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Administrative data

Description of key information

NOEL (30d) (male and female) ≥ 1000 mg/kg bw/day (nominal)

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
From January 15th to February 28th, 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:
The read across approach is detailed into the document attached to the IUCLID section 13.
Reason / purpose:
read-across source
Principles of method if other than guideline:
The toxicity of the substance was investigated in male and female rats. The substance was daily administered by gavage at the dose level of 1000 mg/kg bw; the total duration of dosing was of 30 day; two additional weeks were dedicated to recovery.
GLP compliance:
no
Limit test:
yes
Species:
rat
Strain:
other: Tif: RAIf (SPF)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: 4 weeks.
- Weight at study initiation: 128-129 g, female; 113-115 g males.
- Housing: the animals were housed individually in Macrolon cages type 3 with standardized granulated soft wood bedding (Socicté Parisicnne des sciures Pantin).
- Diet: pelleted standard diet (Nafag. No. /890) ad Libitum.

Neither insecticides nor chemicals were applied in the animal room with the exceptic of a desinfectant: Fungitex SB (Prod. Nr. 30071).

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 1°C
- Humidity: 55 ± 5 %
- Air changes: 15-17 per hr
- Photoperiod: 10 hrs light/day
Route of administration:
oral: gavage
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS
- Volume: 10 ml/kg of a daily prepared suspension (10 % compound in Carboxymethyl-cellulose 2 %).
Duration of treatment / exposure:
4 weeks and 2 days of the 5th week (30 days)
Frequency of treatment:
5 times at week (in total 22 doses); 1 dose per day for 4 weeks plus in the 5th week on Monday to Tuesday.
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
20 males, 20 females per experimental group; 10 male and 10 female animals had to undergo a two weeks recovery period.
Control animals:
yes, concurrent vehicle
Details on study design:
- Recovery: two weeks of recovery period.
Observations and examinations performed and frequency:
MORTALITY
Recorded daily (a.m. and p.m.).

SYMPTOMS
Recorded daily.

BODY WEIGHT
Recorded weekly.

FOOD CONSUMPTION
Recorded weekly.

EYE EXAMINATION
Eye were examined by observation prior to dosing and after the treatment period.

HEARING TEST
Hearing test was conducted prior to dosing and after the treatment period (by hand clapping).

MEAN FOOD CONSERVATION
It was calculated according the following formula: MFC = (weekly food consumption in g / midweek body weight) x (1000/7)

HAEMATOLOGY and CLINICAL CHEMISTRY
Haematologic and blood chemistry measurements were carried out by standard methods on 20 randomized rats (5 males, 5 females per group) from the control and one test group at test day 28.
To reduce the biologic variability due to circadian rhythms, blood sampling for haematology and blood chemistry was between the hours of 8.00 and 9.00 a.m. For blood chemistry measurements food was withheld overnight prior to blood removal. The site of blood removal was the orbital sinus and a micro-
haematocrit glass capillary tube was used.
Blood samples from each animal with the respective anticoagulant (EDTA for performing haematologleal and heparin for blood chemistr measurements) were aliquoted into individual vials.
No anaesthesia was used to restrain the animals. All blood collection was by manual restraint only. The quantitative assay of all blood parameters was completed within a 8 h. period under "Quality Control" conditions.
- Parameters haematology: haemoglobin, erythrocytes, leucocytes total count, leucocytes differential count, asp. aminotransferase, ala aminotransferase.

URINALYSIS
Urinalysis measurements were carried out by standard methods on 20 randomized rats (5 males, 5 females per group) from the control and one test group at test day 28.
Urine for analysis was collected overnight. The individual rats were housed in special metabolism cages.
Food and water was withheld during the time of urine collection.
- Parameters: pH, protein, glucose, urobilinogen, urine sediment.
Sacrifice and pathology:
GROSS PATHOLOGY
The following organ weights were taken of all kill.ed aniinals: adrenals, kidneys, liver. Organ weights and organ to bodyweight ratlos revealed no statistically significant differences between treated and control animals.

