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Description of key information

BASF, 2009. Methylaminoethanol. Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test in Wistar Rats Oral Administration (Gavage). According to the GLP and OECD guideline 422.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2008-11-25 to 2009-01-2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study
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
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:
Route of administration:
oral: gavage
Vehicle:
water
Details on oral 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:
Analytical verification of doses or concentrations:
yes
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
Remarks:
Doses / Concentrations:
0, 50, 150 and 450 mg/kg bw/day
Basis:
nominal in water
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
- Rationale for animal assignment (if not random): randomized
- Rationale for selecting satellite groups: no sattelite groups
- Post-exposure recovery period in satellite groups: not applicable
- Section schedule rationale (if not random): randomized

After the acclimatization period, at least 13 days after the beginning of treatment, males and females from the same test group were mated overnight in a ratio of 1:1.
On study day 32, a functional observational battery and motor activity measurement were
carried out in the first five male animals per group.
The females were allowed to litter and rear their pups until day 4 after parturition. On PND 4, all pups were sacrificed and examined.
On study day 53, a functional observational battery and motor activity measurement was carried out in the first five female animals (with litter) per group.
From the first 5 male animals and the first 5 female animals (with litter) urinalysis were carried out on study days 34 (males) and 50 (females). Clinicochemical and hematological examinations were carried out on study days 35 (males) and 55 (females).
At the end of the study (males: study day 35, females: study day 55), the animals were sacrificed after a fasting period (withdrawal of food) for at least 16-20 hours.
Positive control:
no
Observations and examinations performed and frequency:
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 :
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).

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data


OPHTHALMOSCOPIC EXAMINATION: No data
- Time schedule for examinations:
- Dose groups that were examined:

HAEMATOLOGY: Yes
- Time schedule for collection of blood:In the morning
- Anaesthetic used for blood collection: Yes (isoflurane (Isoba®, Essex GmbH Munich, Germany)).
- Animals fasted: No data
- How many animals: 5 per sex and group
- Parameters checked in table [No.4] were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: In the morning
- Animals fasted: No data
- How many animals: 5 per sex and group
- Parameters checked in table [No. 4] were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: overnight
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes
- Parameters checked in table [No. 5] were examined.

NEUROBEHAVIOURAL EXAMINATION: Yes. Functional Observation Battery
- Time schedule for examinations: A functional observational battery was performed at the end of the administration period starting at about 10:00 h.
- Dose groups that were examined: in the first five animals per sex and group.
- Battery of functions tested:
- Home cage observations: The animals were observed in their closed home cages; any disturbing activities (touching
the cage or rack, noise) were avoided during these examinations in order not to influence the
behavior of the animals. Attention was paid to:
1. posture
2. tremors
3. convulsions
4. abnormal movements
5. impairment of gait
6. other findings
- Open field observations: the parameters examined are listed in the Table 2.
- Sensory activity / grip strength are listed in the Table 3
-Motor activity:
The motor activity (MA) was measured on the same day as FOB was performed in 5 parental
males and females (with litter) per group. The examinations were performed using the Multi-
Varimex system supplied by Columbus Instruments Int. Corp., Ohio, U.S.A. For this purpose,
the animals were placed in cages for the time of measurement. Four beams were allocated
per cage. The number of beam interrupts was counted over 12 intervals for 5 minutes in each
case. The sequence at which the animals were placed in the cages was selected at random.
The measurement was started at about 14:00 h. On account of the measuring variant
"staggered", the starting time was varied by the time needed to place the animals in the
cages. For each animal, measurement was started individually when the 1st beam was
interrupted and ended exactly 1 hour later. The animals received no food or water during the
measurements. After the transfer of the last animal in each case, the room where the
measurements were carried out was darkened.

OTHER: reproduction indices, clinical observation of litters/pups and their necropsy findings are recorded in the endpoint 7.8.
Sacrifice and pathology:
GROSS PATHOLOGY: Yes. 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 Table6).
HISTOPATHOLOGY: Yes (see table 7)
Other examinations:
Reproductive organs of perental animals and litters/pups (described in the endpoint 7.8)
Statistics:
Food consumption, body weight and body weight change (parental animals and pups, 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 : 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).
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Mortality:
mortality observed, treatment-related
Description (incidence):
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
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
treatment-related but not adverse effects
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
treatment-related but not adverse effects
Urinalysis findings:
effects observed, treatment-related
Description (incidence and severity):
see details on results
Behaviour (functional findings):
no effects observed
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
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY

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 WEIGHT GAIN
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.


