Registration Dossier

Administrative data

Description of key information

Oral route: The acute oral median lethal dose (LD50) of the target substance in the female Wistar strain rat was found to be > 2000 mg/kg bw (OECD 420 and EU Method B.1).

 

Dermal route: The LD50 of an analogue test item was > 2000 mg/kg when administered once for 24 hours to the clipped, unabraded skin of male and female albino rats (OECD 402 and OPPTS 870.1200).

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
19 June 2017 to 10 July 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
fixed dose procedure
Limit test:
yes
Species:
rat
Strain:
Wistar
Remarks:
RccHan:WIST
Sex:
female
Details on test animals or test system and environmental conditions:
ANIMAL INFORMATION
- Female Wistar (RccHan:WIST) strain rats were supplied by Envigo RMS (UK) Limited, Oxon, UK.
- On receipt the animals were randomly allocated to cages.
- The female animals were nulliparous and non-pregnant.
- After an acclimatisation period of at least five days, animals were selected at random and given a number unique within the study by indelible ink-marking on the tail and a number written on the cage card.
- At the start of the study the animals were 8 to 12 weeks of age.
- Body weight variation did not exceed ± 20 % of the mean body weight at the start of treatment.

ANIMAL CARE AND HUSBANDRY
- Animals were housed in groups of up to four in suspended solid-floor polypropylene cages furnished with woodflakes.
- With the exception of an overnight fast immediately before dosing, and for approximately 3 to 4 hours after dosing, free access to mains drinking water and food (2014C Teklad Global Rodent diet supplied by Envigo RMS (UK) Limited, Oxon, UK) was allowed throughout the study.
- Diet and drinking water were routinely analysed and were considered not to contain any contaminants that would reasonably be expected to affect the purpose or integrity of the study.
- Temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70 % respectively.
- Rate of air exchange was at least 15 changes per hour.
- Lighting was controlled by a time switch to give 12 hours continuous light and 12 hours darkness.
- Animals were provided with environmental enrichment items that were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.
Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on oral exposure:
TEST ITEM PREPARATION AND ANALYSIS
- The test item was freshly prepared, as required, as a suspension in arachis oil. Arachis oil BP was used because the test item did not dissolve/suspend in distilled water.
- The test item was formulated within 2 hours of being applied to the test system. It is assumed that the formulation was stable for this duration.
- No analysis was conducted to determine the homogeneity, concentration or stability of the test item formulation. This is an exception with regard to GLP and was reflected in the GLP compliance statement.

EXPOSURE TO TEST ITEM
- All animals were dosed once only by gavage, using a metal cannula attached to a graduated syringe.
- The volume administered to each animal was calculated according to the fasted body weight at the time of dosing.
- Treatment of animals was sequential.
- Sufficient time was allowed between each dose group to confirm the survival of the previously dosed animals.
- Clinical observations were made 0.5, 1, 2 and 4 hours after dosing and then daily for 14 days.
- Morbidity and mortality checks were made twice daily (early and late) during normal working days and once daily at weekends and public holidays.
- Individual body weights were recorded on Day 0 (the day of dosing) and on Days 7 and 14.
- At the end of the observation period animals were killed by cervical dislocation.
- All animals were subjected to gross necropsy consisting of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.


Doses:
Single dose of 2000 mg/kg bw
No. of animals per sex per dose:
Five
Control animals:
no
Details on study design:
STUDY DESIGN
- Using available information on the toxicity of the test item, 2000 mg/kg was chosen as the
starting dose (see Annex 2, attached).
- A single female animal was treated at a dose level of 2000 mg/kg (concentration 200 mg/mL; dose volume 10 mL/kg).
- In the absence of toxicity at a dose level of 2000 mg/kg, an additional four animals were treated at a dose level of 2000 mg/kg (concentration 200 mg/mL; dose volume 10 mL/kg).

DATA EVALUATION
- The test item was evaluated according to Annex 3 of the OECD Guidelines for Testing of Chemicals No 420 “Acute Oral Toxicity – Fixed Dose Method” (adopted 17 December 2001) as shown in the flow chart in Annex 3 (attached).
- Evaluation of data included identification of the number of animals that died during the study (or that were killed for humane reasons) plus determination of the nature, severity, onset and duration of toxic effects. If possible, the signs of evident toxicity were described. Evident toxicity refers to the toxic effects of sufficient severity that administration of the next higher dose level could result in development of severe signs of toxicity and probable mortality. Effects on body weights and abnormalities noted at necropsy were also identified.

