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EC number: 207-529-8 | CAS number: 479-27-6
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
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- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Toxicity to reproduction
Administrative data
- Endpoint:
- screening for reproductive / developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 12 March 2018 to 6 June 2018
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 019
- Report date:
- 2019
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
- GLP compliance:
- yes
Test material
- Reference substance name:
- 1,8-naphthylenediamine
- EC Number:
- 207-529-8
- EC Name:
- 1,8-naphthylenediamine
- Cas Number:
- 479-27-6
- Molecular formula:
- C10H10N2
- IUPAC Name:
- naphthalene-1,8-diamine
- Details on test material:
- purity: 99.5%
Constituent 1
Test animals
- Species:
- rat
- Strain:
- Sprague-Dawley
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- A total of 120 Hsd: Sprague Dawley SD rats (55 males and 65 virgin females), 7 to 8 weeks old and weighing 200 to 225 g for males and 175 to 200 g for females, were ordered from Charles River Italia S.p.A., Calco (Lecco), Italy. After arrival the weight range for each sex was determined and found to be 217-236 g for males and 190-217 g for females and the animals were temporarily identified within the cage by means of a coloured mark on the tail. A health check was then performed by a veterinarian.
An acclimatisation period of 26 days was allowed before the start of treatment, during which time the health status of the animals was assessed by thorough observations.
The animals were housed in a limited access rodent facility. Animal room controls were set to maintain temperature and relative humidity at 22°C ± 2°C and 55% ± 15%, respectively; actual conditions were monitored, recorded and the records retained. There were approximately 15 to 20 air changes per hour and the rooms were lit by artificial light for 12 hours each day. No relevant deviations from these ranges were recorded during the study.
From arrival to mating, animals were housed up to 5 of one sex to a cage, in polysulfone solid bottomed cages measuring 59.5 × 38 × 20 cm (Tecniplast Gazzada S.a.r.l., Buguggiate, Varese). Nesting material was provided inside suitable bedding bags and changed at least twice a week.
During mating, animals were housed one male to one female in clear polysulfone cages measuring 42.5 × 26.6 × 18.5 cm with a stainless steel mesh lid and floor (Tecniplast Gazzada S.a.r.l., Buguggiate, Varese). Each cage tray held absorbent material which was inspected and changed daily.
After mating, the males were re-caged as they were before mating. The females were transferred to individual solid bottomed cages for the gestation period, birth and lactation (measuring 42.5 × 26.6 × 18.5 cm). Nesting material was provided inside suitable bedding bags. In addition, suitable nesting material (Scobis 0 Mucedola) was provided as necessary and changed at least 2 times a week.
Drinking water was supplied ad libitum to each cage via water bottles.
A commercially available laboratory rodent diet (4 RF 21, Mucedola S.r.l., Via G. Galilei, 4, 20019 Settimo Milanese (MI), Italy) was offered ad libitum throughout the study, except as noted in section 4.4.
There was no information available to indicate that any non-nutrient substance likely to influence the effect of the test item was present in the drinking water or the diet. Records of analyses of water and diet are kept on file at RTC.
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on exposure:
- The test item was administered orally by gavage at a dose volume of 5 mL/kg body weight in corn oil.
Control animals received the vehicle alone at the same dose volume. The dose was administered to each animal on the basis of the most recently recorded body weight and the volume administered was recorded for each animal. The required amount of 1,8-Diaminonaphthalene chip was suspended in the vehicle. The
formulations were prepared daily (concentrations of 7.5, 15, 30 and 60 mg/mL). - Details on mating procedure:
- Mating was monogamous (one male to one female). A vaginal smear was taken from the day after the start of pairing until positive identification of copulation (sperm identification, vaginal plug in situ or copulation plugs found in the cage tray). The female was paired with the same male until positive identification occurred or 14 days had elapsed.
- Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Analyses were performed in a separate study in order to validate the analytical method and the formulation procedure and to verify the stability of the formulations (RTC Study no. A3058).
