Registration Dossier

Administrative data

Description of key information

For the whole category of alcohol ethoxysulfates (AES) a oral NOAEL of 300 mg/kg bw/d was established.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Version / remarks:
adopted 12 May 1981
Deviations:
yes
Remarks:
Urinalysis and neurobehaviour were not examined.
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Hsd/Win:Wu
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann, Borchen, Germany
- Age at study initiation: approx. 5 weeks
- Weight at study initiation: 49 - 62 g (males); 52 - 74 g (females)
- Housing: 2 or 3 animals in Makrolon Type M 5 cages (E. Becker & Co. GmbH, Castrop-Rauxel, Germany) with standard softwood bedding (ARWI-Center, Essen, Germany)
- Diet: pelletised Altromin Maintenance Diet 1324 (Altromin GmbH, Lage, Germany), ad libitum
- Water : Community tap water, ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 12 - 23
- Humidity (%): 30 - 73
- Air changes (per hr): 10 - 15
- Photoperiod (hrs dark / hrs light): 12 / 12
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The dosing solutions of the test article and vehicle were prepared daily by dissolving an appropriate amount of the test material in water yielding final concentrations of 2.5, 7.5 and 22.5 mg/mL, respectively. The solutions were administered to the animals within approx. 2 - 3 h of preparation.

VEHICLE
- Concentration in vehicle: 2.5, 7.5 and 22.5 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg bw
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The dosing solutions of the test substance in water were analysed once to confirm the concentrations achieved. The formulations were analysed by "Henkel TTA-Zentrale Analytik" in the time from 09/15 - 09/17/1993 (study number TTA 93-9571).
Duration of treatment / exposure:
90 days
Frequency of treatment:
once daily, 5 days/week
Dose / conc.:
25 mg/kg bw/day (actual dose received)
Remarks:
corrected for a.i. content of test material
Dose / conc.:
75 mg/kg bw/day (actual dose received)
Remarks:
corrected for a.i. content of test material
Dose / conc.:
225 mg/kg bw/day (actual dose received)
Remarks:
corrected for a.i. content of test material
No. of animals per sex per dose:
10 (main study)
5 (satellite control, mid- and high-dose groups)
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose selection is based on the results of an earlier toxicoligical investigation. Rats (10 animals/dose/sex) were orally exposed to 100, 500 and 1000 mg/kg bw/day for 28 days (BASF, 1985, TBD870334). Adverse effects (lesions in the mucosa of the forestomach) were observed at 500 and 1000 mg/kg bw/day. In addition, alterations in hematology and clinical chemistry parameters were observed in the mid- and high-dose group animals when compared to the controls. Therefore, 25, 75 and 225 mg/kg bw/day were selected as the dose levels for the main sub-chronic (90-day) study. Moreover, the test substance is classified for its skin irritation and eye corrosion properties. The dose levels, therefore, needed to account for local irritating or corrosive effects in the gastro-intestinal tract of the test animals. A maximum dose of 225 mg/kg bw/day was found to be tolerated by the test animals for administration over a period of 90 days.

- Fasting period before blood sampling for clinical biochemistry: yes, over night
- Rationale for selecting satellite groups: In order to determine the reversibility of possible test substance-related alterations.
- Post-exposure recovery period in satellite groups: 28 days
Positive control:
Not necessary
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Acclimatisation period: once daily; treatment period: twice daily; weekend and public holidays: once daily
- Cage side observations: Mortality and morbidity

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Acclimatisation period: once daily; treatment period: twice daily; weekend and public holidays: once daily

BODY WEIGHT: Yes
- Time schedule for examinations: Acclimatisation periode: once; treatment period and treatment free-period: once weekly

FOOD AND WATER CONSUMPTION AND COMPOUND INTAKE:
- Time schedule: weekly, weighing per cage with the exeption of the dates for clinical chemistry

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: At beginning and end of treatment
- Dose groups that were examined: all

