Registration Dossier

Administrative data

Description of key information

The acute toxicity of the test item was tested in differents levels (acute oral toxicity, acute inhalation toxicity and acute dermal toxicity in rats).

All experiments were performed according OECD GLP Guideline.

- Acute oral toxicity was performed according to the OECD 420: LD50 > 2000 mg/kg bw

- Acute dermal toxicty was performed according to the OECD 402: LD50 > 2000 mg/kg bw

- Acute inhalation toxicty was performed according to the OECD 403: LC 50 (4 hours, nose only) = 5,03mg/L (5030 mg/m3)

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:
The study was conducted between 27 October 2015 and 24 November 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP 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
Qualifier:
according to guideline
Guideline:
other: Japanese MAFF, 2000
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
fixed dose procedure
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals or test system and environmental conditions:
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 females were nulliparous and non pregnant. After an acclimatization period of at least 5 days the 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 a cage card. At the start of the study the animals were 8 to 12 weeks of age. The body weight variation did not exceed ±20% of the mean body weight at the start of treatment.
The 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. The diet, drinking water and bedding were routinely analyzed and were considered not to contain any contaminants that would reasonably be expected to affect the purpose or integrity of the study.

The temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give 12 hours continuous light and 12 hours darkness.

The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.


Justification
Rats are the preferred species of choice as historically used for safety evaluation studies and are specified in the appropriate test guidelines.
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
Test Item Formulation and Experimental Preparation
For the purpose of the study the test item was freshly prepared, as required, as a solution 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 has been reflected in the GLP compliance statement.
Doses:
A single animal was treated at 300 mg/kg at a concentration of 30 mg/mL.
In the absence of toxicity at a dose level of 300 mg/kg, an additional animal was treated at 2000 mg/kg at a concentration of 200 mg/mL.
In the absence of toxicity at a dose level of 2000 mg/kg, an additional group of 4 animals were treated at 2000 mg/kg.
No. of animals per sex per dose:
A single animal was treated at 300 mg/kg.
Five animals were treated at 2000 mg/kg.
Control animals:
no
Details on study design:
Procedure
In the absence of data regarding the toxicity of the test item, 300 mg/kg was chosen as the starting dose. A single female animal was treated with 300 mg/kg at a concentration of 30 mg/mL (dose volume 10 mg/kg).

In the absence of toxicity at a dose level of 300 mg/kg, an additional animal was treated at 2000 mg/kg at a concentration of 200 mg/mL (dose volume 10 mg/kg). In the absence of toxicity at a dose level of 2000 mg/kg, an additional group of 4 female animals were treated at 2000 mg/kg at a concentration of 200 mg/mL (dose volume 10 mg/kg).

A total of five animals were therefore treated at a dose level of 2000 mg/kg in the study.

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 30 minutes, 1, 2, and 4 hours after dosing and then daily for 14 days. Morbidity and mortality checks were made twice daily.

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 the animals were killed by cervical dislocation. All animals were subjected to gross necropsy. This consisted of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.


Evaluation of Data
Evaluation of data included identification of the number of animals that died during the study (or that were killed for humane reasons), and determination of the nature, severity, onset and duration of the 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.

Using the mortality data obtained, an estimate of the acute oral median lethal dose (LD50) of the test item was made.
Statistics:
No data
Preliminary study:
Dose Level - 300 mg/kg

Mortality
There was no mortality.

Clinical Observations
No signs of systemic toxicity were noted during the observation period. Orange colored stained urine was noted during the day of dosing.

Body Weight
The animal showed expected gains in body weight over the observation period.

Necropsy
No abnormalities were noted at necropsy.
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
Dose Level - 2000 mg/kg: There were no deaths.
Clinical signs:
Dose Level - 2000 mg/kg: No signs of systemic toxicity were noted during the observation period.
Body weight:
Dose Level - 2000 mg/kg: All animals showed expected gains in body weight over the observation period.
Gross pathology:
Dose Level - 2000 mg/kg: No abnormalities were noted at necropsy.

