Registration Dossier

Administrative data

Endpoint:
toxicity to reproduction
Remarks:
other: combined study (repeated dose and reproduction/developmental toxicity screening)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Qualifier:
according to
Guideline:
other: EPA OPPTS 870.3650 Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Methylaminoethanol (Test item No.07/0540-2)
- Substance type: organic
- Physical state: colorless liquid
- Analytical purity: 99.7 area-% (Analytical Report: 07L00323)

- Purity test date: November 06-07, 2007
- Lot/batch No.: from continuous production
- Expiration date of the lot/batch: 24 Oct 2007


- Stability under test conditions: The stability of the test item under storage conditions over the test period was guaranteed by the sponsor, and the sponsor holds this responsibility
- Storage condition of test material: Room temperature, under N2
- Other:

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH
- Age at study initiation: 10-11 weeks
- Weight at study initiation: The weight variation of the animals used did not exceed 20 percent of the mean weight of each sex.
- Fasting period before study: no data
- Housing: individually in type M III polycarbonate cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: yes

ENVIRONMENTAL CONDITIONS
- Temperature (°C):20-24 °C,
- Humidity (%):30-70%
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: To:

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: The test substance was applied as a solution. To prepare the solution, the appropriate
amount of test substance was weighed out depending on the desired concentration. Then the vehicle (highly deionized water) was filled up to the desired volume, subsequently mixed using a magnetic stirrer.
The test-substance solutions were prepared in such intervals that the stability was guaranteed.

VEHICLE
- highly deionized water
- Concentration in vehicle:0.5, 1.5 and 4.5 g/100 mL
- Amount of vehicle (if gavage): 100 mL
- Lot/batch no. (if required):
- Purity:
Details on mating procedure:
- M/F ratio per cage: 1/1
- Length of cohabitation: overnight
- Proof of pregnancy: [sperm in vaginal smear] referred to as [day 0 ] of pregnancy

- Further matings after two unsuccessful attempts: [no]
- After successful mating each pregnant female was caged ( Pregnant females were provided with nesting material (cellulose wadding) toward the end of pregnancy.):
- Any other deviations from standard protocol: no
Details on analytical verification of doses or concentrations:
The analyses of the test-substance preparations were carried out at the Analytical Chemistry Laboratory of the Experimental Toxicology and Ecology of BASF SE. The stability of the test substance in highly deionized water at room temperature for a period of 10 days was proven before the start of the administration period (Project No.: 01Y0540/078008). The concentration control analyses revealed that the values were in the expected range of
the target concentration, i.e. were in a range of about 90.1-102.2% of the nominal concentration.
Duration of treatment / exposure:
The duration of treatment covered a 2-week pre-mating and mating period in both sexes, approximately 1 week post-mating in males, and the entire gestation period as well as 4 days of lactation in females (35 days for males and 55 days for females).
Frequency of treatment:
daily at the same time in the morning
Details on study schedule:

- Age at mating of the mated animals in the study: [13-14] weeks
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 50, 150 and 450 mg/kg bw/day
Basis:
nominal conc.
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: no data
- Rationale for animal assignment (if not random): randomized
- Other:
Positive control:
no

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
A check for moribund and dead animals was made twice daily on working days and once daily on Saturdays, Sundays and public holidays. If animals were in a moribund state, they were sacrificed and necropsied.
- Time schedule: A cageside examination was conducted before and after treatment for any signs of morbidity, pertinent behavioral changes and signs of overt toxicity. Abnormalities and changes were documented for each animal.
- Cage side observations checked in table [No.1] were included.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Detailed clinical observations were performed in all animals prior to the administration period and thereafter at weekly intervals. The findings were ranked according to the degree of severity, if applicable. The animals were transferred to a standard arena (50 x 37.5 cm with side borders of 25 cm high).
- The parameters examined are listed in the Table 1

BODY WEIGHT: Yes
- Time schedule for examinations: once a week at the same time of the day (in the morning).


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): no

Generally, food consumption was determined once a week (in a period of 7 days) for male
and female parental animals, with the following exceptions:
• Food consumption was not determined during the mating period (male and female F0
animals).
• Food consumption of the F0 females with evidence of sperm was determined on GD 0,
7, 14 and 20.
• Food consumption of F0 females, which gave birth to a litter, was determined on PND 0
and 4.
Food consumption was not determined in females without positive evidence of sperm (during
the mating period of dams used in parallel) and females without litter (during the lactation
period of dams used in parallel).

