effects observed:

no

The high dose showed clear effects related to palatability (esp. low food intake first week, clear increase after lowering dose level, lower BW gain), and clinical signs of discomfort starting after about 2 weeks leading to hunched posture of all animals, and red/brown staining of fur and piloerection in most day. This improved a little with lowering of dose. No specific toxicity was observed in haematology and clinical chemistry, terminal necropsy (besides stained fur in half of the animals and irritation stomach in one female), and organ weight (high dose are affected in a pattern consistent with lower BW, not considered indicative for organ toxicity). Histopathology only reported possible treatment related effects of haemosiderin accumulation in spleen and kidney. Generally, increased deposition of haemosiderin is followed from haemolytic effects.

Typically in cases of haemolytic anaemia higher grades of severity for both splenic extramedullary haemopoiesis and for splenic haemosiderin accumulation would be expected and this is often associated with accumulations of haemosiderin in the renal tubules. However, haemosiderin does not usually appear in excess in the renal tubules until a threshold increase in the spleen has been exceeded.

In this case, there was no evidence of increased haemolysis from the blood haematology parameters, and also no indication for an increased extramedullary haemopoiesis. Additionally, there was no Kupffer cell hemosiderin pigmentation of the liver. Haemosiderin is normally observed in kidneys of aged rats, where it accumulates in tubular epithelium. However, there are no indications of bleeding (black faeces, increase spleen weight, accelerated erythropoiesis). Other study parameters essentially did not show treatment-related effects. Also, as far as it is known, for haemosiderin deposition no significant difference is generally seen between males and females.

So in this case, there was no indication of extramedullary haemopoiesis, no effects in blood haematology, and if anything, the haemosiderin deposition seemed to decrease with dose in both males and females. In view that all reported levels are only minimal or incidental slight, levels in the males remain below levels in female, and normally there is no difference between male and females, it can be concluded that the reported statistical difference between control and dosed groups in the males are of no toxicological relevance. Therefore, in line with the study authors the NOAEL can be established at 500 ppm (i.e., equivalent to 40 mg a.i./kg bw/day).

Conclusions:

Based on the results of the read across study, a similar NOAEL of 40.3 mg/kg bw/day can be considered for the test substance, Quaternary ammonium compounds, C12-14 alkyltrimethyl, Me sulfates.

Executive summary:

A 90-day oral repeat dose study was conducted to determine the oral repeated dose toxicity of the read across substance, Coco TMAC (active ingredient: 35.5%), according to OECD Guideline 408 and EU Method B.26, in compliance with GLP. Sprague-Dawley rats were administered the test substance at dose levels of 0, 100, 500 or 2000 ppm (corresponding to 0, 22, 113 and 273 mg/kg bw/day in males and 0, 25, 121, 297 mg/kg bw/day in females) through the diet for 90 d. The active ingredient equivalence of the doses were calculated to be 0, 7.9, 40.3 and 96.9 mg a.i./kg bw/day in males and 8.8, 42.9, 105.3 mg a.i./kg bw/day in females. The highest dose of 2000 ppm was reduced to 1000 ppm from Day 29 onwards due to deterioration of health of the test animals. At the highest dose, the treatment-related findings were clinical signs of toxicity, reduced body weight gain and food efficiency, organ weight changes and microscopic changes in the spleen and kidneys. At the mid dose, reduced body weight gain (males) and reduced food consumption, reduced absolute heart weight and higher incidence of haemosiderin accumulation in the kidneys of males was observed. No treatment-related effects were observed at the lowest dose.Based on the results of the study, dietary administration of the test substance to rats for a period of 90 d at levels up to 273 mg/kg bw/day resulted in toxicologically significant effects at the high dose and marginal effects at the next lower dose of 113 mg/kg bw/day (500 ppm). No such effects were demonstrated at the lowest dose of 22 mg/kg bw/day (100 ppm).The changes observed at the mid dose (500 ppm) were considered to be minor, isolated effects associated with the reduced palatability of the test substance and were considered not to represent an adverse health effect.Therefore, based on effects on body weight, food efficiency and clinical signs the study authors established the NOAEL at the mid dose level of 500 ppm (equivalent to 40.3 mg ai./kg bw/day) (Jones, 2002).