Individual Clinical Observations and Mortality Data -300mg/kg

Dose Level mg/kg

Animal Number and Sex

Effects Noted After Dosing
(Hours)

Effects Noted During Period After Dosing
(Days)

½

1

2

4

1

2

3

4

5

6

7

8

9

10

11

12

13

14

300

1-0

Female

0U

0U

0U

0U

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0 = No signs of systemic toxicity

U = Orange colored stained urine

Individual Body Weights and Body Weight Changes -300mg/kg

Dose Level

mg/kg

Animal Number
and Sex

Body Weight (g) at Day

Body Weight Gain (g)
During Week

0

7

14

1

2

300

1-0 Female

190

207

226

17

19


 Individual Necropsy Findings -300 mg/kg

Dose Level
mg/kg

Animal Number
and Sex

Time of Death

Macroscopic Observations

300

1-0 Female

Killed Day 14

No abnormalities detected

Individual Clinical Observations and Mortality Data -2000mg/kg

Dose Level mg/kg

Animal Number and Sex

Effects Noted After Dosing
(Hours)

Effects Noted During Period After Dosing
(Days)

½

1

2

4

1

2

3

4

5

6

7

8

9

10

11

12

13

14

2000

2-0

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-0

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-1

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-2

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-3

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0 = No signs of systemic toxicity

Individual Body Weights and Body Weight Changes -2000mg/kg

Dose Level

mg/kg

Animal Number
and Sex

Body Weight (g) at Day

Body Weight Gain (g) During Week

0

7

14

1

2

2000

2-0 Female

150

166

176

16

10

3-0 Female

164

181

202

17

21

3-1 Female

179

197

214

18

17

3-2 Female

178

199

219

21

20

3-3 Female

176

192

207

16

15

Individual Necropsy Findings -2000mg/kg

Dose Level
mg/kg

Animal Number
and Sex

Time of Death

Macroscopic Observations

2000

2-0 Female

Killed Day 14

No abnormalities detected

3-0 Female

Killed Day 14

No abnormalities detected

3-1 Female

Killed Day 14

No abnormalities detected

3-2 Female

Killed Day 14

No abnormalities detected

3-3 Female

Killed Day 14

No abnormalities detected

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was estimated to be greater than 2000 mg/kg body weight (Globally Harmonized Classification System - Unclassified).
Executive summary:

Introduction

The study was performed to assess the acute oral toxicity of the test item in the female Wistar strain rat.

 

Methods

Following a sighting test at dose levels of 300 mg/kg and 2000 mg/kg, a further group of four fasted females was given a single oral dose of test item, as a solution in distilled water, 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

Mortality. There were no deaths.

Clinical Observations. There were no signs of systemic toxicity. Orange colored stained urine was noted in the initial animal, treated at a dose level of 300 mg/kg body weight.

Body Weight. All animals showed expected gains in body weight.

Necropsy. 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 estimated to be greater than 2000 mg/kg body weight (Globally Harmonized Classification System-Unclassified).

 

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
2 000 mg/kg bw
Quality of whole database:
Study performed according to GLP Guideline and the score of the study is Klimisch 1.

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
The study was conducted between 25 September 2015 and 26 November 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
Animals and Animal Husbandry
Male and female RccHan™ : WIST strain rats were supplied by Envigo RMS (UK) Limited, Oxon, UK. On receipt the animals were randomly allocated to cages. After an acclimatization period of at least five days the animals were given a number unique within the study by ear punching and a number written on a color coded cage card. At the start of the study the animals were approximately eight to twelve weeks old and within the weight range of 200g to 350g. The females were nulliparous and non-pregnant.