Samples of the formulations prepared during the current study (the first and the last week of treatment) were analysed to check the homogeneity and concentration. Results of the analyses were within the acceptability limits stated in RTC SOPs for suspensions (85-115% for concentration and CV < 10% for homogeneity). Chemical analysis was carried out by the Analytical Chemistry Department at RTC. The software used for this activity was Empower® 2 Build No. 2154. Results of formulation analyses are presented in Addendum 3 of this report - Duration of treatment / exposure:
- Males
Animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing and thereafter through the day before necropsy, for a total of 34-35 days. Dose volumes were adjusted once per week for each animal according to the last recorded body weight.
Females
Animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing and thereafter during pairing, post coitum and post partum periods until Day 13 post partum, for a total of 51-64 days.
Dose volumes were adjusted once per week for each animal according to the last recorded body weight up to mating. During the gestation and lactation periods, dose volumes were calculated according to the last recorded body weight. - Frequency of treatment:
- Animals were dosed once a day, 7 days a week
- Details on study schedule:
- Males were treated for 14 days prior to pairing and during pairing until the day before necropsy, for a total of 34/35 days. Females were treated for 14 days prior to pairing, during pairing and throughout the gestation and lactation periods until Day 13 post partum, for a total of 51 to 64 days.
Doses / concentrationsopen allclose all
- Dose / conc.:
- 0 mg/kg bw/day (nominal)
- Dose / conc.:
- 37.5 mg/kg bw/day (nominal)
- Dose / conc.:
- 75 mg/kg bw/day (nominal)
- Dose / conc.:
- 150 mg/kg bw/day (nominal)
- Dose / conc.:
- 300 mg/kg bw/day (nominal)
- No. of animals per sex per dose:
- 10 Males and 10 Female
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- The following investigations were performed: body weight, clinical signs (including neurotoxicity assessment, motor activity and sensory reactivity to stimuli),
food consumption, clinical pathology investigations (haematology and clinical chemistry in five animals/sex/group randomly selected), macroscopic observations, organ weights.
In addition, oestrous cycle evaluation for parental females (2 weeks before dosing, during pre-mating and mating phases, prior to necropsy), mating performance, thyroid hormone determination and litter data were performed. Clinical signs, anogenital distance, external and/or internal examination were recorded for pups. Thyroid hormone levels were also determined in 1 pup/sex/group randomly selected at Day 14 post partum. Histopathological evaluation was performed in the first instance on five randomly selected control and high dose males and females, as well as on all abnormalities detected during post mortem observation. Subsequently, on the basis of treatment-related findings, the histopathological evaluation was extended to thyroid, kidneys, liver and stomach of all the remaining control and treated animals of both sexes, and on spleen, thymus and adrenals of all the remaining control and treated females. The identification of the stages of the spermatogenic cycle was also performed in five randomly selected males of the control and high dose groups.
Examinations
- Parental animals: Observations and examinations:
- The following investigations were performed: body weight, clinical signs (including neurotoxicity assessment, motor activity and sensory reactivity to stimuli), food consumption, clinical pathology investigations (haematology and clinical chemistry in five animals/sex/group randomly selected), macroscopic observations, organ weights.
In addition, oestrous cycle evaluation for parental females (2 weeks before dosing, during pre-mating and mating phases, prior to necropsy) - Oestrous cyclicity (parental animals):
- A parturition check was performed from Day 20 to Day 25 post coitum.
Females which did not give birth after 25 days of post coitum period were sacrificed shortly after. Female nos. X0890007 and X0890011 (Group 1), X0890051 and X0890053 (Group 3) which did not give birth after 25 days of post coitum period were sacrificed on Days 27 and 26 post coitum, respectively. These animals were found not pregnant at necropsy.
Gestation length was calculated as the time between the day of successful mating (Day 0 post coitum) and the day of birth. The day of birth (when parturition was complete) was defined as Day 0 post partum. - Litter observations:
- On Day 4 post partum, the size of each litter was adjusted by eliminating extra pups by random selection to yield, as nearly as possible, four pups per sex per litter. Partial adjustment (for example, 5 males and 3 females) was acceptable. At least one culled male and one culled female were selected for hormone determination. For litters without extra pups to be culled (i.e.: litters with 8 pups or less), at least 1 female pup was sacrificed for hormone determination in order to retain more male pups for nipple retention on Day 14 post partum.
- Postmortem examinations (parental animals):
- The clinical history of the males and females of the parental generation was studied and a detailed post mortem examination was conducted (including examination of the external surface and orifices). Changes were noted, the requisite organs weighed and the required tissue samples preserved in fixative and processed for histopathological examination.
