Octanoic acid, mixed tetraesters with 2-ethylhexanoic acid, heptanoic acid and pentaerythritol

Endpoint summary

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Reference 1

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:

from November 1992 to December 1992

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

RA study

Justification for type of information:

The supporting repeat dose study is considered sufficient to fulfil the information requirement.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

other: Alpk:APfSD

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Zeneca Pharmaceuticals, Alderly Park, Macclesfield, Cheshire, UK
- Age at study initiation: 28 d
- Weight at study initiation: 148.45 g (males); 122.6 g (females)
- Housing: sexes separately, five per cage, cages had measurements of 26.5 x 50.0 x 20.0 cm and were constructed of stainless steel mesh with one solid side
- Diet: CT1 diet; Special Diets Services Limited, Witham, Essex, UK
- Water: tap water, ad libitum
- Acclimation period: approx. 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23
- Humidity (%): 45-65
- Air changes (per hr): 25-30
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: From: November 1992 To: December 1992

Route of administration:

oral: feed

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: All diet preparations were based on CT1 diet (Special Diets Services Limited, Witham, Essex, UK). They were prepared by grinding the appropriate amount of test substance with 1 kg of milled CT1 diet. This premix was then added to 14 kg of diet and mixed thoroughly with a Pharma Blender Model PMA 100S (T K Filder).

DIET PREPARATION
- Rate of preparation of diet (frequency): 15 kg batches
- Mixing appropriate amounts with (Type of food): CT1 diet (Special Diets Services Limited, Witham, Essex, UK)
- Storage temperature of food: - 20°C, stored at room temperature for usage up to 14 days

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Chemical stability was determined for diet preparations over a period of 5 weeks following storage at room temperature or at -20°C. Samples were extracted with ethyl acetate. The supernatant was diluted with ethyl acetate to give solutions containing appropriate concentrations of the test substance. Extracts were analysed by gas chromatography using flame ionisation detection. The extract concentration was calculated by reference to data from a standard containing a known concentration.

Duration of treatment / exposure:

28 days

Frequency of treatment:

Daily

Dose / conc.:

1 000 mg/kg diet

Remarks:

112 and 119 mg/kg bw/day for males and females resp. (actual dose received)

Dose / conc.:

5 000 mg/kg diet

Remarks:

562 and 586 mg/kg bw/day for males and females resp (actual dose received)

Dose / conc.:

12 500 mg/kg diet

Remarks:

1450 and 1613 mg/kg bw/day for males and females resp. (actual dose received)

No. of animals per sex per dose:

5

Control animals:

yes

Details on study design:

- Dose selection rationale: Based on results of preliminary feeding studies
- Rationale for animal assignment (if not random): The sexes were randomised separately.

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS:
Yes- Time schedule: Daily
- Cage side observations checked: changes in clinical condition and behaviour and significant changes were recorded.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: on Days 8, 15, 22, 29- observations included, but were not limited to the assessment of autonomic function (e.g. lacrimation, salivation, piloerection, exophthalmus, urination, defecation, pupillary function, ptosis); description, incidence and severity of any convulsions, tremors, abnormal motor function, alteration in respiration, reactivity to stimuli, changes in the level of arousal, sensorimotor responses

BODY WEIGHT: Yes, measurement in replicate order immediately before feeding and at the same day once a week until termination.

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as mg food/kg body weight: Yes, on a weekly basis
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

HAEMATOLOGY: Yes At termination, all rats were bled by cardiac puncture and samples were collected. Parameters determined: Hemoglobin, red cell count, haematocrit, mean cell volume, mean cell haemoglobin, mean cell haemoglobin concentration, platelet count, white blood cell count, neutrophil count, lymphocyte count, monocyte count, eosinophil count, prothrombin time and kaolin-cephalin time

CLINICAL CHEMISTRY: Yes At termination, all rats were bled by cardiac puncture and samples were collected. Parameters determined: Albumin, total protein, cholesterol, triglycerides, urea, creatinine, glucose, total bilirubin and alkaline phosphatase, plasma gamma-glutamyl transferase, plasma alanine aminotransferase, plasma aspartate aminotransferase, plasma creatine kinase, plasma sodium, plasma potassium, plasma chloride, plasma calcium and plasma phosphorus