Reference 2

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

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

Adequacy of study:

supporting study

Study period:

1991

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

Refer to section 13 for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.

Reason / purpose for cross-reference:

read-across source

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Version / remarks:

see 'Principles of method if other than guideline'

Deviations:

no

Principles of method if other than guideline:

Groups of 10 male and 10 female rats received test substance by oral gavage at the dose levels of 0, 30, 100 and 300 mg/kg bw/day for 28d. In addition, 5 male and 5 female animals were included in control and high dose group (recovery group). Animals were observed for clinical signs of toxicity, body weight changes, food and water consumption, haematological and biochemical parameters. Animals were necropsied and analysed for any visible abnormalities. Several tissues/organs were subjected to histopathological investigations in control and high dose group. Ophthalmological examination was also performed.

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Route of administration:

oral: gavage

Vehicle:

water

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

28 days

Frequency of treatment:

Once daily

Remarks:

Doses / Concentrations:
0, 30, 100 and 300 mg/kg bw/day
Basis: other: The test substance used was a 24-26% aqueous solution but it is not clear if the doses mentioned are corrected for undiluted test substance.

No. of animals per sex per dose:

10 per sex per dose in main groups; 5 per sex in recovery groups (vehicle and highest dose group)

Control animals:

yes, concurrent vehicle

Details on study design:

- Rationale for selecting satellite groups: To check the reversibility of the effects, if any
- Post-exposure recovery period in satellite groups: at least 27d

Observations and examinations performed and frequency:

Animals were observed for clinical signs of toxicity. Body weight, food and water consumption were recorded. Haematological and biochemical parameters were analysed. All animals were necropsied and checked for macroscopically visible abnormalities. Several tissues/organs were preserved and processed for histopathological investigations in control and high dose group. Additionally, eyes were examined with slit lamp microscope for any treatment-related abnormality.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Other examinations:

Animals in the recovery groups were observed for at least 27d after termination of the treatment.

Statistics:

Yes, no detail available in the summary.

Clinical signs:

no effects observed

Description (incidence and severity):

Symptoms of local irritation were observed in the high dose group during the last week of treatment.

Mortality:

no mortality observed

Description (incidence):

No deaths occurred throughout the study.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Total body weight gain was comparable to control in test groups.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

The mean food consumption in all treated groups was comparable to the control.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

Water consumption was significantly increased in all animals of the high dose group.

Ophthalmological findings:

no effects observed

Description (incidence and severity):

The examination of the eyes by slit lamp microscope showed no treatment-related effects.

Haematological findings:

no effects observed

Description (incidence and severity):

The haematological examinations revealed no substance related variation from the control values.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

The highly significant increase in ALT-activity in the high dose group. The observed increase of the ALT-activity is considered to be an isolated finding as no other parameter is correlated with this deviation. Therefore, this finding is interpreted as incidental.

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

The absolute and relative organ weights such as spleen and adrenals showed some substance related changes in the males of high dose group.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

The macroscopical examination of the organs displayed substance related effects such as oedema of the mucosa of the forestomach, thickening of the mucosa and indication of ulceration in the animals of high dose group. Some additional observations like hydronephrosis, hydrometra and discolouration of the thymus showed no dose dependence and were therefore considered to be spontaneous. The observed irritative effects at the mucosa of the forestomach have been disappeared in the male and female animals of the recovery group 27d after termination of the treatment.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

The forestomach of the male and female animals of the high dose group showed effects indicating local irritation like inflammatory oedema of the submucosa, sporadic ulceration and acanthosis of the mucosa up to papillomatous hyperplasia. These observations were considered to be due to the irritating properties of test substance and were considered not to be symptoms of a systemic toxicity. The animals of the recovery group of high dose showed a complete and regular regeneration of the forestomach mucosa.

Histopathological findings: neoplastic:

not examined

Key result

Dose descriptor:

NOAEL

Effect level:

300 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

the test substance used was a 24-26% aqueous solution but it is not clear if the doses mentioned were corrected for undiluted test substance.

