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Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
year of publication: 1978
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
documentation insufficient for assessment
Reason / purpose for cross-reference:
reference to same study
Objective of study:
other: excretion of possible metabolites
Principles of method if other than guideline:
investigation of excretion of possible metabolites in the urine
GLP compliance:
no
Radiolabelling:
no
Species:
rat
Strain:
not specified
Sex:
male/female
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Duration and frequency of treatment / exposure:
23 month
Dose / conc.:
9 000 ppm
Remarks:
Doses / Concentrations:
9000 ppm in diet
No. of animals per sex per dose / concentration:
10
Control animals:
yes, plain diet
Details on dosing and sampling:
- 2-h urine samples were collected (no further information)
Type:
excretion
Results:
no 3,3'-dichlorobenzidine was detected in the urine
Details on excretion:
- no 3,3'-dichlorobenzidine was detected in the urine
Key result
Test no.:
#1
Toxicokinetic parameters:
other: no 3,3'-dichlorobenzidine was detected in the urine
Conclusions:
The test item is not absorbed after oral application and there is no evidence for a metabolic splitting of the test item to 3,3'-dichlorobenzidine
The test item seems not to be absorbed after oral application and there is no evidence for a metabolic splitting of the test item to 3,3'-dichlorobenzidine.
Executive summary:

Rat were exposed to 9000 ppm test item in diet for two years. Two-hour urine samples were collected after 6 and 23 months from 10 male and female rats and were analysed by thin layer chromatography for the presence of the hypothetical cleavage product 3,3'-dichlorobenzidine. No 3,3'-dichlorobenzidine was detectable in urine of treated and control animals.

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
Year of publication: 2001
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Remarks:
There was 3,3'-dichlorbenzidine deteted in the untreaded control.
Objective of study:
other: bioavailability of cleavage products
Qualifier:
equivalent or similar to guideline
Guideline:
other: OECD Guideline 412 (Repeated Dose Inhalation Toxicity)
Deviations:
yes
Remarks:
e.g. no data on haematology, because the main subject of the study was the investigation of the bioavailability and not the investigation of toxicity after repeated exposure
GLP compliance:
yes
Remarks:
Performance and documentation was on the basis of GLP-procedures according to the German Chemikaliengesetz, but the test was not under supervision of the quality assurance unit
Radiolabelling:
no
Species:
rat
Strain:
Wistar
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River, Germany
- Age at study initiation: about 3 months
- Weight at study initiation: about 430 g
- Fasting period before study: no data
- Housing: Makrolon cages
- Individual metabolism cages: yes
- Diet: Altromin Pellet-Feed (1324N) or Altromin no Pellet-Feed (1321N) ad libitum
- Water: drinking water ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 40-70%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 h/12 h


Route of administration:
other: intratracheal instillation
Vehicle:
other: suspension in isotonic saline containing 1% (w/v) Tween 80
Details on exposure:
- intratracheal instillation of 0.3 ml of suspension
Duration and frequency of treatment / exposure:
once per week for 4 consecutive weeks (day 1, 8, 15, 22 and 29 of the experiment)
Dose / conc.:
0 other: mg/animal/application
Dose / conc.:
10 other: mg/animal/application
Dose / conc.:
20 other: mg/animal/application
No. of animals per sex per dose / concentration:
- 12 males per group in the control and test substance group (6 of the animals were sacrificed on day 31 and 6 animals survived till the end of the 4 weeks post exposure period)
- 6 males in the positive control group
Control animals:
yes, concurrent vehicle
Positive control reference chemical:
5 times 7.31 mg 3,3'-dichlorobenzidine (CAS 91-94-1) or 5 times 5,6 mg Direct Red 46 (CAS 6548-29-4) per animal
Details on study design:
- Dose selection rationale: ascertainment of the maximum tolerable dose in a pretest
Details on dosing and sampling:
BIOAVAILABILITY STUDY
- Tissues and body fluids sampled: urine, faeces, blood, organs (lung, liver, spleen, kidney, urinary bladder, gastro intestinal tract, mamma)
- Time and frequency of sampling of urine and faeces: on day 1, 2, 3, 7, 8, 9, 14, 15, 16, 21, 22, 23, 28, 29, 30 for the animals which were sacrificed on day 31
- Time and frequency of sampling of urine and faeces: on day 28, 29, 30, 35, 42, 49, 56 for the animals which were sacrificed on day 57 at the end of the post exposure period
- Time and frequency of sampling of blood and organs: after sacrifice on day 31 or 57



Statistics:
no data
Type:
absorption
Results:
very limited absorption
Type:
excretion
Results:
3,3'-Dichlorobenzidine was detectable in the urine, and partially in the faeces, likely due to contamination of analytical instruments
Metabolites identified:
not measured

 

One animal of the negative control group had to be killed in a moribund condition on day 4. There was no direct correlation to the treatment obvious.