MAJOR COMPUTERISED SYSTEMS
- Delta Controls: ORCAview
Statistics:
STATISTICS
- Using the mortality data obtained, an estimate of the acute oral median lethal dose (LD50) of the test item was made.
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
- Individual mortality data are given in Appendix 1 (attached).
- No unscheduled animal deaths took place.
Clinical signs:
- Individual clinical observations are presented in Appendix 1 (attached).
- No signs of systemic toxicity were noted during the observation period.
Body weight:
- Individual body weight and body weight changes are given in Appendix 2 (attached).
- All animals showed expected gains in body weight over the observation period.
Gross pathology:
- Individual necropsy findings are given in Appendix 3 (attached).
- No abnormalities were noted at necropsy.
Interpretation of results:
GHS criteria not met
Conclusions:
The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was found to be > 2000 mg/kg bw.
Executive summary:

GUIDELINE

The study was performed to assess acute oral toxicity of the test item in the Wistar strain rat in compliance with the OECD Guideline for Testing of Chemicals No 420 “Acute Oral Toxicity – Fixed Dose Method” (2001) and Method B.1 bis Acute Toxicity (Oral) of Commission Regulation (EC) No 440/2008.

 

METHODS

Following a sighting test at 2000 mg/kg, an additional four fasted female animals were given a single oral dose of test item, as a suspension in arachis oil BP, at a dose level of 2000 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

 

RESULTS

No unscheduled animal deaths took place during the study and no signs of systemic toxicity were reported. All animals showed expected gains in body weight and no abnormalities were noted at necropsy.

 

CONCLUSION

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was found to be > 2000 mg/kg bw.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Acute toxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no study available

Acute toxicity: via dermal route

Link to relevant study records

Referenceopen allclose all

Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
05 January 2015 to 20 January 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1200 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes
Test type:
fixed dose procedure
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Remarks:
Crl:CD(SD)
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST SYSTEM, ANIMAL RECEIPT AND ACCLIMATION
- Crl:CD(SD) rats from Charles River Laboratories, Inc., Raleigh, NC were used as the test system on this study. The animal model, the Crl:CD(SD) albino rat, is generally recognised as appropriate for acute dermal toxicity studies. The number of animalssselected was the minimum required to satisfy regulatory guidelines. The experimentalsdesign used the procedures and standards required by the current federal and internationalstest guidelines.
- The albino rats utilized for this study were received in good health from Charles River Laboratories, Inc., Raleigh, NC, on 30 December 2014 The rats were inspected by a qualified technician upon receipt, weighed, and uniquely identified by a subcutaneous microchip (BMDS) implanted in the dorso-scapular area. The rats were acclimated to laboratory conditions for a minimum of 5 days. During this period, each animal was observed twice daily for mortality and changes in general appearance or behaviour.

ANIMAL HOUSING
- Upon arrival, all animals were housed individually in clean, stainless steel, wire-mesh cages suspended above cage-board. The animals were maintained by the animal husbandry staff of WIL Research in accordance with SOPs. The animal facilities at WIL Research are accredited by AAALAC International.
- Enrichment devices were provided to all animals as appropriate throughout the study for environmental enrichment and to aid in maintaining oral health of the animals, and were sanitised weekly.

DIET, DRINKING WATER AND MAINTENANCE
- The basal diet used in this study, PMI Nutrition International, LLC, Certified Rodent LabDiet 5002, was a certified feed with appropriate analyses performed by the manufacturer and provided to WIL Research.
- Municipal water supplying the facility was analysed for contaminants according to WIL Research SOPs. The results of the diet and water analyses are maintained at Charles River.
- No contaminants were present in animal feed or water at concentrations sufficient to interfere with the objectives of this study.
- The basal diet and municipal water, delivered by an automatic watering system, were provided ad libitum throughout the acclimation period and during the study.

ENVIRONMENTAL CONDITIONS
- All animals were housed throughout the acclimation period and during the study in an environmentally controlled room. The room temperature and humidity controls were set to maintain environmental conditions of 71 ± 5 °F (22 ± 3 °C) and 50 ± 20 %, respectively.
- Room temperature and relative humidity data were monitored continuously and were scheduled for automatic collection on an hourly basis. Actual mean daily temperature ranged from 70.4 °F to 70.5 °F (21.3 °C to 21.4 °C) and mean daily relative humidity ranged from 32.0 % to 43.0 % during
the study.
- Fluorescent lighting provided illumination for a 12-hour light (0600 hours to 1800 hours) and 12-hour dark photoperiod. Lighting conditions were recorded every 15 minutes.
- Air handling units were set to provide a minimum of 10 fresh air changes per hour.
Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
PREPARATION
- Prior to use, the bulk test substance container was inverted and/or swirled.
- A sufficient amount of test substance was transferred into a storage container for dispensation.

