Registration Dossier

Administrative data

Description of key information

Acute toxicity testing has been conducted via the oral, dermal and inhalation routes.
Acute oral toxicity: The acute median lethal dose (LD50) to rats of D.I.X.T. was estimated to be estimated to be 1.5 g/kg (1500 mg/kg) bodweight (for males and females combined).
Acute dermal toxicity: The acute lethal dermal dose to rats of D.I.X.T was found to be greater than 2.0 g/kg (>2000 mg/kg) bodyweight.
Acute inhalation toxicity: AS100 is not sufficiently volatile to warrant an inhalation toxicity. An inhalation study is

enclosed but is for information only as the mechanical needs of aspiration and low vapour pressure warrant the results void.

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 undertaken between 16 January and 13 February 1986.
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study conducted in accordance with generally accepted scientific principles, possibly with incomplete or methodological deficiencies, which do not affect the quality of relevant results.
Qualifier:
equivalent or similar to
Guideline:
EU Method B.1 (Acute Toxicity (Oral))
Deviations:
no
Qualifier:
according to
Guideline:
EPA OTS 798.1175 (Acute Oral Toxicity)
Deviations:
no
GLP compliance:
not specified
Test type:
standard acute method
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Obtained from Charles River U.K. Limited, Margate, Kent, England.
- Age at study initiation: Four to six weeks of age.
- Weight at study initiation: 98 to 149 g.
- Fasting period before study: Access to food was prevented overnight prior to and approximately 4 hours after dosing.
- Housing: Rats were housed in groups by sex in metal cages with mire mesh floors.
- Diet (e.g. ad libitum): A standard laboratory rodent diet (Labsure LAD 1) was provided ad libitum.
- Water (e.g. ad libitum): Water was provided ad libitum.
- Acclimation period: Minimum period of 6 days prior to the start of the main study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): The mean daily minimum and maximum temperatures were 20°C and 22°C.
- Humidity (%): The mean daily relative humidity value was 60%.
- Air changes (per hr): The rate of air exchange was maintained at approximately 15 air changes/hour.
- Photoperiod (hrs dark / hrs light): Lighting was controlled by means of a time switch to give 12 hours artificial light in each 24 hour period.
Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
MAXIMUM DOSE VOLUME APPLIED: 4.92 ml/kg


Doses:
Main study: 1.0, 1.6, 2.5, 4.0 and 6.4 g/kg
No. of animals per sex per dose:
5 males and 5 females per dose.
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days after dosing.

- Frequency of observations and weighing: Animals were observed soon after dosing, then at frequent intervals for the remainder of Day 1. On subsequent days the animals were observed once in the morning and once at the end of the experimental day. Clinical signs were recorded at each observation.
Individual bodyweights of rats on Days 1 (day of dosing), 8, 15 and at death were recorded.

- Necropsy of survivors performed: Surviving animals on the main study were killed on Day 15 by cervical dislocation. All animals that died during the study and those killed on Day 15 were subjected to a macroscopic post mortem examination which consisted of opening the abdominal and thoracic cavities. The macroscopic appearance of abnormal organs when present was recorded.

- Other examinations performed: The following were recorded during the main study:-
- Approximate time of death of individual rats
- The nature, severity, approximate time of onset and duration of each toxic sign.

Statistics:
The acute median lethal oral dose (LD50) to male and female rats was calculated using the method of:
Finney (1971) Probit Analysis (3rd Edition) Cambridge University Press.

Separate LD so values for males and females were estimated by undertaking probit analysis on the mortality data by fitting two parallel lines on the data (males only and females only) using the technique described by Finney (1978, Statistical Method in Biological Assay, 3rd Edition, Charles Griffin, London). A chi-squared test was carried out to check that the data did not contain any evidence for non-parallelism.
Preliminary study:
The results of the preliminary study indicated that the LD50 was between 2.5 - 5.0 g/kg bodweight (see Table 1 - attached background material).
Sex:
male/female
Dose descriptor:
LD50
Effect level:
1 500 mg/kg bw
Based on:
test mat.
95% CL:
400 - 2 400
Mortality:
Mortalities occurred amongst rats dosed at 1.0 g/kg and above within two and 71 hours of dosing (see table 2 - attached background material).