Sex:

male/female

Basis for effect level:

other: overall effects: no dose-dependent toxicologically significant effects up to the highest dose tested.

Key result

Critical effects observed:

no

Lowest effective dose / conc.:

300 mg/kg bw/day (nominal)

Conclusions:

Based on the results of the read across study, the rat 28d NOAEL for systemic effects was determined to be 300 mg/kg bw/day.

Executive summary:

A study was conducted to determine the oral repeated dose toxicity of the read across substance, C16 TMAC, according to a method similar to OECD Guideline 407, in compliance with GLP. Groups of 10 male and female rats were administered 0, 30, 100 and 300 mg/kg bw/day of the test substance by oral gavage for 28 d. The test substance used was a 24-26% aqueous solution but it is not clear if the doses mentioned were corrected. There were no treatment-related changes at the 30 and 100 mg/kg bw/day. In the high dose there was an increase in water consumption, changes in the absolute and relative weights of the adrenals and spleens (in males) without corresponding changes in haematology, clinical chemistry and lesions in histopathology. The forestomach of the high-dose group showed few microscopic changes, however animals in the high-dose recovery group showed a complete and regular regeneration of the forestomach mucosa. Hence, the forestomach effects were considered to be due to the irritating properties of the test substance rather than symptoms of systemic toxicity. Based on the results of the read across study, the 28 d NOAEL was determined to be 300 mg/kg bw/day (Potokar, 1991).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

40.3 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

The information requirements for this tonnage band is sufficiently met with the available data.

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: inhalation

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Reason / purpose for cross-reference:

data waiving: supporting information

Reason / purpose for cross-reference:

data waiving: supporting information

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: inhalation

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Reason / purpose for cross-reference:

data waiving: supporting information

Reason / purpose for cross-reference:

data waiving: supporting information

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Reference

Endpoint:

short-term repeated dose toxicity: dermal

Type of information:

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

Adequacy of study:

key study

Study period:

1979

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

Refer to section 13 for details on the read-across justification. The study with the read across substance is considered sufficient to fulfil the information requirements as further explained in the provided endpoint summary.

Reason / purpose for cross-reference:

read-across source

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 410 (Repeated Dose Dermal Toxicity: 21/28-Day Study)

GLP compliance:

not specified

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Sex:

male/female

Type of coverage:

open

Vehicle:

water

Details on exposure:

TEST SITE
- Area of exposure:
- % coverage: 25% of the body surface area.
- Time intervals for shavings or clipplings: all rabbits was abraded with a clipper head prior to the start of each application

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Following the exposure period, the treated skin surface was cleaned with water

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0 or 10 mg/kg bw/day
- Concentration (if solution): 0 or 0.5% aqueous solutions, respectively. The dosage volume was 2.0 mL/kg bw

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes, The animals were restrained with collars during the exposure period

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

6.5 to 7 hours

Frequency of treatment:

5 days/week for 4 wks

Remarks:

Doses / Concentrations: 0 or 10 mg/kg/day
Basis: no data

No. of animals per sex per dose:

5 New Zealand albino rabbits/sex/group

Control animals:

yes, concurrent vehicle

Details on study design:

- Duration of observation period following administration: All rabbits were examined daily for clinical signs and mortality. Dermal irritation readings were recorded daily. The animals were weighed weekly during the exposure period. Blood was collected for haematology measurements before initiation of dosing and prior to termination. Liver and kidneys were weighed at necropsy. A complete list of tissues
was collected for histopathological evaluation
- Frequency of observations and weighing: weekly
- Necropsy of survivors performed: yes

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes / No / No data
- Time schedule: twice daily

DERMAL IRRITATION (if dermal study): Yes
- Time schedule for examinations: Daily

BODY WEIGHT: Yes
- Time schedule for examinations: Weekly

HAEMATOLOGY: Yes
- Time schedule for collection of blood: Blood was collected for haematology measurements before initiation of dosing and prior to termination

OTHER:
Mortality : twice daily

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes Liver and kidneys were weighed at necropsy. A complete list of tissues was collected for histopathological evaluation