HAEMATOLOGY AND CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: After 6 weeks of administration (intermediate analysis) and at the end of treatment (final analysis)
- Anaesthetic used for blood collection: Yes (ether)
- Animals fasted: Yes
- Haematology parameters checked: Red blood cell count (RBC), Hematocrit (HCT), Mean corpuscular volume (MCV), Hemoglobin (HOB), White blood cell count (HBC), Platelet count (PLT), Differential white blood cell count (Bands, polymorphnuclear neutrophils (Seg), lymphocytes (Lymph), polymorphnuclear eosinophils (Eo), monocytes (Mono), polymorphnuclear basophils (Basa), myelocytes (My), juvenil (Ju), diverse (Div))
- Clinical chemistry parameters checked: Gamma-glutamyl-transferase (GGT), Aspartate-amino-transferase (AST), Alanine-amino-transferase (ALT), Alkaline phosphatase (AP), Sodium (Na), Potassium (K), Glucose, Urea, Total protein, Calcium (Ca), Creatinine, Total cholesterol, Chloride, Total bilirubin

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
After the last application all animals (main groups and satellite groups) were sacrificed by an overdose of ether and were exsanguinated by cardial puncture after opening of the thorax. All animals were submitted to full necrospsy procedures: external surface, all orifices, cranial cavity, external surface of the brain, thoracic, abdominal and pelvis cavities and viscera, carcass.
- Organ weights (paired organs were weighed separately): brain, testes, heart, liver, spleen, adrenals, kidneys, thymus
- Organs/tissues: adrenals, aorta, bone marrow, brain, caecum, colon, duodenum, epididymis, eyes, heart, ileum, jejunum, kidneys, liver, lungs, lymph node (submandibularis), lymph node (mesenteric), lymph node (inguinalis), mammary gland, oesophagus, ovaries, pancreas, parathyroids, prostate, rectum, salivary glands, skeletal muscle, skin, spinal cord (lumbar region), spleen, stomach (fundus, pylorus), testes, thymus, thyroids, trachea, urinary bladder, uterus (horn, cervix), lumbar vertebrae, all gross lesions
- Fixative: 10% neutral formalin except eyes fixed and preserved in Davidson's fluid

HISTOPATHOLOGY: Yes
Histopathological examinations were performed on all organs of male and female rats of all groups.
Statistics:
t-Test for determination of significant differences between the groups for body weight gains;
t-Test for determination of significant differences between the groups for hematological and clinical chemistry examinations;
Steel-Test for determination of significant differences between the groups for organ weight gains.

Evaluation:
- significance on 95% level: slight in-/decrease
- significance on 99% level: in-/decrease
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
One male in the mid-dose group showed a slight emaciation of short duration in the week 13 and 14.
In some females of all groups and only one male of the low-dose group slight to severe alopecia or thin fur were observed and considered as not treatment-related symptoms. The male showed also a skin incrustation on its back. Slight to severe acute swelling of the right ear forming a hematoma was observed in 3 males of the mid-dose group, one male of the high-dose group, one male of the satellite control group and one male of the the satellite mid-dose group. The swelling of the ears was not histologically examined because in an experiment before no treatment-relationship could be found. The exact etiology of this finding is unknown and may be presumably connected with the ear tattoo.
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Three females died due to bloodletting. One female was sacrificed after 66 applications after observing a traumatic fracture of the mandibula/maxilla. The mortalities were not treatment-related.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
The total body weight gain of the animals of the dosing groups and the males of the high-dose satellite group showed no deviations and was similar to the control groups. The total body weight gain of the females in the high-dose satellite group showed a slight decrease in weeks 2, 4, 5, 7, 8, 9, 11 and 12.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
The mean food consumption of the animals of the dosing groups showed some deviations which were in the normal range of historical values and were therefore not considered treatment-related. The severe increase of the females in the mid-dose group in weeks 8 to 10 was due to the female no. 64 which was considered as a pellet biting animal. The mean food consumption of males of the high-dose satellite group showed a slight increase in weeks 4 to 8 and the females of the high-dose satellite group exhibited a slight decrease from the beginning until the end of treatment. The reason for these not consistent variations is unknown.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
effects observed, non-treatment-related
Description (incidence and severity):
The mean water consumption of the animals of the dosing groups and the high-dose satellite group showed severe but not dose-related deviations which were therefore not considered treatment-related. The reason for these not consistent variations is unknown.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
The examination of the eyes by slit lamp showed no treatment-related effects.
Haematological findings:
no effects observed
Description (incidence and severity):
There were no significant deviations in any parameter and the values were similar in the treatment and control groups.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
Low-dose (25 mg/kg bw/day) group: slight increase of creatinine in males
Mid-dose (75 mg/kg bw/day) group: slight increase of creatinine and glucose in males

