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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Not stated
Reliability:
1 (reliable without restriction)
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1997
Report Date:
1997

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Version / remarks:
(1981)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPP 82-1 (90-Day Oral Toxicity)
Version / remarks:
(1984)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OTS 798.2650 (90-Day Oral Toxicity in Rodents)
Version / remarks:
(1989)
Deviations:
no
Qualifier:
according to
Guideline:
other: JMAFF 59 NohSan No. 4200 (Guidance on toxicology study data for application of agriculture chemical registration) (1985)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Technical Grade MKH 3586
- Physical state: beige powder
- Analytical purity: 97.8 - 98.2%
- Impurities (identity and concentrations):
- Purity test date: June 1994, Dec 1994, Aug 1995
- Lot/batch No.: 17004/93
- Expiration date of the lot/batch: Stability of lot confirmed for the duraton of the study

Test animals

Species:
rat
Strain:
Fischer 344
Sex:
male/female

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
90 days / dietary
Frequency of treatment:
Continuous treatment
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
M & F: 0, 100, 250, 500, 1000, 2500, 5000 ppm
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
M/F:0, 6.9/7.9, 17.8/20.5, 32.9/38.0, 67.2/77.7, 181.8/200.6, 353.8/396.8 mg/kg/day
Basis:
actual ingested
No. of animals per sex per dose:
30 animals/sex/dose for control and high dose group
15 animals/sex/dose for other groups
Control animals:
yes, plain diet

Results and discussion

Effect levels

Dose descriptor:
NOAEL
Effect level:
ca. 500 ppm
Based on:
other: liver, thyroid, pancreatic and haematological changes at >1000 ppm
Sex:
male/female
Basis for effect level:
other: (equivalent to 32.9/38.0 mg/kg/day [M/F])

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

OBSERVATIONS:

Clinical signs of toxicity:

With the exception of increased frequency of food spillage noted in 5000 ppm males and females no other signs were related to exposure to the test material:

 

Table 7.5.1.2: Incidence of food spillage

Dose

(ppm)

Males

Females

0

0

0

100

0

0

250

0

1

500

0

1

1000

0

1

2500

0

1

5000

7*

4*

*Statistically significant

 

Mortality:

Mortality in treatment groups was comparable to the concurrent control group.

 

Bodyweight and bodyweight gain:

Bodyweight gain was unaffected up to 500 ppm, at doses above this a decline in both genders was observed:

 

Food consumption:

Refer to Table 7.5.1.1 for actual ingested levels of test material. Food consumption and utilisation was unaffected in either sex

 

Table 7.5.1.3: Bodyweight gain %relative change respective to control group

Dose

(ppm)

Males

Females

1000

6%

9%

2500

16%

14%

5000

19%

19%

 

Opthalmoscopic examination:

No test material related effects were observed.

 

Urinalysis:

Treatment related increases in urinary pH (500 – 5000 ppm, F), urinary leukocyte numbers (2500 and 5000 ppm, M &F) and urinary protein concentration (5000 ppm, M). The actual toxicological significance of these parameter changes was unknown.

 

Haematology & clinical chemistry:

Haematology:

 

Table 7.5.1.4: Summary of haematological changes, gain %relative change respective to control group

Parameter

Males

(ppm)

Females

(ppm)

increased reticulocyte number

2500, 5000

2500, 5000

increased platelet count

2500, 5000

-

decreased erythrocyte count

1000, 2500, 5000

5000

Decreased haemoglobin and haemocrit

1000, 2500, 5000

1000, 2500, 5000

Decreased MCV, MCHb

2500, 5000

1000, 2500, 5000

 

These changes observed were consistent with haemolytic anaemia, which was confirmed at necropsy manifest as increased incidence of pigmentation (suggestive of increased destruction of RBC rather than a direct effect on RBC production).

