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Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
yes
Remarks:
additional histopathological examinations were performed
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: liquid
Details on test material:
- Name of test material sec-butyl aminoethanol
- Physical state: colorless to yellow liquid
- Analytical purity: 99.76%
- Lot/batch No.: A2053H010101
- Expiration date of the lot/batch: January 2013
- Storage conditions of test material: at room temperature (< 50°C) and protected from humidity.

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Breeder: Charles River Laboratories France, L’Arbresle, France
- Age at study initiation: at the beginning of the treatment period, the males were approximately 10 weeks old
- Mean body weight at study initiation: the males had a mean body weight of 397 g (range: 354 g to 427 g) and the females were approximately
9 weeks old and had a mean body weight of 213 g (range: 183 g to 234 g)
- Fasting period before study: no
- Housing: individually housed, except during pairing, in wire-mesh cages
- Diet: SSNIFF R/M-H pelleted diet (free access)
- Water: tap water filtered with a 0.22 µm filter (free access)
- Acclimation period: at least 7 days before the beginning of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): approximately 12 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 h/12 h (7:00 - 19:00).

IN-LIFE DATES: 05 April 2011 to 17 October 2011.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: water drinking water (treated by reverse osmosis)
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
Dose-levels were calculated in terms of 2-[(1-methylpropyl)amino]ethanol (CAS No. 35265-04-4) which is the form supplied by the Sponsor: there was no correction
factor.

The test item was administered as a solution in the vehicle. The formulations were stirred for 30 minutes in order to ensure the solubilisation of the
test item in the vehicle. No aluminium equipment was used for the preparation of the dosage forms.
The test item dosage forms were prepared on a weekly basis, stored at room temperature prior to use and delivered in brown flasks.

VEHICLE
- Concentration in vehicle: 1, 5 and 10 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg/day.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Type of method: High Performance Liquid Chromatography with tandem Mass Spectrometry detection (LC/MS MS) analytical method for the determination of 2-[(1-methylpropyl)amino]ethanol in dosage form samples was used.
Test item concentrations: the test item concentrations in the administered dosage forms analyzed in weeks 1, 5 and 7 remained within an acceptable range of -1.2% to +4.2% when compared to the nominal values (± 10%) on a weekly basis according to a stability (CIT/Study No. 37512 AHS).
Duration of treatment / exposure:
In the males:
− 2 weeks before mating,
− during the mating period (up to 3 weeks),
− until sacrifice (i.e. at least 5 weeks in total),

In the females:
− 2 weeks before mating,
− during the mating period (up to 3 weeks),
− during pregnancy,
− during lactation until day 4 post-partum inclusive,
− until sacrifice for females which had not delivered (day 25 p.c.).
Frequency of treatment:
Daily.
Doses / concentrations
Remarks:
Doses / Concentrations:
10, 50 and 100 mg/kg/day
Basis:
actual ingested
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
The dose-levels were selected in agreement with the Sponsor based on the results of the CIT/Study No. 37513 TSR.

- Rationale for animal assignment: computerized stratification procedure.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS:
- Time schedule: at least twice a day during the treatment period.

DETAILED CLINICAL OBSERVATIONS:
- Time schedule: once a day during the treatment period.

BODY WEIGHT (GAIN):
- Time schedule: Males: on the first day of treatment, then once a week until sacrifice. Females: on the first day of treatment, then once a week until mating (or until sacrifice), on Days 0, 7, 14 and 20 post-coitum and days 1 and 5 post-partum.

FOOD CONSUMPTION:
- Time schedule: once a week until sacrifice, with the exception of the mating period (not recorded).

FOOD EFFICIENCY: No

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

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
SACRIFICE
The males were sacrificed after at least 5 weeks of treatment. The body and selected organs were weighed and a complete macroscopic post-mortem examination was performed. A microscopic examination was performed on the kidneys, stomach with forestomach, epididymis, testes and thymus
from the males in the control- and high dose groups, liver from all groups and and on all macroscopic lesions.

The dams were sacrificed on day 5 p.p., the body and selected organs were weighed and a complete macroscopic examination was performed.
A microscopic examination was performed on the kidneys and the stomach with forestomach, in the control and high-dose groups and, on the liver, ovaries (with oviducts) and thymus in all groups and on all macroscopic lesions.

