Registration Dossier

Administrative data

Description of key information

- Oral LD50: >2000 mg/kg bw (combined M+F) (OECD TG 420)
- Inhalation LD50: >12000 mg/m3 (route-to-route extrapolation from acute oral toxicity study)
- Dermal LD50: > 2000 mg/kg bw (OECD 402)

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:
Between 10th January 1995 and 9th February 1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted according to OECD guidelines and in compliance with GLP.
Reference:
Composition 0
Qualifier:
according to
Guideline:
EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)
Deviations:
no
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
GLP compliance:
yes (incl. certificate)
Test type:
fixed dose procedure
Limit test:
no
Test material information:
Composition 1
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
Equal numbers of healthy male and female CD rats of Sprague-Dawley origin (Hsd/Ola:SpragueDawley(CD)) were obtained from Harlan Olac Ltd. , Bicester, axon, England.
They were in the weight range of 92 to 126 g and approximately four to seven weeks of age prior to dosing (Day 1) in the main study. All the rats were acclimatised to the experimental environment for a minimum period of five days prior to the start of the main study.
The rats were allocated without conscious bias to cages within the treatment groups. They were housed in groups of up to five rats of the same sex in metal cages with wire mesh floors in Building R14 Room 6.
A standard laboratory rodent diet (Special Diet Services LAD 1) and drinking water were provided ad libitum. Access to food only was prevented overnight prior to and approximately 4 hours after dosing.
The batch(es) of diet used for the study was analysed for certain nutrients, possible contaminants and micro-organisms.
Results of routine physical and chemical examination of drinking water at source, as conducted. usually weekly by the supplier, are made available to Huntingdon Research Centre Ltd. (as quarterly summaries).
Animal room temperature was set to achieve a temperature of 22 ± 3°C. Relative humidity was not controlled but was anticipated to be in the range 30 - 70 % RH. Permanent daily recordings of these parameters were made and these are archived with other Department raw data. Any slight deviation in temperature and humidity that may have occurred was not considered to have affected the integrity or validity of the study. Air exchange was maintained at 10 to 15 air changes per hour and lighting controlled by means of a time switch to provide 12 hours of artificial light (0700 - 1900 hours) in each 24-hour period.
Each animal was identified by cage number and ear punching. Each cage was identified by a coloured label displaying the dose level, study schedule number, animal mark and the initials of the Study Director and Home Office licensee.
Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
The appropriate dose volume of the test substance was administered to each rat by oral gavage using a syringe and plastic catheter (8 choke).
The day of dosing was designated Day 1 .
Doses:
A preliminary study was carried out by dosing one female rat at 500 mg/kg bodyweight.

A group of ten rats (five males and five females) was treated at 2000 mg/kg bodyweight. Additional groups of ten rats (five males and five females) were similarly treated at 500 and 50 mg/kg bodyweight to further define the acute oral toxicity and establish the discriminating dosage of Salicynalva.
No. of animals per sex per dose:
A preliminary study was carried out by dosing one female rat at 500 mg/kg bodyweight.

A group of ten rats (five males and five females) was treated at 2000 mg/kg bodyweight.

Additional groups of ten rats (five males and five females) were similarly treated at 500 and 50 mg/kg bodyweight.
Control animals:
no
Details on study design:
TREATMENT PROCEDURE
Preliminary study
A preliminary study was carried out by dosing one female rat at 500 mg/kg bodyweight.

Main study
A group of ten rats (five males and five females) was treated at 2000 mg/kg bodyweight. Additional groups of ten rats (five males and five females) were similarly treated at 500 and 50 mg/kg bodyweight to further define the acute oral toxIcity and establish the LC50 of Salicynalva.

Control animal
No control animals were included in this study.

ADMINISTRATION OF TEST SUBSTANCE
The appropriate dose volume of the test substance was administered to each rat by oral gavage using a syringe and plastic catheter (8 choke).
The day of dosing was designated Day 1 .

OBSERVATIONS
Mortality
Cages of rats were checked at least twice daily for any mortalities.

Clinical signs
Animals were observed soon after dosing and at frequent intervals for the remainder of Day 1 (a minimum period of five hours). On subsequent days animals were observed once in the morning and again at the end of the experimental day (with the exception of Day 15 - morning only). This latter
observation was at approximately 16:30 hours on week days or 11:30 hours on Saturdays and Sundays.
The nature and severity of the clinical signs and time were recorded at each observation.
The animals in the preliminary study and those surviving treatment in the main study were observed for 7 and 14 days respectively after dosing.