Sex: Male; Dose: 1.0 g/kg; Number of deaths in group of 5; 1
Sex: Male; Dose: 1.6 g/kg; Number of deaths in group of 5; 4
Sex: Male; Dose: 2.5 g/kg; Number of deaths in group of 5; 2
Sex: Male; Dose: 4.0 g/kg; Number of deaths in group of 5; 5
Sex: Male; Dose: 6.4 g/kg; Number of deaths in group of 5; 5
Sex: Female; Dose: 1.0 g/kg; Number of deaths in group of 5; 4
Sex: Female; Dose: 1.6 g/kg; Number of deaths in group of 5; 0
Sex: Female; Dose: 2.5 g/kg; Number of deaths in group of 5; 3
Sex: Female; Dose: 4.0 g/kg; Number of deaths in group of 5; 4
Sex: Female; Dose: 6.4 g/kg; Number of deaths in group of 5; 4
Clinical signs:
Signs of reaction to treatment observed shortly after dosing amongst rats of all groups were pilo-erection, abnormal body carriage (hunched posture), abnormal gait (waddling), lethargy, decreased respiratory rate, pallor of the extremities and diuresis. These were accompanied by:
ptosis in all rats dosed at 1.6 g/kg and above,
increased salivation in all rats dosed at 1.6 g/kg,
abdominal distension in one male rat dosed at 2.5 g/kg,
a coma-like condition in one male rat dosed at 2.5 g/kg.

Recovery, as judged by external appearance and behaviour, was apparently complete by Days 7 to 11.

See table 4 (attached background material) for results.
Body weight:
Bodyweight losses were recorded for all but one of the rats that died (Table 5 - attached background material).
Bodyweighy gains were recorded for all surviving rats on Days 8 and 15 (Table 5 - attached background material).
Gross pathology:
Animals that died during study:
Autopsy revealed pallor of the liver, spleen and kidneys and congestion of the blood vessels of the small and large intestine amongst rats that died. Perforations of the duodenum were seen amongst rats at all dose levels. A light yellow fluid was found in the peritoneal cavity of three rats dosed at 4.0 g/kg and one female dosed at 6.4 g/kg ( seeTable 3 – attached background material).

Terminal autopsy:
Terminal autopsy revealed perforations in the duodenum in a small number of animals. Enlargement of the spleen was seen in one female rat dosed at 2.5 g/kg (see Table 6 - attached background material).

Estimation of the LD50 value

The acute median lethal oral dose of D.I.X.T. and its 957% confidence limits were estimated to be:

Males and females combined: 1.5 (0.40 to 2.4) g/kg bodyweight.

The slope of the probit line was 1.9 with a standard error of 0.71 using log. transformation of dose. The heterogeneity factor was not significant.

 

When probit analysis was carried out by fitting two parallel lines the values were:

Males only: 1.3 (0.22 to 2.5) g/kg bodyweight.

Females only: 1.8 (0.50 to 3.5) g/kg bodyweight.

The slope of the parallel probit lines was 1.9 with a standard error of 0.71 using log. transformation of dose. The heterogeneity factor was significant.

 

The mortality response curves are presented in Figure 1 (see attached background material).

 

The chi-squared test for parallelism gave no evidence of non-parallelism.

Interpretation of results:
harmful
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The acute median lethal dose (LD50) and their 95% confidence limits to rats of D.I.X.T. were estimated to be:
Males and females combined: 1.5 (0.40 to 2.4) g/kg bodyweight.
Males only: 1.3 (0.22 to 2.5) g/kg bodyweight.
Females only: 1.8 (0.50 to 3.5) g/kg bodyweight.
Executive summary:

Introduction:

The experimental procedure used followed the Testing Guidelines as described in the Federal Register, Vol 50, No.188, Part II of 27 September 1985, Section 798.1175 - Acute Oral Toxicity.

Method:

A preliminary test was carried out to establish a dosing regimen using groups of two males and two female rats at three dose levels of 1.0, 2.5 and 5.0 g/kg bodyweight.

Based on the findings of the preliminary test, further groups of rats (5 male and 5 female per dose group) were dosed at dose levels of 1.0, 1.6, 2.5, 4.0 and 6.4 g/kg bodyweight. The appropriate dose volume of the test substance was administered to each rat using a syringe and plastic catheter or a glass microlitre syringe and metal cannula.

Animals were observed soon after dosing, then at frequent intervals for the remainder of Day 1. On subsequent days the animals were observed once in the morning and once at the end of the experimental day. Clinical signs were recorded at each observation.

Individual bodyweights of rats on Days 1 (day of dosing), 8, 15 and at death were recorded.

Surviving animals on the main study were killed on Day 15 by cervical dislocation. All animals that died during the study and those killed on Day 15 were subjected to a macroscopic post mortem examination which consisted of opening the abdominal and thoracic cavities. The macroscopic appearance of abnormal organs when present was recorded.