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study) No

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.


HAEMATOLOGY

At the end of the administration period red blood cell counts (RBC), hemoglobin
concentrations and hematocrit values were decreased in rats of both sexes in test groups 2
(150 mg/kg bw/d) and 3 (450 mg/kg bw/d).
Additionally, the hematocrit values were significantly decreased in females and males of test
group 1 (50 mg/kg bw/d). This decrease compared to the controls was below 10% (males:
5%; females 7%), and it was the only dose-dependently changed red blood cell parameter in
this test group. Therefore, the hematocrit decrease in rats of test group 1 (50 mg/kg bw/d)
was regarded as treatment-related but not adverse (reference 1 and 2).
The mean corpuscular volume (MCV) was decreased in male rats of all treatment groups (not
significantly changed in test group 3 [450 mg/kg bw/d]). The measured MCV and RBC values
were used to calculate the hematocrit values. In male rats of test group 1 (50 mg/kg bw/d) the
MCV reflected the decreased hematocrit value because the RBC was not changed.
Therefore, the decreased MCV in these rats was regarded as treatment-related, but not
adverse as mentioned above.
In female rats of test group 3 (450 mg/kg bw/d) the relative reticulocyte counts were
increased.
No significant change was observed in the total white blood cell counts (WBC) of treated rats.
However, some changes in the relative and absolute differential blood cell counts were
measured (males: increased relative neutrophil counts and decreased relative eosinophil
counts in test group 3 [450 mg/kg bw/d], decreased relative monocyte counts in test group 2
[150 mg/kg bw/d]; females: decreased absolute eosinophil counts in test group 3 [450 mg/kg
bw/d], decreased relative neutrophil counts and increased relative lymphocyte counts in test
group 2 [150 mg/kg bw/d]).
These changes were regarded as being incidental and not treatment-related because they
were not dose-dependently changed and not consistent in both sexes.
The prothrombin time was shortened in rats of both sexes of test group 3 (450 mg/kg bw/d)
and, additionally, in females of test group 2 (150 mg/kg bw/d).

CLINICAL CHEMISTRY
Liver enzyme activity was not changed in male and female rats of any test substance-treated
group.
The urea levels were increased in males of test group 2 (150 mg/kg bw/d) and in rats of both
sexes in test group 3 (450 mg/kg bw/d).
The total bilirubin concentrations were significantly higher in rats of both sexes in test groups
2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d).
The total protein and the albumin levels were increased in females of test group 1 (50 mg/kg
bw/d) and higher (total protein level was not significantly increased in test group 3 [450 mg/kg
bw/d]), although the increases were not dose-dependent.
In males the total protein levels were significantly increased in test groups 1 (50 mg/kg bw/d)
and 2 (150 mg/kg bw/d) and the albumin concentrations in test group 2 (150 mg/kg bw/d),
only. These parameters were not changed dose-dependently, and the deviated values were
within the historical control ranges (total protein: 62.45-69.74 g/L; albumin 36.12-39.76 g/L).
Therefore, these deviations were regarded as non-adverse effects.
The sodium concentrations were increased in rats of both sexes in test groups 2 (150 mg/kg
bw/d) and 3 (450 mg/kg bw/d) and, additionally, in males of test group 1 (50 mg/kg bw/d).
The sodium mean in males at least of the low dose group was within the historical control
range (140.9-147.1 mmol/L). Apart from this, only this electrolyte level was deviated in test
group 1 (50 mg/kg bw/d). Therefore, the sodium levels increase at least in males of the low
dose group was regarded as a non-adverse effect.
In males of test groups 1 (50 mg/kg bw/d) and 2 (150 mg/kg bw/d) the cholesterol levels were
decreased. The parameter was not changed dose-dependently, and such deviation was not
observed in females. Therefore, the cholesterol levels decrease in males of test groups 1 (50
mg/kg bw/d) and 2 (150 mg/kg bw/d) was regarded as non-adverse.
In treated females the potassium concentrations were significantly higher in test group 1 (50
mg/kg bw/d), the creatinine levels were higher in test group 2 (150 mg/kg bw/d) and the
magnesium concentrations were increased in test groups 1 and 2. These values were not
changed dose-dependently, and the deviations of these parameters were not measured in
male rats. Therefore, these changes were regarded as incidental rather than treatmentrelated.