Bodyweight
The bodyweight of each rat in the main study was recorded on Days 1 (prior to dosing), 2, 3, 4, 8 and 15 or at death. Individual weekly bodyweight change and group mean body weight data were calculated.

TERMINAL STUDIES
Termination
All surviving animals on the main study were killed on Day 15 by cervical dislocation.
Macroscopic pathology
All animals were subjected to a macroscopic examination which consisted of opening the abdominal and thoracic cavities. The macroscopic appearance of all tissues was recorded.
Preliminary study:
A preliminary study was carried out by dosing one female rat at 500 mg/kg bodyweight. This dosage was selected to give an initial indication of test substance toxicity. A minimal systemic response to treatment (piloerection only) was seen within five minutes of dosing.
There were no other clinical signs and recovery was complete by Day 7. This animal achieved a satisfactory bodyweight gain for a study of this duration.
Macroscopic examination on Day 8 revealed a thickening of the glandular region of the stomach but otherwise no abnormalities were observed.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Sex:
male/female
Dose descriptor:
discriminating dose
Effect level:
>= 50 mg/kg bw
Based on:
test mat.
Mortality:
One male at 500 mg/kg and two males and two females at 2000 mg/kg died during the study. All deaths occurred between 24 and 48 hours of dosing.
Clinical signs:
Piloerection was observed in all rats within five minutes of dosing and persisted in the majority of animals throughout the remainder of Day 1. This sign was accompanied on Day 1 and/or at later intervals by abnormal body carriage (hunched posture) in all rats at 500 and 2000 mg/kg and
abnormal gait (waddling) in all rats at 500 and 2000 mg/kg, lethargy in four males and four females at 2000 mg/kg; decreased respiratory rate in one male at 500 mg/kg and one male and one female at 2000 mg/kg; pallor of the extremities in all rats at 500 and 2000 mg/kg; increased urine production in one male and one female at 2000 mg/kg, increased salivation in two males at 500 mg/kg and two females at 2000 mg/kg;
walking on toes in three males at 500 mg/kg and two males at 2000 mg/kg, unsteadiness in one male at 500 mg/kg and three males and three females at 2000 mg/kg, hair loss in three males and one female at 2000 mg/kg.

Recovery of surviving rats was complete within two hours of dosing (females, 50 mg/kg), Day 3 (males, 50 mg/kg), Day 5 (500 mg/kg) or by Day 9 (2000 mg/kg) with the exception of hair loss which had resolved in the majority of surviving rats dosed at 2000 mg/kg by Day 10 or Day 11 but persisted in one surviving rat dosed at 2000 mg/kg through to the end of the observation period (Day 15).
Body weight:
Slightly low bodyweight gains were recorded on Day 8 for three males and one female dosed at 2000 mg/kg bodyweight; these animals achieved the anticipated bodyweight gains on Day 15. All other surviving animals achieved satisfactory bodyweight gains throughout the study.

Slight bodyweight losses were recorded for some decedents.
Gross pathology:
Macroscopic examination of decedents revealed:

Male - 500 mg/kg
Congestion (characterised by dark tissue) in the lungs, liver, spleen and kidneys. In addition, congestion (characterised by dark tissue and fluid contents) was seen in the stomach and small intestine.

Males and females - 2000 mg/kg
Congestion (characterised by dark tissue) in the thymus, heart, lungs, liver and kidneys with splenic pallor and atrophy. In addition, congestion (identified by gaseous distension, fluid contents, thinning/thickening/bleaching and prominent blood vessels) was seen in the stomach and along the alimentary tract.

No macroscopic abnormalities were observed for animals killed on Day 15.
Interpretation of results:
not classified
Remarks:
Migrated information based on LD50 Criteria used for interpretation of results: EU
Conclusions:
The LD50 of the substance when administered orally to rats was established to be >= 2000 mg/kg bodyweight and the discriminating dose >= 50 mg/kg.
Executive summary:

A study was performed to assess the acute oral toxicity of Salicynalva to the rat according to the fixed dose procedure, similar to OECD TG 420. Groups of ten fasted rats (five males and five females) were given a single dose by oral gavage of the test substance, as supplied, at dosages of 500 and 2000 mg/kg bodyweight. A further group was treated using the test substance formulated in distilled water at a dosage of 50 mg/kg body weight. All animals surviving treatment were killed and examined macroscopically on Day 15, the end of the observation period.