Results:

The results of the preliminary study indicated that the acute median lethal oral dose of D.I.X.T was between 2.5 and 5.0 g/kg bodyweight.

Main test:

Mortalities:

Mortalities occured amonhst rats dosed at 1.0 g/kg and above within two and 71 hours of dosing. Autopsy revealed pallor of the liver, spleen and kidneys and congestion of the blood vessels of the small and large intestine amongst rats that died. Perforations of the duodenum were seen amongst rats at all dose levels. A light yellow fluid was found in the peritoneal cavity of three rats dosed at 4.0 g/kg and one female dosed at 6.4 g/kg. Bodyweighy losses were recorded for all but one of the rats that died.

Clinical signs:

Signs of reaction to treatment observed shortly after dosing amongst rats of all groups were pilo-erection, abnormal body carriage (hunched posture), abnormal gait (waddling), lethargy, decreased respiratory rate, pallor of the extremities and diuresis. These were accompanied by:

- ptosis in all rats dosed at 1.6 g/kg and above,

- increased salivation in all rats dosed at 1.6 g/kg,

- abdominal distension in one male rat dosed at 2.5 g/kg,

- a coma-like condition in one male rat dosed at 2.5 g/kg.

Recovery, as judged by external appearance and behaviour, was apparently complete by Days 7 to 11.

Bodyweight:

Bodyweight gains were recorded for all surviving rats on Days 8 and 15.

Terminal autopsy:

Terminal autopsy revealed perforations in the duodenum in a small number of animals. Enlargement of the spleen was seen in one female rat dosed at 2.5 g/kg.

Conclusion:

The acute median lethal dose (LD50) and their 95% confidence limits to rats of D.I.X.T. were estimated to be:

Males and females combined: 1.5 (0.40 to 2.4) g/kg bodyweight.

Males only: 1.3 (0.22 to 2.5) g/kg bodyweight.

Females only: 1.8 (0.50 to 3.5) g/kg bodyweight.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LD50
Value:
1 500 mg/kg bw
Quality of whole database:
Study conducted in accordance with generally accepted scientific principles, possibly with incomplete or methodological deficiencies, which do not affect the quality of relevant results.

Acute toxicity: via inhalation route

Link to relevant study records
Reference
Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
disregarded due to major methodological deficiencies
Study period:
The study was performed between 14 February 2011 and 15 March 2011.
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of relevant results.
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.2 (Acute Toxicity (Inhalation))
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method
Limit test:
yes
Species:
rat
Strain:
other: RccHanTM : WIST
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories UK
Ltd, Oxon, UK
- Age at study initiation: 8 to 12 weeks
- Weight at study initiation: 200g to 350 g
- Fasting period before study: 2 hours
- Housing: 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.
- Diet: With the exception of the exposure period, free access to food (Harlan 2014C Rodent Diet, Harlan Laboratories UK Ltd, Oxon, UK) was allowed throughout the study.
- Water (e.g. ad libitum): With the exception of the exposure period, free access to mains drinking water was allowed throughout the study
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 25 deg C
- Humidity (%): between 30 and 70%
- Air changes (per hr): at least 15
- Photoperiod (hrs dark / hrs light): the
lighting was controlled to give twelve hours continuous light and twelve hours darkness.

IN-LIFE DATES: From: Day 0 To: End of Study
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: The test item was aerosolised using a glass concentric jet nebuliser (Radleys, Saffron Walden, Essex, UK) located at the top of the exposure chamber.
- Exposure chamber volume: The cylindrical exposure chamber had a volume of approximately 30 litres (dimensions: 28 cm diameter x 50 cm high).
- Exposure Chamber Oxygen Concentration: Oxygen levels within the exposure chamber were measured by an electronic oxygen analyser (Servomex (UK) Ltd, Crowborough, East Sussex) located in a sampling port in the animals breathing zone during each exposure period. The test atmosphere was generated to contain at least 19% oxygen.

- Source and rate of air: 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 nebuliser.
air flow (L/MIN): 60


The chamber was maintained under negative pressure, the temperature and humidity were measured by an electronic thermometer/humidity meter. The temperature and humidity were recorded every 30 minutes throughout the four hour exposure period.