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OTHER:
Oestrous cyclicity (parental animals):
not examined
Sperm parameters (parental animals):
Parameters examined in [P] male parental generations:
[testis weight, epididymis weight, other:]
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: [no data]
- If yes, maximum of [all] pups/litter ; excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in [F1] offspring:
[number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, other:] viability index was calculated as follows: (number of live pups on PND4/number of liveborn pups on the day of birth)x100. The same for sex ratio: (number of live male or female pups on day 0/ 4/number of live male and female pups on day 0/ 4)x100
The live pups were examined daily for clinical symptoms (including gross-morphological findings) during the clinical inspection of the dams.
The pups were weighed one day after birth (PND 1) and on day 4 after birth.

GROSS EXAMINATION OF DEAD PUPS:
[ yes, for external and internal abnormalities; possible cause of death was determined for pups born or found dead.]
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals [after approximately 1 week post-mating period]
- Maternal animals: All surviving animals [after PND 4]

GROSS NECROPSY
- Gross necropsy consisted of [external and internal examinations including the cervical, thoracic, and abdominal viscera.] All parental animals were sacrificed by decapitation using isoflurane anesthesia. The exsanguinated animals were necropsied and assessed by gross pathology; special attention was given to the reproductive organs. The animals, which died intercurrently or were sacrificed in a moribund state, were necropsied as soon as possible after their death and assessed by gross pathology. Organ weights were recorded (see Table 2).

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in Table [3] were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring not selected as parental animals and were sacrificed at [4] days of age. All surviving pups (after sacrifice on PND 4 by means of CO2), all stillborn pups and those pups that died before schedule, were examined externally, eviscerated and their organs were assessed macroscopically. All pups without any notable findings or abnormalities were discarded after their macroscopic
evaluation.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows: all gross lesions, lungs and spinal cord (cervical, thoracic and lumbar cord) were preserved in neutrally buffered 4 % formaldehyde solution and then analyzed.

GROSS NECROPSY
- Gross necropsy consisted of [external and internal examinations including the cervical, thoracic, and abdominal viscera.]

HISTOPATHOLOGY / ORGAN WEIGTHS
The tissues indicated in Table [#] were prepared for microscopic examination and weighed, respectively.
Statistics:
Food consumption, body weight and body weight change (parental animals and pups (for the pup weights,
the litter means were used) number of mating days, duration of gestation, number of pups delivered per litter, implantation sites, post implantation loss: DUNNETT-test (two-sided)
Reproduction indices and urinalysis, except color, turbidity, volume and specific gravity, females with stillborn pups, females with all stillborn pups, live birth index, pups stillborn, pups died, pups cannibalized, pups sacrificed moribund, viability index, number of litters with affected pups at necropsy : FISHER'S EXACT test
Proportions of affected pups per litter with necropsy observations: WILCOXON-test (one-sided)
Feces, rearing, grip strength of forelimbs and hindlimbs, landing foot-splay test, motor activity, clinical pathology parameters, urine volume,urine specific gravity and organ weights : KRUSKAL-WALLIS test (two-sided).
Reproductive indices:
Male mating index %: (number of males with confirmed mating* /number of males placed with females)x100; *- defined by a female with vaginal sperm or with implants in utero;
Male fertility index (%): (number of males proving their fertility */number of males placed with females)x100; * - defined by a female with implants in utero;
Female mating index (%): (number of females mated */ number of females placed with males)x100; * - defined as the number of females with vaginal sperm or with implants in utero;
Female fertility index (%): (number of females pregnant */number of females mated **)x100; * defined as the number of females with implants in utero; ** defined as the number of females with vaginal sperm or with implants in utero.
Gestation index (%): (number of females with live pups on the day of birth/number of females pregnant *); * - defined as the number of females with implants in utero;
Live birth index(%): (number of liveborn pups at birth/total number of pups born)x100;
Post implantation loss (%): (number of implantations number of pups delivered/number of implantations)x100
Offspring viability indices:
Viability index (%): (number of live pups on PND4/number of liveborn pups on the day of birth)x100. The same for sex ratio: (number of live male or female pups on day 0/ 4/number of live male and female pups on day 0/ 4)x100

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
see details on results
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Other effects:
not examined

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related
Description (incidence and severity):
see details on results

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)

In test group 3 (450 mg/kg bw/d) one male animal (animal no. 37) was found dead within the first week of the study. One male animal (animal no. 32) of test group 3 (450 mg/kg bw/d) was sacrificed in a moribund state in study week 2. In addition, one female animal (animal no. 126) of test group 2 (150 mg/kg bw/d) was sacrificed on GD 23 because of an inability to deliver.