Clinical signs:

no effects observed

Dermal irritation:

effects observed, treatment-related

Description (incidence and severity):

Treated areas of the skin that showed mild to marked acanthosis with active mitosis, hyperkeratosis, and partial to extensive necrosis of the epidermis and hair follicles, partly with encrustation and exudate

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not examined

Details on results:

- Two control group animals died during the study.
- Slight to moderate erythema was observed in all treated rabbits between days 4 and 8, but disappeared in 4 rabbits by day 17. Very slight to slight oedema was observed between days 6 and 12 in 4 rabbits and subsided by day 17. Two rabbits had intermittent slight oedema during week 4, and one rabbit developed oedema on day 20. No evidence of desquamation or leather-like skin was present in these animals. In the other rabbits, slight atonia occurred up to week 4 in 3 animals. Slight skin fissuring was observed in most of the rabbits but typically disappeared by the end of the study. There were no treatment-related effects on body weight, haematology, organ weight, gross necropsy findings or histopathology, except for treated areas of the skin that showed mild to marked acanthosis with active mitosis, hyperkeratosis, and partial to extensive necrosis of the epidermis and hair follicles, partly with encrustation and exudate.

Key result

Dose descriptor:

NOAEL

Effect level:

10 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

dermal irritation

other: see 'Remark'

Key result

Critical effects observed:

no

Conclusions:

Based on the results of the read across study, the NOAEL for test substance, Quaternary ammonium compounds, C12-14 alkyltrimethyl, Me sulfates, can be considered to be 10 mg/kg bw/day.

Executive summary:

A 28-day repeat dose study was conducted to determine the repeated dose dermal toxicity of the read across substance, C16 TMAC, in New Zealand albino rabbits (both sexes)according to a method similar to OECD Guideline 410. The purity was not specified and the study included a lower than recommended number of animals (10/group rather than 20/group as per the guideline) and histopathology was performed only on limited organs. The test substance (at 0 and 10 mg/kg bw/day) was applied to the shaved, intact skin of groups of five New Zealand albino rabbits/sex/group up to7 h, 5 days/week for 4 weeks. Dermal irritation effects were recorded daily. The animals were weighed weekly during the exposure period. Blood was collected for haematology measurements before initiation of dosing and prior to termination. Liver and kidneys weights were recorded at necropsy and limited histopathology was conducted. There were no systemic treatment-related effects on body weights, haematology, organ weights, gross necropsy findings or histopathology. Treated areas of the skin showed mild to marked acanthosis with active mitosis, hyperkeratosis, and partial to extensive necrosis of the epidermis and hair follicles, partly with encrustation and exudate.Under the study conditions, the 28 d NOAEL for male and female rabbits was found to be 10 mg/kg bw/day (Spicer, 1979).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

10 mg/kg bw/day

Study duration:

subacute

Species:

rabbit

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Oral

Study 1:

A 90-day oral repeat dose study was conducted to determine the oral repeated dose toxicity of the read across substance, Coco TMAC (active ingredient: 35.5%), according to OECD Guideline 408 and EU Method B.26, in compliance with GLP. Sprague-Dawley rats were administered the test substance at dose levels of 0, 100, 500 or 2000 ppm (corresponding to 0, 22, 113 and 273 mg/kg bw/day in males and 0, 25, 121, 297 mg/kg bw/day in females) through the diet for 90 d. The active ingredient equivalence of the doses were calculated to be 0, 7.9, 40.3 and 96.9 mg a.i./kg bw/day in males and 8.8, 42.9, 105.3 mg a.i./kg bw/day in females. The highest dose of 2000 ppm was reduced to 1000 ppm from Day 29 onwards due to deterioration of health of the test animals. At the highest dose, the treatment-related findings were clinical signs of toxicity, reduced body weight gain and food efficiency, organ weight changes and microscopic changes in the spleen and kidneys. At the mid dose, reduced body weight gain (males) and reduced food consumption, reduced absolute heart weight and higher incidence of haemosiderin accumulation in the kidneys of males was observed. No treatment-related effects were observed at the lowest dose. Based on the results of the study, dietary administration of the test substance to rats for a period of 90 d at levels up to 273 mg/kg bw/day resulted in toxicologically significant effects at the high dose and marginal effects at the next lower dose of 113 mg/kg bw/day (500 ppm). No such effects were demonstrated at the lowest dose of 22 mg/kg bw/day (100 ppm). The changes observed at the mid dose (500 ppm) were considered to be minor, isolated effects associated with the reduced palatability of the test substance and were considered not to represent an adverse health effect. Therefore, based on effects on body weight, food efficiency and clinical signs the study authors established the NOAEL at the mid dose level of 500 ppm (equivalent to 40.3 mg ai./kg bw/day) (Jones, 2002).