TEST ATMOSPHERE
Exposure Chamber Atmosphere Concentration
Prior to the start of the study, the non-volatile component of the test item was determined by adding a small, known amount of test item to glass fibre filters and recording their weights. The filters were then dried in a desiccator between 19 and 20°C for approximately 24 hours and then weighed again. The difference in the two weights was taken as the volatile content of the test item and the non-volatile component was calculated as a percentage. The mean non-volatile component of the batch used during the study was found to be 98.48% (n=10).

The test atmosphere was sampled at regular intervals during the exposure period. A weighed glass fibre filter was placed in a filter holder and temporarily sealed in a vacant port of the exposure chamber in the animals’ breathing zone. A known quantity of the exposure chamber atmosphere was drawn through the filter using a vacuum pump. After sampling, the filter was dried, under similar conditions as those previously described, and weighed again 24 hours later. The difference in the pre and post sampling weights, divided by the volume of atmosphere sampled, was the chamber concentration in terms of non-volatile component.

Based on the results of the preliminary work, these figures were adjusted to obtain a true figure for the test item concentration in the chamber. The nominal chamber concentration was calculated by dividing the mass of test item disseminated into the chamber by the total volume of air that flowed through the chamber during the exposure.


TEST ATMOSPHERE (if not tabulated)
- Particle size distribution:
The particle size of the generated atmosphere inside the exposure chamber was
determined three times during each exposure period using a Marple Personal Cascade
Impactor (Westech IS Ltd, Beds., UK).
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric):
Group 1:
Mean Mass Median Aerodynamic Diameter (MMAD) = 1.64 μm
Geometric Standard Deviation (GSD) = 2.33
Group 2:
Mean Mass Median Aerodynamic Diameter (MMAD) = 2.40 μm
Geometric Standard Deviation (GSD) = 1.98
CLASS METHOD (if applicable)
- Rationale for the selection of the starting concentration:
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
Dose Group 1 –2.11 mg/L

Dose Group 2 –5.05 mg/L
No. of animals per sex per dose:
Sighting exposure: 2 rats one male and 1 female were exposed to an atmosphere concentartion of 2.06 mg/L for 4 hours.

5 males and 5 females were subjected to a single exposure to the test item for a period of 4 hours. A concentration of 2.11 mg/L was achieved for the first exposure, a target concentration of 5 mg/L was utilised for the second exposure.
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days

- Frequency of observations and weighing: Individual bodyweights were recorded on arrival, prior to treatment on the day of
exposure and on Days 1, 3, 7 and 14 or at death.

- Necropsy of survivors performed: yes

At the end of the fourteen day observation period, the surviving animals were killed by intravenous overdose of sodium pentobarbitone. All animals, including those that died during the study, 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.

- Other examinations performed: clinical signs, body weight,organ weights, histopathology, 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 up to fourteen days. Any deaths or evidence of overt toxicity were recorded at each observation.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 5.05 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Sex:
male
Dose descriptor:
LC50
Effect level:
> 5.05 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Sex:
female
Dose descriptor:
LC50
Effect level:
4.56 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
Group Number Mean Achieved Atmosphere Concentration (mg/L) Male Deaths Female Deaths Total
1 2.11 1/5 0/5 1/10
2 5.05 2/5 3/5 5/10
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. These observations a
Body weight:
Group 1 – All surviving animals exhibited a bodyweight loss on Day 1, one surviving male and three females exhibited a bodyweight loss or showed no bodyweight gain from Days 1 to 3. Weight gains were noted in all surviving animals for the remainder of the recovery period.

Group 2 – All surviving animals exhibited a bodyweight loss on Day 1, two surviving male animals and all surviving females exhibited a bodyweight loss or exhibited no bodyweight gain from Days 1 to 3. Reasonable bodyweight development was noted in all surviving animals during the remainder of the recovery period.
Gross pathology:
With the exception of one instance of dark patches on the lungs no macroscopic abnormalities were detected at necropsy for animals that survived until the end of the recovery period.

Haemorrhagic or abnormally dark lungs were detected at necropsy amongst animals that died during the course of the study. Due to lung observations noted in the animals it is considered that the deaths noted during this study may have been attributable to local toxicity caused by the test item.

Exposure Chamber Concentration

The actual concentration of the test item was measured seventeen times during each exposure period. The mean values obtained were:

Group number

Atmospheric concentration

Mean Achieved (mg/L)

Standard deviation

Nominal (mg/L)

1

2.11

0.05

5.72

2

5.05

0.16

20.9

Chamber flow rate was maintained at 60 L/min providing 120 air changes per hour.

