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Administrative data

Description of key information

Oral (OECD 401), rat: LD50 >5000 mg/kg bw 
Inhalation (OECD 436), rat: LC50 >3 mg/L air (maximum attainable concentration)
Dermal (OECD 402), rat: LD50 >2000 mg/kg bw

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:
17 Dec - 31 Dec 1987
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions (purity of test substance not specified).
Qualifier:
according to
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
Version / remarks:
adopted 12. May, 1981
Deviations:
yes
Remarks:
purity of test substance not given
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI, USA
- Age at study initiation: approximately 8 weeks
- Weight at study initiation: 240 - 299 g
- Fasting period before study: 17 to 20 hours (over night)
- Housing: separated by sex in groups of 5 in screen-bottom stainless steel cages (heavy gauge)
- Diet: Purina Rodent Chow, ad libitum
- Water: ad libitum
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 23
- Humidity (%): 21 - 31
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 17 Dec To 31 Dec 1987
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 0.5 g/mL
- Amount of vehicle (if gavage): 10 mL/kg bw
- Justification for choice of vehicle: vehicle was chosen due to good solubility of test substance in this vehicle

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw
Doses:
5000 mg/kg bw
No. of animals per sex per dose:
5
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Each animal was observed for clinical signs and mortality at 1, 2.5 and 4 hours following administration, and daily thereafter for 14 days, once daily for clinical signs and twice daily for mortality, weighing was done just before dosing, on day 7 and at study termination
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight
Statistics:
Average body weights were calculated.
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 5 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortalities were observed during the study period.
Clinical signs:
1 male animal had diarrhea 4 hours after administration. 2 male animals had miosis starting at 2.5 hours following administration until day 1 and day 2, respectively. Some female animals showed miosis (1, 2.5 and 4 hours after administration and on day 1), diarrhea (1, 2.5 and 4 hours post administration), hypoactivity and lacrimation (2.5 and 4 hours post administration) and hypersensitivity to touch (one animal 4 hours after administration).
Body weight:
Body weight gain during the study was normal.
Gross pathology:
One female animal had multiple red pinpoint foci in the right salivary gland and diffusely dark red submandibular bilateral lymph nodes. A second female had an enlarged pelvis in the right kidney. The findings in the two female rats were considered to be incidental and unrelated to exposure. All other animals were without visible lesions.

Table 1: Clinical Signs (Number of animals affected)

  Hours         Days      
   1.0  2.5  4.0  1  ...  14
 Males (5.0 g/kg)
Appeared Normal  3  2  4  ...  5
 Diarrhea 0  0  1  0  ...  0
 Miosis 0  2  2  1  ...  0
 Females (5.0 g/kg)
 Appeared Normal  2  0  0  4  ...  5
 Miosis  2  3  3  1  ...  0
 Diarrhea  2  3  4  0  ...  0
 Hypoactivity  0  1  2  0  ...  0
 Lacrimation  0  3  3  0  ...  0
 Hypersensitivity to touch  0  0  1  0  ...  0

Table 2: Average body weights (g)

   Dose Level  Average Body Weights (g)      
   (g/kg)  Initial  Day7  Terminal
 Male  5.0  280  344  367
 Female  5.0  253  279  288
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
DSD: not classified
CLP: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study (Klimisch score 2), and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

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:
10 - 24 November 2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.
Qualifier:
according to
Guideline:
OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)
Deviations:
no
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: Young adult animals (approximately 10 weeks old) were selected.
- Weight at study initiation: Body weight variation did not exceed +/- 20% of the sex mean.
- Housing: Group housing of three animals per sex per cage in labelled Makrolon cages (type IV; height 18 cm) containing sterilised sawdust as bedding material (Lignocel S 8-15, JRS - J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
- Diet (e.g. ad libitum): Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany) except during exposure to the test substance.
- Water (e.g. ad libitum): Free access to tap water except during exposure to the test substance.
- Acclimatization period: at least 5 days before start of treatment under laboratory conditions.
- Health inspection: A health inspection was performed prior to commencement of treatment, to ensure that the animals were in a good state of health.

ENVIRONMENTAL CONDITIONS
Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, at least 10 air changes/hour and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study.

IN-LIFE DATES: 10 - 24 November 2014
Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
air
Details on inhalation exposure:
The design of the exposure chamber is based on the flow past nose-only inhalation chamber (Am. Ind. Hyg Assoc. J. 44(12): 923-928, 1983). The chamber consisted of three animal sections with eight animal ports each. Each animal port had its own atmosphere inlet and exhaust outlet. The animals were placed in restraining tubes and connected to the animal ports. The number of animal sections and number of open inlets were adapted to the air flow in such a way that at each animal port the theoretical air flow was at least 1 L/min, which ensures an adequate oxygen supply to the animals. The main inlet of the test atmosphere was located at the top section and the main outlet was located at the bottom section. The direction of the flow of the test atmosphere guaranteed a freshly generated atmosphere for each individual animal.
All components of the exposure chamber in contact with the test material were made of stainless steel, glass, rubber or plastic. To avoid exposure of the personnel and contamination of the laboratory the exposure chamber was placed in a fume hood, which maintained at a slight negative pressure.
Analytical verification of test atmosphere concentrations:
no
Duration of exposure:
4 h
Concentrations:
3 mg/L
No. of animals per sex per dose:
3
Control animals:
no
Details on study design:
Test substance preparation: The test substance was used as delivered by the sponsor.