MACROSCOPICAL EXAMINATION
At the end of the 30 day toxicity study or after an additional recovery period of two weeks the treated and control rats were bled under ether anaesthesia and complete autopsy was carried out.
The total weight of each animal was deterimined and following organs were weighed: liver, kidneys and adrenals.
The mean organ weights and mean organ to body weight ratios of each of these organs were calculated for all treated and control groups.

MICROSCOPICAL EXAMINATION
Tissue portions of trachea, lungs, liver, pancreas, salivary gland, stomach, small and large intestine, heart, spleen, lymphnodes, thymus, kidneys, urinary bladder, testicles or ovaries, prostate or uterus, pituitary, thyroid, adrenals, skeletal muscle, skin, mammary glands, bone marrow, eyes, brain, spinal cord, sciatic nerve, oesophagus and all other organs and tissues which showed macroscopical changes were fixed in buffered 10 % formalin.
The fixed tissues samples of liver, kidneys and adrenals were embedded in paramat and cut at 3- 5 p. The routine stain was haematoxylin and eosin. Sections from liver, kidneys and adrenals were also stained by Perl's method for iron.
Additional frozen sections from liver were specifically stained fan fat with Sudan III. Only the liver, kidneys and adrenals wene histologically examined.
Statistics:
For each time point and parameter a uni-variate statistical analysis was conducted. Due to the routine manner of the analysis system parameter free methods were applied. The treated group was compared to the control group in respect of dispersion and displacement.
Clinical signs:
no effects observed
Description (incidence and severity):
No clinical symptoms and no signs of local and/or systemic toxicity were observed.
Mortality:
no mortality observed
Description (incidence):
No animal died during the treatment and/or the recovery period.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
The body weight gain of all treated and control rats was comparable during the test and the recovery period.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
The mean food consumption of all treated animals was comparable to the controls during the 30 days of administration and during the recovery period.
Food efficiency:
no effects observed
Description (incidence and severity):
The mean food conversion of the males and females of the treated group was comparable to the controls.
Description (incidence and severity):
Eye examinatian did not reveal any ocular changes.
Haematological findings:
no effects observed
Description (incidence and severity):
The observed haematalogical findings between treated rats and controls were unremarkable.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
In the assessment of blood chemistry values the findings were unreinarkable and camparable to those of the controls.
Urinalysis findings:
no effects observed
Description (incidence and severity):
The findings in the urine were unremarkable. Individual rats revealed same degree of physiolagical proteinuria inclucling those og the control group. This is considered normal in laboratary rats.
Description (incidence and severity):
No loss of hearing ability was registered.
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
Organ weights and organ to bodyweight ratlos revealed no statistically significant differences between treated and control animals.
Gross pathological findings:
no effects observed
Description (incidence and severity):
No compound related gross anatomical changes were noted.
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
All minor changes seen in some control and treated rats and described within the microscopical findings in individual rats were only incidental in nature and not related to the test-item treatment.
Dose descriptor:
NOEL
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Remarks:
at the end of the study period or after the recovery period the treated rats showed neither macroscopical nor microscopical changes which could ha related to the test-item treatment
Critical effects observed:
no
Conclusions:
NOEL (30d) (male and female) ≥ 1000 mg/kg bw/day (nominal)
Executive summary:

The toxicity of the substance was investigated in male and female rats. The substance was daily administered by gavage at the dose level of 1000 mg/kg bw; the total duration of dosing was of 30 day; two additional weeks were dedicated to recovery.

During the experiment no treatment realted reactions were observed with respect to clinical syrnptoms, mortality, food consumption, body weight, clinical laboratory investigations and pathology. Eye examination did not reveal any ocular changes. No loss of hearing ability was registered.

It can be inferred from the observations made during the above study that 1000 mg/kg/day of test item produced no observable effects in male and female rats when administered daily by gavage over a period of 30 days.