Particle Size Distribution

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

Group number

Mean Achieved Atmosphere Concentration (mg/L)

Mean Mass Median Aerodynamic Diameter (µm)

Inhalable Fraction (% < 4 µm)

Geometric

Standard

Deviation

1

2.11

1.64

85.6

2.33

2

5.05

2.40

77.5

1.98

Interpretation of results:
Toxicity Category IV
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The acute inhalation median lethal concentrations (LC50) (it was not possible to calculate 95% confidence limits as only two groups were exposed) of AS 100 (Di-isopropyl xanthogen polysulphide) in the RccHanTM : WIST strain rat, were calculated to be:
All animals : >5.05 mg/L
Males only : >5.05 mg/L
Females only : 4.56 mg/L
Executive summary:

Introduction.

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

Methods.

Groups of ten RccHanTM : WIST strain rats (five males and five females) were exposed to an aerosol atmosphere. The animals were exposed for four hours using a nose only exposure system, followed by a fourteen day observation period.

Results:

The mena achieved atmosphere concentrations were as follows:

Group number

Atmospheric concentration

Mean Achieved (mg/L)

Standard deviation

Nominal (mg/L)

1

2.11

0.05

5.72

2

5.05

0.16

20.9

The characteristics of the achieved atmospheres were as follows:

Group number

Mean Achieved Atmosphere Concentration (mg/L)

Mean Mass Median Aerodynamic Diameter (µm)

Inhalable Fraction (% < 4 µm)

Geometric

Standard

Deviation

1

2.11

1.64

85.6

2.33

2

5.05

2.40

77.5

1.98

The mortality data were summarised as follows:

Group Number

Mean Achieved Atmosphere Concentration (mg/L)

Deaths

Male

Female

Total

1

2.11

1/5

0/5

1/10

2

5.05

2/5

3/5

5/10

Clinical Observations.

Common abnormalities noted during the study included increased respiratory rate, ataxia, hunched posture, pilo-erection and wet fur. There were frequent instances of a heightened sensitivity to audio stimuli and occasional instances of decreased respiratory rate, isolated instances of laboured respiration, noisy respiration, gasping respiration and prostration were also noted. Surviving animals from Group 1 appeared normal from Days 5 to 6 post-exposure, surviving Group 2 animals appeared normal from Days 6 to 7 post-exposure.

Bodyweight.

Group 1 – All surviving animals exhibited a bodyweight loss on Day 1, one surviving male and three females exhibited a bodyweight loss or showed no bodyweight gain from Days 1 to 3. Weight gains were noted in all surviving animals for the remainder of the recovery period.

Group 2 – All surviving animals exhibited a bodyweight loss on Day 1, two surviving male animals and all surviving females exhibited a bodyweight loss or exhibited no bodyweight gain from Days 1 to 3. Reasonable bodyweight development was noted in all surviving animals during the remainder of the recovery period.

Necropsy.

With the exception of one instance of dark patches on the lungs no macroscopic abnormalities were detected at necropsy for animals that survived until the end of the recovery period. Haemorrhagic or abnormally dark lungs were detected at necropsy amongst animals that died during the course of the study. Due to lung observations noted in the animals it is considered that the deaths noted during this study may have been attributable to local toxicity caused by the test item.

Conclusion:

The acute inhalation median lethal concentrations (LC50) (it was not possible to calculate 95% confidence limits as only two groups were exposed) of AS 100 (Di-isopropyl xanthogen polysulphide) in the RccHanTM : WIST strain rat, were calculated to be:

All animals : >5.05 mg/L

Males only : >5.05 mg/L

Females only : 4.56 mg/L

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LC50
Value:
4 560 mg/m³
Quality of whole database:
The study has been conducted according to OECD Guideline 403 and GLP and is adequately reported. The study has been assigned a reliability 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 undertaken between 1 and 15 September 1987
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study conducted in accordance with generally accepted scientific principles, possibly with incomplete or methodological deficiencies, which do not affect the quality of relevant results.
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
no
GLP compliance:
not specified
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
other: CD rats [Crl: COBS CD(SD)BR]
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Obtained from Charles River U.K. Limited, Margate, Kent, England.
- Age at study initiation: Seven to ten weeks of age.
- Weight at study initiation: 200 to 240 g.
- Fasting period before study: None.
- Housing: Rats were housed individually in metal cages with wire mesh floors.
- Diet (e.g. ad libitum): A standard laboratory rodent diet (Labsure LAD 1) was provided ad libitum.
- Water (e.g. ad libitum): Water was provided ad libitum.
- Acclimation period: At least twenty days prior to the start of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): The mean daily minimum and maximum temperatures were 22°C and 25°C.
- Humidity (%): The mean daily relative humidity value was 55%.
- Air changes (per hr): The rate of air exchange was maintained at approximately 15 air changes/hour.
- Photoperiod (hrs dark / hrs light): Lighting was controlled by means of a time switch to give 12 hours artificial light in each 24 hour period.
Type of coverage:
occlusive
Vehicle:
unchanged (no vehicle)
Remarks:
None
Details on dermal exposure:
TEST SITE
One day prior to treatment hair was removed from the dorso-lumbar region of each rat with electric clippers exposing an area equivalent to 10% of the total body surface.

