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Diss Factsheets

Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted and reported according to the OECD combined repeat dose and reproductive/developmental toxicity test 422 and principles of GLP.
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
2003
Report date:
2003

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)
Deviations:
no
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2'-methylacetoacetanilide
EC Number:
202-267-0
EC Name:
2'-methylacetoacetanilide
Cas Number:
93-68-5
Molecular formula:
C11H13NO2
IUPAC Name:
N-(2-methylphenyl)-3-oxobutanamide
Details on test material:
TS: Mitsuboshi Chemical Co., Ltd.
Purity: 99.9%

Test animals

Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ORGANISMS
Age: 9 weeks for male, 8 weeks for female
Weight at initiation: 343-391 g for male, 211-241 g for female
Number of animals: 10 per sex per dose
Pellet food and water: free take
MATING PROCEDURE
one by one in each cage
(All of those 10 pairs had finished mating by Day 4.)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: 1% Methylcellulose water solution
Details on oral exposure:
ADMINISTRATION
Vehicle: 1% methylcellulose water solution, 0.5mL/100g body weight
Type of administration: gavage, once a day
Duration of administration:
male; 44 days (including 14 days before mating)
female: 41-45 days (from 14 days before mating to 3 days after
parturition)
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
no data available.
Duration of treatment / exposure:
males: 44days, females: from 14days before mating to Day 3 of lactation
(41 - 45days)
Frequency of treatment:
one administration per day
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 8, 25, 80, 250 mg/kg/day
Basis:
other: 1% Methylcellulose water solution
No. of animals per sex per dose:
10 animals per sex per dose
Control animals:
yes, concurrent vehicle
Details on study design:
AAOT was administered to Sprague-Dawley rats (10/sex/dose) at doses of 0, 8, 25, 80, 250 mg/kg/day
by oral gavage. The dosing period was 44 days for males and 41 - 45 days (including 14 days before mating
and 3 days after pregnancy) for females.
Positive control:
No data available.

Examinations

Observations and examinations performed and frequency:
CLINICAL OBSERVATIONS AND FREQUENCY
Clinical signs and mortality: every day
Body weight: once a week, and the time of termination
Food consumption: at every body weight check (24hr consumption)
Water consumption: not checked
Sacrifice and pathology:
Organs: by male after extraction of blood, and by female at day 4 after (estimated) pregnant;
for weight check; brain, liver, kidney, spleen, heart, thymus, thyroid, pituitary, adrenals, testes and epididymides
for observation; above mentioned ones plus, lung, stomach, bladder,
medulla, spinal cord, sciatic nerve, etc.
Other examinations:
HISTOPATHOLOGICAL OBSERVATIONS
Urinalysis: by male at Day 39 - 43; pH, blood, protein, ketones, bilirubin, urobilinogen, specific gravity, deposit and appearance
Hematology: by male at day 45 (stopped feeding at 17:00 on the day before terminal kill); erythrocyte count, hemoglobin, hematocrit, MCV,
MCH, mean corpuscular hemoglobin(MCHC), leukocyte count, platelet count, reticulocyte count, Heinz-body and methemoglobin
Blood biochemical: Same sample as hematology was used.; total protein, albumin, albumin/globulin(A/G) ratio, glucose, triglyceride,
total cholesterol, total bilirubin, nitrogen of urea, creatinine, GOT, GPT, gamma-GTP, lactate dehydrogenase(LDH),
alkaline phosphatase, cholin esterase, calcium, phosphate, sodium and potassium
Statistics:
No data available.

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Description (incidence and severity):
No change in mortality and behavior were observed in any groups
Mortality:
no mortality observed
Description (incidence):
No change in mortality and behavior were observed in any groups
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No toxicological effect was observed in any groups
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No toxicological effect was observed in any groups
Food efficiency:
not examined
Description (incidence and severity):
No findings
Water consumption and compound intake (if drinking water study):
not examined
Description (incidence and severity):
not checked
Ophthalmological findings:
not examined
Description (incidence and severity):
not checked
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
see table under section "any other information on results inc. tables"
Clinical biochemistry findings:
effects observed, treatment-related
Description (incidence and severity):
see table under section "any other information on results inc. tables"
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
Male: Increases of specific gravity was observed in 250mg/kg group. It's likely within normal range, and no related change was observed in another check items.
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
no findings
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
see table under section "any other information on results inc. tables"
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
see table under section "any other information on results inc. tables"
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
see table under section "any other information on results inc. tables"
Histopathological findings: neoplastic:
no effects observed
Description (incidence and severity):
No findings
Details on results:
In the 250 mg/kg/day group the following effects were observed: decreases of erythrocyte count, hemoglobin concentration
and hematocrit value in males; increases of mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH),
reticulocyte count, methemoglobin concentration, bilirubin and potassium in males; increase of pituitary weight in males;
increases of weight of spleen, weight of liver, extramedullary hematopoiesis and congestion in spleen in both sexes; and
blackening of spleen and hemosiderin deposit in liver and spleen in both sexes.
In the 80 mg/kg/day group the following effects were observed: decrease of erythrocyte count and increases of MCV and
bilirubin in male; increase of congestion in spleen in female; and blackening of spleen and hemosiderin deposit in liver and spleen in both sexes.
No changes in mortality, behavior or toxic effect on the body weight and food consumption were observed in any groups.
Increase of specific gravity of urine was observed in males of the 250 mg/kg group. However no related changes were observed in other findings.