ASSIGNMENT OF ANIMALS TO TREATMENT GROUPS
- Animals used in the study were randomly selected based on health and body weight from available stock and assigned to groups by use of WTDMS.
- The selected animals were approximately 9 weeks old at initiation of dosing; body weight values ranged from 295 g to 302 g for males and from 200 g to 215 g for females (± 20 % of the mean for each sex).

TEST SUBSTANCE ADMINISTRATION
- One group of 5 male and 5 female rats were dermally administered a single dose (24-hour, semi-occluded exposure) at a dose level of 2000 mg/kg (limit dose).
- The test substance was dosed undiluted based on its specific gravity. The dose volume was determined by dividing the dose levels, expressed as g/kg, by the density (0.98 g/mL, as provided by the safety data sheet). Individual doses were calculated based on body weights taken just prior to dosing and a dose volume of 2.04 mL/kg.
- On the day prior to dosing, the hair was removed from the backs and flanks of the rats using a small animal clipper. Individual doses of the test substance were applied to the maximum area possible on the dorsal skin. Doses covered at least 10 % of the total body surface. Each dose was applied to the skin and overwrapped with gauze bandages (< 8 ply) secured with non-irritating tape. Twenty-four hours following test substance application, the bandages were removed, and the sites were wiped with disposable paper towels moistened with tepid tap water.
Duration of exposure:
24 hours
Doses:
Single dose
No. of animals per sex per dose:
Five males and five females
Control animals:
no
Details on study design:
MORTALITY
- The rats were observed at approximately 1, 2, and 4 hours post-application on study day 0 and twice daily, once in the morning and once in the afternoon, thereafter for 14 days.

CLINICAL OBSERVATIONS
- The rats were observed at approximately 1, 2, and 4 hours post-application on study day 0 and once daily thereafter for 14 days.
- Observations included, but were not limited to, evaluation for changes in appearance of skin and fur, eyes, mucous membranes, respiratory and circulatory systems, autonomic effects, and central nervous system effects.

DERMAL OBSERVATIONS
- The application sites were examined for erythema, edema, and other dermal findings (see Appendix C, attached) beginning 30–60 minutes after bandage removal and daily thereafter through Study Day 14.
- The areas of application were clipped free of hair on the day prior to dosing and as needed to facilitate accurate dermal observations.

BODY WEIGHTS
- Body weights were obtained and recorded on Study Days 0 (initiation), 7, and 14 (termination).

NECROPSY
- Upon termination, all rats were euthanized by carbon dioxide inhalation.
- The major organ systems of the cranial, thoracic, and abdominal cavities were examined for all animals.
- Tissues were not collected.

DATA ACQUISITION AND ANALYSIS
- The major computer systems used on this study include, but are not limited to, those listed in the attached table.
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
- There were no deaths during the study.
Clinical signs:
- Summary data are shown in Table 1 (attached) and individual data are given in Table 5 (attached).
- There were no clinical observations noted during the study.
Body weight:
- Summary data are shown in Tables 2 and 3 (attached) and individual data are given in Tables 7 and 8 (attached).
- There were no remarkable body weight changes noted during the study.
Gross pathology:
- Summary data are shown in Table 4 (attached) and individual data are given in Table 9 (attached).
- There were no macroscopic findings at the scheduled necropsy.
Other findings:
DERMAL OBSERVATIONS
- Summary data are shown in Table 1 (attached) and individual data are given in Table 6 (attached).
- Dermal findings noted during the study consisted of very slight (grade 1) to slight (grade 2) erythema and desquamation for all 5 males and 5 females.
- The erythema subsided by study day 6, and desquamation subsided by study day 11. In addition, female numbers 5126 and 5129 had scabbing within the dose site on study day 2 and 3, respectively.
- There was no oedema observed at any dose site.
Interpretation of results:
GHS criteria not met
Conclusions:
The LD50 of the test item was greater than 2000 mg/kg when administered once for 24 hours to the clipped, unabraded skin of male and female albino rats.
Executive summary:

GUIDELINE

The study was designed to be in general compliance with the EPA OPPTS Guideline 870.1200 (1996):Acute Dermal Toxicityand the OECD Guidelines for Testing of Chemicals, Section 402 (1987):Acute Dermal Toxicity”. The objective was to determine the acute dermal median lethal dose (LD50), evaluate potential systemic toxicity, and evaluate the local irritative effects of the test substance when applied once to the skin of albino rats.