The test substance was applied by spreading it evenly over the prepared skin. The treated area was then promptly covered with gauze which was held in place with an impermeable dressing encircled firmly around the trunk.



REMOVAL OF TEST SUBSTANCE
At the end of the 24-hours exposure period, the dressings were carefully removed and the treated area of skin decontaminated by washing in warm water and blotting dry with absorbent paper.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): The rats were treated at 2.0 g/kg bodyweight.


Duration of exposure:
24 hours.
Doses:
2.0 g/kg bodyweight.
No. of animals per sex per dose:
A group of ten rats (five males and five females) was treated at 2.0 g/kg bodyweight.
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days

- Frequency of observations and weighing: Animals were observed soon after dosing and at frequent intervals for the remainder of Day 1. On
subsequent days the animals were observed once in the morning and again at the end of the experimental day. Clinical signs were recorded at each observation.

The treated areas of skin were examined daily for signs of dermal irritation and assessed according to the below scoring system (see Erythema and Eschar Formation).

A separate record was kept of dermal changes other than erythema and oedema.

The following were recorded on the study:
- The nature, severity approximate time of onset and duration of each toxic sign.
- Individual bodyweights of rats on Days 1 (day of dosing), 8 and 15



- Necropsy of survivors performed: All animals on the study were killed on Day 15 by cervical dislocation and were subjected to a macroscopic post mortem examination, which consisted of opening the abdominal and thoracic cavities. The macroscopic appearance of abnormal organs when present was recorded.

Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
There were no deaths among ten rats subjected to a single occluded dermal application of D.I.X.T at 2.0 g/kg bodyweight.
Clinical signs:
There were no clinical signs of systemic reaction to treatment.
Body weight:
Low bodyweight gains were recorded on Day 8 for four female rats and on Day 15 for three of the same animals and one other female (see Table 2 - attached background material).
Gross pathology:
Terminal autopsy findings were normal.
Other findings:
Dermal Responses (see Table 1 - attached background material):
Slight or well-defined erythema was apparent on Days 2, 3 and 4 at the sites of application of the test substance. No other irritation reactions or dermal changes were observed.
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The acute lethal dermal dose to rats of D.I.X.T was found to be greater than 2.0 g/kg bodyweight.
Executive summary:

Introduction:

The experimental procedure was based on that recommended under Annex V of EEC directive 79/831/EEC, Part B Methods for determination of toxicity. Method B3, Acute Dermal Toxicity, and the OECD guideline for Testing of Chemicals No. 402 "Acute Dermal Toxicity".

Method:

A group of ten rats (five males and five females) were treated at 2.0 g/kg bodyweight for a 24 hour exposure period.

Clinical signs, dermal irritation, bodyweight development were monitered during the study. All animals were killed on Day 15 and subjected to a macroscopic post mortem examination.

Results:

Mortality:

There were no deaths among ten rats subjected to a single occluded dermal application of D.I.X.T at 2.0 g/kg bodyweight.

Clinical signs:

There were no clinical signs of systemic reaction to treatment.

Dermal responses:

Slight or well-defined erythema was apparent on Days 2, 3 and 4 at the sites of application of the test substance. No other irritation reactions or dermal changes were observed.

Bodyweight:

Low bodyweight gains were recorded on Day 8 for four female rats and on Day 15 for three of the same animals and one other female.

Terminal autopsy:

Terminal autopsy findings were normal.

Conclusion:

The acute lethal dermal dose to rats of D.I.X.T was found to be greater than 2.0 g/kg (>2000 mg/kg) bodyweight.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
LD50
Value:
2 000 mg/kg bw
Quality of whole database:
Study conducted in accordance with generally accepted scientific principles (OECD Guideline 402), possibly with incomplete or methodological deficiencies, which do not affect the quality of relevant results.

Additional information

Acute Oral Toxicity:

Introduction:

The experimental procedure used followed the Testing Guidelines as described in the Federal Register, Vol 50, No.188, Part II of 27 September 1985, Section 798.1175 - Acute Oral Toxicity.