In test group 3 (450 mg/kg bw/d), salivation after treatment was observed in study week 1 in one male animal (animal no. 36) and in study weeks 1, 6 and 7 in six female animals. Poor general state was observed in test group 3 (450 mg/kg bw/d) in study weeks 1 and 2 in two male animals (animal nos. 32 and 36) and in study weeks 1, 6 and 7 in two female animals (animal nos. 132 and 135). In test group 3 (450 mg/kg bw/d), apathy was observed in study week 2 in one male animal (animal no. 32). Clonic convulsion was observed in test group 3 (450 mg/kg bw/d) in study week 1 in one
male animal (animal no. 39).

The detailed clinical observations on study days 0, 7, 13, 21, 28 in males and females and
additionally day 35, 42 and 49 in female animals did not reveal any additional abnormalities
in animals of test groups 0-3 (0, 50, 150 and 450 mg/kg bw/d).


BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)

In test group 3 (450 mg/kg bw/d) male animals’ body weight was significantly lower in week 4
and body weight change was already significantly lower between weeks 1-2 and in summary
between weeks 0-4. In test group 2 (150 mg/kg bw/d) male animals’ body weight change was
significantly lower between weeks 3-4
Body weights and body weight changes of all female animals treated with 50, 150 or 450
mg/kg bw/d were not significantly changed during premating.
During gestation body weights of female animals of test group 2 (150 mg/kg bw/d) were
significantly lower on GD 14 and 20 and of test group 3 (450 mg/kg bw/d) body weight was
even decreased on GD 20.
Body weight changes of female animals during gestation were significantly lower between
GD 0-7 in test group 1 (50 mg/kg bw/d) as well as between GD 0-7 and GD 7-14 in test group
2 (150 mg/kg bw/d). A body weight loss could be detected between GD 14-20 in test groups
2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d). Consequently, the overall body weight change
between GD 0-20 was also significantly lower for these test groups.
Body weights and body weight changes of female animals treated with 50 mg/kg bw/d were
not significantly changed during lactation. During lactation, a comparison of body weight data
of test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d) to the control were not meaningful
as only one litter consisting of one stillborn pup existed in test group 2 (150 mg/kg bw/d) and
no pups were alive in test group 3 (450 mg/kg bw/d).
During the post-weaning period female body weights were significantly lower in test groups 2
(150 mg/kg bw/d) and 3 (450 mg/kg bw/d) in study week 6 and 7. The same was true for
females of test group 1 (50 mg/kg bw/d) in study week 7. As the terminal mean body weight
in this test group was unaffected (see section 4.4.1.1. Absolute organ weights) this change
was assessed as incidental and not related to treatment.

Significantly decreased food consumption of the male animals of test group 3 (450 mg/kg
bw/d) was observed during the first two study weeks.
Food consumption of the female rats of test group 3 (450 mg/kg bw/d) was significantly
decreased during the first study week.
During gestation the food consumption in test group 2 (150 mg/kg bw/d) was significantly
decreased between GD 14 and 20.
During lactation food consumption in test group 2 (150 mg/kg bw/d) was significantly lower
compared to the control.