- Postmortem examinations (offspring):
- All pups found dead in the cage were examined for external and internal abnormalities.
All culled pups sacrificed at Day 4 post partum were subjected to an external examination.
Sex was determined by internal gonads inspection.
All live pups sacrificed on Day 14 post partum were killed and examined for external abnormalities and sex confirmation by gonads inspection. - Statistics:
- Standard deviations were calculated as appropriate. For continuous variables the significance of the differences amongst group means was assessed by Dunnett’s test or a modified t test, depending on the homogeneity of data. The non-parametric Kruskal-Wallis analysis of variance was used for the other parameters and for the evaluation of the hormone thyroid data in pups. Intergroup differences between the control and treated groups were assessed by the non-parametric version of the Williams test. The mean values, standard deviations and statistical analysis were calculated from actual values in the computer without rounding off. Statistical analysis of histopathological findings was carried out by means of the nonparametric Kolmogorov-Smirnov test if ‘n’ was more than 5.
- Reproductive indices:
- All females were examined also for the following:
– number of visible implantation sites (pregnant animals)
– number of corpora lutea (pregnant animals)
Results and discussion
Results: P0 (first parental generation)
General toxicity (P0)
- Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- Semiclosed eyes, lethargy, decreased activity, kyphosis and/or ataxia were the most severe clinical signs recorded during the first days of treatment in both sexes at the dosage of 300 mg/kg/day. Some of these signs, such as decreased acitivity, kyphosis and semiclosed eyes were occasionally recorded also in animals receiving 150 mg/kg/day.
In addition, minor clinical signs such as piloerection, salivation and staining were also present in both sexes at the dosages of 75, 150 and 300 mg/kg/day.
At the dosage of 37.5 mg/kg/day only piloerection was noted in 5 out of 10 males, ocular discharge in 3 out of 10 females and semiclosed eyes in 1 out of 10 females. - Mortality:
- mortality observed, non-treatment-related
- Description (incidence):
- One case of premature death occurred during the study. A single male animal from the high dose group (no. X0890082) was found dead on Day 7 of study. Due to the cannibalization of most organs and tissues, only brown staining of tail was observed. At histopathology, the most relevant findings observed were chronic inflammation, alveolar septal thickening and emphysema of the lungs. The cause of death of this animal was probably due to a mis-dosing.
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- In males, no significant differences were found in body weight between control and treated groups during the study. Body weight gain was statistically significantly decreased before pairing in the mid-low group (on Day 8 of treatment) and high dose group (on Days 8 and 15 of treatment), with respect to controls. From pairing to sacrifice, the body weight gain of treated males was comparable with controls.
In females, no significant differences were found in body weigth between control and treated groups before pairing. During post coitum and post partum periods, statistically significant reductions in body weight were noted in mid-low (up to approximately 12% on Day 1 post partum), mid-high (up to approximately 12% from Day 20 post coitum to Day 4 post partum) and high dose (up to 15% on Day 20 post coitum and Day 4 post partum) groups compared to controls.
Body weight gain was statistically significantly decreased before pairing in mid-high and high dose groups on Days 15 and 8 of treatment, respectively. During post coitum and post partum periods, statistically significant reductions in body weight gain were recorded in the high dose group starting from Day 20 post coitum. This reduction disappeared starting from Day 7 post partum, with values comparable to the controls.
No differences in body weight and body weight gain were recorded in low dose treated animals of both sexes compared to controls. - Food efficiency:
- effects observed, treatment-related
- Description (incidence and severity):
- In males, no significant differences in food consumption were observed during the study.
In females, statistically significant decreases (up to approximately 31% in the high dose group) were noted in all treated groups compared to controls, during the post coitum period.
In the post partum period, a slight but statistically significant decrease was still present (up to 17%) in high dose females up to the end of the study, whereas no differences were recorded in the other treated groups. - Ophthalmological findings:
- not examined
- Haematological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Slight reticulocytosis was recorded in some males dosed at 75, 150 and 300 mg/kg/day.