One male at 500 mg/kg and two males and two females at 2000 mg/kg died during the study. All deaths occurred between 24 and 48 hours of dosing. A slight body weight loss was recorded for some decedents. Macroscopic examination revealed congestion in the majority of major organs and tissues. Clinical signs of reaction to treatment were confined to piloerection in rats dosed at 50 mg/kg. Among rats treated at 500 and 2000 mg/kg, piloerection, abnormal body carriage, abnormal gait, decreased respiratory rate, pallor of the extremities, increased salivation, walking on toes and unsteadiness were observed. In addition, lethargy, increased urine production and hair loss were evident among rats dosed at 2000 mg/kg only. Recovery of surviving rats was complete in all instances by Day 9 with the exception of hair loss which had resolved in the majority of surviving rats dosed at 2000 mg/kg by Day 10 or Day 11 but persisted in one surviving rat dosed at 2000 mg/kg through to the end of the observation period (Day 15). Slightly low body weight gains were recorded on Day 8 for three males and one female dosed at 2000 mg/kg bodyweight; these animals achieved the anticipated bodyweight gains on Day 15. All other surviving animals achieved satisfactory bodyweight gains throughout the study. No abnormalities were recorded at the macroscopic examination on Day 15. The LD50 of

Salicynalva when administered orally to rats was established to be > 2000 mg/kg bodyweight and the discriminating dose >= 50 mg/kg because one animal died at 500 mg/kg and evident toxicity was seen at that dose.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The one study available is of adequate quality.

Acute toxicity: via inhalation route

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The information available is of adequate quality.

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:
Between 17 January 1995 and 31 January 1995
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted according to OECD guidelines and in compliance with GLP.
Reference:
Composition 0
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
standard acute method
Limit test:
yes
Test material information:
Composition 1
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
Equal numbers of healthy male and female CD rats of Sprague-Dawley origin (Hsd/Ola:SpragueDawley(CD)) were obtained from Harlan Olac Ltd., Bicester, Oxon, England.
They were in the weight range of 215 to 241 g and approximately seven to ten weeks of age prior to dosing (Day 1). All the rats were acclimatised to the experimental environment for a period of five days prior to the start of the study.
The rats were allocated without conscious bias to cages within the treatment groups. They were housed individually in metal cages with wire mesh floors in Building R14 Room 6.
A standard laboratory rodent diet (Biosure LAD 1) and drinking water were provided ad libitum.
Each batch of diet used for the study was analysed for certain nutrients, possible contaminants and microorganisms .
Results of routine physical and chemical examination of drinking water at source as conducted, usually weekly by the supplier, are made available to Huntingdon Research Centre Ltd. as quarterly summaries.
Animal room temperature was set to achieve a temperature of 22 ± 3°C. Relative humidity was not controlled but was anticipated to be in the range 30 - 70%. Permanent daily recordings of these parameters were made and these are archived with other Department raw data. Any slight deviation in temperature and humidity that may have occurred had no impact on the study in the opinion of the Study Director. Air exchange was maintained at 10 to 15 air changes per hour and lighting controlled by means of a time switch to provide 12 hours of artificial light (0700 - 1900 hours) in each 24-hour period.
Each animal was identified by cage number and ear punching. Each cage was identified by a coloured label displaying the dose level, study schedule number, animal mark and the initials of the Study Director and Home Office license.
Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
A group of ten rats (five males and five females) was treated at 2.0 g/kg bodyweight. The test substance was applied by spreading it evenly over the prepared skin
Duration of exposure:
24 hours
Doses:
2.0 g/kg bodyweight.
No. of animals per sex per dose:
five males
five females
Control animals:
not required
Details on study design:
TREATMENT PROCEDURE
A group of ten rats (five males and five females) was treated at 2.0 g/kg bodyweight.
One day prior to treatment, hair was removed from the dorso-lumbar region of each rat with electric clippers exposing an area equivalent to approximately 10% of the total body surface.
The test substance was applied by spreading it evenly over the prepared skin. The treated area (approximately 50 mm x 50 mm) was then promptly covered with gauze which was held in place with a non-irritative dressing encircled firmly around the trunk.
At the end of the 24 hours exposure period, the dressings were carefully removed and the treated area of skin was washed with warm (30° to 40°C) water and blotted dry with absorbent paper.
The day of dosing was designated Day 1.

Control animals
No control animals were included in this study.

OBSERVATIONS
Mortality
Cages of rats were checked at least twice daily for any mortalities.
Clinical signs
Animals were observed soon after dosing and at frequent intervals for the remainder of Day 1 (a period of six hours). On subsequent days animals were observed once in the morning and again at the end of the experimental day (with the exception of Day 15 - morning only). This latter observation was at approximately 16: 30 hours on week days or 11: 30 hours on Saturdays and Sundays. The nature and severity of the clinical signs and time were recorded at each observation.
All animals were observed for 14 days after dosing.