Conclusion

NOEL (30d) (male and female) ≥ 1000 mg/kg bw/day (nominal)

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

There are no information about the repeated dose toxicity potential of Direct Orange 118, thus the available data on structural analogous Similar Substance 01 have been taken into consideration. The read across approach can be considered as reliable and suitable for the purpose; details and explanations are detailed in the report attached to the IUCLID section 13.

The toxicity of the Similar Substance 01 was investigated in male and female rats, over a period of 30 days. The substance was daily administered by gavage at the dose level of 1000 mg/kg bw. During the experiment no treatment realted reactions were observed with respect to clinical syrnptoms, mortality, food consumption, body weight, clinical laboratory investigations and pathology. Eye examination did not reveal any ocular changes. No loss of hearing ability was registered. 1000 mg/kg/day was indicated as No Observable Effect Level for both male and female rats.

The same results were obtained into a second sub-acute experiment, described in literature. The substance was administered to female and male rats as 22 daily doses, over a period of 30 days, at 1000 mg/kg body weight/day by gavage. No mortality or signs of toxicity were observed during the study period; food and water consumption were within the normal range following treatment with the test item. Test compound did not affect body weights. Comparison of the urinalysis of test and control animals and bilirubin and urobilinogen tests were negative. The clinical examinations of liver function revealed no signs of toxicity following treatment and recovery. No overall effects were observed on the hematological profiles of the test animals compared with the controls. Organ weights (liver, kidneys, adrenals) were subject to analysis of variance and analysis of covariance on final body weight. Histopathological examinations of selected tissues showed no significant effects related to the treatment with the substance. It was concluded that the oral dose level of 1000 mg/kg body weight/day was tolerated either without toxicity at all or without irreversible signs of toxicity in female and male rats, when administered over a period of 30 days.

The toxicity of the Similar Substance 01 was also examined in 13- and 28- weeks feeding studies with weanling albino rats as the experimental animals. General condition, behaviour, survival, growth, food intake and food efficiency were not adversely affected in any of the groups. Haematological and blood biochemichal parameters did not indicate a treatment-related effect of the dye. The percentage of eosinophils was slightly increased at 3000 ppm in females only. There were no changes in the composition of the urine which could be attributed to the treatment with the substance. Organ weights did not show distinct differences between the test groups and the controls. Gross pathology failed to reveal changes that could be attributed to the test substance. Microscopically, there were minimal liver changes, viz. pronounced lobular pattern of the liver in the 1500 and 3000 ppm groups in males of the 6-month study only. It was concluded that the no toxic effect level of test item when fed to rats for 6 months was 750 ppm in the diet, or 37.5 mg/kg body weight/day, with the understanding that the effects found at the 1500 and 3000 ppm levels were only weak, aspecific and of doubtful, if any biological significance.

Justification for classification or non-classification

According to the CLP Regulation (EC) No 1272/2008, 3.9 Specific target organ toxicity - repeated exposure section, substances are classified as specific target organ toxicants following repeated exposure by the use of expert judgement, on the basis of the weight of all evidence available, including the use of recommended guidance values, which take into account the duration of exposure and the dose/concentration, which produced the effect(s), and are placed in one of two categories, depending upon the nature and severity of the effect(s) observed.

In order to help reach a decision about whether a substance shall be classified or not, and to what degree it shall be classified (Category 1 or Category 2), dose/concentration ‘guidance values’ are provided for consideration of the dose/concentration which has been shown to produce significant health effects. The guidance values refer to effects seen in a standard 90-day toxicity study conducted in rats. Nevertheless, they can be used as a basis to extrapolate equivalent guidance values for toxicity studies of greater or lesser duration (the assessment shall be done on a case-by- case basis). For example, for 28-day study the guidance values are increased by a factor of three, thus the limit for sub-acute studies should be 300 mg/kg bw/day.

 

In the specific evaluation, the No Observed Effect Level was established at 1000 mg/kg bw/day, on the basis of the results obtained in the subacute study on rats.

 

In conclusion, the substance does not meet the criteria to be classified for repeated dose toxicity, according to the CLP Regulation (EC) No 1272/2008.