The animals were housed in groups of up to five by sex in solid-floor polypropylene cages with stainless steel lids, furnished with softwood flakes (Datesand Ltd., Cheshire, UK) and provided with environmental enrichment items: wooden chew blocks and cardboard “fun tunnels” (Datesand Ltd., Cheshire, UK). With the exception of the exposure period, free access to mains drinking water and food (Rodent 2014C Teklad Global Certified Diet, Envigo RMS (UK) Limited, Oxon, UK) was allowed throughout the study. The diet, drinking water, bedding, chew blocks and cardboard “fun tunnels” are routinely analyzed and are considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.

The temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (06:00 to 18:00) and twelve hours darkness. The animals were retained in this accommodation at all times except during the exposure period.
Route of administration:
inhalation: dust
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
Inhalation Exposure
Atmosphere Generation
A dust atmosphere was produced from the test item using a SAG 410 Solid Aerosol Generator (TOPAS GmbH, Dresden, Germany) located adjacent to the exposure chamber. The SAG 410 was connected to a metered compressed air supply.

Compressed air was supplied by means of an oil free compressor and passed through a water trap and respiratory quality filters before it was introduced to the SAG 410.

The cylindrical exposure chamber had a volume of approximately 30 liters (dimensions: 28 cm diameter x 50 cm high). The concentration within the chamber was controlled by adjusting the test item feed rate from the SAG 410. The extract from the exposure chamber passed through a ‘scrubber’ trap and was connected with a high efficiency filter to a metered exhaust system. The chamber was maintained under negative pressure.
Homogeneity of the test atmosphere within the chamber was not specifically determined during this study. Chambers of the same design (ADG Developments Ltd, Hitchin, Herts, UK) have been fully validated and shown to produce evenly distributed atmospheres in the animals’ breathing zone with a wide variety of test items (Green J D et al, 1984).

Prior to the start of the study, test item atmospheres were generated within the exposure chamber. During this characterization period test item input rates and grinding times were varied in an attempt to achieve the required atmospheric conditions.


Exposure Procedure
One day prior to the day of exposure, each rat was acclimatized (for approximately 2 hours) to a tapered polycarbonate restraining tube. During the exposure period, each rat was individually held in a tapered, polycarbonate restraining tube fitted onto a single tier of the exposure chamber and sealed by means of a rubber ‘O’ ring. Only the nose of each animal was exposed to the test atmosphere.

Following an appropriate equilibration period a single group of ten rats (five males and five females) was exposed to an atmosphere of the test item for a period of four hours. A target concentration of 5.0 mg/L was used for the exposure. As the mean achieved concentration was 101 % of target and no deaths occurred, no further levels were required.


Exposure Chamber Temperature and Relative Humidity
The temperature and relative humidity inside the exposure chamber were measured by an electronic thermometer/humidity meter (Hanna Instruments Ltd, Beds., UK) located in a vacant port in the animals’ breathing zone of the chamber and recorded every thirty minutes throughout the four-hour exposure period.


Exposure Chamber Oxygen Concentration
Oxygen levels within the exposure chamber were measured by an electronic oxygen analyzer (Servomex (UK) Ltd, Crowborough, East Sussex) located in a port in the animals breathing zone during the four-hour exposure period.


Exposure Chamber Atmosphere Concentration
During the characterization phase of the study the test atmosphere was sampled twice and filter samples were then submitted for chemical analysis to determine if the original test item was similar to the composition of the airborne test item.
The actual chamber concentration was measured at regular intervals during the exposure period. The gravimetric method used glass fiber filters placed in a filter holder. The holder was temporarily sealed in a vacant port in the exposure chamber in the animals’ breathing zone and a suitable, known volume of exposure chamber air was drawn through the filter using a vacuum pump.

Each filter was weighed before and after sampling in order to calculate the weight of collected test item. The difference in the two weights, divided by the volume of atmosphere sampled, gave the actual chamber concentration.

The nominal chamber concentration was calculated by dividing the mass of test item used by the total volume of air passed through the chamber.

The nominal concentration was 198 % of the actual mean achieved atmosphere concentration and shows that keeping the aerosol airborne was relatively straightforward.