Study 2:

A study was conducted to determine the oral repeated dose toxicity of the read across substance, C16 TMAC, according to a method similar to OECD Guideline 407, in compliance with GLP. Groups of 10 male and female rats were administered 0, 30, 100 and 300 mg/kg bw/day of the test substance by oral gavage for 28 d. The test substance used was a 24-26% aqueous solution but it is not clear if the doses mentioned were corrected. There were no treatment-related changes at the 30 and 100 mg/kg bw/day. In the high dose there was an increase in water consumption, changes in the absolute and relative weights of the adrenals and spleens (in males) without corresponding changes in haematology, clinical chemistry and lesions in histopathology. The forestomach of the high-dose group showed few microscopic changes, however animals in the high-dose recovery group showed a complete and regular regeneration of the forestomach mucosa. Hence, the forestomach effects were considered to be due to the irritating properties of the test substance rather than symptoms of systemic toxicity. Based on the results of the read across study, the 28 d NOAEL was determined to be 300 mg/kg bw/day (Potokar, 1991).

 

As per the Biocides assessment report on Coco TMAC, which was published by the Italian authorities in April 2016, similar results were obtained from subchronic feeding study with the read across substance, DDAC in rats where the critical effect upon oral exposure was identified as decreased body weight gain, leading to a NOAEL of 42 mg/kg bw/day. Further, results from repeated dose stduies in dogs with DDAC do not indicate species differernce towards this mechanism of toxicity by quaternary ammonium compounds. The 90-day dog study with DDAC resulted in a NOAEL of 15 mg ai./kg bw/day, the highest tested dosage tested (ECHA biocides assessment report, 2016).

Overall, considering the available studies with read across substances, Coco TMAC and C16 TMAC, which revealed NOAELs based on reduction in body weight and body weight gain (consistent with decreased food consumption), it can be concluded that all effects are due to local gastrointestinal irritaton/corrosion and consequent reduced food intake without observing any primary systemic effect. Therefore, the derivation of a DNEL for systemic effects was deemed inappropriate.

Dermal

A 28-day repeat dose study was conducted to determine the repeated dose dermal toxicity of the read across substance, C16 TMAC, in New Zealand albino rabbits (both sexes) according to a method similar to OECD Guideline 410. The purity was not specified and the study included a lower than recommended number of animals (10/group rather than 20/group as per the guideline) and histopathology was performed only on limited organs. The test substance (at 0 and 10 mg/kg bw/day) was applied to the shaved, intact skin of groups of five New Zealand albino rabbits/sex/group up to 7 h, 5 days/week for 4 weeks. Dermal irritation effects were recorded daily. The animals were weighed weekly during the exposure period. Blood was collected for haematology measurements before initiation of dosing and prior to termination. Liver and kidneys weights were recorded at necropsy and limited histopathology was conducted. There were no systemic treatment-related effects on body weights, haematology, organ weights, gross necropsy findings or histopathology. Treated areas of the skin showed mild to marked acanthosis with active mitosis, hyperkeratosis, and partial to extensive necrosis of the epidermis and hair follicles, partly with encrustation and exudate. Under the study conditions, the 28 d NOAEL for male and female rabbits was found to be 10 mg/kg bw/day (Spicer, 1979).

Justification for classification or non-classification

Based on the effects observed in the repeat dose toxicity studies with the read across substances, the test substance Quaternary ammonium compounds, C12-14 alkyltrimethyl, Me sulfates, does not warrant classification for STOT RE according to EU CLP (EC/1272/2008) criteria.