GROSS NECROPSY
On completion of the treatment period all surviving F0 males and females were deeply anesthetized by an intraperitoneal injection of sodium
pentobarbital and sacrificed by exsanguination.
- males: after the end of the pairing period (at least 5 weeks of treatment in total),
- females: on day 5 p.p.,
- females which had not delivered: on day 25 p.c. (after a body weight recording to check for a possible un-noticed delivery),
- mothers with litter dying entirely: as appropriate.

HISTOPATHOLOGY
A microscopic examination was performed on:
- all tissues listed in the tissue procedure table in the males and females of the control- and high-dose groups (groups 1 and 4) sacrificed at the end of the treatment period and for female that were sacrificed prematurely,
- all macroscopic lesions of all the animals of the low- and intermediate-dose groups (groups 2 and 3) sacrificed on completion of the treatment
period,
- thymus of males from control and high-dose groups (groups 1 and 4) and thymus of females from control, low-, mid- and high-dose groups
(groups 1 to 4),
- all females sacrificed because of no delivery to investigate possible causes.

Special emphasis was paid to the stages of spermatogenesis in the male gonads and histopathology of interstitial testicular cell structure.

At the request of by the Sponsor and based upon the microscopic results of the high-dose group, liver and ovaries (with oviducts) of the low- and
intermediate-dose groups were examined.

ORGAN WEIGHTS:
The body weight of each animal sacrificed as scheduled was recorded before sacrifice, and the organs specified in the Tissue Procedure Table were
weighed (wet) as soon as possible after dissection.
The ratio of organ weight to body weight (recorded immediately before sacrifice) was calculated.
Statistics:
Data are compared by one-way analysis of variances and Dunnett test (mean values being considered as normally distributed, variances being considered as homogenous) or by Fischer exact probability test (proportions).

Results and discussion

Results of examinations

Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
at 100 mg/kg bw/d in females during pregnancy period
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
at 100 mg/kg bw/d in females during pregnancy period
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
at 100 mg/kg bw/d in females
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
at 100 mg/kg bw/d in females
Histopathological findings: neoplastic:
not examined
Details on results:
MORTALITY:
There were no unscheduled deaths.

CLINICAL SIGNS:
Males
There were no treatment-related clinical signs.
The few isolated observations (soft feces, chromodacryorrhea, cutaneous lesion or area of air loss) recorded in controls or treated groups are commonly observed in this species and strain.

Females (premating period)
There were no findings during the premating period.

Females (pregnancy period)
There were no treatment-related clinical signs.
One out of 10 and 6/10 females in the 50 and 100 mg/kg/day groups, respectively, were sacrificed for absence of delivery on day 25 p.c..
All females that did not deliver were pregnant with the exception of one high-dose animal.
An isolated observation (soft feces) recorded in the 10 mg/kg/day group is commonly observed in this species and strain.

Females (lactation period)
There were no treatment-related clinical signs at 10 and 50 mg/kg/day.
One out of four females in the 100 mg/kg/day group with a dead litter was prematurely sacrificed on day 3 p.p..

BODY WEIGHT (GAIN):
Males
There were no treatment-related effects on mean body weights or mean body weight changes.
Isolated reductions in mean body weight were recorded on study day 15 (-7.0% vs. controls, p<0.05) and study day 22 (-6.7% vs. controls, p<0.05) in animals treated at 10 mg/kg/day. In absence of a dose-relationship, an association with the treatment by the test item was considered unlikely.
Isolated reductions or increases in mean body weight change were recorded on periods of days 1 to 8 (-31.0% vs. controls, p<0.05) and 8 to 15
(-41.4% vs. controls, p<0.05) at 10 mg/kg/day or on period of days 22 to 29 at 50 or 100 mg/kg/day (+64.7% and +52.9% vs. controls, p<0.05).
In absence of a dose-relationship, an association with the treatment by the test item was considered unlikely.

Females (premating period)
There were no effects on mean body weight or mean body weight change during the premating period.

Females (pregnancy period)
On day 20 p.c. and at 100 mg/kg/day, there was a statistically significant reduction in mean body weight (-21.3% vs. controls, p<0.001).
Mean body weight change was also markedly reduced at 100 mg/kg/day on periods of days 7 to 14 (-33.3% vs. controls, p<0.05) and on periods of days 14 to 20 p.c. ( 92.4% vs. controls, p<0.001), resulting in an overall reduction of -55.6% vs. controls (p<0.001) on period of days 0 to 20 p.c..
Taking into account both the amplitude of the effects and their presence in the high-dose group only, these finding were considered to be related to the treatment by the test item.