Particle Size Distribution
The particle size of the generated atmosphere inside the exposure chamber was determined three times during the exposure period using a Marple Personal Cascade Impactor (Westech IS Ltd, Beds., UK). This device consisted of six impactor stages (8.4, 7.3, 3.6, 1.3, 0.94 and 0.43 µm cut points) with stainless steel collection substrates and a backup glass fiber filter, housed in an aluminum sampler. The sampler was temporarily sealed in a sampling port in the animals’ breathing zone and a suitable, known volume of exposure chamber air was drawn through it using a vacuum pump.

The collection substrates and backup filter were weighed before and after sampling and the weight of test item, collected at each stage, calculated by difference.

The mean amount for each stage was used to determine the cumulative amount below each cut-off point size. In this way, the proportion (%) of aerosol less than 8.4, 7.3, 3.6, 1.3, 0.94 and 0.43 µm was calculated.

The resulting values were converted to probits and plotted against Log10 cut-point size. From this plot, the Mass Median Aerodynamic Diameter (MMAD) was determined (as the 50 % point) and the geometric standard deviation was calculated. In addition the proportion (%) of aerosol less than 4 µm (considered to be the inhalable fraction) was determined.
Analytical verification of test atmosphere concentrations:
yes
Remarks:
See below
Duration of exposure:
4 h
Concentrations:
5.0 mg/L
No. of animals per sex per dose:
Five males and five females
Control animals:
no
Details on study design:
Observations
Clinical Signs
All animals were observed for clinical signs at hourly intervals during exposure, immediately on removal from the restraining tubes at the end of exposure, one hour after termination of exposure and subsequently once daily for fourteen days. Any evidence of overt toxicity was recorded at each observation.

Body Weight
Individual body weights were recorded on arrival, prior to treatment on the day of exposure and on Days 1, 3, 7 and 14.

Necropsy
At the end of the fourteen day observation period the animals were killed by intravenous overdose of sodium pentobarbitone. All animals were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded. The respiratory tract was subjected to a detailed macroscopic examination for signs of irritancy or local toxicity.
Statistics:
No data
Preliminary study:
No data
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5.03 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
There were no deaths
Clinical signs:
other: Signs of hunched posture and pilo-erection are commonly seen in animals for short periods on removal from the chamber following 4-Hour inhalation studies. Wet fur is commonly recorded both during and for a short period after exposure. Generalized fur sta
Body weight:
All animals exhibited body weight losses or showed no body weight gains on the first day post-exposure. With the exception of three female animals which showed further body weight losses from Days 1 to 3 post-exposure, all animals exhibited body weight gains during the remainder of the recovery period.
Gross pathology:
No macroscopic abnormalities were detected amongst animals at necropsy.
Other findings:
Exposure Chamber Concentration
The chamber flow rate was maintained at 60 L/min providing 120 air changes per hour.
The theoretical chamber equilibration time (T99) was 3 minutes* (Silver, 1946).
* Test atmospheres were generated for a total of 14 minutes prior to animal insertion to ensure the target test item concentration was being achieved.

Exposure Chamber Concentration

The test atmosphere was sampled seventeen times during the exposure period and the actual concentration of the test item calculated. The mean values obtained were:

 

Atmosphere Concentration

Mean Achieved (mg/L)

Standard Deviation

Nominal (mg/L)

5.03

0.12

9.96

Particle Size Distribution

The particle size analysis of the atmosphere drawn from the animals’ breathing zone, was as follows:

 

Mean Achieved Atmosphere Concentration (mg/L)

Mean Mass Median Aerodynamic Diameter (µm)

Inhalable Fraction

(% <4 µm)

Geometric Standard Deviation

5.03

3.66

54.1

2.36

Exposure Chamber Atmosphere Concentrations

Duration of Exposure (minutes)

Net Weight of Sample (mg)

Volume of Air Sampled (L)