TEST SUBSTANCE INTAKE (PARENTAL ANIMALS) not applicable

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS) not examined

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS) not examined

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)

The male mating index was 100% in all test groups. Fertility was proven for most of the F0 parental males of test groups 0 (control) and 1 (50
mg/kg bw/d) within the scheduled mating interval for the F1 litter. One control male and one male of test group 1 did not generate F1 pups. Furthermore, six males of test group 2 and nine males of test group 3 did not generate F1 pups. Thus, the male fertility index ranged between 11% and 90% (see Tab.4 ). For test groups 0 (control) and 1 (50 mg/kg bw/d) these findings reflected the normal range of biological variation inherent in the strain of rats used for this study as all respective values were within the range of the historical control data (see PART III, Supplement). With regard to pathological findings in epididymidis and testis (see section 4.4. Pathology) the test substance did adversely affect reproduction of the F0 males in test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d).
The female mating index calculated after the mating period for F1 litter was 100% for all test groups. The mean duration until sperm was detected (GD 0) amounted to 2.4, 1.4, 2.5, and 2.9 days (0, 50, 150 and 450 mg/kg bw/d, respectively). Consequently, the differences between the test groups were assessed as being spontaneous in nature and without biological relevance. All sperm-positive rats of test groups 0 (control) and 1 (50 mg/kg bw/d) delivered pups or had implantations in utero with the following exceptions: one female (test group 0) and one female (50 mg/kg bw/d) did not become pregnant. 6 females of test group 2 (150 mg/kg bw/d), and 9 females of test group 3 (450 mg/kgbw/d) did not become pregnant. The fertility index varied between 10% and 90% (Tab. 5).
Implantation was not affected by the treatment in test group 1 (50 mg/kg bw/d) since the
mean number of implantation sites was comparable test group 0 (0 mg/kg bw/d). In test
groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d) a significant reduction with only 5 and 1
implantation sites was found.The mean duration of gestation, i.e. 22.1 and 22.2 days, was similar in test groups 0 (control)
and 1 (50 mg/kg bw/d). No parturition was seen in test group 2 (150 mg/kg bw/d) except of
female No. 126 which was sacrificed on GD 23 because of an inability to deliver. Gestation
length was not calculable for test group 3 (450 mg/kg bw/d).
The gestation index varied between 89% (control group) and 100% (50 mg/kg body
weight/day). All values seen in test groups 0 (control) and 1 (50 mg/kg bw/d) reflect the normal range of
biological variation inherent in the strain of rats used for this study. All respective values were
within the range of the historical control data (PART III, Supplement) and did not show a
relation to dosing. However, a clear relation to dosing was obtained for test groups 2 (150
mg/kg bw/d) and 3 (450 mg/kg bw/d).
The mean number of F1 pups delivered per dam was not affected in test group 1 (50 mg/kg
bw/d) whereas only one pup was delivered in test group 2 (150 mg/kg bw/d) and none in test
group 3 (450 mg/kg bw/d).
The rate of liveborn pups was unaffected in test group 1 (50 mg/kg bw/d) and the live birth
index was 96%. The rate of stillborn pups was not significantly different compared to the
control group and within the range of the historical control data (PART III, Supplement).
In test group 2 (150 mg/kg bw/d) the live birth index was 0 because only one stillborn pup
was delivered.

ORGAN WEIGHTS (PARENTAL ANIMALS)

Absolute organ weights: When compared to control group 0 (set to 100%), the mean absolute weights of the organs listed in the Table 8 were significantly increased or decreased. All other mean absolute weight parameters did not show significant differences when
compared to test group 0 (control).
Relative organ weights: The terminal body weight was significantly decreased in males of test group 3 (450 mg/kg
bw/d) and in females of test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d) resulting in
significant, secondary weight changes in various organs (Table 9)


GROSS PATHOLOGY (PARENTAL ANIMALS)

Three males of test group 3 (450 mg/kg bw/d) showed erosions or ulcers in the glandular
stomach.
The liver was enlarged in three males and one female of test group 2 (150 mg/kg bw/d) as
well as in three males and five females of test group 3 (450 mg/kg bw/d). Four males of test
group 1 (50 mg/kg bw/d) and four males of test group 2 (150 mg/kg bw/d) showed a
prominent acinar pattern of the liver.
The mesenteric lymph nodes were red discolored in one female of test group 2 (150 mg/kg
bw/d) and in two females of test group 3 (450 mg/kg bw/d).
All other gross lesions occurred either singly or were biologically equally distributed over the
control group and the treatment groups. They were considered to be incidental.