Body-weight development was normal.

The test substance was deposited in the lung: The test substance was detectable in macrophages, in the alveolar and intrabronchiolar region. Lung weights were elevated at the end of the exposure period and still elevated at the end of the post exposure period. Bronchio-alveolar hyperplasia and interstitial fibrosis were detected and persisted till the end of the observation period. Pigment loaded macrophages were also detected in lung associated lymph nodes, which showed increased weight and reactive lymphoid hyperplasia. Effects persisted till the end of the observation time.

3,3'-Dichlorobenzidine was detectable in the hydrolysed urine of treated animals during the exposure period. 3,3'-Dichlorobenzidine excretion via urine was in the range of 3,9 and 4.2 ng/24 hours in the low dose group and about 4.6 ng/24 h on day 2 and 8.8 ng/24 hours on day 30 in the high dose group. 3,3'-Dichlorobenzidine was detectable in two of 30 faeces samples (2.2 ng/24 h and 4.4 ng/24 h). No 3,3'-dichlorobenzidine was verifiable in the faeces and urine during the post exposure period. No 3,3'-dichlorobenzidine was detectable in urine and faeces during the post exposure observation period (measurement on day 42 and 56).

3,3'-Dichlorobenzidine bound to haemoglobin was detectable in 3 out of 5 probes of the low dose group (maximal value: 0.24 ng/g) and in 4 out of 5 probes of the high dose group (maximal value: 6.66 ng/g) at the end of the exposure period. Haemoglobin bound 3,3'-dichlorobenzidine was detectable in 2 out of 5 probes of the high dose group at the end of the post-exposure period (maximal value: 0.13 ng/g).

3,3'-Dichlorobenzidine was also traceable in the urine of several animals of the control group (2 animals sacrificed on day 31 and 5 animals sacrificed on day 57). The authors of the report concluded that this was probably due to a contamination of the collecting containers.

Pigment content of the lungs determined at the end of the exposure period was about 72 -76% of the applied dose. The pigment content of the lungs did not decline during the post exposure period.

The authors calculated that about 0.001 - 0.005% of the theoretically possible 3,3'-dichlorobenzidine was liberated within 24 hours from the applied test substance. Concerning the pigment content of the lung at the end of the exposure period of the high dose group a bioavailability of 0.0012 Mol%/24 hours can be calculated (0.00006 Mol%/24 hours at the end of the post exposure period).

3,3'-dichlorobenzidine treatment:

Animals treated with 3,3'-dichlorobenzidine excreted about 3% of the applied dose via urine (about 194000 ng on day 1 and 226000 ng on day 29) during the first 24 h after dosing. Urinary excretion declined rapidly thereafter: about 10400 ng/24 h during the second and 1030 ng/24 h during the third 24 hours after dosing. The average urinary excretion on the day before the 5th application was 59 ng/24 h. Excretion of 3,3'-dichlorobenzidine via faeces was delayed in comparison to excretion via urine: About 3.6% ( 262 µg; day 1 and 2) or 2.6% (190 µg, day 29 and 30) were excreted within 48 hours after application.

At sacrifice about 1496 ng 3,3'-dichlorobenzidine were bound per g haemoglobin.