Chamber Flow Rate (L/min)

Atmosphere Concentration (mg/L)

5

10.26

2

60

5.13

15

10.11

2

60

5.06

30

9.51

2

60

4.76

45

9.75

2

60

4.88

60

10.26

2

60

5.13

75

10.00

2

60

5.00

90

10.29

2

60

5.15

105

10.22

2

60

5.11

120

9.72

2

60

4.86

135

10.32

2

60

5.16

150

10.16

2

60

5.08

165

10.12

2

60

5.06

180

9.90

2

60

4.95

195

10.05

2

60

5.03

210

9.89

2

60

4.95

225

10.22

2

60

5.11

235

10.21

2

60

5.11

Mean achieved atmosphere concentration (mg/L) =5.03

Standard deviation =0.12

 

Nominal concentration:

Test item used (g)

152

Air Flow (L/min)

60

Total Generation Time (mins)

254[1]

Nominal Concentration (mg/L)

9.96

[1]= Test atmospheres were generated for a total of 14 minutes prior to animal insertion to ensure the target test item concentration was being achieved.

Particle Size Distribution

Cascade Impactor Data 

Impactor Stage Number

Cut Point

(µm)

Amount Collected (mg) per Sample Number

Mean Amount Collected (mg)

1

2

3

3

8.4

0.35

0.29

0.33

0.32

4

7.3

0.36

0.32

0.34

0.34

5

3.6

0.66

0.53

0.62

0.60

6

1.3

0.75

0.73

0.65

0.71

7

0.94

0.22

0.25

0.22

0.23

8

0.43

0.11

0.16

0.15

0.14

Back-up Filter

<0.43

0.01

0.01

0.01

0.01

Total Mean Amount of Test Item Collected

2.35

 

Calculation 

Cut Point

(µm)

Log10

Cut Point

Mean Cumulative Amount Less Than Cut Point

(mg)

(%)

Probit

8.4

0.924

2.03

86.4

6.10

7.3

0.863

1.69

71.9

5.58

3.6

0.556

1.09

46.4

4.91

1.3

0.114

0.38

16.2

4.01

0.94

-0.027

0.15

6.38

3.48

0.43

-0.367

0.01

0.43

2.37

Results 

Mean Mass Median Aerodynamic Diameter (MMAD) =3.66µm

Geometric Standard Deviation (GSD) =2.36

Predicted amount less than 4 µm =54.1%

 

Mortality Data

MeanAchievedAtmosphere Concentration (mg/L)

Sex

Deaths During Exposure

Deaths Post Exposure

(1 Hour)

Deaths During Day of Observation

Total Deaths

1

2

3

4

5

6

7

8-14

5.03

Male

0

0

0

0

0

0

0

0

0

0

0/10

Female

0

0

0

0

0

0

0

0

0

0

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
No deaths occurred in a group of ten rats exposed to a mean achieved atmosphere concentration of 5.03 mg/L for four hours. It was therefore considered that the acute inhalation median lethal concentration (4 hr LC50) of FAT 40871/A TE, in the RccHanTM : WIST strain rat, was greater than 5.03 mg/L.
Executive summary:

Introduction

A study was performed to assess the acute inhalation toxicity of the test item.The method used was designed to be compatible with that described in the OECD Guideline for Testing of Chemicals No. 403 “Acute Inhalation Toxicity” (2009) and Method B2. Acute Inhalation Toxicity, 2014, of Commission Regulation (EC) No. 440/2008.

 

Methods.......

A group of ten RccHan™ : WIST strain rats (five males and five females) was exposed to a dust atmosphere. The animals were exposed for four hours using a nose only exposure system, followed by a fourteen day observation period.

 

Results…..