HISTOPATHOLOGY (PARENTAL ANIMALS) (see Table 8)

Kidneys: The graded severity of tubular degeneration was dose-related increased. The statistically
significant increase of the relative kidney weights in animals of test groups 2 (150 mg/kg
bw/d) and 3 (450 mg/kg bw/d) was considered to be caused by the tubular degeneration/
regeneration process.
Testes: The decrease of the absolute testes weight in males of test group 3 (450 mg/kg bw/d) was
related to the diffuse tubular degeneration.
Ovaries: In ovaries, vacuoles of different size were observed in the sex cord stroma in females of test
groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d). Incidence and severity was dose-related
increased (see Table 10). In addition, one female of test group 1 (50 mg/kg bw/d), one female of test group 2 (150
mg/kg bw/d) and all females of test group 3 (450 mg/kg bw/d) showed ovarian cysts. The
occurrence of cysts in females of test group 3 (450 mg/kg bw/d) was assessed as treatmentrelated.
The cysts in each one female of test groups 1 (50 mg/kg bw/d) and 2 (150 mg/kg
bw/d) were considered to be rather incidental.
Although there was no clear histopathological correlate for the decreased absolute and
relative ovarian weights in females of test group 3 (450 mg/kg bw/d), a test substance-related
effect cannot be ruled out.
Spleen: Incidence and graded severity of extramedullary hematopoiesis were dose-related increased
in males and females of test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d). The increased relative spleen weights in males of test group 3 (450 mg/kg bw/d) as well as in
females of test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d) were associated with
these findings.

OTHER FINDINGS (PARENTAL ANIMALS) clinical chemistry, haematology, urinalysis, neurobehavioral observations (see under endpoint 7.5.1.)

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (nominal)
Based on:
act. ingr.
Sex:
female
Basis for effect level:
other: general, systemic toxicity
Dose descriptor:
NOAEL
Remarks:
less than
Effect level:
50 mg/kg bw/day (nominal)
Based on:
act. ingr.
Sex:
male
Basis for effect level:
other: general, systemic toxicity
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (nominal)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: ferility, reproductive performance

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
meets only control and low dose group
Mortality / viability:
mortality observed, treatment-related
Description (incidence and severity):
see details on results
Body weight and weight changes:
no effects observed
Description (incidence and severity):
meets only control and low dose group
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
meets only control and low dose group
Histopathological findings:
no effects observed
Description (incidence and severity):
meets only control and low dose group

Details on results (F1)

VIABILITY (OFFSPRING)

The mean number of delivered pups per dam and the rate of liveborn and stillborn pups were
evenly distributed among test groups 0 (control) and 1 (50 mg/kg bw/d). The respective
values reflect the normal range of biological variation inherent in the strain used in this study.

The viability index as indicator for pup mortality between PND 0-4 was 100% for test groups
0 (control) and 1 (50 mg/kg bw/d). No viable pups were observed in test group 2 (150 mg/kg
bw/d) and test group 3 (450 mg/kg bw/d).

CLINICAL SIGNS (OFFSPRING)

The F1 pups did not show adverse clinical signs up to scheduled sacrifice on PND 4. In one
litter (dam No. 112 of test group 1) one pup showed a papilloma-like a skin flap. This single
observation was considered to be spontaneous in nature and not to be adverse.

BODY WEIGHT (OFFSPRING)

Mean pup body weights/pup body weight changes of all pups in test group (50 mg/kg bw/d)
were comparable to the concurrent control values. The observable differences between the
groups were assessed as being spontaneous in nature and without biological relevance.
One runt of each gender was seen in test group 0 (control) and 5 female runts were seen in
test group 1 (50 mg/kg bw/d). Both values were within the range of the biological variation
inherent in the strain of rats used for this study.

SEXUAL MATURATION (OFFSPRING) not applicable

ORGAN WEIGHTS (OFFSPRING) not examined

GROSS PATHOLOGY (OFFSPRING)

One stillborn pup of test group 1 (50 mg/kg bw/d) showed post mortem autolysis. In 3 pups of
test group 1 (50 mg/kg bw/d) and in the single stillborn pup of test group 2 (150 mg/kg bw/d)
the stomach was found empty.