URINALYSIS
In rats of both sexes in test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d) the incidence
of blood (haemoglobin) was found higher compared to the controls (in females of test group 3
not significant). Additionally, the incidence of higher leucocyte counts in the urine sediment
was significantly increased in males of test group 2 (150 mg/kg bw/d). However, no
significantly higher leucocyte counts were found in the urine sediment of rats of both sexes of
test group 3 (450 mg/kg bw/d).
In males of test group 3 (450 mg/kg bw/d), the incidence of higher transitional cell counts was
increased.

The urine was discolored almost all in the males and females of test group 3 (450 mg/kg bw/d) from study week 1 onwards.

NEUROBEHAVIOUR

Home cage observations: No test substance-related or spontaneous findings in male and female animals of all test
groups during the home cage observation were observed.

Open field observations: The open field observations did not reveal any test substance-related findings in male and
female animals of all test groups.

Sensorimotor tests/reflexes: There were no test substance-related findings in male and female animals of all test groups.
Any deviations from "zero values" were equally distributed between test substance-treated
groups and controls or occurred in single animals only. Therefore, these observations were
considered as being incidental.

Motor activity measurement: There were no significant deviations concerning the overall motor activity (summation of all
intervals) in the male and female animals of all test groups in comparison to the concurrent
control group.
Regarding single intervals, in males of test group 1 and 2 (50 and 150 mg/kg bw/d) two
isolated significantly increased value was measured at interval 4. These findings were
considered as being incidental since the overall motor activity was not changed and no
findings were observed for female animals.

ORGAN WEIGHTS

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

Pups
A papilloma was macroscopically observed in the skin of one male pup of test group 1 (50
mg/ kg bw/d; dam No. 112). All other treated males and females as well as all control animals
did not show gross lesions.
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: NON-NEOPLASTIC (see Table 10)
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.
HISTOPATHOLOGY: NEOPLASTIC (if applicable) No

HISTORICAL CONTROL DATA (if applicable) No

OTHER FINDINGS
- Clinical observations for females during gestation of F1 litter:
Findings were only seen in test group 3 (450 mg/kg bw/d), i.e. poor general state was
observed in 4 female animals (animal nos. 132, 135, 136 and 140) from GD 11 onwards,
salivation after treatment was observed in 2 female animals (animal nos. 136 and 139) from
GD 2 onwards and discolored urine was observed in all animals during the whole gestation
period.
- Clinical observations for females during lactation of F1 litter:
No test substance-related clinical findings occurred in the female animals. Only one pup
(dam no. 112) from test group 1 (50 mg/kg bw/d) showed a papilloma-like skin flap from day
of birth (PND 0) onwards until the end of study (Detailed findings see in the endpoint 7.8)

DECEDENTS
One male (No. 32) of test group 3 (450 mg/kg bw/d) was sacrificed in a moribund state.
Severe meningitis was observed in the brain, a spermatogenic granuloma was noted in the
right epididymis, and an erosion/ ulcer occurred in the glandular stomach. Another male (No.
37) of test group 3 (450 mg/kg bw/d) died prematurely. This male showed a severe alveolar
histiocytosis in the lungs, a moderate purulent inflammation of the trachea and an erosion/
ulcer in the glandular stomach. The premature death of these two males was considered to
be incidental.
One female of test group 2 (No. 126, 150 mg/kg bw/d) was sacrificed during parturition. One
dead fetus was found in the right uterus horn and birth channel. No more fetuses were
detected. An influence of the test substance cannot be ruled out.
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: reproductive performance and fertility
Critical effects observed:
not specified

Table 8: Absolute organ weights

 

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%

* : p ≤ 0.05; **: p ≤ 0.01

Table 9: Relative organ weights

 

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 10: Histopathology

 

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

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.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
50 mg/kg bw/day
Study duration:
subchronic
Species:
rat

Additional information

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 (BASF, 2010).

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.

In conclusion, 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 males based on the tubular degeneration in the kidneys of six male animals.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
Only one reliable study available.

