Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Toxicological information

Repeated dose toxicity: oral

Currently viewing:

Administrative data

short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study without detailed documentation

Data source

Referenceopen allclose all

Reference Type:
An antioxidant, N,N 0 -diphenyl-p-phenylenediamine (DPPD), affects labor and delivery in rats: A 28-day repeated dose test and reproduction/developmental toxicity test
Matsumoto M. et al.
Bibliographic source:
Food and Chemical Toxicology 56 (2013) 290–296
Reference Type:
other: tables from study report

Materials and methods

Test guidelineopen allclose all
according to guideline
other: the notice on the test method concerning new chemical substances ( No. 1121002, Pharmaceutical and Food Safety Bureau, MHLW; No.2, Manufacturing Industries Bureau, METI; No. 031121002, Environmental Policy Bureau, MOE)
Version / remarks:
November 21, 2003
according to guideline
other: see version/remarks
Version / remarks:
the standard for the test facility conducting tests concerning new chemical substances, etc. (November 21, 2003, No. 1121003 Pharmaceutical and Food Safety Bureau, MHLW; No. 3 Manufacturing Industries Bureau, METI; No. 031121004 Environmental Policy Bureau, MOE)
GLP compliance:
not specified

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Test material form:
Specific details on test material used for the study:
- Source and lot No. of test material: Seiko Chemical (Tokyo, Japan), 307605R
- Purity test date: 99.87%
- Name (as cited in article): DPPD

Test animals

other: [Crl:CD(SD)] SPF
Details on test animals or test system and environmental conditions:
- Source: Atsugi Breeding Center, Charles River Japan, Inc., (Kanagawa, Japan)
- Age at study initiation: Five weeks
- Weight at study initiation: male, 152-172 g; female, 130-147 g
- Housing: individually
- Diet: Basal diet, Labo MR Stock; NOSAN corporation (Tokyo, Japan), ad libitum
- Water: ad libitum

- Temperature (°C): 21.9–23.0
- Humidity (%): 55–61
- Air changes (per hr): 10 and more
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Details on oral exposure:
Dosing volume: 5 mL/kg bw
Duration of treatment / exposure:
28 days
Frequency of treatment:
Once daily
Doses / concentrationsopen allclose all
Dose / conc.:
100 other: mg/kg bw/day
Dose / conc.:
300 other: mg/kg bw/day
Dose / conc.:
1 000 other: mg/kg bw/day
No. of animals per sex per dose:
- control, 1000 mg/kg bw/day: 10
- 100, 300 mg/kg bw/day: 5
Control animals:
yes, concurrent vehicle
Details on study design:
- After the dosing period, five rats per each sex at 0 and 1000 mg/kg bw/day were reared for 14 days without administration of DPPD as the recovery groups. The volume of each dose was adjusted to 5 mL/kg body weight based on the latest body weight.


Observations and examinations performed and frequency:
- Time schedule: Weekly

- Time schedule: Clinical signs in detailed observation in all animals were recorded one day before the administration period and once a week during the administration period.

- Time schedule for examinations: days 1, 7, 14, 21, and 28 of the administration period, days 7 and 14 of the recovery period, and on the day of necropsy.

- Time schedule for examinations: Once a week during both administration and recovery periods.
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: yes

- Time schedule for collection of blood: day of euthanizing (1 day after final administration or 1 day after completion of the recovery period)
- Anaesthetic used for blood collection: Yes, for euthanizing by exsanguination
- Animals fasted: Yes
- Collected blood samples were examined for haematology by an automated haematology analyzer (XT-2000i, Sysmex Co., Kobe, Japan) and automatic coagulometer (KC-10A, Amelung, US). Serum biochemistry was tested by an automatic analyzer (JCA-BM8, JEOL, Tokyo, Japan) and automated electrolyte analyzer (NAKL-132, TOA electronics Ltd., Tokyo, Japan)

- Time schedule for collection of urine: Fresh urine was sampled from animals on day 22 of the administration period and on day 8 of the recovery period.
- Urine samples were tested for colour, pH, protein, glucose, ketone bodies, bilirubin, occult blood, and urobilinogen.