Dermal responses
Local dermal irritation at the treatment site was assessed daily using the following numerical system:

Erythema and Eschar Formation Value

No erythema 0
Slight erythema 1
Well-defined erythema 2
Moderate to severe erythema 3
Severe erythema (beet redness) to slight eschar formation (injuries in depth) 4

Oedema Formation

No oedema 0
Slight oedema 1
Well-defined oedema (edges of area well-defined by definite raising) 2
Moderate oedema (raised approximately 1 millimetre) 3
Severe oedema (raised more than 1 millimetre and extending beyond the area of exposure) 4

Bodyweight
Individual bodyweights were recorded on Days 1 (prior to dosing), 8 and 15. Individual weekly bodyweight changes were calculated.

Macroscopic examination
All animals were killed on Day 15 by cervical dislocation and were subjected to a macroscopic examination which consisted of opening the abdominal and thoracic cavities. The macroscopIc appearance of all examined tissues was recorded.

Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
There were no deaths following a single dermal application of Salicynalva to a group of ten rats (five males and five females) at a dosage of 2.0 g/kg bodyweight.

Clinical signs:
There were no signs of systemic reaction to treatment.
Body weight:
Slightly low bodyweight gains were recorded for three males and three females on Day 8 and in one male and two females on Day 15. In addition, two females showed a slight bodyweight loss on Day 8 and one female showed no body weight gain at Day 15. Such changes are not unusual and are commonly associated with the treatment procedure. Remaining males achieved anticipated bodyweight gains throughout the study.
Gross pathology:
No macroscopic abnormalities were observed for animals killed on Day 15.
Other findings:
With the exception of a residual black staining from the test substance on the treatment sites of some animals there were no other dermal changes observed in any animal throughout the dose period (scores of zero for erythema and oedema were recorded for all animals).
Interpretation of results:
not classified
Remarks:
Migrated information LD50 >=2000 mg/kg bw Criteria used for interpretation of results: EU
Conclusions:
The acute lethal dermal dose to rats of Salicynalva was found to be >= 2000 mg/kg bodyweight.
Executive summary:

A study was performed to assess the acute dermal toxicity of Salicynalva to the rat. The method followed was that described in EEC Methods for the determination of toxicity, Annex to Directive 92/69/EEC (OJ No. L383A, 29. 12. 92), Part B, Method B.3. Acute toxicity (dermal) and OECD Guideline for Testing of Chemicals No. 402 "Acute Dermal Toxicity". Adopted: 24 February 1987. A group of ten rats (five males and five females) was given a single occlusive dermal application of the test substance, as supplied by the sponsor and administered at a dosage of 2.0 g/kg bodyweight, no vehicle was used. All animals were killed and examined macroscopically on Day 15, the end of the observation period. There were no deaths and no signs of systemic reaction to treatment. With the exception of a residual black staining from the test substance on the treatment sites of some animals, there were no other dermal changes observed in any animal throughout the dose period (scores of zero for erythema and oedema were recorded for all animals). Slightly low bodyweight gains were recorded for three males and three females on Day 8, with a similar trend noted for one male and two females on Day 15. In addition, two females showed a slight bodyweight loss on Day 8 and one female showed no bodyweight gain on Day 15. The remaining animals achieved anticipated bodyweight gains throughout the study. No abnormalities were recorded at the macroscopic examination on Day 15.

The acute lethal dermal dose to rats of Salicynalva was found to be greater than 2000 mg/kg bodyweight.

 

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The one study available is of adequate quality.

Additional information

Acute oral toxicity: A study was performed to assess the acute oral toxicity of Salicynalva to the rat according to the fixed dose procedure, similar to OECD TG 420. Groups of ten fasted rats (five males and five females) were given a single dose by oral gavage of the test substance, as supplied, at dosages of 500 and 2000 mg/kg bodyweight. A further group was treated using the test substance formulated in distilled water at a dosage of 50 mg/kg body weight. All animals surviving treatment were killed and examined macroscopically on Day 15, the end of the observation period. One male at 500 mg/kg and two males and two females at 2000 mg/kg died during the study. All deaths occurred between 24 and 48 hours of dosing. A slight body weight loss was recorded for some decedents. Macroscopic examination revealed congestion in the majority of major organs and tissues. Clinical signs of reaction to treatment were confined to piloerection in rats dosed at 50 mg/kg. Among rats treated at 500 and 2000 mg/kg, piloerection, abnormal body carriage, abnormal gait, decreased respiratory rate, pallor of the extremities, increased salivation, walking on toes and unsteadiness were observed. In addition, lethargy, increased urine production and hair loss were evident among rats dosed at 2000 mg/kg only. Recovery of surviving rats was complete in all instances by Day 9 with the exception of hair loss which had resolved in the majority of surviving rats dosed at 2000 mg/kg by Day 10 or Day 11 but persisted in one surviving rat dosed at 2000 mg/kg through to the end of the observation period (Day 15). Slightly low body weight gains were recorded on Day 8 for three males and one female dosed at 2000 mg/kg bodyweight; these animals achieved the anticipated bodyweight gains on Day 15. All other surviving animals achieved satisfactory bodyweight gains throughout the study. No abnormalities were recorded at the macroscopic examination on Day 15. The LD50 of Salicynalva when administered orally to rats was established to be > 2000 mg/kg bodyweight and the discriminating dose >= 50 mg/kg bw because one animal died at 500 mg/kg an evident toxicity was seen at that dose.