The mean achieved atmosphere concentration was as follows: 

Atmosphere Concentration

Mean Achieved (mg/L)

Standard Deviation

Nominal (mg/L)

5.03

0.12

9.96

 

The characteristics of the achieved atmosphere were as follows: 

Mean Achieved Atmosphere Concentration (mg/L)

Mean Mass Median Aerodynamic Diameter (µm)

Inhalable Fraction

(% <4 µm)

Geometric Standard Deviation

5.03

3.66

54.1

2.36

 

The mortality data were summarized as follows: 

Mean Achieved Atmosphere Concentration (mg/L)

Deaths

Male

Female

Total

5.03

0/5

0/5

0/10

 

Clinical Observations

Common abnormalities noted during the study included decreased respiratory rate, hunched posture, pilo-erection, generalized orange fur staining caused by the test item and wet fur. Occasional instances of increased respiration were also noted. All animals recovered such that no significant observations were apparent from Days 2 to 3 post-exposure.

 

Body Weight

All animals exhibited body weight losses or showed no body weight gains on the first day post-exposure. With the exception of three female animals which showed further body weight losses from Days 1 to 3 post-exposure, all animals exhibited body weight gains during the remainder of the recovery period. 

 

Necropsy

No macroscopic abnormalities were detected amongst animals at necropsy.

 

Conclusion

No deaths occurred in a group of ten rats exposed to a mean achieved atmosphere concentration of 5.03mg/L for four hours. It was therefore considered that the acute inhalation median lethal concentration (4 hr LC50) of FAT40871/A TE, in the RccHanTM: WIST strain rat, was greater than 5.03mg/L.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LC50
Value:
5 030 mg/m³
Quality of whole database:
Study performed according to GLP Guideline and the score of the study is Klimisch 1.

Acute toxicity: via dermal route

Link to relevant study records
Reference
Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
The study was conducted between 29 October 2015 and 19 November 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Japanese MAFF, 2000
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1200 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
Animals and Animal Husbandry
Five male and five 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 females were nulliparous and non pregnant. After an acclimatization period of at least 5 days the 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 a cage card. At the start of the study the animals weighed at least 200 g, and were 8 to 12 weeks of age. The weight variation did not exceed ±20% of the mean weight for each sex.

The animals were housed in suspended solid floor polypropylene cages furnished with woodflakes. The initial two animals were housed individually throughout the study. The further group of eight animals (four male and four female) were housed individually during the 24 Hour exposure period and in groups of four, by sex, for the remainder of the study. 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. The diet, drinking water and bedding were routinely analyzed and were considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.

The temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give 12 hours continuous light and 12 hours darkness.

The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.


Justification
Rats are the preferred species of choice as historically used for safety evaluation studies and are specified in the appropriate test guidelines.
Type of coverage:
semiocclusive
Vehicle:
water
Details on dermal exposure:
Test Item Formulation and Experimental Preparation
For the purpose of the study the test item was weighed out according to each animal’s individual body weight and moistened with distilled water prior to application.
The absorption of the test item was not determined.

Procedure
On the day before treatment the back and flanks of each animal were clipped free of hair.

In the absence of data suggesting the test item was toxic, one male and one female rat were initially treated with the test item at a dose level of 2000 mg/kg.

The appropriate amount of test item, moistened with distilled water, was applied as evenly as possible to an area of shorn skin (approximately 10% of the total body surface area). A piece of surgical gauze was placed over the treatment area and semi occluded with a piece of self adhesive bandage. Shortly after dosing the dressings were examined to ensure that they were securely in place.
After the 24 Hour contact period the bandage was carefully removed and the treated skin and surrounding hair wiped with cotton wool moistened with distilled water to remove any residual test item.

As no mortalities were noted a further group of animals (four males and four females) was similarly treated with the test item at a dose level of 2000 mg/kg body weight to give a total of five males and five females. The animals were caged individually for the 24 Hour exposure period. After the 24 Hour contact period the bandages were carefully removed and the treated skin and surrounding hair wiped with cotton wool moistened with distilled water to remove any residual test item. These animals were returned to group housing for the remainder of the test period.
Duration of exposure:
24 hours
Doses:
2000 mg/kg
No. of animals per sex per dose:
Five males and five females
Control animals:
no
Details on study design:
The animals were observed for deaths or overt signs of toxicity 30 minutes, 1, 2 and 4 hours after dosing and subsequently once daily for 14 days.