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: on Days 8, 15, 22, 29
- Dose groups that were examined: All
- Battery of functions tested: sensory activity / grip strength

Sacrifice and pathology:

All rats were killed by exsangination under terminal anaesthesia induced by halotane and subjected to a full post mortem examination. Brain, adrenal glands, kidneys, liver, ovaries and testes were weighed, paired organs were weighed together.The following tissues were removed during the post mortem examination and fixed in 10% neutral buffered formol saline and fixed in 10% neutral buffered formol saline (except eyes and Harderian glands which were fixed in Davidson's solution and skin, mammary gland and testes/epididymides which were fixed in Bouin's solution): adrenal gland, aorta, bladder, bone (femur including knee joint), bone marrow (femur), brain, caecum, cervical lymph node, cervix, colon, duodenum, epididymis, eye and Harderian gland, heart, ileum, jejunum, kidney, liver, lung, mammary gland (inginal - female only) , mesenteric lymph node, nasal passages, oesophagus, oral cavity, ovary, pancreas, parathyroid gland, pituitary gland, prostate gland, rectum, salivary gland, sciatic nerve, seminal vesicle, skin (right flank), spinal cord, spleen, sternum, stomach, testis, thymus, thyroid gland, trachea, uterus, voluntary muscle and abnormalities.Adrenal gland, brain, heart, kidney, liver, lung, ovary, spleen, testis and abnormalities were processed and embedded in paraffin wax.All other tissues were stored. 5 micrometer sections of all processed tissues from control and the highest dose were cut, stained with haematoxylin and eosin and examined under the light microscope. Subsequently, liver and kidneys of male rats from the other dose groups (1000 and 5000 mg/kgbw/day) were also examined histopathologically.

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes (adrenals, aorta, bladder, bone and bone marrow (femur), brain, caecum, colon, cervical lymph node, cervix, colon, duodenum, epididymis, eye and harderian gland, heart, ileum, jejunum, kidney, liver, lungs, mammary gland, mesenteric lymph node, nasal passages, oesophagus, oral cavity, ovaries, pancreas, parathyroid glad, pituary gland, prostate gland, rectum, salivary glands, sciatic nerve, seminal vesicles, skin, spinal chord, spleen, sternum, stomach, testes, thymus, thyroid gland, trachea, uterus, voluntary muscle)

Statistics:

Bodyweights were considered by analysis of covariance on initial body weight, separately for males and females. Time to tail flick and fore and hindlimb grip strength at weeks 2, 3, 4 and 5 were considered by analysis of variance, separately for both sexes. Haematological and clinical blood parameters were considered by analysis of variance.Organ weights were considered by analysis of variance and covariance on final body weight separately for both sexes.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

Reduction in haemoglobin and haematocrit (males), reductions in haemoglobin, haematocrit and in white blood cell count (females). There were statistically significant reductions in haemoglobin and haematocrit at the highest dose in male rats. Statistically significant reductions in haemoglobin and haematocrit were seen in females at low and mid doses and in white blood cell count at low dose. In the absence of a coherent dose-response relationship, these differences were considered incidental to treatment. Any other statistically significant changes were considered spurious and unrelated to treatment with the test substance.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

There were minor reductions in plasma cholesterol, triglyceride and total protein levels and plasma alanine transferase activities in males at the highest dose compared to controls. Any other statistically significant changes were considered spurious and unrelated to treatment with the test substance.

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

A slightly reduced splay reflex was observed in one female of the 1000 ppm group (on days 29 and 30), in one male of the 5000 ppm group (on day 29) and in one male of the 12.500 ppm group (on day 29). As isolated observations, these were considered to be incidental.There were no differences in time to tail flick in either sex which could be attributed to treatment. The statistically significant increase in time to response observed on day 22 for males (5000 ppm) and day 8 for females (1000 ppm) were considered to be incidental to treatment in the absence of similar changes at higher dose levels. There was no evidence of any treatment related effects on forelimb or hindlimb grip strength. Any other statistically significant changes were considered spurious and unrelated to treatment with the test substance.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Kidney weights adjusted for body weight were statistically significant increased in males at 5000 and 12500 ppm. All the females in the treatment groups had slightly raised kidney weights compared to control, but none achieved statistical significance, and there was no evidence of a coherent dose response relationship.Liver weights adjusted for body weight were statistically significant increased in both sexes at 12500 ppm and in males at 5000 ppm. Any other statistically significant changes were considered spurious and unrelated to treatment with the test substance.