Method:

A preliminary test was carried out to establish a dosing regimen using groups of two males and two female rats at three dose levels of 1.0, 2.5 and 5.0 g/kg bodyweight.

Based on the findings of the preliminary test, further groups of rats (5 male and 5 female per dose group) were dosed at dose levels of 1.0, 1.6, 2.5, 4.0 and 6.4 g/kg bodyweight. The appropriate dose volume of the test substance was administered to each rat using a syringe and plastic catheter or a glass microlitre syringe and metal cannula.

Animals were observed soon after dosing, then at frequent intervals for the remainder of Day 1. On subsequent days the animals were observed once in the morning and once at the end of the experimental day. Clinical signs were recorded at each observation.

Individual bodyweights of rats on Days 1 (day of dosing), 8, 15 and at death were recorded.

Surviving animals on the main study were killed on Day 15 by cervical dislocation. All animals that died during the study and those killed on Day 15 were subjected to a macroscopic post mortem examination which consisted of opening the abdominal and thoracic cavities. The macroscopic appearance of abnormal organs when present was recorded.

Results:

The results of the preliminary study indicated that the acute median lethal oral dose of D.I.X.T was between 2.5 and 5.0 g/kg bodyweight.

Main test:

Mortalities:

Mortalities occurred amongst rats dosed at 1.0 g/kg and above within two and 71 hours of dosing. Autopsy revealed pallor of the liver, spleen and kidneys and congestion of the blood vessels of the small and large intestine amongst rats that died. Perforations of the duodenum were seen amongst rats at all dose levels. A light yellow fluid was found in the peritoneal cavity of three rats dosed at 4.0 g/kg and one female dosed at 6.4 g/kg. Bodyweighy losses were recorded for all but one of the rats that died.

Clinical signs:

Signs of reaction to treatment observed shortly after dosing amongst rats of all groups were pilo-erection, abnormal body carriage (hunched posture), abnormal gait (waddling), lethargy, decreased respiratory rate, pallor of the extremities and diuresis. These were accompanied by:

- ptosis in all rats dosed at 1.6 g/kg and above,

- increased salivation in all rats dosed at 1.6 g/kg,

- abdominal distension in one male rat dosed at 2.5 g/kg,

- a coma-like condition in one male rat dosed at 2.5 g/kg.

Recovery, as judged by external appearance and behaviour, was apparently complete by Days 7 to 11.

Bodyweight:

Bodyweight gains were recorded for all surviving rats on Days 8 and 15.

Terminal autopsy:

Terminal autopsy revealed perforations in the duodenum in a small number of animals. Enlargement of the spleen was seen in one female rat dosed at 2.5 g/kg.

Conclusion:

The acute median lethal dose (LD50) and their 95% confidence limits to rats of D.I.X.T. were estimated to be:

Males and females combined: 1.5 (0.40 to 2.4) g/kg bodyweight.

Males only: 1.3 (0.22 to 2.5) g/kg bodyweight.

Females only: 1.8 (0.50 to 3.5) g/kg bodyweight.

Acute Dermal Toxicity:

Introduction:

The experimental procedure was based on that recommended under Annex V of EEC directive 79/831/EEC, Part B Methods for determination of toxicity. Method B3, Acute Dermal Toxicity, and the OECD guideline for Testing of Chemicals No. 402 "Acute Dermal Toxicity".

Method:

A group of ten rats (five males and five females) were treated at 2.0 g/kg bodyweight for a 24 hour exposure period.

Clinical signs, dermal irritation, bodyweight development were monitored during the study. All animals were killed on Day 15 and subjected to a macroscopic post mortem examination.

Results:

Mortality:

There were no deaths among ten rats subjected to a single occluded dermal application of D.I.X.T at 2.0 g/kg bodyweight.

Clinical signs:

There were no clinical signs of systemic reaction to treatment.

Dermal responses:

Slight or well-defined erythema was apparent on Days 2, 3 and 4 at the sites of application of the test substance. No other irritation reactions or dermal changes were observed.

Bodyweight:

Low bodyweight gains were recorded on Day 8 for four female rats and on Day 15 for three of the same animals and one other female.

Terminal autopsy:

Terminal autopsy findings were normal.

Conclusion:

The acute lethal dermal dose to rats of D.I.X.T was found to be greater than 2.0 g/kg (>2000 mg/kg) bodyweight.

Acute Inhalation Toxicity:

Introduction.

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

Methods.