Repeated dose toxicity: via oral route - systemic effects (target organ) cardiovascular / hematological: spleen; digestive: liver; urogenital: epididymides; urogenital: kidneys; urogenital: ovaries; urogenital: testes

Justification for classification or non-classification

N-methylethanolamine caused severely impaired fertility in rats treated at dose levels of 150 and 450 mg/kg bw/d in the Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test (OECD 422; BASF, 2010). Although mating (male and female mating indices) was not influenced no lifeborn pups were delivered for both test groups. Signs of general toxicity were observed in animals of only the highest dose group (450 mg/kg bw). Changed levels of clinical chemistry parameters together with histopathological findings in organs of treated animals points to the systemic toxicity hazard by prolonged exposure. This is a summary of the most relevant findings which are very likely accountable for these effects:

Test group 3 (450 mg/kg body weight/day):

Males (xout of 10 animals):

Target: gonades

·        Testes: tubular degeneration (8 out of 10 animals)

·        Epidymides: oligospermia (9)

 

Target: kidney

·        Kidneys: tubular degeneration (10)

·        Decreased urea clearance (mean of test group)

·        Blood in urine (mean of test group)

 

Target: liver

·        Liver: central fatty change (5)

·        Liver: peripheral fatty change (2)

·        Increased albumin level (mean of test group)

 

Target: blood

·        Spleen: extramedullar haematopoiesis (8)

·        Haemolytic anaemia (mean of test group)

 

Target: stomach

·        Forestomach: erosion/ulceration (3)

·        Glandular stomach: erosion/ulceration (2)

 

Dams (xout of 10 animals):

Target: gonades

·        Ovaries: vacuolization of sex cord stroma (10 out of 10 animals)

 

Target: kidney

·        Kidneys: tubular degeneration (9)

·        Decreased urea clearance (mean of test group)

·        Blood in urine (mean of test group)

 

Target: liver

·        Liver: central hypertrophy (9)

·        Increased albumin level (mean of test group)

 

Target: blood

·        Spleen: extramedullar haematopoiesis (8)

·        Haemolytic anaemia (mean of test group)

 

Target: stomach

·        Forestomach: erosion/ulceration (1)

 

Additional finding

·        Mesenteric lymph nodes: sinus erythrocytosis (5)

 

Test group 2 (150 mg/kg body weight/day):

Males (xanimals):

Target: kidney

·        Kidneys: tubular degeneration (10)

·        Decreased urea clearance (mean of test group)

·        Blood in urine (mean of test group)

 

Target: liver

·        Liver: peripheral fatty change (5 out of 7 animals)

 

Target: blood

·        Haemolytic anaemia (mean of test group)

 

Dams (xanimals):

Target: gonades

·        Ovaries: vacuolization of sex cord stroma (4 out of 7 animals)

 

Target: kidney

·        Kidneys: tubular degeneration (9)

·        Blood in urine (mean of test group)

 

Target: blood

·        Spleen: extramedullar haematopoiesis (1)

·        Haemolytic anaemia (mean of test group)

 

Additional finding

·        Mesenteric lymph nodes: sinus erythrocytosis (1 out of 2 animals)

 

Test group 1 (50 mg/kg body weight/day):

Males (xanimals):

Target: kidney

·        Kidneys: tubular degeneration (6)

 

The kidneys of males of all treatment groups as well as in females of test groups 2 (150 mg/kg bw/d) and 3 (450 mg/kg bw/d) revealed a minimal to severe tubular degeneration (see next table) which was regarded to be treatment-related. The severity increased dose-dependently:

Tubular degeneration in the kidney:

 

Male animals

Female animals

Test group
(mg / kg bw / day)

0

 

1

(50)

2

(150)

3

(450)

0

 

1

(50)

2

(150)

3

(450)

Number of animals

10

10

10

10

10

10

10

10

Degeneration, tubular

-

6

10

10

-

-

9

9

minimal

-

3

1

1

-

-

9

7

slight

-

3

4

1

-

-

-

2

moderate

-

-

5

6

-

-

-

-

severe

-

-

-

2

-

-

-

-

 

Tubular degeneration in the testes / epididymides:

 

Maleanimals

Test group

(mg / kg bw / day)

0

1 (50)

2 (150)

3 (450)

Number ofanimals

10

10

10

10

Diffuse tubular degeneration

-

 

7

10

minimal

-

-

7

1

slight

-

-

-

1

moderate

-

-

-

5

severe

-

-

-

3

 

Tubular degeneration in kidneys and testes was dose dependent and assessed as an adverse effect. This is probably a key factor in the fertility effects. Furthermore, the effects in kidneys are probably related to anaemia. Since serum bilirubin was high, some amount of haemoglobin in the urine can be present, especially if the kidneys are damaged. It was manifested in discoloured urine of the treated animals.