Mean group data were 33% to 41% above controls, with dose-relation only in animals dosed at 75 and 150 mg/kg/day. Due to the absence of related findings of the other red blood cell parameters (e.g. erythrocytes, haemoglobin, haematocrit), this finding was not considered to be adverse. The statistically significant difference of neutrophils recorded between controls and males dosed at 150 mg/kg/day was not dose-related, therefore it was considered unrelated to treatment. No changes were recorded in treated females.
Coagulation No changes were recorded. - Clinical biochemistry findings:
- effects observed, treatment-related
- Description (incidence and severity):
- Cholesterol was increased in animals of both sexes dosed at 300 mg/kg/day (approximately 43%) and in females receiving 150 mg/kg/day (35%). Protein, albumin and calcium were also increased in animals dosed at 300 mg/kg/day (5% to 11%); albumin was statistically significantly higher than controls also in males receiving 150 mg/kg/day (8%) and calcium was also increased in females dosed at 37.5 mg/kg/day (8%). The statistically significant difference of sodium recorded between controls and males dosed at 75 mg/kg/day was not dose-related, therefore it was considered incidental. The severity of the findings observed was not considered to be suggestive of tissue/organ injury.
- Urinalysis findings:
- not examined
- Behaviour (functional findings):
- no effects observed
- Description (incidence and severity):
- Observation of animals at removal from the cage and in an open arena (neurotoxicity assessment) did not show relevant changes.
- Immunological findings:
- effects observed, non-treatment-related
- Organ weight findings including organ / body weight ratios:
- effects observed, treatment-related
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- At microscopic examination, treatment-related findings were seen in the kidneys (increased hyaline droplet accumulation of marked degree and yellow/brown pigmentation), stomach (hyperplasia of the non glandular squamous epithelium and mucosal erosion), liver (centrilobular hepatocytic hypertrophy associated with minimal to mild degree of inflammatory cell foci) and thyroid (yellow/brown pigmentation) of treated animals of both sexes and in the spleen (yellow/brown pigmentation), thymus (lymphoid atrophy) and adrenals (cortical hypertrophy) of treated females.
No treatment related changes were noted in the evaluation of the seminiferous tubules respect to their stage in the spermatogenic cycle. - Histopathological findings: neoplastic:
- no effects observed
- Other effects:
- not specified
Reproductive function / performance (P0)
- Reproductive function: oestrous cycle:
- no effects observed
- Reproductive function: sperm measures:
- no effects observed
- Reproductive performance:
- no effects observed
Effect levels (P0)
- Key result
- Dose descriptor:
- NOAEL
- Remarks:
- Reproductive and developmental toxicity
- Effect level:
- 300 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- reproductive function (oestrous cycle)
- reproductive function (sperm measures)
- reproductive performance
Results: F1 generation
General toxicity (F1)
- Clinical signs:
- no effects observed
- Mortality / viability:
- mortality observed, non-treatment-related
- Body weight and weight changes:
- not examined
- Food efficiency:
- not examined
- Ophthalmological findings:
- not examined
- Haematological findings:
- not examined
- Clinical biochemistry findings:
- no effects observed
- Urinalysis findings:
- not examined
- Sexual maturation:
- not examined
- Anogenital distance (AGD):
- effects observed, non-treatment-related
- Description (incidence and severity):
- The slight but statistically significant increase in anogenital distance detected in male (4%) and female (1.4%) pups was not considered related to treatment.
- Nipple retention in male pups:
- no effects observed
- Description (incidence and severity):
- No nipples were observed in male pups on Day 14 post partum.
- Organ weight findings including organ / body weight ratios:
- not examined
- Gross pathological findings:
- no effects observed
- Histopathological findings:
- no effects observed
Effect levels (F1)
- Key result
- Dose descriptor:
- other:
- Remarks:
- No effects
- Generation:
- F1
- Basis for effect level:
- other: No effects
- Remarks on result:
- not determinable due to absence of adverse toxic effects
Overall reproductive toxicity
- Key result
- Reproductive effects observed:
- no
Applicant's summary and conclusion
- Conclusions:
- In conclusion, considering the treatment-related changes observed in both sexes at all dose levels together with the large number of target organs involved and the effect considered adverse on the thyroid, it is not possible to establish a NOAEL for general toxicity. Regarding the reproductive and developmental toxicity no adverse effects were observed at all dose levels and the dose of 300 mg/kg/day could be considered the NOAEL for this endpoint.