Gross pathological findings:

no effects observed

Neuropathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Treatment related findings were present in the kidney of male rats from all dose groups. In the 5000 and 12500 ppm dose group these comprised increased tubular hyaline droplet formation and tubular basophilia in all animals, and granular cast formation in four of the 5000 ppm animals and all of the 12500 ppm animals; the latter occurring at the cortico-medullary injection. In the 1000 ppm group, increased renal hyaline droplet formation and/or tubular basophilia were seen, but not granular cast formation. In the liver, there was minimal hepatocyte hypertrophy in 4/5 male rats in the 12500 ppm group.

Histopathological findings: neoplastic:

no effects observed

Details on results:

The increased kidney weights and microscopic findings of renal tubular basophilia, granular cast formation and increased hyaline droplet formation present in male rats at 5000 and 12.500 ppm are clearly treatment related. These findings are consistent with the well characterized light hydrocarbon nephropathy described for male rats, following to a variety of chemicals including light hydrocarbons such as unleaded gasoline and trimethyl pentane. The characteristics include an increased accumulation of hyaline droplets in male rat kidneys, the main constituent of which is alpha 2µ-globulin. It is widely accepted that this phenomenon is specific to male rat and as such appears to have no relevance for man.

Key result

Dose descriptor:

NOAEL

Effect level:

1 613 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: no treatment-related effects

Key result

Dose descriptor:

NOAEL

Effect level:

1 450 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: no treatment-related effects

Key result

Critical effects observed:

no

All diets prepared were found to be within 4% of the target concentration. The homogeneity of the test material in the diet, determined at 1000 and 12500 ppm inclusion levels was within 2% of the overall mean concentration for both levels. Chemical stability of the test substance, assessed at the 1000 and 12500 ppm inclusion levels stored at room temperature or at -20 °C was satisfactory over the period of use.

Conclusions:

Under the study conditions, the systemic NOAEL of the substance in rats were determined to be 1450 and 1613 mg/kg bw/day for males and females respectively.

Executive summary:

A study was conducted to determine the repeated dose toxicity of the read-across substance CAS 68424 -31-7 in rats according to OECD Guideline 407, in compliance with GLP. The Alpk:APfSD (Wistar derived) strain of rat was used in the study. The dose levels were selected on the basis of the results from preliminary feeding studies. Groups of five male and female rats were administered the test substance through diet at nominal dose levels of 1000, 5000 and 12500 ppm for 28 days (equivalent to actual dose levels of 112, 562 and 1450 mg/kg bw/day for males and 119, 586 and 1613 mg/kg bw/day for females). The analyzed concentrations of the test substance in the diet were within acceptable limits and the homogeneity and chemical stability of the formulation were satisfactory over the dosing period. The treated animals were then killed and subjected to a full post mortem examination. During the study, animals were observed daily for any changes in clinical condition and detailed clinical observations, including quantitative assessments of sensory perception and muscle weakness, were performed at weekly intervals. Bodyweights and food consumption were also recorded each week. Haematological and blood chemistry parameters were measured at study termination. Selected organs and tissues were removed and weighed at necropsy, processed and examined microscopically. There were no clinical signs indicative of neurological dysfunction in any treatment group nor was there any evidence of neuropathological changes in the brain. A minimal hepatocyte hypertrophy, present in the males of high dose group is considered to be evidence of an adaptive response. Microscopic examination of the kidneys from male animals from all treated groups revealed an increase in hyaline droplet formation and tubular basophilia; this phenomenon is specific to the male rat and as such appears to have no relevance to man. Under the study conditions, the systemic NOAEL of the substance in rats were determined to be 1450 and 1613 mg/kg bw/day respectively (Brammer, 1993).

Reference 2

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

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

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

RA study

Justification for type of information:

Refer to the section 13 of IUCLID dataset for details on the read across justification. The combined repeat dose-reproductive developmental toxicity 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 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

GLP compliance:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc., Atsugi breeding center
- Age at study initiation: (P) 10 wks
- Weight at study initiation: (P) Males: 330.8-420.4 g; Females: 207.2-245.0 g
- Housing: 2 animals per cage at mating
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0-25.0
- Humidity (%): 40.0-75.0
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 hrs dark/12 hrs light

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS: Doses were based on a 14 d range-finding study.