 

METHODS

The test substance was administered once dermally for a 24-hour period under semi-occlusive dressing to Crl:CD(SD) albino rats to perform the limit test. The test substance was administered to one group of 5 male and 5 female rats at a dose level of 2000 mg/kg. Mortality, clinical observations, dermal findings and body weight changes were evaluated over a 14-day observation period. All animals were subjected to a gross necropsy.

 

There were no deaths, clinical observations, remarkable body weight changes, or test substance-related gross necropsy findings. Dermal findings noted during the study consisted of very slight (grade 1) to slight (grade 2) erythema and desquamation for all 5 males and 5 females and scabbing within the dose site for 2 females.

 

CONCLUSION

The LD50 of the test item was greater than 2000 mg/kg when administered once for 24 hours to the clipped, unabraded skin of male and female albino rats.

Endpoint:
acute toxicity: dermal
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Justification for type of information:
See read-across justification attached in Section 13.
Reason / purpose for cross-reference:
read-across source
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed

Additional information

Oral route

The key study was performed to assess acute oral toxicity of the registered substance in the Wistar strain rat in compliance with the OECD Guideline for Testing of Chemicals No 420 “Acute Oral Toxicity – Fixed Dose Method” (2001) and Method B.1 bis Acute Toxicity (Oral) of Commission Regulation (EC) No 440/2008.

 

Following a sighting test at 2000 mg/kg, an additional four fasted female animals were given a single oral dose of test item, as a suspension in arachis oil BP, at a dose level of 2000 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

 

No unscheduled animal deaths took place during the study and no signs of systemic toxicity were reported. All animals showed expected gains in body weight and no abnormalities were noted at necropsy.

 

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was found to be > 2000 mg/kg bw.

Dermal route

The key study was designed to be in general compliance with the EPA OPPTS Guideline 870.1200 (1996) “Acute Dermal Toxicity” and the OECD Guidelines for Testing of Chemicals, Section 402 (1987):“Acute Dermal Toxicity”. The objective was to determine the acute dermal median lethal dose (LD50), evaluate potential systemic toxicity, and evaluate the local irritative effects of the test substance when applied once to the skin of albino rats.

 

An analogue test substance was administered once dermally for a 24-hour period under semi-occlusive dressing to Crl:CD(SD) albino rats to perform the limit test. The test substance was administered to one group of 5 male and 5 female rats at a dose level of 2000 mg/kg. Mortality, clinical observations, dermal findings and body weight changes were evaluated over a 14-day observation period. All animals were subjected to a gross necropsy.

 

There were no deaths, clinical observations, remarkable body weight changes, or test substance-related gross necropsy findings. Dermal findings noted during the study consisted of very slight (grade 1) to slight (grade 2) erythema and desquamation for all 5 males and 5 females and scabbing within the dose site for 2 females.

 

The LD50 of the test item was greater than 2000 mg/kg when administered once for 24 hours to the clipped, unabraded skin of male and female albino rats.

Inhalation route

The registered substance has a high onset boiling point (decomposition from approximately 150 °C at 101 kPa) and the determined vapour pressure for a close analogue was shown to be 6.27 x 10E-03 Pa at 25 °C. It is therefore expected that inhalation exposure will be low under general use conditions at ambient temperature. Lack of systemic toxicity when the target substance is administered via the oral route and the analogue is administered via the dermal route suggest that determined vapour pressures of 1.51 x 10E-02 Pa at 55 °C and 7.24 x 10E-02 Pa at 85 °C for the analogue substance also gives no cause for concern. The inhalation route is therefore not the most applicable method of investigating acute toxicity.

Justification for classification or non-classification

The registered substance exhibits low acute toxicity via the oral route (LD50 > 2000 mg/kg bw) and an analogue substance was shown to exhibit similar low toxicity via the dermal route (LD50 > 2000 mg/kg bw) in test animals. Based on these data, classification of the target substance for acute toxicity is not required under the terms of Regulation (EC) No 1272/2008.