Effect levels

Dose descriptor:
NOAEL
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see 'Remark'

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

HEMATOLOGICAL AND BLOOD CHEMICAL FINDINGS IN MAL

Dose (mg/kg/day)

0

8

25

80

250

erythrocyte count:

mean corpuscular volume

(MCV):

hemoglobin concentration:

hematocrit value:

mean corpuscular hemoglobin

(MCH):

reticulocyte count:

methemoglobin concentration:

Heinz-body in erythrocytes:

bilirubin:

potassium:

-

 

-

-

-

 

-

-

-

-

-

-

-

 

-

-

-

 

-

-

-

-

-

-

-

 

-

-

-

 

-

-

-

-

-

-

D

 

I

-

-

 

-

-

-

-

-

-

D

 

I

D

D

 

I

I

I

O

I

I

-: normal or nothing, I: increased, D: decreased, O: observed

HISTOPATHOLOGICAL FINDINGS, ETC. IN MALE

Dose (mg/kg/day)

0

8

25

80

250

blackening of spleen:

enlargement of spleen:

weight of spleen:

weight of pituitary:

weight of liver:

hemosiderin deposit in

liver and spleen:

extramedullary hematopoiesis:

congestion in spleen:

eosinophilic body in

tuble of kidneys:

-

-

-

-

-

 

-

-

-

 

-

-

-

-

-

-

 

-

-

-

 

-

-

-

-

-

-

 

-

-

-

 

-

O

I

-

-

-

 

O

-

-

 

-

O

-

I

I

-

 

O

I

I

 

I

-: normal or nothing, I: increased, D: decreased, O: observed

HISTOPATHOLOGICAL FINDINGS, ETC. IN FEMALE

Dose (mg/kg/day)

0

8

25

80

250

blackening of spleen:

enlargement of spleen:

weight of spleen:

weight of pituitary:

weight of liver:

hemosiderin deposit in

liver and spleen:

extramedullary hematopoiesis:

congestion in spleen:

eosinophilic body in

tuble of kidneys:

-

-

-

-

-

 

-

-

-

 

-

-

-

-

-

-

 

-

-

-

 

-

-

-

-

-

-

 

-

-

-

 

-

O

-

-

-

-

 

O

-

I

 

-

O

O

I

-

I

 

O

I

I

 

-

-: normal or nothing, I: increased, D: decreased, O: observed

Applicant's summary and conclusion

Conclusions:
Toxicological effects and the target organs are hemolytic anemia and the related changes on the blood, spleen, liver and kidney,
including male kidney (increasing of eosinophilic bodies) and female liver (increasing of the weight).
The NOAEL for repeat dose toxicity to rats is 25 mg/kg/day in both sexes.
Executive summary:

The Combined Repeat Dose and Reproduction/Developmental Toxicity Screening Test (MHW Japan 1999b) was well conducted according to OECD TG422, following GLP and published in the OECD SIDS Dossier in year 2003.

The test results are described as follows. AAOT was administered to Sprague-Dawley rats (10/sex/dose) at doses of 0, 8, 25, 80, 250 mg/kg/day by oral gavage. The dosing period was 44 days for males and 41 - 45 days (including 14 days before mating and 3 days after pregnancy) for females.

The blood findings in males in the 250 mg/kg/day group were: decreases of erythrocyte count, hemoglobin concentration and hematocrit value, also increases of mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), reticulocyte count, methemoglobin concentration, bilirubin and potassium. Other findings in the 250 mg/kg/day group were: increase of pituitary weight in males; increases of weight of spleen, weight of liver, extramedullary hematopoiesis and congestion in spleen, also blackening of spleen and hemosiderin deposit in liver and spleen in both sexes. The blood findings in males in the 80 mg/kg/day group were: decrease of erythrocyte count and increase of MCV and bilirubin. Other findings in the 80 mg/kg/day group were: increase of congestion in spleen in females, blackening of spleen and hemosiderin deposit in liver and spleen in both sexes. In all dose groups up to 250 mg/kg/day, no changes in mortality, behavior or toxic effects on the body weight and food consumption were observed in any sexes. No toxic effects were observed in any dose groups up to 25 mg/kg/day.

The NOAEL for repeat dose toxicity to rats is 25 mg/kg/day in both sexes.