DIET PREPARATION
- Rate of preparation of diet (frequency): Prepared up to 8 days before dosing
- Storage temperature of food: Room temperature and shade

VEHICLE
- Justification for use and choice of vehicle (if other than water): For solubility
- Concentration in vehicle: 5, 15, 50% w/v
- Lot/batch no. (if required): V2P1825

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Concentration of test substance was analyzed by GC method

Duration of treatment / exposure:

42 days for males and 42-54 days for females

Frequency of treatment:

Daily once

Dose / conc.:

100 mg/kg bw/day (actual dose received)

Dose / conc.:

300 mg/kg bw/day (actual dose received)

Dose / conc.:

1 000 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

12 animals (5 animals for satelite groups)

Control animals:

yes, concurrent vehicle

Positive control:

Not applicable

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily during breeding period and recovery period, twice daily during dosing

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once a week

Functional behavior: Day 42 in all groups and Day 14 of recovery for satellite groups

BODY WEIGHT: Yes
- Time schedule for examinations: for males weekly and at necropsy, for females weekly during dosing and on Days 0, 7, 14 and 20 of gestation and on Days 0 and 4 of lactation and at necropsy

FOOD CONSUMPTION: weekly

WATER CONSUMPTION: No data

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at necropsy
- Anaesthetic used for blood collection: No
- Animals fasted: No data
- How many animals: 5 animals/sex/dose including the satellite groups

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at necropsy
- Animals fasted: No data
- How many animals: 5 animals/sex/dose including the satellite groups

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Statistics:

Fisher’s exact test (significance level: 5%) was performed on the function test results. Among the histopathological findings in the test substance treatment groups, the graded data was tested for significance in relation to the control group using Mann-Whitney U-testing (significance level: 5%), while the total values for positive grades were tested for significance with the control group using Fisher’s exact one-sided testing (significance level: 5%). For the other data, the values obtained for each individual or the mean values for each litter were used as single samples, and these samples were compared with the values within the satellite group or within the other groups. Where two groups were used in the analysis, F-testing was performed first, followed by Student t-testing if no significant differences were observed. If a significant difference was observed in the F-testing, Aspin-Welch testing was performed. When three or more groups were used in the analysis, the Bartlett method was used first in order to test for uniformity of variance in each group (significance level: 5%).If the variance was uniform, one-way analysis of variance (significance level: 5%) was performed. If a significant difference was observed between groups, the Dunnett method was used to perform a multiple comparison (significance level: 5%). On the other hand, if the variance in any of the groups was 0 and if the variance was not uniform, Kruskal-Wallis rank testing (significance level: 5%) was performed. If a significant difference was observed between groups, the Dunnett testing method was used to perform a multiple comparison (significance level: 5%).

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

A statistically significant increase in haemoglobin and haematocrit in females at 100 mg/kg bw/day and increased prothrombin time in females at 100 mg/kg bw/day were not considered to be treatment related as no effects at higher doses were noted. No effects were observed at the end of the recovery period.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

No effects in males or females in main group; statistically significant increase in urea nitrogen (121% of control) in females at 1000 mg/kg bw/day at the end of the recovery period; no effects in males at the end of the recovery period.

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

Statistically significant increase in absolute brain weight of females at 300 mg/kg bw/day was not considered to be treatment related as no effect at 1000 mg/kg bw/day was noted. A statistically significant increase of relative liver weight (105% of control) in females at 1000 mg/kg bw/day at the end of recovery, was not considered to be treatment related as it was only seen at the end of recovery, the increase was slight and no histopathological effect was found. No effects in males during main test or at end of recovery were noted.

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

Testes: focal seminiferous tubule atrophy: 3/12 M at 100 and 1/12 M at 300 mg/kg bw/day; no findings at 1000 mg/kg bw/day and no findings at the end of the recovery period; therefore not treatment related.
Kidney: very slight basophilic tubules in cortex in 2/5 control males and 4/5 males at 1000 mg/kg bw/day and in 1/5 female at 1000 mg/kg bw/day; no findings at end of recovery period.
Ovary: follicle cysts and increased number of follicles in closed state and a corresponding marked reduction in corpus luteum in 1/12 female at 1000 mg/kg bw/day; at end of recovery follicle cysts in 1/5 female at 1000 mg/kg bw/day. Other findings were comparable to the control group in incidence and severeness.