Conclusions:
The test substance has a high potential to persist in the lung due to the significantly limited clearance capacity of the lung as a consequence of the particle overload.
Executive summary:

Male Wistar rats were exposed for five times to 10 or 20 mg test item/animal by intratrachel instillation at weekly intervals. Urine and faeces were collected during the exposure and 28 days observation period, blood was collected at the end of the exposure and post observation period. Urine, faeces and haemoglobin were analysed for the presence of 3,3'-dichlorobenzidine after acid hydrolysis, the pigment content of the lungs was determined gravimetrically and the lungs and lung associated lymph nodes were investigated histopathologically. The pigment was deposited in the lung and persisted in the lung during the observation period. 3,3'-Dichlorobenzidine was detectable in the urine, and partially in the faeces and haemoglobin of treated rats during the exposure period. Nevertheless, no proof of bioavailability of the pigment was obtained since 3,3'-Dichlorobenzidine was still detectable in the blood at the end of the post exposure period, but not in the urine and faeces during the post exposure period.

Based on this observation a constant absorption and metabolization of the applied pigment is excluded.

Traces of 3,3'-dichlorobenzidine were also detected in control animals, probably due to impurities of the analytical instruments. Concerning the pigment content of the lung at the end of the exposure period a bioavailability of 0.0012 Mol% / 24 hours can be calculated for the high exposure group. At the end of the observation period the bioavailability was only 0.00006 Mol% / 24 hours for the high exposure group.

However, since traces of 3,3'-dichlorobenzidine were also found in blood, urine and feces of control animals, it cannot be concluded if the detection of 3,3'-dichlorobenzidine is due to the absorption and biotransformation of the applied pigment or due to the contamination of analytical instruments.

Endpoint:
basic toxicokinetics, other
Type of information:
other: Expert statement
Adequacy of study:
key study
Study period:
2020
Reliability:
2 (reliable with restrictions)
Details on absorption:
A prerequisite for a relevant absorption is that the substance can be dissolved in either aque-ous (e.g., gastrointestinal fluid, blood plasma, sweat) or lipophilic (e.g., lipoproteins, lipid membranes, triglycerides) media or in both. C.I. Pigment Yellow 83 can be considered insol-uble because it has an extremely low solubility in water and n-octanol. Therefore, it is unlike-ly that C.I. Pigment Yellow 83 becomes systemically bioavailable after oral, dermal or inhala-tion exposure.
Based on the chronic oral toxicity study with C.I. Pigment Yellow 83 absorption of toxico-logically significant amounts via the gastrointestinal tract is considered unlikely, since C.I. Pigment Yellow 83 did not show any effects on inner organs and blood or urine. No metabo-lites could be detected.
The skin sensitisation studies with C.I. Pigment Yellow 83 indicate no local dermal bioavail-ability. Systemic availability also seems to be negligible after dermal exposure since no sys-temic signs of intoxication were seen after occlusive administration of 500 mg C.I. Pigment Yellow 83 per kg body weight in rabbits in the acute dermal irritation study.
Dermal absorption is, therefore, considered unlikely.
In the unlikely event of exposure to aerosolized pigment in respirable form, the substance is considered to behave like an inert dust. Therefore, the deposited pigment particles will mostly be cleared from the lung via the mucocilliary transport. As the pigment will not dis-solve in the lung surfactant, the only way the pigment can enter the body is via phagocyto-sis of pigment particles by lung macrophages followed by migration of the macrophages into the interstitium and into the draining lymph nodes. However, the internal dose deliv-ered via this mechanism can be considered negligible.
Details on distribution in tissues:
The Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxici-ty Screening Test as well as all older studies employing repeated exposure with C.I. Pigment Yellow 83 did not indicate any relevant histopathological changes in any of the investigated organs. This may indicate that the pigment either does not affect specific organs as targets, i.e., is non-toxic, or is not distributed within the body in significant amounts. As indicated above, the physico-chemical parameters of the pigment support the conclusion that the pig-ment is not absorbed into the body and thus does not become systemically available. There were also no other signs of deposition of the pigment in any organ including excretory or-gans, like the kidney, indicating that even exposure to high doses of the pigment does not lead to bioaccumulation in special compartments of the body.
Based on the available information on absorption distribution of the test material in the body in significant amounts is unlikely and specific hotspots of distribution cannot be identified.
Thus, it is concluded, that C.I. Pigment Yellow 83 is not systemically available at relevant concentrations within the organism.
There were no signs of bioaccumulation of the test material. This view is supported by the physical-chemical properties (solubility in water and octanol)
Details on excretion:
Considering the physico-chemical properties and the molecular structure and size of the mol-ecules and the absence of any indication of absorption and/or metabolism it is assumed that excretion, if any, is likely to occur via faeces. This notion is confirmed by the discoloration of faeces observed in the oral studies as the only alteration.