Females (lactation period)
There were no significant effects on mean body weight during the lactation period. However, mean body weight change was reduced in the three remaining dams treated at 100 mg/kg/day on periods of days 1 to 5 p.p. ( 68.2% vs. controls, p<0.01). It was correlated with lower food consumption at this dose-level.

FOOD CONSUMPTION:
Males
There were no treatment-related effects on mean food consumption.

Females (premating period)
There were no effects on mean food consumption during the premating period.

Females (pregnancy period)
There were no effects on mean food consumption during the pregnancy period up to 50 mg/kg/day.
At 100 mg/kg/day, mean food consumption was reduced on days 14 to 20 (-16.1 vs. controls, p<0.05). This effect was considered to be related to the treatment by the test item.

Females (lactation period)
There was a lower mean food consumption in the three remaining dams treated at 100 mg/kg/day on period of days 1 to 5 p.p. (37.2% vs. controls,
p<0.001). This effect was probably secondary to the low number of suckling pups.

ORGAN WEIGHTS:
Treatment-related lower mean absolute and relative liver weights were noted at 100 mg/kg/day, females being more affected than males. It correlated microscopically with lower glycogen content in hepatocytes of treated animals.
High absolute and relative mean thymus weights were noted in females given the test item at 100 mg/kg/day. In the absence of microscopic correlates, a relationship to treatment was ruled out.

GROSS PATHOLOGY:
No treatment-related macroscopic changes were noted in males and females given the test item.
The few macroscopic findings noted at the end of the treatment period were of those commonly recorded in the Sprague Dawley rat and none were
considered to be related to the test item administration.

HISTOPATHOLOGY:
In the liver, a lower mean glycogen content was observed in males and females given the test item at 100 mg/kg/day. There was a trend to dose-related decreased glycogen storage in females from 10 mg/kg/day. The effect was more severe in males than in females. Taking into account the amplitude of the change, this effect was not considered to be adverse.
Focal to multifocal subcapsular necrosis were observed in the liver of some treated females without dose relationship. As the change was often focal, at minimal severity, and could be observed spontaneously, a relationship to treatment was ruled out.

Reproductive organs
Males
Testes and epididymides were observed microscopically. Qualitative testis staging did not indicate any abnormalities in the integrity of the various cell types present within the different stages of the spermatogenic cycle.

Females
Treatment-related microscopic change was observed in the ovaries of animals given 100 mg/kg/day. It consisted of a higher incidence and severity
of increased size of follicular structures. Follicles were larger, with constant internal cavities (late stage follicles). It was observed in 1/10 control females, 1/10 females given 10 mg/kg/day, 2/10 animals treated at 50 mg/kg/day and 7/10 females given 100 mg/kg/day. Although marginal, the increased incidence and severity of the change in females given 100 mg/kg/day was considered to be likely related to the test item.

Effect levels

open allclose all
Dose descriptor:
NOAEL
Effect level:
50 mg/kg bw/day (actual dose received)
Sex:
female
Basis for effect level:
other: Reduced body weight gain in females during gestation at 100 mg/kg bw/d
Dose descriptor:
NOAEL
Effect level:
>= 100 mg/kg bw/day (actual dose received)
Sex:
male
Basis for effect level:
other: No treatment-related effect was observed in male rats.

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
The test item, Sec-Butyl aminoethanol (99.7% purity), was administered daily by oral gavage to male and female Sprague Dawley rats, for 2 weeks before mating, during mating, gestation and until day 5 post partum, at dose-levels of 10, 50 or 100 mg/kg/day.
At the dose-levels of 10 and 50 mg/kg/day, there was no treatment-related effect.
At the dose-level of 100 mg/kg/day, no treatment-related effect was observed in male rats. Mean body weight gain was markedly reduced during the whole gestation and mean food consumption was reduced on days 14 to 20 of gestation. High absolute and relative mean thymus weights and a higher incidence and severity of increased size of follicular structures in ovaries were noted in females.
Based on the results of this study:
- the No Observed Adverse Effect Level (NOAEL) for parental toxicity was considered to be 50 mg/kg/day (based on the reduced body weight gain in females during gestation).
Executive summary:

The potential toxic effects of the test item, sec-butyl aminoethanol was evaluated following daily oral administration (gavage) to male and female rats from before mating, through mating and gestation and until day 4post-partum (p.p.)(Spezia, 2011a). This study was conducted as a reproduction/developmental screening test according to OECD 421 guideline and GLP.