Vacuolization of sex cord stroma

 

Femaleanimals

Test group

(mg / kg bw / day)

0

1 (50)

2 (150)

3 (450)

Number ofanimals

10

10

10

10

Occurrence of cysts

-

-

-

10

Vacuolization of sex cord stroma

-

-

4

10

 

The occurrence of cysts and vacuolization of sex cord stroma in ovaries was related to treatment and was considered to be adverse. This is another key factor to fertility effects.

Conclusion on classification and labelling:

Based on the findings observed in the Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test (OECD 422; BASF, 2010) classification with a “hazard category 2” for “Specific Target Organ Toxicity Repeated Exposure” (STOT RE Cat. 2) is warranted for Methyl-Monoethanolamine (MMEA; CAS 109-83-1) according to the criteria of EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulations No 1272/2008. The substance will be labelled accordingly with “H373: May cause damage to organs through prolonged or repeated exposure”. Target organs are the kidney, testes, epididymides, ovaries, liver, and spleen.

The rationale is as follows:       

1. Substance and treatment-related adverse effects on kidneys (i.e. tubular degeneration) after repeated dose exposure occur in the range of the low dose (i.e. 50 mg/kg bw/day).

2. Substance and treatment-related adverse effects on blood (i.e. haemolytic anaemia) occurred in the range of the mid and high doses (150 and 450 mg/kg bw/day).

3. MMEA is not subject for classification as being toxic to male and female fertility. Adverse effects on fertility are considered to occur secondary to the specific target organ toxicity (i.e. adverse effects on kidney and anaemia).  

 

Details on the rationale:

Ad 1)MMEA exerted treatment-related and adverse effects on the kidneys in males and females (i.e. tubular degeneration was observed at all doses tested. Additionally, changes in urine parameters were reported that corroborate the impaired functionality of the renal system at 150 and 450 mg/kg bw/day (e.g. occurrence of blood in the urine, reduction in urea clearance). The effects may be secondary to anaemia.

Ad 2)Haemolytic anaemia has been reported to be caused substance- and treatment-related by MMEA. A significant decrease in red blood cell count and in total haemoglobin content could be observed in the mid and high doses. In consequence, a decrease in haematocrit was observed in line with the before mentioned findings representing anaemia. Further indications for a haemolytic anaemia can be derived from the fact that an increase in hemosiderin storage in the spleens has been reported. Additionally, an increase in bilirubin levels was reported to occur in the urine indicative for the haemolytic character of the anaemia.

Ad 3)MMEA is not subject for classification for adverse effects on fertility, as the effects on testes and epididymides, as well as effects on the ovaries did not occur isolated and in the absence of other toxic effects. The adverse effects on fertility were reported to occur in the range of other significant toxic effects (i.e. kidney toxicity and haemolytic anaemia as described above). Thus, the adverse effects on reproduction affecting male and female fertility are considered to be secondary to the general toxicity effects observed and to result from a non-specific mechanism. The adverse and significant toxic effects on the kidneys occurs in the lowest dose of 50 mg/kg bw/day whereas fertility was impaired only at higher dose levels (= 150 and 450 mg/kg bw/day. Furthermore, haemolytic anaemia represents an adverse and significant effect which occurs in the same range as effects on fertility are being reported.

Reproduction and fertility have been linked with kidney failure in male rats and kidney functional impairment (Nazian and Dietz, 1987, Menjívar et al., 2000). In these available studies male fertility was reported to be impaired by liver insufficiency as a secondary consequence (e.g. chronic nephrosis, uraemia in consequence of partial nephrectomy). Especially chronic nephrosis as an umbrella term for degenerative tubular kidney disease represents a comparable situation to what has been observed for MMEA. Ortiz et al. (1999) reported decreased male fertility in consequence of chronic nephrosis.