Metabolites identified:
no
Details on metabolites:
Since the solution of the substance in cellular fluid or cellular membranes is a prerequisite for its metabolism, it is unlikely that the insoluble pigment becomes accessible for metabo-lizing systems in relevant amounts.
Diarylide yellow pigments are not or only to a negligible extend cleaved, taking into ac-count the negative results obtained with C.I. Pigment Yellow 83 in investigations on car-cinogenicity.
The results of the mutagenicity tests provide qualitative information on the metabolic fate of C.I. Pigment Yellow 83. In the mutagenicity tests, the pigment proved to be non-mutagenic in the absence as well as in the presence of an exogenous metabolizing system, indicating that the pigment, even directly exposed to metabolizing enzymes, is not convert-ed into toxic or genotoxic metabolites. This conclusion is also supported by the lack of any morphological and histopathological changes of organs involved in xenobiotic metabolism, such as the liver, in the Chronic Toxicity Study with C.I. Pigment Yellow 83. Furthermore, the missing skin or eye irritating or skin sensitizing properties argue against any interaction with biological material.
Therefore, C.I. Pigment Yellow 83 is considered to just pass through the intestinal tract without significant metabolism.

Bioaccessibility (or Bioavailability) testing results:
Bio-accessability is not likely
Conclusions:
Based on all available data, C.I. Pigment Yellow 83 does not exhibit conspicuous toxicoki-netic behaviour in the sense of accumulative and/or delayed effects with regard to the indi-vidual parameters absorption, distribution, metabolism and excretion.
The results from studies with dermal exposure indicate that C.I. Pigment Yellow 83 has a no relevant dermal absorptive potential. C.I. Pigment Yellow 83 is most probably not absorbed from the gastrointestinal tract in significant amounts.
Indications of an intense metabolism or a bio-accumulative potential do not exist as no tox-icity occurred even after chronic exposure, which points to no bio-accumulation potential and complete excretion of all possibly available C.I. Pigment Yellow 83 and/or metabolites.
Executive summary:

Based on the available data base on C.I. Pigment Yellow 83 relevant information exists to make a qualitative evaluation of the toxicokinetic profile of this compound. This is in line with animal welfare considerations because additional animal tests can be avoided by such an evaluation. The substance is available in bulk- as well as nano-form. The available data have mostly been generated with non-nano material, but the conclusions drawn are considered valid for the nano form as well, because the material is chemically identical, and the physical properties are widely overlapping.

 

The results of basic toxicity testing give no reason to anticipate unusual characteristics regarding the toxico-kinetics of C.I. Pigment Yellow 83. C.I. Pigment Yellow 83 is not absorbed from the gastro-intestinal tract in toxicologically significant amounts. The data indicate that there is no relevant dermal absorption. After inhalation unspecific reactions to unreactive dust can be expected. Indications of a bio-accumulative potential as well as metabolism towards genotoxic sub-structures do not exist. Excretion of traces of possibly systemically available C.I. Pigment Yellow 83 and/or metabolites via faeces is likely.

Description of key information

Based on the available data base on C.I. Pigment Yellow 83 relevant information exists to make a qualitative evaluation of the toxicokinetic profile of this compound. This is in line with animal welfare considerations because additional animal tests can be avoided by such an evaluation. The substance is available in bulk- as well as nano-form. The available data have mostly been generated with non-nano material, but the conclusions drawn are considered valid for the nano form as well, because the material is chemically identical, and the physical properties are widely overlapping.

 

The results of basic toxicity testing give no reason to anticipate unusual characteristics regarding the toxico-kinetics of C.I. Pigment Yellow 83. C.I. Pigment Yellow 83 is not absorbed from the gastro-intestinal tract in toxicologically significant amounts. The data indicate that there is no relevant dermal absorption. After inhalation unspecific reactions to unreactive dust can be expected. Indications of a bio-accumulative potential as well as metabolism towards genotoxic sub-structures do not exist. Excretion of traces of possibly systemically available C.I. Pigment Yellow 83 and/or metabolites via faeces is likely.

Key value for chemical safety assessment

Bioaccumulation potential:
no bioaccumulation potential

Additional information