Three groups of 10 male and 10 female Sprague-Dawley rats received the test item, Sec-Butyl aminoethanol (purity 99.7%), by oral (gavage) administration once a day, 2 weeks before mating, through mating and gestation and until day 4p.p..The dose-levels were 10, 50 or 100 mg/kg/day. Another group of 10 males and 10 females received the vehicle, drinking water treated by reverse osmosis, alone, under the same experimental conditions and acted as a control group. The dosing volume was 10 mL/kg/day. Clinical signs and mortality were checked once and twice a day during the treatment period, respectively. Body weight and food consumption were recorded weekly and at determined intervals for females during gestation and lactation.

The males were sacrificed after at least 5 weeks of treatment. The body and selected organs were weighed and a complete macroscopicpost-mortemexamination was performed. A microscopic examination was performed on the kidneys, stomach with forestomach, epididymis, testes and thymus from the males in the control- and high‑dose groups, liver from all groups and on all macroscopic lesions. The dams were sacrificed on day 5p.p.,the body and selected organs were weighed and a complete macroscopic examination was performed. A microscopic examination was performed on the kidneys and the stomach with forestomach, in the control and high-dose groups and, on the liver, ovaries (with oviducts) and thymus in all groups and on all macroscopic lesions.

There were no unscheduled deaths in males. There were no treatment-related clinical signs in males and in females during the premating, pregnancy or lactation periods. There were no treatment-related effects on mean body weights or mean body weight changes in males and in females during the premating period. In females at 100 mg/kg/day only and on day 20p.c.,there was a markedly and statistically significant reduction in mean body weight (-21.3%vs.controls, p < 0.001). Mean body weight change was also markedly reduced at 100 mg/kg/day on periods of days 7 to 14 (-33.3%vs.controls, p > 0.05) and days 14 to 20p.c.(‑92.4%vs.controls, p < 0.001), resulting in an overall reduction of -55.6%vs.controls (p < 0.001) on period of days 0 to 20p.c.Taking into account both the amplitude of the effects and their presence in the high-dose group only, these finding were considered to be related to the treatment by the test item. There were no treatment-related effects on mean food consumption in male. At 100 mg/kg/day, in females and during the pregnancy period only, mean food consumption was moderately reduced on days 14 to 20 (-16.1vs.controls, p < 0.05). This effect was considered to be related to the treatment by the test item. Treatment-related lower liver weights were noted in males and females at 100 mg/kg/day and correlated microscopically with lower glycogen content in hepatocytes of treated animals. There was a trend to dose-related decreased glycogen storage in females from 10 mg/kg/day. This change was not considered to be adverse. No treatment-related macroscopic changes were noted in males and females (including the prematurely sacrificed one) given the test item. Qualitative testis staging did not indicate any abnormalities in the integrity of the various cell types present within the different stages of the spermatogenic cycle. Treatment-related microscopic increased size of follicular structures was observed in the ovaries of animals given 100 mg/kg/day. Although marginal, the increased incidence and severity of the ovarian change in females given 50 mg/kg/day was considered to be likely related to the test item.

The test item, Sec-Butyl aminoethanol (99.7% purity), was administered daily by oral gavage to male and female Sprague‑Dawley rats, for 2 weeks before mating, during mating, gestation and until day 5post‑partum, at dose-levels of 10, 50 or 100 mg/kg/day.  

At the dose-levels of 10 and 50 mg/kg/day, there was no treatment-related effect.

At the dose-level of 100 mg/kg/day, no treatment-related effect was observed in male rats. Mean body weight gain of females was markedly reduced during the whole gestation and mean food consumption was reduced on days 14 to 20 of gestation. High absolute and relative mean thymus weights and a higher incidence and severity of increased size of follicular structures in ovaries were noted in females.

Based on the results of this study:

. the No Observed Adverse Effect Level (NOAEL) for parental toxicity was considered to be higher than 100 mg/kg/day in males and 50 mg/kg/day in females (based on the reduced body weight gain during gestation).