Considering the possible mechanism, effects on the choline-homeostasis could play a role. Various alkanolamines are known to produce choline-deficiency (e.g. diethanolamine DEOA CAS 111-42-2). Choline is a vitamin-like compound with various physiological functions (i.e. building block of phospholipids and acetyl-choline, one-carbon-metabolism and DNA-methylation etc.). It could be demonstrated that certain alkanolamines exert an inhibitory effect on either choline-uptake and/or choline-metabolism. Thereby, alkanolamines cause a choline-depletion. A hallmark of choline-depletion is a fatty liver change (Zeisel, 1994). In line with this, liver enlargement concurrent with an increase in absolute absolute and relative liver weight has been reports in all dose groups after MMEA-treatment. Furthermore, minimal fatty changes and central hepatocellular hypertrophy have been observed in parallel. Similar effects have also been reported for DEOA, where choline-deficiency caused liver and kidney effects (Melnick, 1992) in repeated dose toxicity tests. In long-term studies with DEOA, liver and kidney tumours developed in mice but not in rats (NTP, 1992). In depth investigation on the possible mode-of-action revealed that the liver tumours formation could be attributed to an increase in hepatocellular proliferation probably due to a DEOA-induced choline-depletion (Lehman-McKeeman and Gamsky, 2000; Lehman-McKeeman et al., 2002). However, an increase in hepatocellular proliferation has been reported for rodent hepatocytesin vitroonly whereas human primary hepatocytes did not respond. This indicates that rodent cell might be more sensitive and prone towards choline-depletion than human hepatocytes are. Thus, the human relevancy of the findings is questionable as no increase in proliferation was observed in the human hepatocytes (Stott, 2000; Kamendulis and Klaunig, 2005). Therefore, the mode-of-action of MMEA causing kidney lesions might rely on choline-depletion (as reported for various alkanoalmines as well). This mode-of-action has been demonstrated in the context of liver tumour formation to lack human relevance. It is thus concluded that MMEA-induced adverse kidney effects might arise in addition from a toxicokinetic difference with rodents being most sensitive species.This “choline” issue may also be causing some specific effect on membrane integrity that is resulting in a spectrum of toxic effects (anaemia, testes tubule degeneration, and possibly kidney tubule degeneration). This was considered to be part of the DEA toxicity spectrum as well, since some anemia was noted at the high dose levels of the DEA chronic studies.

Moreover, the dramatic reduction in male and female fertility (10 and 11%, respectively) occurred in the range of morbidity already in the high dose of 450 mg/kg bw/day. Twenty per cent of the males (2/10) were either found dead (1/10) or had to be sacrificed due a poor general status (1/10). In the mid dose (150 mg/kg bw/day) reduction in fertility was still evident in the presence of adverse and severe kidney effects and haemolytic anaemia. However, no animal died treatment related in the mid dose group.

Taken together, MMEA should be classified for specific target organ toxicity affecting kidney and blood. Effects on fertility are considered to occur as a secondary consequence and are thus not subject to classification.

 

 

References

 

Lehman-McKeeman LD, Gamsky EA (2000). Choline supplementation inhibits diethanolamine-induced morphological transformation in Syrian hamster embryo cells: evidence for a carcinogenic mechanism. Toxicol Sci, 55, 303-10

 

Lehman-McKeeman, L. D. et al.(2002). Diethanolamine Induces Hepatic Choline Deficiency in Mice.Toxicological sciences 67, 39-45

Kamendulis LM and Klaunig JE (2005). Species differences in the induction of hepatocellular DNA synthesis by Diethanolamnin. Toxicological Sciences 87(2),328-336.

Melnick R (1992). NTP technical report on the toxicity studies of Diethanolamine (CAS No. 111-42-2) Administered Topically and in Drinking Water to F344/N Rats and B6C3F1 Mice. Toxic Rep Ser. 20:1-D10.

Menjíar M, Cárdenas M, Ortiz G, Pedraza-Chaverrí J(2000). Fertility Diminution in Female Rats with Experimental Chronic Nephrosis. Biol reproduction 63, 1549–1554

Nazian SJ, Dietz JR (1987). Reproductive Changes during the Early Stages of Chronic Renal Insufficiency in the Mal Rat. Biol of Reproduction 37: 105-111

National Toxicology Program (1992). Toxicity Studies of Diethanolamine (CAS No. 111-42-2) Administered Topically and in Drinking Water to F344/N Rats and B6C3F1 Mice. Tech. Rep. Ser. No. 20; NIH Publication No. 92-3343), Department of Health and Human Services, Research Triangle Park, NC. Report no.: TR20.

Ortiz G, Vilchis F, Cárdenas M, Cruz C, Pedraza-Cahverris J, Menjívar M (1999). Reproduciton : Function in Male Rats with Chronic Nephrosis. Journal Reprod and Fertility 117: 223-228

Stott WT, Bartels MJ, Brzak KA, Mar M, Markham DA, Thornton CM, Zeisel SH (2000).Potential mechanisms of tumorigenic action of diethanolamine in mice. Toxicol. Lett., 114, 67-75.

Zeisel SH and Blasztajn JK (1994). Cholin and human nutrition. Ann. Rev. Nutr. 14: 269-296