An acute inhalation toxicity study is not needed for substances for which acute oral and dermal toxicity values are available. Using route to route extrapolation the inhalation toxicity can be derived as follows: an oral LD50 of > 2000 mg/kg bw can be roughly converted into 120000 mg/per person by multiplying it with a person’s weight: (2000 x 60). An inhalation volume for one person during 4 h (standard exposure time in the OECD TG for acute inhalation is 5m3 (assuming 10m3/8h for workers). This means that an LC50 concentration in 1 m3 and 4 hours exposure is 24000 mg/m3 (120000 mg/5m3). Taking into account that during that the absorption via the inhalation route is twice the oral route the LC50, the LC50 for inhalation could be 12000 mg/m3. The maximum saturated vapour pressure for the substance is 455 mg/m3 (6.4 Pa (Vap Pr. Salicynalva) x 173(MW) / 8.3 (R, gas constant) x 293 (°K)). This means that Salicynalva cannot reach a concentration higher than 455 mg/m3. Therefore an LC50 for inhalation cannot be reached and no classification and labeling is needed for the acute inhalation route.

Acute dermal toxicity: A study was performed to assess the acute dermal toxicity of Salicynalva to the rat. The method followed was that described in OECD Guideline for Testing of Chemicals No. 402 "Acute Dermal Toxicity". Adopted: 24 February 1987.A group of ten rats (five males and five females) was given a single occlusive dermal application of the test substance, as supplied by the sponsor and administered at a dosage of 2.0 g/kg bodyweight, no vehicle was used. All animals were killed and examined macroscopically on Day 15, the end of the observation period.There were no deaths and no signs of systemic reaction to treatment. With the exception of a residual black staining from the test substance on the treatment sites of some animals, there were no other dermal changes observed in any animal throughout the dose period (scores of zero for erythema and oedema were recorded for all animals). Slightly low bodyweight gains were recorded for three males and three females on Day 8, with a similar trend noted for one male and two females on Day 15. In addition, two females showed a slight bodyweight loss on Day 8 and one female showed no bodyweight gain on Day 15. The remaining animals achieved anticipated bodyweight gains throughout the study. No abnormalities were recorded at the macroscopic examination on Day 15. The acute lethal dermal dose to rats of Salicynalva was found to be greater than 2000 mg/kg bodyweight.


Justification for selection of acute toxicity – oral endpoint
One acute oral toxicity study is available, which is performed according to OECD guidelines and under GLP conditions. This study is adequate for covering this endpoint.

Justification for selection of acute toxicity – inhalation endpoint
The acute inhalation toxicity is derived using route to route extrapolation from the acute toxicity results, using 100% absorption. This assessment is considered to be sufficient adequate for covering this endpoint.

Justification for selection of acute toxicity – dermal endpoint
One acute dermal toxicity study is available, which is performed according to OECD guidelines and under GLP conditions. This study is adequate for covering this endpoint.

Justification for classification or non-classification

As Salicynalva has an oral LD50 value, dermal LD50 value and inhalatory LC50 value of >2000 mg/kg, >2000 mg/kg and >12000 mg/m3 respectively it does not have to be classified as acutely toxic by the oral, dermal and inhalation route according to the criteria outlined in Annex I of Regulation (EC) 1272/2008 (CLP). According to the criteria outlined in Annex VI of EU Directive 67/548/EEC (DSD) it must be classified as harmful for the oral route (Xn, R22) based on the discriminating dose of 50 mg/kg bw. This criterion is no longer applicable in CLP. Under REACH the CLP is leading and therefore the absence of classification and labelling for acute toxicity will be taken forward to the risk characterisation. It is noted that Salicynalva has a harmonized classification as H302 in table 3.1 and R22 in table 3.2 of Annex VI of CLP for this endpoint. A request is in preparation to remove the H302 classification from table 3.1 of the Annex VI entry.