The test substance was administered as an aerosol by inhalation for 4 hours to two groups of three male and three female Wistar rats each. Animals were subjected to daily observations and determination of body weights on Days 1, 2, 4, 8 and 15 and at death. Macroscopic examination was performed on the day of death or after terminal sacrifice (day 15).

The actual time weighted mean concentration was 3 ± 0.1 mg/L. The nominal concentration (amount of test substance used divided by the volume of pressurized air used) was 44 mg/L. The generation efficiency (ratio of actual and nominal concentration) was 8%.

The concentration measurements (n=15) equally distributed over time showed that it was difficult to maintain a stable level at this technically maximum attainable concentration. The generation was interrupted in order to remove test substance deposits from the system at four occasions with a total interruption time of approximately 30 minutes. The generation time was elongated (total generation time 253 minutes) to compensate for these interruptions, resulting in an actual exposure time of at least 227 minutes. Since the interruptions and variations in concentration were taken into account by calculation a time weighted mean concentration, it was considered that the results of this study are representative for a single exposure of 3 mg/L for 4 hours. This was the technically maximum attainable concentration.
Statistics:
No statistical analysis was performed.
Sex:
male/female
Dose descriptor:
LC50
Effect level:
> 3 mg/L air
Based on:
test mat.
Exp. duration:
4 h
Mortality:
No mortality occurred.
Clinical signs:
other: Rapid breathing was seen for the animals during exposure. No clinical signs were noted following the exposure.
Body weight:
Body weight loss was seen for all females during the first week post-exposure (up to Day 8). Two females regained weight during the second week resulting in an overall body weight gain during the complete observation period, while one female remained below the initial body weight at the start of the study. Overall body weight gain in males was within the range expected for rats of this strain and age used in this type of study.
Gross pathology:
No abnormalities were found at macroscopic post mortem examination of the animals.
Interpretation of results:
practically nontoxic
Remarks:
Migrated information Criteria used for interpretation of results: OECD GHS
Conclusions:
The LC50, 4h value of 4,4’-(9H-fluoren-9-ylidene)bis(2-chloroaniline) in Wistar rats value was established to exceed the maximum obtainable concentration of 3 mg/L.
Based on these results 4,4’-(9H-fluoren-9-ylidene)bis(2-chloroaniline) does not have to be classified and has no obligatory labelling requirement for acute inhalation toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) (including all amendments) and Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures (including all amendments).
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.5, of Regulation (EC) No 1907/2006.

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:
3 Dec - 17 Dec 1990
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP - Guideline study.
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Version / remarks:
adopted 1984
Deviations:
no
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes
Species:
rat
Strain:
other: Crl. (CD) SD BR VAF plus
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River UK Limited, Kent, England
- Age at study initiation: 7 to 10 weeks
- Weight at study initiation: 205 - 243 g
- Housing: individually in metal cages with wire mesh floors
- Diet: standard laboratory rodent diet (Biosure, LAD 1), ad libitum
- Water: ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 22
- Humidity (%): 46
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 3 Dec To: 17 Dec 1990
Type of coverage:
occlusive
Vehicle:
water
Details on dermal exposure:
TEST SITE
- Area of exposure: 5 x 5 cm clipped skin of the dorso-lumbar region
- % coverage: 10% of total body surface
- Type of wrap if used: gauze which was held in place by an impermeable dressing

REMOVAL OF TEST SUBSTANCE
- Washing: in warm water (30-40°C) and blotting dry with absorbent paper
- Time after start of exposure: 24 hours

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 3.0 mL/kg bw
- Concentration (if solution): 66.7% (w/v) in distilled water
Duration of exposure:
24 hours
Doses:
2000 mg/kg bw
No. of animals per sex per dose:
5
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: animals were observed twice daily and weighing was carried out on day1, 8 and 15 (day1 = day of dosing)
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight, dermal irritation (treated areas of skin were observed daily for signs of irritation and assessed according to the Draize scoring system)
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortalities were observed during the study period.
Clinical signs:
No clinical sigsn of systemic effects were seen.
Body weight:
Slightly low body weight gains were noted for one female on Day8 and a second one on Day15 but these rats achieved anticipated gains on Day15 and Day8, respectively. All other rats achieved anticipated body weight gains during the study period.
Gross pathology:
No abnormalitites were found at terminal necropsy.
Other findings:
- Other observations: No dermal reactions at the site of application were observed.