After removal of the dressings and subsequently once daily for 14 days, the test sites were examined for evidence of primary irritation. Any other skin reactions, if present were also recorded.
Individual body weights were recorded prior to application of the test item on Day 0 and on Days 7 and 14.
At the end of the study the animals were killed by cervical dislocation. All animals were subjected to gross necropsy. This consisted of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.

Evaluation of Data
Data evaluations included the relationship, if any, between the exposure of the animal to the test item and the incidence and severity of all abnormalities including behavioral and clinical observations, gross lesions, body weight changes, mortality and any other toxicological effects.
Using the mortality data obtained, an estimate of the acute dermal median lethal dose (LD50) of the test item was made.
Statistics:
No data
Preliminary study:
No data
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
There were no deaths.
Clinical signs:
No signs of systemic toxicity were noted during the observation period.
Body weight:
Two females showed no gain in body weight during the first week with expected gain in body weight during the second week. One other female showed expected gain in body weight during the first week but body weight loss during the second week. The remaining seven animals showed expected gains in body weight over the study period.
Gross pathology:
No abnormalities were noted at necropsy.
Other findings:
Dermal Reactions
Dark orange colored staining, which prevented evaluation of erythema, was noted 1 day after dosing at the test sites of the two initial treated animals. Pale orange/yellow colored staining, not preventing evaluation of skin responses, was noted at the test sites the initial two treated animals 2 days after dosing and up to 11 Days after dosing.

Very slight to well-defined erythema and very slight edema were noted at the test site of the initial treated male with very slight erythema and very slight edema noted at the test site of initial treated female.

Pale orange/yellow colored staining, not preventing evaluation of skin responses, was noted at the test sites of the four additional treated males and four additional treated females during the study.

Very slight erythema, with or without very slight edema, was noted at the test sites of the four additional treated females up to 6 Days after dosing. There were no signs of dermal irritation noted at the test sites of the four additional treated males.

Individual Clinical Observations and Mortality Data

Dose Level mg/kg

Animal Number and Sex

Effects Noted After Dosing
(Hours)

Effects Noted During Period After Dosing
(Days)

½

1

2

4

1

2

3

4

5

6

7

8

9

10

11

12

13

14

2000

1-0

Male

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-0

Male

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-1

Male

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-2

Male

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-3

Male

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

2-0

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

4-0

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

4-1

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

4-2

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

4-3

Female

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0 = No mortality or signs of systemic toxicity

Individual Dermal Reactions - Males

Dose Level mg/kg

Animal Number and Sex

Observation

Effects Noted After Initiation of Exposure (Days)

1

2

3

4

5

6

7

8

9

10

11

12

13

14

2000

1-0

Male

Erythema

?s

2

1

1

1

1

1

0

0

0

0

0

0

0

Edema

1

1

1

1

1

1

1

0

0

0

0

0

0

0

Other

0

STA

STA

STA

STA

STA

STA

STA

STA

STA

0

0

0

0

3-0

Male

Erythema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Edema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Other

0

0

0

0

0

0

0

0

0

0

0

0

0

0

3-1

Male

Erythema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Edema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Other

STA

STA

STA

STA

0

0

0

0

0

0

0

0

0

0

3-2

Male

Erythema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Edema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Other

STA

STA

STA

STA

STA

0

0

0

0

0

0

0

0

0

3-3

Male

Erythema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Edema

0

0

0

0

0

0

0

0

0

0

0

0

0

0

Other

STA

STA

STA

STA

STA

0

0

0

0

0

0

0

0

0

0= No reactions                   

?s = Dark orange colored staining preventing evaluation of erythema                   

STA = Pale orange/yellow colored staining

Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The acute dermal median lethal dose (LD50) of the test item in the Wistar strain rat was found to be greater than 2000 mg/kg body weight.
Executive summary:

Introduction

The study was performed to assess the acute dermal toxicity of the test item in the Wistar strain rat.