Groups of ten RccHanTM : WIST strain rats (five males and five females) were exposed to an aerosol atmosphere. The animals were exposed for four hours using a nose only exposure system, followed by a fourteen day observation period.

Results:

The mena achieved atmosphere concentrations were as follows:

Group number

Atmospheric concentration

Mean Achieved (mg/L)

Standard deviation

Nominal (mg/L)

1

2.11

0.05

5.72

2

5.05

0.16

20.9

The characteristics of the achieved atmospheres were as follows:

Group number

Mean Achieved Atmosphere Concentration (mg/L)

Mean Mass Median Aerodynamic Diameter (µm)

Inhalable Fraction (% < 4 µm)

Geometric

Standard

Deviation

1

2.11

1.64

85.6

2.33

2

5.05

2.40

77.5

1.98

The mortality data were summarised as follows:

Group Number

Mean Achieved Atmosphere Concentration (mg/L)

Deaths

Male

Female

Total

1

2.11

1/5

0/5

1/10

2

5.05

2/5

3/5

5/10

Clinical Observations.

Common abnormalities noted during the study included increased respiratory rate, ataxia, hunched posture, pilo-erection and wet fur. There were frequent instances of a heightened sensitivity to audio stimuli and occasional instances of decreased respiratory rate, isolated instances of laboured respiration, noisy respiration, gasping respiration and prostration were also noted. Surviving animals from Group 1 appeared normal from Days 5 to 6 post-exposure, surviving Group 2 animals appeared normal from Days 6 to 7 post-exposure.

Bodyweight.

Group 1 – All surviving animals exhibited a bodyweight loss on Day 1, one surviving male and three females exhibited a bodyweight loss or showed no bodyweight gain from Days 1 to 3. Weight gains were noted in all surviving animals for the remainder of the recovery period.

Group 2 – All surviving animals exhibited a bodyweight loss on Day 1, two surviving male animals and all surviving females exhibited a bodyweight loss or exhibited no bodyweight gain from Days 1 to 3. Reasonable bodyweight development was noted in all surviving animals during the remainder of the recovery period.

Necropsy.

With the exception of one instance of dark patches on the lungs no macroscopic abnormalities were detected at necropsy for animals that survived until the end of the recovery period. Haemorrhagic or abnormally dark lungs were detected at necropsy amongst animals that died during the course of the study. Due to lung observations noted in the animals it is considered that the deaths noted during this study may have been attributable to local toxicity caused by the test item.

Conclusion:

The acute inhalation median lethal concentrations (LC50) (it was not possible to calculate 95% confidence limits as only two groups were exposed) of AS 100 (Di-isopropyl xanthogen polysulphide) in the RccHanTM : WIST strain rat, were calculated to be:

All animals : >5.05 mg/L

Males only : >5.05 mg/L

Females only : 4.56 mg/L


Justification for selection of acute toxicity – oral endpoint
The acute oral toxicity study is assigned as reliability study 2 and is the only acute oral toxicity study available.

Justification for selection of acute toxicity – inhalation endpoint
The acute inhalation toxicity study is assigned as reliability study 1 and is the only acute oral inhalation study available.

Justification for selection of acute toxicity – dermal endpoint
The acute dermal toxicity study is assigned as reliability study 2 and is the only acute dermal toxicity study available.

Justification for classification or non-classification

Acute Oral Toxicity:

The acute median lethal dose (LD50) to rats of D.I.X.T. was estimated to be estimated to be 1.5 g/kg (1500 mg/kg) bodweight (for males and females combined).

Therefore, according to the CLP regulation, the substance shall be classifed for acute toxicity by the oral route as Acute Tox Category 4 (for substances with oral LD50 determined to be >300 - ≤2000 mg/kg bodyweight). The hazard statement H302: Harmful if swallowed, will apply to this substance.

Acute Dermal Toxicity:

The substance does not meet the criteria for classification, under the CLP regulations, for acute toxicity via the dermal route based on the results of an acute dermal toxicity study, which both gave a LD50 results of >2000 mg/kg bodyweight, which is greater than the cut-off criteria for classification.

Acute Inhalation Toxicity:

The acute inhalaton LC50 for females only was found to be 4.56 mg/L.

Therefore, according to the CLP regulation, the substance shall be classified for acute toxicity by the inhalation route as Acute Tox Category 4 (for substances with LC50 for dust/mist determined to be >1 -≤5 mg/L)). The hazard statement H332: Harmful if inhaled, will apply to this substance as a worst case assessment based on the female animals.