Table 1: Body weights

 Sex  Dose  Animal number Bodyweight in g (individual gain weekly) at 
   (mg/kg)    Day1  Day8  Day15
 Male  2000  1  243  320 (77)  384 (64)
   2  206  269 (63)  333 (64)
     3  232  310 (78)  378 (68)
     4  234  317 (83)  387 (70)
     5  231  306 (75)  377 (71)
   Mean    229  304  372
 Female  2000  6  214  238 (24)  247 (9)
     7  225  259 (34)  276 (17)
     8  205  209 (4)  225 (16)
     9  217  233 (16)  260 (27)
     10  205  221 (16)  238 (17)
   Mean    213  232  249
Interpretation of results:
not classified
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
DSD: not classified
CLP: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.5, of Regulation (EC) No 1907/2006.

Additional information

Acute oral toxicity

An acute oral toxicity study (limit test) was performed with 4,4'-(9H-fluoren-9-ylidene)bis(2-chloroaniline) (CAS 107934-68-9) in Sprague-Dawley rats, according to OECD 401 (Glaza, 1988a). 5 rats per sex received a single dose of 5000 mg/kg bw by gavage. There were no mortalities. Clinical signs that were reported included diarrhea observed in 1 male animal 4 hours after administration and miosis starting at 2.5 hours following administration until day 1 and day 2 in 2 male animals. Some female animals showed miosis (1, 2.5 and 4 hours after administration and on day 1), diarrhea (1, 2.5 and 4 hours post administration), hypoactivity and lacrimation (2.5 and 4 hours post administration) and hypersensitivity to touch (one animal 4 hours after administration). There was no effect on the body weight gain of the animals. Necropsy revealed multiple red pinpoint foci in the right salivary gland and diffusely dark red submandibular bilateral lymph nodes in one female animal. A second female had an enlarged pelvis in the right kidney. The findings in the two female rats were considered to be incidental and unrelated to exposure. All other animals were without visible lesions. The acute oral LD50 in rats was found to be > 5000 mg/kg bw.

Acute inhalation toxicity

The acute inhalation toxicity of 4,4'-(9H-fluoren-9-ylidene)bis(2-chloroaniline) (CAS 107934-68-9) was assessed in a study performed according to OECD Guideline 436 in compliance with GLP (WIL Research Europe B.V., 2015). Three Wistar rats/sex were administered 3 ± 0.1 mg/L (actual concentration) of the test substance as an aerosol via nose-only exposure for 4 hours. The applied test concentration was the technically maximum attainable concentration. The nominal concentration was 44 mg/L. Concentration measurements were equally distributed over time and showed that it was difficult to maintain a stable test concentration at the technically maximum attainable concentration The generation was interrupted 4-times in order to remove test substance deposits from the system. The generation time was elongated to compensate for these interruptions. The mass median aerodynamic diameter (MMAD) and geometric standard deviation (gsd) was determined to be 3.2 µm (gsd 2.1) and 3.9 µm (gsd 2.3) in two independent measurements during the exposure period.

No mortality occurred up to the end of the 14-day observation period. Rapid breathing was noted for the animals during exposure. No clinical signs were observed following the inhalative exposure. All females lost body weight during the first week up to Day 8. Two females regained weight during the second week resulting in an overall body weight gain during the 14-day observation period, while one female remained below the initial body weight at the start of the study. Overall body weight gain in male rats was considered normal. No abnormalities were reported during the macroscopic post mortem examination. Based on the results of the conducted study, the LC50 value for the test substance is considered to be > 3 mg/L in male and female rats.

Acute dermal toxicity

An acute dermal toxicity study (limit test) was performed with 4,4'-(9H-fluoren-9-ylidene)bis(2-chloroaniline) (CAS 107934-68-9) in Crl. (CD) SD BR VAF plus rats, equivalent to OECD 402 (Baldrick, 1991). 2000 mg/kg bw of the test substance was applied to the clipped skin of 5 rats/sex/dose under an occlusive dressing for 24 hours. No mortality occurred and no clinical signs were observed. Slightly lower body weight gains were noted for one female on Day 8 and a second one on Day 15 compared to the others, but these rats achieved anticipated gains on Day 15 and Day 8, respectively. All other rats achieved anticipated body weight gains during the study period. No abnormalitites were found at terminal necropsy. No dermal reactions were observed at the site of application. The LD50 is considered to be > 2000 mg/kg bw.


Justification for selection of acute toxicity – oral endpoint
There is only one study available.

Justification for selection of acute toxicity – inhalation endpoint
There is only one study available.

Justification for selection of acute toxicity – dermal endpoint
There is only one study available.

Justification for classification or non-classification

The available data on acute oral, inhalation and dermal toxicity of 4,4´-(9H-fluoren-9-ylidene)bis(2-chloroaniline) does not meet the criteria for classification of Acute Toxicity according to Regulation (EC) 1272/2008 and according to Directive 67/54 and is therefore conclusive but not sufficient for classification.