 

Methods

Initially, two animals (one male and one female) were given a single, 24 hour, semi‑occluded dermal application of the test item to intact skin at a dose level of 2000 mg/kg body weight. Based on the results of the initial test, a further group of eight animals (four males and four females) was similarly treated. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

 

Results

Mortality. There were no deaths.

Clinical Observations. There were no signs of systemic toxicity.

Dermal Irritation. Very slight to well‑defined erythema and very slight edema were noted at the test sites of one male and all females. There were no signs of dermal irritation noted at the test sites of four males.

Body Weight. Two females showed no gain in body weight during the first week with expected gain in body weight during the second week. One other female showed expected gain in body weight during the first week but body weight loss during the second week. The remaining seven animals showed expected gains in body weight over the study period.

Necropsy. No abnormalities were noted at necropsy.

 

Conclusion

The acute dermal median lethal dose (LD50) of the test item in the Wistar strain rat was found to be greater than 2000 mg/kg body weight.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
2 000 mg/kg bw
Quality of whole database:
Study performed according to GLP Guideline and the score of the study is Klimisch 1.

Additional information

Oral:

The test substance was assessed for acute oral toxicity according to OECD Test Guideline 420 and EU Method B.1 bis using a fixed dose method. There were no deaths during the study. No signs of systemic toxicity were noted during the observation period and all animals showed expected gains in body weight over the observation period. No abnormalities were noted at necropsy. Hence, the acute oral LD50 dose of the test item in female Wistar rats was found to be greater than 2000 mg/kg body weight.

Inhalation:

Acute inhalation toxicity of FAT 40871/A TE was studied in rats according to OECD 403 and Method B2. Acute Inhalation Toxicity, 2014, of Commission Regulation (EC) No. 440/2008. Five rats/sex were nose-only exposed for 4 hours to 5.03 mg/L (Mean Achivied Atmosphere Concentration), the maximum attainable concentration, with a Mean Mass Median Aerodynamic Diameter of 3.6 µm (Inhable fraction % < 4

µm is 54.1%.

Rats were observed for a 14 days period.

No mortality was observed. Clinical observations included decreased respiratory rate, hunched posture, pilo-erection, generalized orange fur staining caused by the test item and wet fur. Occasinal instances of increased respiration were also noted. All animals recovered such that no significant observationswere apparent from Days 2 to 3 post exposure.

Body weight loss or no body weight gains was observed on the first day post exposure. With the exception of three female animals which showed further body weight losses from days 1 to 3 post-exposure, all animals exhibited body weught gains during the remainder of the recovery period.

No macroscopic findings at necropsy were detected. The LC 50 (4h nose-only exposure) of FAT 40871/A TE in rats is >5.03 mg/L.

Dermal:

The acute dermal toxicity of the test item was studied in the Wistar strain rats. A group of ten animals (five males and five females) was given a single, 24 hour, semi‑occluded dermal application of the test item to intact skin 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. Neither deaths nor any signs of systemic toxicity were recorded.

Very slight to well-defined erythema and very slight edema were noted at the test sites of one male and all females. There were no signs of dermal irritation noted at the test sites of four males.

Regardinf the body weight, two females showed no gain in body weight during the first week with expected gain in body weight during the second week. One other female showed expected gain in body weight during the first week but body weght loss during the second week. The remaining seven animals showed expected gains in body weight over the study period.

Justification for classification or non-classification

Based on the oral and dermal LD50 determined and in the inhalation test, the LC 50 determined, no classification is required.

Based on the above mentioned results the substance does not need to be classified according to CLP regulation (Regulation EC No.1272/2008) and DSD (Directive 67/548/EEC).