 

Clinical chemistry: decrease in hepatic aminopyrine N-demethylase activityin males at 1000, 2500 and 5000 ppm. An increase in hepatic p-nitroanisole O-demethylase activity in females (1000, 2500 and 5000ppm) and males (2500 and 5000 ppm) was observed. In light of the findings at necropsy (hepatocytomegaly and karyomegaly) these alterations may have been an adaptive response by the liver to increased need to facilitate metabolism and excrete exogenously administered test material.

An increase in serum thyroxine concentration was observed in males at 1000 ppm and greater, with increases in serum T3in males (1000, 2500, 5000 ppm) and females (2500, 5000 ppm). Refer to sub-chronic 90 day mechanistic study for mode of action.

 

Sacrifice and Gross Pathology:

No gross lesions observed at necropsy.

 

Histopathology:

Liver:Increased incidence of hepatocellular changes observed in males (1000, 2500, 5000 ppm) and females (2500, 5000 ppm). The lesion was characterised by both cytomegaly and enlarged cells with granular eosinophilic to occasionally vacuolated cytoplasm and karyomegaly, prominent nuclear enlargement which increased in severity with increasing dose.

 

Thyroid:Increased incidence of follicular cell hyperplasia was observed at 2500 and 5000 ppm in both genders. The lesion was characterised as minimally enlarged cells, more columnar in type, with increased numbers of active follicles within the central area of the gland as evidenced by increased colloid secretion.

 

Adrenal: increased incidence of diffuse cytoplasmic vacuolisation of the adrenal cortical epithelial cells observed in males at 5000 ppm, characterised by multiple small vacuoles that gave the cells an overall pale appearance.

 

Pancreas: increased incidence of cytoplasmic vacuolisation was observed at 5000 ppm in both genders. The lesion was characterised by vacuolar change often associated with dense bodies representative of subtle degeneration or apoptosis in the acinar cells.

 

Spleen:increased incidence of pigmentation was observed in males (1000, 2500, 5000 ppm) and females (2500, 5000 ppm). special stains suggested deposition of haemosiderin (iron containing pigment derived from Hb of disintegrated RBC).

 

Bone marrow: atrophy of bone marrow observed in 2500 and 5000 ppm females. Additionally, an increased incidence of atrophy or inactivity of the growth plate was observed in 2500 and 5000 ppm females.

 

Organ weights:

Test material related increases in liver weights were observed in both males and females of the high dose group where liver weights were 43 (M) and 41% (F) higher than respective control groups.

Applicant's summary and conclusion

Conclusions:
Based on the results of this study, the NOAEL was 500 ppm (32.9 mg/kg/day for males and 38.0 mg/kg/day for females) based on liver, thyroid, pancreatic and haematological changes observed at doses of 1000 ppm and greater.
Executive summary:

Seven groups, 6 treatment and 1 control group of 30 animals/dose/sex, for the control and high dose groups or 15 animals/sex/group for all other groups were fed MKH 3586 in the diet for 13 consecutive weeks. Dietary concentrations of 0, 100, 250, 500, 1000, 2500 or 5000 ppm were administered, equivalent to compound intakes of 0, 6.9, 178, 32.9, 67.2, 181.8 and 353.8 mg/kg/day for males and 0, 7.9, 20.5, 38.0, 77.7, 200.6, 396.8 mg/kg/day for females respectively. Following the exposure period of the study, 15 animals/sex of the control and high dose groups were placed on control diet (recovery period) for a further 4 weeks to assess any reversible effects.

 

Test material related changes were in the liver (hepatocytomegaly and karyomegaly), pancreas (cytoplasmic vacuolisation), spleen (pigmentation due to deposition of haemosiderin) and bone marrow (atrophy of the marrow along with inactivity of the growth plate). These changes occurred at doses of 1000 ppm and greater. Haematological changes were representative of haemolytic anaemia, suggestive of increased destruction of RBC, rather than a direct effect on RBC production.

 

Based on the results of this study, the NOAEL was 500 ppm (32.9 mg/kg/day for males and 38.0 mg/kg/day for females) based on liver, thyroid, pancreatic and haematological changes observed at doses of 1000 ppm and greater.