These significant findings are to be considered as within the normal range of historical values.
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
Absolute organ weights:
There were no significant deviations for absolute organ weights and the values were similar for the treatment and the control groups.

Relative organ weights:
Mid-dose (75 mg/kg bw/day): decrease of the heart in females
This finding is considered incidental and within the normal range of historical control data.
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
The macroscopical findings consisted of various spontaneous findings in males and females of all groups such as hydrometra of the uterus and various other diagnoses. Substance-related findings of the forestomach such as mucosal edema were observed in 4 males of the high-dose group (225 mg/kg bw/day). The satellite groups did not show substance-related findings.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
The microscopical findings consisted of spontaneous lesions in males and females of all groups such as hyperplasia of the mammary gland and hydrometriosis of the uterus and other spontaneous lesions. Acute and chronic lesions of the eyes due to bloodletting were observed occasionally.
The forestomach of the animals of the high-dose group (225 mg/kg bw/day) showed some lesions such as a hyperplasia, submucosal edema and ulceration. The morphology was consistent with the macroscopical findings. The lesions were observed in a higher frequency and higher degree in males than in females. The forestomach was evaluated as the target organ and examined in the low-dose (25 mg/kg bw/day) and mid-dose (75 mg/kg bw/day) groups and in all satellite groups. In the low- and mid-dose groups, 3 out of 10 animals showed small eosinophilic foci in the stratified epithelium of the forestomach. In some of the foci a small number of acute cells were observed. Beside these findings the forestomach of 1 animal showed an inflammatory edema beneath the mucosa. The mucosa of the forestomach of the mid-dose satellite group (75 mg/kg bw/day) showed no substance-related findings. The tendency of the regeneration of the mucosa was regular but incomplete in some of the male and female animals of the high-dose satellite group. The incomplete regeneration consisted of mild hyperplasia and few inflammatory cells in the submucosa. No signs of chronic ulcerations were observed.
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Key result
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
> 225 mg/kg bw/day (nominal)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: no systemic effects observed
Remarks on result:
other: The NOAEL considers the actual concentration of the registered substance in the test material investigated (70.1%).
Key result
Dose descriptor:
LOAEL
Remarks:
local
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
gross pathology
histopathology: non-neoplastic
Remarks on result:
other: The LOAEL considers the actual concentration of the registered substance in the test material investigated (70.1%).
Key result
Critical effects observed:
yes
Lowest effective dose / conc.:
25 mg/kg bw/day (actual dose received)
System:
gastrointestinal tract
Organ:
other: forestomach
Treatment related:
yes
Dose response relationship:
yes
Relevant for humans:
no
Conclusions:
A daily adminstration of the test substance revealed no systemically toxic effects other than varying degrees of expected treatment-related local effects in the forestomachs of males and females in all treatment groups. For the systemic toxicity after repeated application a No-Observed-Adverse-Effect-Level (NOAEL) of 225 mg/kg bw/day (the highest dose tested) can be deduced. The effects observed in the mucosa of the forestomach have to be interpreted as alterations due to the irritative potential of the test substance and not as signs of a cumulative systemic toxicity of the test substance. The Lowest-Observed-Adverse-Effect-Level (LOAEL) for local effects was 25 mg/kg bw/day.
Executive summary:

No treatment related lethality was found. The animals did not show any significant macroscopical and microscopical systemic organ injury. Food consumption, water intake and body weight gain were normal in all treatment groups. The haematological examination showed no deviation in any parameter. Clinical chemistry, examinations of the eyes by the slit lamp microscopy showed no treatment-related effects. Thus, for the systemic toxicity after repeated application a No-Observed-Adverse-Effect-Level (NOAEL) of 225 mg/kg bw/day can be deduced.