- Time schedule for examinations: Day 27 of the administration period and day 13 of the recovery period.
- Battery of functions tested: Sensory reactions for a sight reaction, hearing reaction, sense of touch reaction, pain reaction, pupil reflex, and righting reflex; grip strength of fore and hind limbs and spontaneous motor activity
Sacrifice and pathology:
Euthanized by exsanguination, 1 day after final administration or 1 day after completion of the recovery period

External surfaces of the rats were examined. Abdomen and thoracic cavities were opened, and gross internal examination was performed. The brain, thymus, heart, liver, spleen, kidney, adrenal gland, thyroid gland, pituitary gland, testis, epididymis, and ovary were isolated and weighed.

Histopathological evaluations were performed on the organs mentioned in ‘gross pathology’ in addition to the eye ball, spinal cord, lung, trachea, stomach, intestines, prostate, seminal vesicle, vagina, uterus, urinary bladder, sciatic nerve, lymph nodes, and bone marrow (femur) in control and highest dose groups.
Other examinations:
- Bilirubin levels significantly increased without being related to toxicological effects in males. Because both bilirubin and the test compound contain – NH substitutes, the interference of DPPD with bilirubin measurements was anticipated.
- The interference of test compound with bilirubin measurements was tested as follows. Serum samples were taken from untreated male rats, and 0.2 mL of test compound at 0.001, 0.01, 0.1, and 1 mg/mL (1:1 aceton and dimethyl sulfoxide) was added to 0.5 mL serum of rats. In addition, rat liver S9 was added to test compound at 0.1 mg/mL to test the interference of test compound metabolites. Bilirubin levels were measured by the diazo method.
To assess the homogeneity of data, parametric data were analysed with Bartlett’s test or the F-test. When homogeneity was recognized, data were analysed using a one-way analysis of variance or the Student’s t-test. Non-homogeneous data were analysed with Kruskal–Wallis’s rank test or the Aspin–Welch t-test. Nonparametric data were analysed with Kruskal–Wallis’s rank test or Mann–Whitney’s U test. The Dunnett test or Dunnett type test was used to assess multiple comparisons. Fisher’s exact test was used to assess categorical data. Five per cent levels of probability were used as the criterion for significance.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
- Food consumption significantly decreased at 300 and 1000 mg/kg bw/day in males
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
- Total bilirubin significantly increased in all treatment groups at the end of the treatment period in males, but it was not observed at the end of the recovery period. When the test compound was added to rat serum, bilirubin levels measured by the diazo method increased in a concentration-related manner with or without the rat S9 mix. Therefore, increased bilirubin levels in this study were considered to be due to interference by the test compound.
- In females, γ-GTP significantly decreased (0.63 IU/L) at 1000 mg/kg bw/day at the end of the administration period, but it was within the background data of the facility (0.31–2.06 IU/L) and was not considered to be toxicologically significant. This change was not observed at the end of the recovery period.
Urinalysis findings:
effects observed, non-treatment-related
Description (incidence and severity):
- Protein levels significantly decreased in all treatment groups, but this was not dose dependent and was considered to be due to spontaneously occurring higher levels in control groups.
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
- Relative and absolute weights of the thyroid gland in males and absolute weight of the kidney in females significantly increased at 100 mg/kg bw/day, but histopathological changes were not significantly different in these organs at the end of the administration period.
- No other effects were observed in organ weights in both sexes.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Slight hydrometra in the uterus was found in one female at 300 mg/kg bw/day, and dilatation of the lumen was histopathologically observed in the uterus of this female at the end of administration period; however, no gross or histopathological effects in the uterus were observed at 1000 mg/kg bw/day.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined

Effect levels

Dose descriptor:
Effect level:
1 000 other: mg/kg bw/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain
clinical biochemistry
clinical signs
gross pathology
histopathology: non-neoplastic
organ weights and organ / body weight ratios

Target system / organ toxicity

Critical effects observed:

Applicant's summary and conclusion