1,2-dichloro-4-nitrobenzene

Toxicity to reproduction: other studies

Administrative data

Endpoint:

toxicity to reproduction: other studies

Type of information:

experimental study

Adequacy of study:

other information

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: OECD TG 407 including examination of reproductive organs

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

other information

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study, GLP

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: approximately 6 week
- Housing: in groups of 5 animals
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22
- Humidity (%): 50
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

other: sesame oil

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
the test compound was suspended homogenousely in the vehicle by means of a magnetic stirrer

VEHICLE
- Justification for use and choice of vehicle (if other than water):
- Concentration in vehicle: 0 - 2 % (w/v)
- Amount of vehicle (if gavage): 5 ml/kg bw

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

the determination of the concentration was performed by photometrical detection after HPLC seperation on a reversed phase column

Duration of treatment / exposure:

28 days

Frequency of treatment:

once per day

Dose / conc.:

0 mg/kg bw/day (nominal)

Dose / conc.:

4 mg/kg bw/day (nominal)

Dose / conc.:

20 mg/kg bw/day (nominal)

Dose / conc.:

100 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5 animals/sex/dose group

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: based on preliminary dose range finding study

Positive control:

not relevant

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: at the start of the study and then twice weekly throughout the study

FOOD CONSUMPTION
twice weekly

WATER CONSUMPTION
once weekly

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: weekly

HAEMATOLOGY: Yes
At the termination of the study, hematological examinations were performed on all animals without previous withdrawl of food. Blood samples were taken from the retrobarbital venous plexus in narcosis. In order to prevent systematic errors, blood sampling was conducted in a randomized order.
hematological parameters:
erythrocytes, hemoglobin, hematocritm mean cellular volumd (MCV), Mean cellular hemoglobin (MCH), Mean cellular hemoglobin concentration(MCHC), Leucocyte count, thrombocyte count, differential leucocyte count, and red cell morphology, reticulocyte count, Heinz bodies, coagulation time

CLINICAL CHEMISTRY: Yes
After blood sampling for hematology the animals were killed by cutting of the vena cava cranialis in deep narcosis and exanguinated. In order to prevent systematic errors, exanguination was conducted in a randomized order.
Parameters:
sodium, potassium, inorganic phosphorus, uric acid, bilirubin total, creatinene, serum-glucose, urea-nitrogen, calcium, chloride, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, gamma-glutamyltransferase, total protein, albumin

URINALYSIS: Yes
urinalysis was performed on all animals a few days before termination of the study. for this purpose, the urine was collected by using metabolism cages (Overnight from day 26 to day 27)
Parameters:
appearance, colour, pH-value, hemoglobin, protein, glucose, ketone bodies, bilirubin, urobilinogen, specific weight, sediment, volume

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: weekly

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
After exanguination all animals were necropsied and checked for macroscopically visible abnormalities. The autopsy included macroscopic examination of the skin, orifices, eyes, teeth, oral mucosa and internal organs.

organ weights (calculation of organ to body weight ratio): heart, lung, liver, kidneys, spleen, ovaries, testes, epididymides, adrenals, brain, thymus

HISTOPATHOLOGY: Yes
Heart, liver, kidneys, adrenals, spleen, lung, brain, thymus, ovaries, testes, epididymides, trachea, stomach, jejunum, colon, urinary bladder, uterus, prostata, seminal vesicles, skeletal muscles, N. ischiadicus, femur with bone marrow, spinal cord (cervical, thoracal and lumbal), lymph nodes (dervical, and iliacal)

Other examinations:

no further data

Statistics:

---One way analyses of variance with sequentially rejective multiple comparison
---One way analyses of variance based on ranks with equentially rejective multiple comparison
---Trend Test analyses for non-neoplastic lesions (ARMITAGE)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

100 mg-group, m/f: irregular respiration, stilted gain
>= 20 mg/kg bw/day, m/f: increased salivation

Mortality:

mortality observed, treatment-related

Description (incidence):

No death occurred throughout the study

Body weight and weight changes:

no effects observed

Description (incidence and severity):

all groups: body weight gain was not impaired

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

food consumption unaffected

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

>=20 mg/kg bw/day(m/f): slightly increased water intake (not significant, not dose dependent)

Ophthalmological findings:

no effects observed

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

4 mg/kg bw/day:
male: significant decrease in erythrocyte counts (6.98 10E12/l versus 7.63 10E12/L [historical control: 6.34-8.95 10E/L])

20 mg/kg bw/day:
male: significant decrease in erythrocyte counts (6.87 10E12/L versus 7.63 10E12/L of concurrent control) and haematocrit (0.42 UNITY versus 0.46 UNITY of control)
female: significant increase in MCV (62 10E-15L versus 58 10E-15L)

100 mg/kg bw/day:
male: decrease in erythrocyte values (6.34 10E12/L versus 7.63 10E12/L of control), decrease in haematocrit (0.42/0.39 UNITY versus 0.46/0.41 UNITY of control), decrease in haemoglobin (140/131 g/L versus 149/138 g/L of control), increase in MCV values 67/66 10E-15L versus 60/58 10E-15L of control), reticulocyte counts (0.077/0.080 UNITY versus 0.011/0.008 UNITY of control)