HISTOPATHOLOGY (OFFSPRING) no treatment -related effects in the low dose group


OTHER FINDINGS (OFFSPRING)

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

Table 4: Reproductive performance (male animals)

 

Test group 0

(0 mg/kg bw/d)

Test group 1

(50 mg/kg bw/d)

Test group 2

(150 mg/kg bw/d)

Test group 3

(450 mg/kg bw/d)

Male fertility

index [%]

90

90

40

11**

Table 5: Reproductive performance (female animals)

 

Test group 0

(0 mg/kg bw/d)

Test group 1

(50 mg/kg bw/d)

Test group 2

(150 mg/kg bw/d)

Test group 3

(450 mg/kg bw/d)

Female fertility

index [%]

90

90

40*

10**

* p ≤ 0.05; ** p ≤ 0.01

Table 6: Absolute organ weight (parental animals)

Male animals

Female animals

Test group (mg/kg bw/day)

1

(50)

2

(150

3

(450)

1

(50)

2

(150

3

(450)

Terminal body weight

101%

96%

86%**

95%

93%**

85%**

Adrenal glands

 

 

 

96%

90%

82%**

Brain

 

 

 

99%

100%

96%*

Epididymides

100%

90%

68%**

 

 

 

Liver

113%*

121%**

129%**

105%

123%**

124%**

Ovaries

 

 

 

97%

99%

74%**

Testes

103%

105%

78%**

 

 

 

Thymus

98%

92%

67%**

88%

83%*

69%

Table 7: Relative organ weight (parental animals)

 

Male animals

Female animals

Test group (mg/kg bw/day)

1

(50)

2

(150

3

(450)

1

(50)

2

(150

3

(450)

Adrenal glands

104%

102%

128%*

 

 

 

Brain

98%

104%

114%*

104%*

107%*

113%**

Epydidymides

94%

98%

80%**

 

 

 

Heart

96%

106%*

122%**

98%

105%*

116%**

Kidney

101%

110%*

126%**

108%

116%**

132%**

Liver

111%**

127%**

150%**

111%**

133%**

146%**

Ovaries

 

 

 

102%

106%

86%*

Seminal vesicle

104%

113%*

117%*

 

 

 

Spleen

102%

111%

144%**

102%

112%*

121%**

Testes

101%

110%*

91%

 

 

 

Thymus

97%

96%

79%*

 

 

 

* : p ≤ 0.05; **: p ≤ 0.01

Table 8: Histopathology (parental animals)

 

Male animals

Female animals

Test group (mg/kg bw/day)

1

(50)

2

(150

3

(450)

1

(50)

2

(150

3

(450)

Kidneys

Multifocal tubular degeneration

 

Multifocal tubular degeneration; increase of the kidney weight

 

increase of the kidney weight

 

 

 

Testes

 

diffuse tubular degeneration

 

 

 

Epididymides

 

Oligospermia

 

 

 

Ovaries

 

 

 

Ovarian cysts incidental

Ovarian cysts

Spleen

 

extramedullary hematopoiesis

 

extramedullary hematopoiesis;

hemosiderin storage

Liver

Fatty change of hepatocytes

 

enlarged livers

Fore- and glandular stomach

 

 

Erosions or ulcers

 

 

Erosions or ulcers

Mesenteric lymph node

 

 

Sinus erythrocytosis

 

Sinus erythrocytosis

Thymus

 

 

reduced cellularity of cortex

 

 

reduced cellularity of cortex

Applicant's summary and conclusion

Conclusions:
Under the conditions of the present reproduction/developmental toxicity screening test the NOAEL (no observed adverse effect level) for reproductive performance and fertility was 50 mg/kg bw/d for the parental rats. The NOAEL for general, systemic toxicity of the test substance was 50 mg/kg bw/d for females and less than 50 mg/kg bw/d for male animals based on the tubular degeneration in the kidneys of six males.
Executive summary:

Methylaminoethanol was administered orally via gavage to groups of 10 male and 10 female Wistar rats (F0 animals) at dose levels of 50, 150 and 450 mg/kg bw/d.

The objective of the study was to detect possible effects of the test substance on the integrity and performance of the reproductive system of both sexes. Furthermore, it was intended to obtain information about the general toxicological profile including target organs and the no observed adverse effect level (NOAEL) after repeated oral administration. Control animals were dosed daily with the vehicle (highly deionized water). The duration of treatment covered a 2-week pre-mating and mating period in both sexes, approximately 1 week post-mating in males, and the entire gestation period as well as 4 days of lactation in females.

Regarding clinical examinations, signs of general systemic toxicity were only observed at a dose level of 450 mg/kg bw/d as there were significantly lower body weights in male and female parental animals accompanied with reduced food consumption and reduced general condition in single animals in several phases of the study. Reduced food consumption and body weights during gestation in females of test group 2 (150 mg/kg bw/d) were most likely related to implantation losses.