- Executive summary:
The toxic effects on Sprague Dawley rats of both sexes after repeated dosing with 1,8-Diaminonaphthalene chip, as well as any effects of the test item on male and female
reproductive performance, such as gonadal function, mating behaviour, conception, development of conceptuses, parturition and lactation of the offspring were investigated in this
study. All doses (37.5, 75, 150 and 300 mg/kg/day) were administered orally by gavage. The control group received corn oil. The dose volume was 5 mL/kg body weight. Males were treated for 2 weeks prior to pairing and during pairing with females until the day before necropsy, for a total of 34-35 days. Females were treated for 2 weeks prior to pairing and thereafter during pairing, post coitum and post partum periods until Day 13 post partum (for at least 51 days).
One male in the high dose group (300 mg/kg) was found dead on Day 7 of treatment. The cause of death of this animal was probably due to a mis-dosing.
Semiclosed eyes, lethargy, decreased activity, kyphosis and/or ataxia were the most severe clinical signs recorded during the first days of treatment in both sexes at the dosages of 150 and 300 mg/kg/day.
In males, no relevant differences were found in body weight and food consumption between control and treated groups during the study.
In females, significant reductions in body weight were noted in mid-low, mid-high and high dose groups compared to controls during post coitum and post partum periods up to
the end of the study. In addition, significant decreases were noted in food consumption in all treated females in the post coitum period and in high dose females in the post partum
period.
No findings observed at clinical pathology investigation were considered to be adverse or suggestive of tissue/organ injury. Thyroxine was decreased in males dosed at 75, 150 and
300 mg/kg/day, but due to the absence of changes of the other hormones, this finding was considered to be of limited toxicological importance.
Concerning the reproductive parameters, no relevant differences were found in terms of mating performance including the pre-coital interval (number of days paired to sperm
positive day), copulatory evidence (the positive identification of mating i.e. the presence of sperm and/or copulation plug in situ or in the cage) or fertility index. All pregnant females
had a comparable length of gestation and gave birth on Day 22/23 post coitum. Litter data and sex ratios were unaffected by treatment. No relevant clinical signs were recorded in
control and treated pups during the lactation period and no relevant differences were seen between controls and treated pups in anogenital distance. Necropsy findings in decedent
pups and in pups sacrificed on Days 4 and 14 post partum did not reveal any treatment-related effect, as well as in thyroid weight and hormones analysis of pups sacrificed on Day
14 post partum.
At post mortem examination of parental animals, decrease in terminal body weight was observed in all treated females, when compared to the controls. Increase in kidneys and
liver weights was observed in all treated animals of both sexes, in adrenals weight of all treated females and in the spleen weight of females treated at ≥ 150 mg/kg/day. These
variations were considered treatment-related, on the basis of the histopathological results.
In addition, decrease in thymus weight was observed in females treated at ≥ 150 mg/kg/day.
Treatment-related changes were noted at macroscopic examination such as abnormal dark colouration in thyroid, skin and tail of treated animals of both sexes and in kidneys, liver,
mesenteric lymph nodes, spleen, stomach and head of treated females. These macroscopic changes correlated with histopathological findings.
At microscopic examination, treatment-related findings were seen in the kidneys (increased hyaline droplet accumulation of marked degree and yellow/brown pigmentation), stomach
(hyperplasia of the non glandular squamous epithelium and mucosal erosion), liver (centrilobular hepatocytic hypertrophy associated with minimal to mild degree of inflammatory
cell foci) and thyroid (yellow/brown pigmentation) of treated animals of both sexes and in the spleen (yellow/brown pigmentation), thymus (lymphoid atrophy) and adrenals (cortical
hypertrophy) of treated females.
No treatment related changes were noted in the evaluation of the seminiferous tubules respect to their stage in the spermatogenic cycle.
In conclusion, considering the treatment-related changes observed in both sexes at all dose levels together with the large number of target organs involved and the effect considered
adverse on the thyroid, it is not possible to establish a NOAEL for general toxicity. Regarding the reproductive and developmental toxicity no adverse effects were observed at all dose levels and the dose of 300 mg/kg/day could be considered the NOAEL for this endpoint.
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