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: There were no effects related to test substance treatment.

Key result

Critical effects observed:

no

Conclusions:

Under the study conditions, the systemic NOAEL of the substance in rats was determined to be greater than 1000 mg/kg bw/day.

Executive summary:

A study was conducted to determine the repeated dose toxicity of the read-across substance CAS 7299 -99 -2 in rats according to OECD Guideline 422, in compliance with GLP. A combined repeated dose-reproductive developmental screening study was conducted in rats in which the test substance was administered to groups of 12 animals per sex via oral gavage at dose levels of 100, 300 and 1000 mg/kg bw/day for 42 days (males) or 42 -54 days (for females). Mortality, clinical signs, food consumption and body weight measurements were performed on a regular basis during the dosing period. Hematological and clinical chemistry examinations were performed before scheduled necropsy. Terminal body weight and organ weights were determined after the necropsy along with gross pathological and histopathological examinations of the major organs. Apart from few non-treatment related effects, no adverse effects were observed in any of the treated groups. Under the study conditions, the systemic NOAEL of the substance in rats was determined to be greater than 1000 mg/kg bw/day (MHLW, 2005).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

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

A study was conducted to determine the repeated dose toxicity of the read-across substance CAS 7299 -99 -2 in rats according to OECD Guideline 422, in compliance with GLP. A combined repeated dose-reproductive developmental screening study was conducted in rats in which the test substance was administered to groups of 12 animals per sex via oral gavage at dose levels of 100, 300 and 1000 mg/kg bw/day for 42 days (males) or 42 -54 days (for females). Mortality, clinical signs, food consumption and body weight measurements were performed on a regular basis during the dosing period. Hematological and clinical chemistry examinations were performed before scheduled necropsy. Terminal body weight and organ weights were determined after the necropsy along with gross pathological and histopathological examinations of the major organs. Apart from few non-treatment related effects, no adverse effects were observed in any of the treated groups. Under the study conditions, the systemic NOAEL of the substance in rats was determined to be greater than 1000 mg/kg bw/day (MHLW, 2005).

A supporting study was conducted to determine the repeated dose toxicity of the read-across substance CAS 68424 -31-7 in rats according to OECD Guideline 407, in compliance with GLP. The Alpk:APfSD (Wistar derived) strain of rat was used in the study. The dose levels were selected on the basis of the results from preliminary feeding studies. Groups of five male and female rats were administered the test substance through diet at nominal dose levels of 1000, 5000 and 12500 ppm for 28 days (equivalent to actual dose levels of 112, 562 and 1450 mg/kg bw/day for males and 119, 586 and 1613 mg/kg bw/day for females). The analyzed concentrations of the test substance in the diet were within acceptable limits and the homogeneity and chemical stability of the formulation were satisfactory over the dosing period. The treated animals were then killed and subjected to a full post mortem examination. During the study, animals were observed daily for any changes in clinical condition and detailed clinical observations, including quantitative assessments of sensory perception and muscle weakness, were performed at weekly intervals. Bodyweights and food consumption were also recorded each week. Haematological and blood chemistry parameters were measured at study termination. Selected organs and tissues were removed and weighed at necropsy, processed and examined microscopically. There were no clinical signs indicative of neurological dysfunction in any treatment group nor was there any evidence of neuropathological changes in the brain. A minimal hepatocyte hypertrophy, present in the males of high dose group is considered to be evidence of an adaptive response. Microscopic examination of the kidneys from male animals from all treated groups revealed an increase in hyaline droplet formation and tubular basophilia; this phenomenon is specific to the male rat and as such appears to have no relevance to man. Under the study conditions, the systemic NOAEL of the substance in rats were determined to be 1450 and 1613 mg/kg bw/day respectively (Brammer, 1993).

Justification for classification or non-classification

Based on the repeated dose studies conducted with the read-across substances, no classification is warranted according to EU CLP (EC 1272/2008) criteria.