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

male: significant increase in sodium- and chlorid-ions when compared to concurrent control, which were, however, within the historical control values of this strain:
control//4/20/100 mg/kg bw/day//historical control range:
Sodium: 135//139/142//142//132-149 mmol/L, Chlorid: 98// 101/102/102//95-106 mmol/L
20 mg/kg bw:
increase in alk. phosphatase (f: 261 U/L versus 175 U/L of control)
100 mg/kg bw/day:
increase in urea values (m/f: 8.44/8.17 mmol/L versus 5.62/6.80 mmol/L of control) )m [indicative for an impaired kidney function; however, no histopathological correlates were found]
increase in ALAT(GPT) (m: 54 U/L versus 44 U/L), increase in alk. phosphatase (f: 267 U/L versus 175 U/L of control)

Endocrine findings:

not examined

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

dark yellow discolouration of urine: male: from 20 mg/kg bw/day onwards; female: at 100 mg/kg bw/day;
pH-Value: female: at 100 mg-group significantly decreased: pH=5.4 versus pH=6.1 (control)

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

absolut organ weights not affected, rel. organ weights:
20 mg/kg bw
increased relative liver weight (m: 4.545 % versus 4.036 % of control)
100 mg/kg bw/day
increase in rel. liver weight (m/f: 4.571/4815 % versus 4.036/3.926 % of control)
increased rel. spleen weight (m/f: 0.321/0.330 % versus 0.189/0.234 % of control)

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

100 mg/kg bw, spleen: 5/5 m and 5/5 f dark discoloration spleen

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

m(grading)-f(grading), low, mid, high dose versus control:
congestion
low: 0/5(0) - 0/5(0), mid: 3/5(slight) - 5/5(slight), high: 5/5(moderate) - 5/5(moderate) versus contr.: 0/5 - 0/5
increased extramedullary haematopoiesis:
low: 2/5(minimal) - 2/5(minimal), mid: 5/5(minimal) - 5/5(slight), high: 5/5(slight) - 5/5(slight) versus contr.: 3/5(minimal) - 0/5
increase in haemosiderosis:
low: 0/5 - 5/5(slight), mid: 5/5(minimal) - 5/5(marked), high: 5/5(moderate) -5/5(marked) versus contr.: 0/5 - 5/5(mild)

Histopathological findings: neoplastic:

no effects observed

Dose descriptor:

NOAEL

Effect level:

4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: based on changes in red blood cell parameters, on hemolytic anemia with microscopic changes in the spleen and increase in relative liver weights in the next higher dose and increase in relative spleen weight at 100 mg/kg bw/d

Critical effects observed:

not specified

Conclusions:

The NOAEL was 4 mg/kg bw/d based on changes in red blood cell parameters, on hemolytic anemia with microscopic changes in the spleen.

Executive summary:

In a study according to OECD Guideline 407 male and female Sprague-Dawley rats received 0, 4, 20, 100 mg/kg bw/d dissolved in sesame oil by gavage for 28 days. The NOAEL was 4 mg/kg bw/d based on changes in red blood cell parameters, on hemolytic anemia with microscopic changes in the spleen and increase in relative liver weights in the next higher dose and increase in relative spleen weight at 100 mg/kg bw/d (Hoechst AG 1993).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: OECD TG 407

Deviations:

not applicable

Principles of method if other than guideline:

examination of reproductive organs in a repeated dose toxicity study according to OECD TG 407

GLP compliance:

yes

Type of method:

in vivo

Test material

Test material information

Constituent 1

Reference substance name:

1,2-dichloro-4-nitrobenzene

EC Number:

202-764-2

EC Name:

1,2-dichloro-4-nitrobenzene

Cas Number:

99-54-7

Molecular formula:

C6H3Cl2NO2

IUPAC Name:

1,2-dichloro-4-nitrobenzene

Details on test material:

1,2-dichloro-4-nitrobenzene, purity 99 %

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: approximately 6 week
- Housing: in groups of 5 animals
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22
- Humidity (%): 50
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: Sesame oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
the test compound was suspended homogenousely in the vehicle by means of a magnetic stirrer

VEHICLE
- Justification for use and choice of vehicle (if other than water):
- Concentration in vehicle: 0 - 2 % (w/v)
- Amount of vehicle (if gavage): 5 ml/kg bw

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

the determination of the concentration was performed by photometrical detection after HPLC seperation on a reversed phase column

Duration of treatment / exposure:

28 d

Frequency of treatment:

daily

Duration of test:

28 days

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

4 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

20 mg/kg bw/day (nominal)

Doses / concentrations 4

Dose / conc.:

100 mg/kg bw/day (nominal)

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: based on preliminary dose range finding study

Statistics:

---One way analyses of variance with sequentially rejective multiple comparison
---One way analyses of variance based on ranks with equentially rejective multiple comparison
---Trend Test analyses for non-neoplastic lesions (ARMITAGE)

Results and discussion

Observed effects

see 'Any other information on results'

Any other information on results incl. tables

OBSERVATIONS:
No death occurred throughout the study
unspecific signs of intoxication: 100 mg-group, m/f: irregular respiration, stilted gain, >= 20 mg/kg bw/day, m/f: increased salivation;
all groups: body weight gain was not impaired, food consumption unaffected, >=20 mg/kg bw/day(m/f): slightly increased water intake (not significant, not dose dependent)
REPRODUCTIVE ORGAN EVALUATION:
no pathologic findings were reported.

Applicant's summary and conclusion

Conclusions:

No pathologic findings on reproductive organs were reported.

Executive summary:

During 28-day-treatment of male and female Wistar rats with 0, 4, 20 or 100 mg/kg bw/d 1,2 -dichloro-4 -nitrobenzene dissolved in sesame oil according to OECD TG 407 reproductive organs were not affected (Hoechst AG 1993).