The necropsy revealed treatment-related lesions of the forestomach in the animals of the high-dose group. The effects were incompletely reversible as observed microscopically in animals of the satellite groups. The animals of the low-dose (25 mg/kg bw/day) and mid-dose (75 mg/kg bw/day) group showed microscopcial lesions of the mucosa of the forestomach. The lesions described were reversible in the animals of the mid-dose satellite group. These effects have to be interpreted as alterations due to the irritative potential of the test substance and not as signs of a cumulative systemic toxicity of the test substance. The Lowest-Observed-Adverse-Effect-Level (LOAEL) for local effects was 25 mg/kg bw/day.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
300 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
The whole data base is conclusive and of high quality.
System:
gastrointestinal tract
Organ:
other: forestomach

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Data on repeated dose toxicity are available for the registered substance C12-14 AES Na (CAS 68991-38-3).

In a subchronic oral gavage study according to OECD Guideline 408, AES (C12-14) Na (CAS 68891-38-3) was tested for systemic toxicity at doses of 25, 75 and 225 mg/kg bw/day on Wistar rats (BASF, 1994e). No clinical signs of systemic toxicity, no mortalities and no relevant treatment-related effects on any other investigated parameter (body weight, body weight gain, food consumption, water consumption, ophthalmoscopic examinations, haematology, clinical chemistry, organ weights, gross pathology and histopathology) were seen at the highest dose level. Apart from the missing systemic toxicity, local treatment-related concentration-dependent irritation to different degrees in the forestomach was seen in all test groups. Since there is no human equivalent to the forestomach in rats, these effects are not considered to be relevant to the human health assessment. Thus, a no adverse effect level (NOAEL) of greater than 225 mg/kg could be established.

In order to derive a reliable NOAEL for the whole AES category, NOAEL values from the repeated oral toxicity studies of other category members are summarised in the following table.

Table 1: Dietary NOAELs and LOAELs (a.i.) for repeated oral toxicity studies of AES

Substance

Duration

(weeks)

NOAEL/NOEL

(mg/kg bw/day)

LOAEL

(mg/kg bw/day)

Reference

AES(C12-14)Na

13

225

> 225

BASF (1994c)

AES(C12-15;3EO)Na

AES(C12;3EO)Na

13

250

> 250

Walker (1967)

AES(C10-16;3EO)Na

13

100

1100

P%G (1977a)

AES(C10-16;3EO)Na

13

600

> 600

P&G (1977b)

AES(C12-14)Na

13

300

> 300

BASF (1999)

AES(C12;3EO)

104

250

> 250

Little (1991)

The reference studies for other AES substances are detailed in the Category Justification document attached to the technical dossier. No LOAEL values could be detected in the most studies. With exception of one study all NOAEL values represent the highest dose levels tested. Therefore an average of all NOAEL values was chosen as basis for the risk assessment. The available oral toxicity studies provide a coherent picture on the subchronic and chronic oral toxicity of AES substances. Based on the described effects, the NOAEL of 300 mg/kg bw/day representing an average of all NOAEL values available, was chosen for the risk assessment of the whole AES category. The NOAEL value of 225 mg/kg bw/day determined with the registered substance C12-14 AES Na (CAS 68891-38-3) fits well within the range of NOAEL values for other AES substances.

Justification for classification or non-classification

The available data on repeated dose toxicity do not meet the criteria for classification according to Regulation (EC) No. 1272/2008, and are therefore conclusive but not sufficient for classification.