Detailed clinical examinations in an open field, detailed observations in a functional observational battery (FOB) and measurements of motor activity did not reveal indications of test substance-induced effects in low, mid and high-dose rats. Therefore, the clonic convulsions were assessed as being incidental.

Salivation was seen after dosing in all high-dose rats. From the temporary, short appearance immediately after dosing it is likely, that this finding was induced by a bad taste of the test substance or local affection of the upper digestive tract. Urine discoloration was also observed for all high-dose rats which was most likely related to the test compound. However, both types of findings were not considered to be adverse, toxicologically relevant effects.

Fertility was severely impaired by test-substance administration at dose levels of 150 and 450 mg/kg bw/d. Although mating (male and female mating indices) was not influenced no lifeborn pups were delivered for both test groups.

The deviated levels of clinical chemistry and haematology parameters pointed to anemia and changed liver cell metabolism. The total protein and the albumin levels were significantly higher in female rats starting at test group 1 (50 mg/kg bw/d). As these were the only deviating parameters in females of this test group the changes were regarded as treatment-related, but non-adverse. The reason for the increase of the sodium concentrations in rats of both sexes in test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d) remains unclear, but a test substance-related effect could not be excluded. The higher incidences of leucocytes in the urine of rats of both sexes in test group 3 (450 mg/kg bw/d) and, additionally, in males of the test group 2 (150 mg/kg bw/d) as well as the increased incidence of higher transitional cell counts in males of test group 3 (450 mg/kg bw/d) can be regarded as signs of an affection of the urinary tract in treated rats.

Regarding pathology, after administration of the test substance the terminal body weight was significantly lower in females of test group 2 (150 mg/kg bw/d) and in males and females of test group 3 (450 mg/kg bw/d). The body weight reduction resulted in weight changes of adrenal glands, brain, heart, seminal vesicle, and thymus. Target organs were the kidney, testes, epididymides, ovaries, liver, and spleen. In kidneys and testes, tubular degeneration was dose dependent and assessed as an adverse effect. In ovaries, the occurrence of cysts and vacuolization of sex cord stroma was related to treatment and was considered to be adverse. In test group 3 (450 mg/kg bw/d), the infertility was linked to the reduced number of sperms (oligospermia) caused by tubular degeneration in testes. In addition, the occurrence of ovarian cysts and vacuolization of the sex cord stroma in females may have influenced the fertility. In test group 2 (150 mg/kg bw/d), the severity of the findings in testes or ovaries was only minimal or slight and the findings did not occur in all infertile animals. Nevertheless, these lesions may have affected fertility. In the spleen, a dose-related increase in incidence and severity of extramedullary hematopoiesis occurred in males and females of test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d). In addition, in females of these test groups the severity of hemosiderin storage was increased. These findings are associated with the increased relative spleen weights in females of test group 2 (150 mg/kg bw/d) as well as in males and females of test group 3 (450 mg/kg bw/d). They were induced in response to anaemia and related to treatment. The liver weights were dose-related increased in males and females of all treatment groups. The liver was enlarged in three males and one female of test group 2 (150 mg/kg bw/d) as well as in three males and five females of test group 3 (450 mg/kg bw/d). In females, the liver enlargement correlated with a minimal central hepatocellular hypertrophy that was observed in five animals of test group 2 (150 mg/kg bw/d) and in 9 animals of test group 3 (450 mg/kg bw/d). In males, mainly a minimal fatty change of hepatocytes was observed in two animals of test group 1 (50 mg/kg bw/d), in 8 animals of test group 2 (150 mg/kg bw/d), and in 7 animals of test group 3 (450 mg/kg bw/d). The liver findings were related to treatment and considered to be adaptive. Although, there were no clear histopathological correlates for the increased liver weights in males of all treatment groups and in females of test group 1 (50 mg/kg bw/d), a test substance-related effect could not be ruled out. There was no correlation between erosion/ ulcer in the stomach and erythrocytosis of the mesenteric lymph node (findings occurred in different animals). However, a treatment-related effect could not be ruled out but was assessed as non-adverse. All further findings occurred either singly or were biologically equally distributed over the control group and the treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.